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Sample records for viable cell yield

  1. Cultivation of murine bone marrow macrophages in sponges: a method that permits recovery of viable cultured cells

    Energy Technology Data Exchange (ETDEWEB)

    Akporiaye, E T; Stewart, S; Stewart, C C

    1984-01-01

    Various investigators have cultured murine bone marrow or peritoneal cells in vitro on glass or plastic surfaces with the ultimate aim of retrieving adherent macrophages for morphologic and functional evaluation. The removal of these adherent macrophages by conventional techniques has been consistently accompanied by low yield and significant cell damage. The authors report here a simple technique for culturing murine bone marrow cells in gelatin sponges (Spongostan and Gelfoam) in growth medium containing 10% fetal bovine serum and 10% L-cell conditioned medium. Viable cells were retrieved from the sponges in 10 min by digestion with collagenase. The in situ growth kinetics were similar to those found for cells cultured on plastic dishes. The recovered cells were adherent, phagocytic, positive for Fc ..gamma.. receptors, and had esterase activity. 23 references, 1 figure, 1 table.

  2. Predictors of viable germ cell tumor in postchemotherapeutic residual retroperitoneal masses

    Directory of Open Access Journals (Sweden)

    Khalid Al Othman

    2014-01-01

    Full Text Available Objective: The aim of this study was to identify predictors of viable germ cell tumor (GCT in postchemotherapeutic residual retroperitoneal masses. Materials and Methods: The pertinent clinical and pathologic data of 16 male patients who underwent postchemotherapeutic retroperitoneal lymph node dissection (PC-RPLND at King Faisal Specialist Hospital and Research Centre between 1994 and 2005 were reviewed retrospectively. It was found that all patients received cisplatin-based chemotherapy for advanced testicular GCT. Results: Out of the 16 male patients, 2 (13%, 8 (50%, and 6 (37% had viable GCT, fibrosis, and teratoma, respectively. Ten (10 of the patients with prechemotherapeutic S1 tumor markers did not have viable GCT, and two of the six patients who had prechemotherapeutic S2 tumor markers have viable GCT. All tumor marker levels normalized after chemotherapy even in patients with viable GCT. Four patients had vascular invasion without viable GCT. Furthermore, four patients had more than 60% embryonal elements in the original pathology, but only 1 had viable GCT at PC-RPLND. Four of the five patients with immature teratoma had teratoma at PC-RPLND but no viable GCT; however, out of the four patients with mature teratoma, one had viable GCT and two had teratoma at PC-RPLND. Of the two patients with viable GCT, one had 100% embryonal cancer in the original pathology, prechemotherapeutic S2 tumor markers, history of orchiopexy, and no vascular invasion; the other patient had yolk sac tumor with 25% embryonal elements and 40% teratoma in the original pathology, and prechemotherapeutic S2 tumor markers. Conclusion: None of the clinical or pathological parameters showed a strong correlation with the presence of viable GCT in PC-RPLND. However, patients with ≥S2 may be at higher risk to have viable GCT. Further studies are needed to clarify this.

  3. Non-viable antagonist cells are associated with reduced biocontrol performance by viable cells of the yeast Papiliotrema flavescens against Fusarium head blight of wheat.

    Science.gov (United States)

    Microbially-based plant disease control products have achieved commercial market success, but the efficacy of such biocontrol products is sometimes deemed inconsistent. Improper processing of harvested microbial biomass or long-term storage can reduce the proportion of viable cells and necessitate t...

  4. Biocatalytically active silCoat-composites entrapping viable Escherichia coli.

    Science.gov (United States)

    Findeisen, A; Thum, O; Ansorge-Schumacher, M B

    2014-02-01

    Application of whole cells in industrial processes requires high catalytic activity, manageability, and viability under technical conditions, which can in principle be accomplished by appropriate immobilization. Here, we report the identification of carrier material allowing exceptionally efficient adsorptive binding of Escherichia coli whole cells hosting catalytically active carbonyl reductase from Candida parapsilosis (CPCR2). With the immobilizates, composite formation with both hydrophobic and hydrophilized silicone was achieved, yielding advanced silCoat-material and HYsilCoat-material, respectively. HYsilCoat-whole cells were viable preparations with a cell loading up to 400 mg(E. coli) · g(-1)(carrier) and considerably lower leaching than native immobilizates. SilCoat-whole cells performed particularly well in neat substrate exhibiting distinctly increased catalytic activity.

  5. Mobilization of Viable Tumor Cells Into the Circulation During Radiation Therapy

    International Nuclear Information System (INIS)

    Martin, Olga A.; Anderson, Robin L.; Russell, Prudence A.; Ashley Cox, R.; Ivashkevich, Alesia; Swierczak, Agnieszka; Doherty, Judy P.; Jacobs, Daphne H.M.; Smith, Jai; Siva, Shankar; Daly, Patricia E.; Ball, David L.

    2014-01-01

    Purpose: To determine whether radiation therapy (RT) could mobilize viable tumor cells into the circulation of non-small cell lung cancer (NSCLC) patients. Methods and Materials: We enumerated circulating tumor cells (CTCs) by fluorescence microscopy of blood samples immunostained with conventional CTC markers. We measured their DNA damage levels using γ-H2AX, a biomarker for radiation-induced DNA double-strand breaks, either by fluorescence-activated cell sorting or by immunofluorescence microscopy. Results: Twenty-seven RT-treated NSCLC patients had blood samples analyzed by 1 or more methods. We identified increased CTC numbers after commencement of RT in 7 of 9 patients treated with palliative RT, and in 4 of 8 patients treated with curative-intent RT. Circulating tumor cells were also identified, singly and in clumps in large numbers, during RT by cytopathologic examination (in all 5 cases studied). Elevated γ-H2AX signal in post-RT blood samples signified the presence of CTCs derived from irradiated tumors. Blood taken after the commencement of RT contained tumor cells that proliferated extensively in vitro (in all 6 cases studied). Circulating tumor cells formed γ-H2AX foci in response to ex vivo irradiation, providing further evidence of their viability. Conclusions: Our findings provide a rationale for the development of strategies to reduce the concentration of viable CTCs by modulating RT fractionation or by coadministering systemic therapies

  6. Mobilization of Viable Tumor Cells Into the Circulation During Radiation Therapy

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Olga A. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Molecular Radiation Biology Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); The Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC (Australia); Anderson, Robin L. [The Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC (Australia); Metastasis Research Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Russell, Prudence A. [Department of Anatomical Pathology, St. Vincent Hospital, Fitzroy, VIC (Australia); Ashley Cox, R. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Ivashkevich, Alesia [Molecular Radiation Biology Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Laboratory of DNA Repair and Genomics, Centre for Innate Immunity and Infectious Disease, Monash Institute for Medical Research, Monash University, Clayton, VIC (Australia); Swierczak, Agnieszka; Doherty, Judy P. [Metastasis Research Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Jacobs, Daphne H.M. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Smith, Jai [Molecular Radiation Biology Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Siva, Shankar; Daly, Patricia E. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Ball, David L. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); The Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC (Australia); and others

    2014-02-01

    Purpose: To determine whether radiation therapy (RT) could mobilize viable tumor cells into the circulation of non-small cell lung cancer (NSCLC) patients. Methods and Materials: We enumerated circulating tumor cells (CTCs) by fluorescence microscopy of blood samples immunostained with conventional CTC markers. We measured their DNA damage levels using γ-H2AX, a biomarker for radiation-induced DNA double-strand breaks, either by fluorescence-activated cell sorting or by immunofluorescence microscopy. Results: Twenty-seven RT-treated NSCLC patients had blood samples analyzed by 1 or more methods. We identified increased CTC numbers after commencement of RT in 7 of 9 patients treated with palliative RT, and in 4 of 8 patients treated with curative-intent RT. Circulating tumor cells were also identified, singly and in clumps in large numbers, during RT by cytopathologic examination (in all 5 cases studied). Elevated γ-H2AX signal in post-RT blood samples signified the presence of CTCs derived from irradiated tumors. Blood taken after the commencement of RT contained tumor cells that proliferated extensively in vitro (in all 6 cases studied). Circulating tumor cells formed γ-H2AX foci in response to ex vivo irradiation, providing further evidence of their viability. Conclusions: Our findings provide a rationale for the development of strategies to reduce the concentration of viable CTCs by modulating RT fractionation or by coadministering systemic therapies.

  7. Modelling the number of viable vegetative cells of Bacillus cereus passing through the stomach

    NARCIS (Netherlands)

    Wijnands, L.M.; Pielaat, A.; Dufrenne, J.B.; Zwietering, M.H.; Leusden, van F.M.

    2009-01-01

    Aims: Model the number of viable vegetative cells of B. cereus surviving the gastric passage after experiments in simulated gastric conditions. Materials and Methods: The inactivation of stationary and exponential phase vegetative cells of twelve different strains of Bacillus cereus, both mesophilic

  8. Selection of viable cell subpopulations from murine tumours using FACS

    International Nuclear Information System (INIS)

    Chaplin, D.J.; Durand, R.E.; Olive, P.L.

    1985-01-01

    The authors developed a technique which enables isolation of viable tumour cells subpopulation as a function of their distance from the blood supply. The basis for this separation procedure is that the fluorochrome, Hoechst 33342, as a result of its high avidity for cellular DNA, exhibits a marked diffusion/consumption gradient when it has to pass through several cell layers. As a result intravenous injection of Hoechst 33342 into tumour bearing animals, results in a heterogeneous straining pattern within the tumour with cells close to blood vessels being brightly fluorescent while those more distant are less intensely stained. Since these differences in staining intensity persist after tumour disaggregation, cells can be sorted into subpopulations on the basis of their fluorescence intensity using a fluorescence activated cell sorter. This technique offers the unique possibility of identifying the location of those cell subpopulations resistant to treatment with either radiation or chemotherapeutic drugs

  9. Deletion of ultraconserved elements yields viable mice

    Energy Technology Data Exchange (ETDEWEB)

    Ahituv, Nadav; Zhu, Yiwen; Visel, Axel; Holt, Amy; Afzal, Veena; Pennacchio, Len A.; Rubin, Edward M.

    2007-07-15

    Ultraconserved elements have been suggested to retainextended perfect sequence identity between the human, mouse, and ratgenomes due to essential functional properties. To investigate thenecessities of these elements in vivo, we removed four non-codingultraconserved elements (ranging in length from 222 to 731 base pairs)from the mouse genome. To maximize the likelihood of observing aphenotype, we chose to delete elements that function as enhancers in amouse transgenic assay and that are near genes that exhibit markedphenotypes both when completely inactivated in the mouse as well as whentheir expression is altered due to other genomic modifications.Remarkably, all four resulting lines of mice lacking these ultraconservedelements were viable and fertile, and failed to reveal any criticalabnormalities when assayed for a variety of phenotypes including growth,longevity, pathology and metabolism. In addition more targeted screens,informed by the abnormalities observed in mice where genes in proximityto the investigated elements had been altered, also failed to revealnotable abnormalities. These results, while not inclusive of all thepossible phenotypic impact of the deleted sequences, indicate thatextreme sequence constraint does not necessarily reflect crucialfunctions required for viability.

  10. Mathematical modelling of the viable epidermis: impact of cell shape and vertical arrangement

    KAUST Repository

    Wittum, Rebecca; Naegel, Arne; Heisig, Michael; Wittum, Gabriel

    2017-01-01

    In-silico methods are valuable tools for understanding the barrier function of the skin. The key benefit is that mathematical modelling allows the interplay between cell shape and function to be elucidated. This study focuses on the viable (living

  11. A combination of direct viable count and fluorescence in situ hybridization for specific enumeration of viable Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus thermophilus.

    Science.gov (United States)

    García-Hernández, J; Moreno, Y; Amorocho, C M; Hernández, M

    2012-03-01

    We have developed a direct viable count (DVC)-FISH procedure for quickly and easily discriminating between viable and nonviable cells of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains, the traditional yogurt bacteria. direct viable count method has been modified and adapted for Lact. delbrueckii subsp. bulgaricus and Strep. thermophilus analysis by testing different times of incubation and concentrations of DNA-gyrase inhibitors. DVC procedure has been combined with fluorescent in situ hybridization (FISH) for the specific detection of viable cells of both bacteria with specific rRNA oligonucleotide probes (DVC-FISH). Of the four antibiotics tested (novobiocin, nalidixic acid, pipemidic acid and ciprofloxacin), novobiocin was the most effective for DVC method and the optimum incubation time was 7 h for both bacteria. The number of viable cells was obtained by the enumeration of specific hybridized cells that were elongated at least twice their original length for Lactobacillus and twice their original size for Streptococcus. This technique was successfully applied to detect viable cells in inoculated faeces. Results showed that this DVC-FISH procedure is a quick and culture-independent useful method to specifically detect viable Lact. delbrueckii subsp. bulgaricus and Strep. thermophilus in different samples, being applied for the first time to lactic acid bacteria. © 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.

  12. Effects of blood transportation on human peripheral mononuclear cell yield, phenotype and function: implications for immune cell biobanking.

    Directory of Open Access Journals (Sweden)

    Anita Posevitz-Fejfár

    Full Text Available Human biospecimen collection, processing and preservation are rapidly emerging subjects providing essential support to clinical as well as basic researchers. Unlike collection of other biospecimens (e.g. DNA and serum, biobanking of viable immune cells, such as peripheral blood mononuclear cells (PBMC and/or isolated immune cell subsets is still in its infancy. While certain aspects of processing and freezing conditions have been studied in the past years, little is known about the effect of blood transportation on immune cell survival, phenotype and specific functions. However, especially for multicentric and cooperative projects it is vital to precisely know those effects. In this study we investigated the effect of blood shipping and pre-processing delay on immune cell phenotype and function both on cellular and subcellular levels. Peripheral blood was collected from healthy volunteers (n = 9: at a distal location (shipped overnight and in the central laboratory (processed immediately. PBMC were processed in the central laboratory and analyzed post-cryopreservation. We analyzed yield, major immune subset distribution, proliferative capacity of T cells, cytokine pattern and T-cell receptor signal transduction. Results show that overnight transportation of blood samples does not globally compromise T- cell subsets as they largely retain their phenotype and proliferative capacity. However, NK and B cell frequencies, the production of certain PBMC-derived cytokines and IL-6 mediated cytokine signaling pathway are altered due to transportation. Various control experiments have been carried out to compare issues related to shipping versus pre-processing delay on site. Our results suggest the implementation of appropriate controls when using multicenter logistics for blood transportation aiming at subsequent isolation of viable immune cells, e.g. in multicenter clinical trials or studies analyzing immune cells/subsets. One important conclusion might

  13. PMA-Linked Fluorescence for Rapid Detection of Viable Bacterial Endospores

    Science.gov (United States)

    LaDuc, Myron T.; Venkateswaran, Kasthuri; Mohapatra, Bidyut

    2012-01-01

    The most common approach for assessing the abundance of viable bacterial endospores is the culture-based plating method. However, culture-based approaches are heavily biased and oftentimes incompatible with upstream sample processing strategies, which make viable cells/spores uncultivable. This shortcoming highlights the need for rapid molecular diagnostic tools to assess more accurately the abundance of viable spacecraft-associated microbiota, perhaps most importantly bacterial endospores. Propidium monoazide (PMA) has received a great deal of attention due to its ability to differentiate live, viable bacterial cells from dead ones. PMA gains access to the DNA of dead cells through compromised membranes. Once inside the cell, it intercalates and eventually covalently bonds with the double-helix structures upon photoactivation with visible light. The covalently bound DNA is significantly altered, and unavailable to downstream molecular-based manipulations and analyses. Microbiological samples can be treated with appropriate concentrations of PMA and exposed to visible light prior to undergoing total genomic DNA extraction, resulting in an extract comprised solely of DNA arising from viable cells. This ability to extract DNA selectively from living cells is extremely powerful, and bears great relevance to many microbiological arenas.

  14. Mathematical modelling of the viable epidermis: impact of cell shape and vertical arrangement

    KAUST Repository

    Wittum, Rebecca

    2017-12-07

    In-silico methods are valuable tools for understanding the barrier function of the skin. The key benefit is that mathematical modelling allows the interplay between cell shape and function to be elucidated. This study focuses on the viable (living) epidermis. For this region, previous works suggested a diffusion model and an approximation of the cells by hexagonal prisms. The work at hand extends this in three ways. First, the extracellular space is treated with full spatial resolution. This induces a decrease of permeability by about 10%. Second, cells of tetrakaidecahedral shape are considered, in addition to the original hexagonal prisms. For both cell types, the resulting membrane permeabilities are compared. Third, for the first time, the influence of cell stacking in the vertical direction is considered. This is particularly important for the stratum granulosum, where tight junctions are present.

  15. Grazing of particle-associated bacteria-an elimination of the non-viable fraction.

    Science.gov (United States)

    Gonsalves, Maria-Judith; Fernandes, Sheryl Oliveira; Priya, Madasamy Lakshmi; LokaBharathi, Ponnapakkam Adikesavan

    Quantification of bacteria being grazed by microzooplankton is gaining importance since they serve as energy subsidies for higher trophic levels which consequently influence fish production. Hence, grazing pressure on viable and non-viable fraction of free and particle-associated bacteria in a tropical estuary controlled mainly by protist grazers was estimated using the seawater dilution technique. In vitro incubations over a period of 42h showed that at the end of 24h, growth coefficient (k) of particle-associated bacteria was 9 times higher at 0.546 than that of free forms. Further, 'k' value of viable cells on particles was double that of free forms at 0.016 and 0.007, respectively. While bacteria associated with particles were grazed (coefficient of removal (g)=0.564), the free forms were relatively less grazed indicating that particle-associated bacteria were exposed to grazers in these waters. Among the viable and non-viable forms, 'g' of non-viable fraction (particle-associated bacteria=0.615, Free=0.0086) was much greater than the viable fraction (particle-associated bacteria=0.056, Free=0.068). Thus, grazing on viable cells was relatively low in both the free and attached states. These observations suggest that non-viable forms of particle-associated bacteria were more prone to grazing and were weeded out leaving the viable cells to replenish the bacterial standing stock. Particle colonization could thus be a temporary refuge for the "persistent variants" where the viable fraction multiply and release their progeny. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  16. Grazing of particle-associated bacteria-an elimination of the non-viable fraction

    Directory of Open Access Journals (Sweden)

    Maria-Judith Gonsalves

    Full Text Available Abstract Quantification of bacteria being grazed by microzooplankton is gaining importance since they serve as energy subsidies for higher trophic levels which consequently influence fish production. Hence, grazing pressure on viable and non-viable fraction of free and particle-associated bacteria in a tropical estuary controlled mainly by protist grazers was estimated using the seawater dilution technique. In vitro incubations over a period of 42 h showed that at the end of 24 h, growth coefficient (k of particle-associated bacteria was 9 times higher at 0.546 than that of free forms. Further, ‘k’ value of viable cells on particles was double that of free forms at 0.016 and 0.007, respectively. While bacteria associated with particles were grazed (coefficient of removal (g = 0.564, the free forms were relatively less grazed indicating that particle-associated bacteria were exposed to grazers in these waters. Among the viable and non-viable forms, ‘g’ of non-viable fraction (particle-associated bacteria = 0.615, Free = 0.0086 was much greater than the viable fraction (particle-associated bacteria = 0.056, Free = 0.068. Thus, grazing on viable cells was relatively low in both the free and attached states. These observations suggest that non-viable forms of particle-associated bacteria were more prone to grazing and were weeded out leaving the viable cells to replenish the bacterial standing stock. Particle colonization could thus be a temporary refuge for the “persistent variants” where the viable fraction multiply and release their progeny.

  17. Sensitive and Specific Biomimetic Lipid Coated Microfluidics to Isolate Viable Circulating Tumor Cells and Microemboli for Cancer Detection.

    Directory of Open Access Journals (Sweden)

    Jia-Yang Chen

    Full Text Available Here we presented a simple and effective membrane mimetic microfluidic device with antibody conjugated supported lipid bilayer (SLB "smart coating" to capture viable circulating tumor cells (CTCs and circulating tumor microemboli (CTM directly from whole blood of all stage clinical cancer patients. The non-covalently bound SLB was able to promote dynamic clustering of lipid-tethered antibodies to CTC antigens and minimized non-specific blood cells retention through its non-fouling nature. A gentle flow further flushed away loosely-bound blood cells to achieve high purity of CTCs, and a stream of air foam injected disintegrate the SLB assemblies to release intact and viable CTCs from the chip. Human blood spiked cancer cell line test showed the ~95% overall efficiency to recover both CTCs and CTMs. Live/dead assay showed that at least 86% of recovered cells maintain viability. By using 2 mL of peripheral blood, the CTCs and CTMs counts of 63 healthy and colorectal cancer donors were positively correlated with the cancer progression. In summary, a simple and effective strategy utilizing biomimetic principle was developed to retrieve viable CTCs for enumeration, molecular analysis, as well as ex vivo culture over weeks. Due to the high sensitivity and specificity, it is the first time to show the high detection rates and quantity of CTCs in non-metastatic cancer patients. This work offers the values in both early cancer detection and prognosis of CTC and provides an accurate non-invasive strategy for routine clinical investigation on CTCs.

  18. Dye Sensitized Solar Cells for Economically Viable Photovoltaic Systems.

    Science.gov (United States)

    Jung, Hyun Suk; Lee, Jung-Kun

    2013-05-16

    TiO2 nanoparticle-based dye sensitized solar cells (DSSCs) have attracted a significant level of scientific and technological interest for their potential as economically viable photovoltaic devices. While DSSCs have multiple benefits such as material abundance, a short energy payback period, constant power output, and compatibility with flexible applications, there are still several challenges that hold back large scale commercialization. Critical factors determining the future of DSSCs involve energy conversion efficiency, long-term stability, and production cost. Continuous advancement of their long-term stability suggests that state-of-the-art DSSCs will operate for over 20 years without a significant decrease in performance. Nevertheless, key questions remain in regards to energy conversion efficiency improvements and material cost reduction. In this Perspective, the present state of the field and the ongoing efforts to address the requirements of DSSCs are summarized with views on the future of DSSCs.

  19. Effects of γ irradiation of hydra: elimination of interstitial cells from viable hydra

    International Nuclear Information System (INIS)

    Fradkin, M.; Kakis, H.; Campbell, R.D.

    1978-01-01

    Hydra attenuata and H. magnipapillata were γ-irradiated from a cesium source. All doses which had any observable effect (3000 rad and above) resulted in a reduction in the number of interstitial cells and of their differentiated product cells, or in the complete elimination of these cells. Interstitial cells were essentially completely eliminated within 5 days after irradiation doses above 5500 rad, and these hydra died. Irradiation doses of 4200 to 5500 rad resulted in a mixture of effects: some hydra recovered completely, some lost all interstitial cells and died, and some lost interstitial cells but could be propagated, as asexually reproducing clones, by hand feeding them. Hydra of some of these hand-fed clones entirely lacked interstitial cells and did not recover interstitial cells during subsequent culturing. Yet when these hydra were repopulated by interstitial cells from a normal hydra, they were restored to normal. Nerve cells became depleted more slowly than interstitial cells following irradiation, so animals can be obtained which possess nerve but no stem (interstitial) cells. The nerve cells and other derivatives of interstitial cells eventually disappear upon prolonged culture of the hydra. Thus γ irradiation can be used to eliminate interstitial cells from hydra, leaving viable polyps composed only of epithelial cells

  20. A reliable protocol for the isolation of viable, chondrogenically differentiated human mesenchymal stem cells from high-density pellet cultures.

    Science.gov (United States)

    Ullah, Mujib; Hamouda, Houda; Stich, Stefan; Sittinger, Michael; Ringe, Jochen

    2012-12-01

    Administration of chondrogenically differentiated mesenchymal stem cells (MSC) is discussed as a promising approach for the regenerative treatment of injured or diseased cartilage. The high-density pellet culture is the standard culture for chondrogenic differentiation, but cells in pellets secrete extracellular matrix (ECM) that they become entrapped in. Protocols for cell isolation from pellets often result in cell damage and dedifferentiation towards less differentiated MSC. Therefore, our aim was to develop a reliable protocol for the isolation of viable, chondrogenically differentiated MSC from high-density pellet cultures. Human bone marrow MSC were chondrogenically stimulated with transforming growth factor-β3, and the cartilaginous structure of the pellets was verified by alcian blue staining of cartilage proteoglycans, antibody staining of cartilage collagen type II, and quantitative real-time reverse-transcription polymerase chain reaction of the marker genes COL2A1 and SOX9. Trypsin and collagenases II and P were tested alone or in combination, and for different concentrations and times, to find a protocol for optimized pellet digestion. Whereas trypsin was not able to release viable cells, 90-min digestion with 300 U of collagenase II, 20 U of collagenase P, and 2 mM CaCl2 worked quite well and resulted in about 2.5×10(5) cells/pellet. The protocol was further optimized for the separation of released cells and ECM from each other. Cells were alcian blue and collagen type II positive and expressed COL2A1 and SOX9, verifying a chondrogenic character. However, they had different morphological shapes. The ECM was also uniformly alcian blue and collagen type II positive but showed different organizational and structural forms. To conclude, our protocol allows the reliable isolation of a defined number of viable, chondrogenically differentiated MSC from high-density pellet cultures. Such cells, as well as the ECM components, are of interest as

  1. PMA-PhyloChip DNA Microarray to Elucidate Viable Microbial Community Structure

    Science.gov (United States)

    Venkateswaran, Kasthuri J.; Stam, Christina N.; Andersen, Gary L.; DeSantis, Todd

    2011-01-01

    Since the Viking missions in the mid-1970s, traditional culture-based methods have been used for microbial enumeration by various NASA programs. Viable microbes are of particular concern for spacecraft cleanliness, for forward contamination of extraterrestrial bodies (proliferation of microbes), and for crew health/safety (viable pathogenic microbes). However, a "true" estimation of viable microbial population and differentiation from their dead cells using the most sensitive molecular methods is a challenge, because of the stability of DNA from dead cells. The goal of this research is to evaluate a rapid and sensitive microbial detection concept that will selectively estimate viable microbes. Nucleic acid amplification approaches such as the polymerase chain reaction (PCR) have shown promise for reducing time to detection for a wide range of applications. The proposed method is based on the use of a fluorescent DNA intercalating agent, propidium monoazide (PMA), which can only penetrate the membrane of dead cells. The PMA-quenched reaction mixtures can be screened, where only the DNA from live cells will be available for subsequent PCR reaction and microarray detection, and be identified as part of the viable microbial community. An additional advantage of the proposed rapid method is that it will detect viable microbes and differentiate from dead cells in only a few hours, as opposed to less comprehensive culture-based assays, which take days to complete. This novel combination approach is called the PMA-Microarray method. DNA intercalating agents such as PMA have previously been used to selectively distinguish between viable and dead bacterial cells. Once in the cell, the dye intercalates with the DNA and, upon photolysis under visible light, produces stable DNA adducts. DNA cross-linked in this way is unavailable for PCR. Environmental samples suspected of containing a mixture of live and dead microbial cells/spores will be treated with PMA, and then incubated

  2. Real-time monitoring of non-viable airborne particles correlates with airborne colonies and represents an acceptable surrogate for daily assessment of cell-processing cleanroom performance.

    Science.gov (United States)

    Raval, Jay S; Koch, Eileen; Donnenberg, Albert D

    2012-10-01

    Airborne particulate monitoring is mandated as a component of good manufacturing practice. We present a procedure developed to monitor and interpret airborne particulates in an International Organization for Standardization (ISO) class 7 cleanroom used for the cell processing of Section 351 and Section 361 products. We collected paired viable and non-viable airborne particle data over a period of 1 year in locations chosen to provide a range of air quality. We used receiver operator characteristic (ROC) analysis to determine empirically the relationship between non-viable and viable airborne particle counts. Viable and non-viable particles were well-correlated (r(2) = 0.78), with outlier observations at the low end of the scale (non-viable particles without detectable airborne colonies). ROC analysis predicted viable counts ≥ 0.5/feet(3) (a limit set by the United States Pharmacopeia) at an action limit of ≥ 32 000 particles (≥ 0.5 µ)/feet(3), with 95.6% sensitivity and 50% specificity. This limit was exceeded 2.6 times during 18 months of retrospective daily cleanroom data (an expected false alarm rate of 1.3 times/year). After implementing this action limit, we were alerted in real time to an air-handling failure undetected by our hospital facilities management. A rational action limit for non-viable particles was determined based on the correlation with airborne colonies. Reaching or exceeding the action limit of 32 000 non-viable particles/feet(3) triggers suspension of cleanroom cell-processing activities, deep cleaning, investigation of air handling, and a deviation management process. Our full procedure for particle monitoring is available as an online supplement.

  3. Viable Cancer Cells in the Remnant Stomach are a Potential Source of Peritoneal Metastasis after Curative Distal Gastrectomy for Gastric Cancer.

    Science.gov (United States)

    Murata, Satoshi; Yamamoto, Hiroshi; Yamaguchi, Tsuyoshi; Kaida, Sachiko; Ishida, Mitsuaki; Kodama, Hirokazu; Takebayashi, Katsushi; Shimizu, Tomoharu; Miyake, Toru; Tani, Tohru; Kushima, Ryoji; Tani, Masaji

    2016-09-01

    The mechanisms underlying peritoneal metastasis (PM) after curative gastrectomy for gastric cancer (GC) are not well elucidated. This study assessed whether viable cancer cells, including cancer stemlike cells (CSCs), were present in the remnant stomach immediately before gastrointestinal (GI) tract reconstruction because these could be a source of PM after gastrectomy. Saline fluid used for remnant stomach lumen irrigation before GI reconstruction was prospectively collected from 142 consecutive patients undergoing distal gastrectomy for GC and cytologically examined. Proliferative activity (Ki67 staining) and stemness (expression of the CSC surface markers CD44s or CD44v6) were evaluated in detected cancer cells. Viable cancer cells were detected in 33 (23.2 %) of the 142 remnant stomachs. These cells formed clusters and stained positively for Ki67, indicating proliferation. Cancer cells in remnant stomachs and surface cancer cells in primary GCs from 10 (30.3 %) of these 33 cases also stained positively for CD44s or CD44v6. In a multiple logistic regression analysis, advanced cancer (odds ratio [OR], 4.65; 95 % confidence interval [CI], 1.32-16.4; P = 0.017), tumor size of 40 mm or larger (OR, 3.78; 95 % CI, 1.12-12.8; P = 0.033), and histologic differentiation (OR, 3.10; 95 % CI, 1.30-7.40; P = 0.011) were associated independently with the presence of cancer cells in the remnant stomach. Viable, proliferative, and clustered cancer cells, including CSCs, were found in remnant gastric lumens immediately before GI reconstruction, indicating a possible cellular source of PM after curative gastrectomy for GC. Dissemination of gastric contents into the peritoneal cavity should be avoided during GI reconstruction.

  4. Tumor-selective replication herpes simplex virus-based technology significantly improves clinical detection and prognostication of viable circulating tumor cells

    DEFF Research Database (Denmark)

    Zhang, Wen; Bao, Li; Yang, Shaoxing

    2016-01-01

    Detection of circulating tumor cells remains a significant challenge due to their vast physical and biological heterogeneity. We developed a cell-surface-marker-independent technology based on telomerase-specific, replication-selective oncolytic herpes-simplex-virus-1 that targets telomerase......-reverse-transcriptase-positive cancer cells and expresses green-fluorescent-protein that identifies viable CTCs from a broad spectrum of malignancies. Our method recovered 75.5-87.2% of tumor cells spiked into healthy donor blood, as validated by different methods, including single cell sequencing. CTCs were detected in 59-100% of 326...

  5. Clinically viable magnetic poly(lactide-co-glycolide) (PLGA) particles for MRI-based cell tracking

    Science.gov (United States)

    Granot, Dorit; Nkansah, Michael K.; Bennewitz, Margaret F.; Tang, Kevin S.; Markakis, Eleni A.; Shapiro, Erik M.

    2013-01-01

    Purpose To design, fabricate, characterize and in vivo assay clinically viable magnetic particles for MRI-based cell tracking. Methods PLGA encapsulated magnetic nano- and microparticles were fabricated. Multiple biologically relevant experiments were performed to assess cell viability, cellular performance and stem cell differentiation. In vivo MRI experiments were performed to separately test cell transplantation and cell migration paradigms, as well as in vivo biodegradation. Results Highly magnetic nano- (~100 nm) and microparticles (~1–2 μm) were fabricated. Magnetic cell labeling in culture occurred rapidly achieving 3–50 pg Fe/cell at 3 hrs for different particles types, and >100 pg Fe/cell after 10 hours, without the requirement of a transfection agent, and with no effect on cell viability. The capability of magnetically labeled mesenchymal or neural stem cells to differentiate down multiple lineages, or for magnetically labeled immune cells to release cytokines following stimulation, was uncompromised. An in vivo biodegradation study revealed that NPs degraded ~80% over the course of 12 weeks. MRI detected as few as 10 magnetically labeled cells, transplanted into the brains of rats. Also, these particles enabled the in vivo monitoring of endogenous neural progenitor cell migration in rat brains over 2 weeks. Conclusion The robust MRI properties and benign safety profile of these particles make them promising candidates for clinical translation for MRI-based cell tracking. PMID:23568825

  6. SALMONELLA SPECIE AND TOTAL VIABLE BACTERIAL LOAD IN ROASTED CHICKENS SOLD IN JOS-NIGERIA

    OpenAIRE

    Carol Okoli; Okonji M.C; Ugoh S.C; Okolo S.N; Okoli A.C; Alu A.J

    2007-01-01

    The study was to investigate for the presence of Salmonella specie and total viable aerobic bacterial load in roasted chickens sold in Jos. The study was carried out on twenty five chicken samples. No salmonella specie was isolated from the samples. However, other bacterial organisms were isolates, viz: 9(36%) of the samples yielded E.coli; 5(20%) yielded Citobacter species; 3(12%) yielded Proteus species and 6(24%) yielded Klebsiella species while 2(8%) showed no growth. An average total via...

  7. Selective detection of viable seed-borne Acidovorax citrulli by real-time PCR with propidium monoazide.

    Science.gov (United States)

    Tian, Qian; Feng, Jian-Jun; Hu, Jie; Zhao, Wen-Jun

    2016-10-14

    In recent years, use of the DNA-intercalating dye propidium monoazide (PMA) in real-time PCR has been reported as a novel method to detect viable bacteria in different types of samples, such as food, environmental, and microbiological samples. In this study, viable cells of Acidovorax citrulli, the causal agent of bacterial seedling blight and fruit blotch, were selectively detected and differentiated from dead cells by real-time fluorescent polymerase chain reaction amplification after the bacterial solution was treated with the DNA-binding dye PMA. The primers and TaqMan probe were based on the A. citrulli genome (Aave_1909, Gene ID: 4669443) and were highly specific for A. citrulli. The detection threshold of this assay was 10 3 colony-forming units per mL (CFU/mL) in pure cell suspensions containing viable and dead cells and infected watermelon seeds. Application of this assay enables the selective detection of viable cells of A. citrulli and facilitates monitoring of the pathogen in watermelon and melon seeds.

  8. Rotary orbital suspension culture of embryonic stem cell-derived neural stem/progenitor cells: impact of hydrodynamic culture on aggregate yield, morphology and cell phenotype.

    Science.gov (United States)

    Laundos, Tiago L; Silva, Joana; Assunção, Marisa; Quelhas, Pedro; Monteiro, Cátia; Oliveira, Carla; Oliveira, Maria J; Pêgo, Ana P; Amaral, Isabel F

    2017-08-01

    Embryonic stem (ES)-derived neural stem/progenitor cells (ES-NSPCs) constitute a promising cell source for application in cell therapies for the treatment of central nervous system disorders. In this study, a rotary orbital hydrodynamic culture system was applied to single-cell suspensions of ES-NSPCs, to obtain homogeneously-sized ES-NSPC cellular aggregates (neurospheres). Hydrodynamic culture allowed the formation of ES-NSPC neurospheres with a narrower size distribution than statically cultured neurospheres, increasing orbital speeds leading to smaller-sized neurospheres and higher neurosphere yield. Neurospheres formed under hydrodynamic conditions (72 h at 55 rpm) showed higher cell compaction and comparable percentages of viable, dead, apoptotic and proliferative cells. Further characterization of cellular aggregates provided new insights into the effect of hydrodynamic shear on ES-NSPC behaviour. Rotary neurospheres exhibited reduced protein levels of N-cadherin and β-catenin, and higher deposition of laminin (without impacting fibronectin deposition), matrix metalloproteinase-2 (MMP-2) activity and percentage of neuronal cells. In line with the increased MMP-2 activity levels found, hydrodynamically-cultured neurospheres showed higher outward migration on laminin. Moreover, when cultured in a 3D fibrin hydrogel, rotary neurospheres generated an increased percentage of neuronal cells. In conclusion, the application of a constant orbital speed to single-cell suspensions of ES-NSPCs, besides allowing the formation of homogeneously-sized neurospheres, promoted ES-NSPC differentiation and outward migration, possibly by influencing the expression of cell-cell adhesion molecules and the secretion of proteases/extracellular matrix proteins. These findings are important when establishing the culture conditions needed to obtain uniformly-sized ES-NSPC aggregates, either for use in regenerative therapies or in in vitro platforms for biomaterial development or

  9. Separation of viable lactic acid bacteria from fermented milk

    Directory of Open Access Journals (Sweden)

    Tomohiko Nishino

    2018-04-01

    Full Text Available Probiotics are live microorganisms that provide health benefits to humans. Some lactic acid bacteria (LAB are probiotic organisms used in the production of fermented foods, such as yogurt, cheese, and pickles. Given their widespread consumption, it is important to understand the physiological state of LAB in foods such as yogurt. However, this analysis is complicated, as it is difficult to separate the LAB from milk components such as solid curds, which prevent cell separation by dilution or centrifugation. In this study, we successfully separated viable LAB from yogurt by density gradient centrifugation. The recovery rate was >90 %, and separation was performed until the stationary phase. Recovered cells were observable by microscopy, meaning that morphological changes and cell viability could be directly detected at the single-cell level. The results indicate that viable LAB can be easily purified from fermented milk. We expect that this method will be a useful tool for the analysis of various aspects of probiotic cells, including their enzyme activity and protein expression. Keywords: Food analysis, Microbiology

  10. On-Chip Dielectrophoretic Separation and Concentration of Viable, Non-Viable and Viable but Not Culturable (VBNC) Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Packard, M M; Shusteff, M; Alocilja, E C

    2012-04-12

    Although bacterial culture remains the gold standard for detection of viable bacteria in environmental specimens, the typical time requirement of twenty-four hours can delay and even jeopardize appropriate public health intervention. In addition, culture is incapable of detecting viable but not culturable (VBNC) species. Conversely, nucleic acid and antibody-based methods greatly decrease time to detection but rarely characterize viability of the bacteria detected. Through selection by membrane permeability, the method described in this work employs positive dielectrophoresis (pDEP) for separation and purification of viable and VBNC species from water and allows concentration of bacteria for downstream applications.

  11. Characterization of the Viable but Nonculturable (VBNC State in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Mohammad Salma

    Full Text Available The Viable But Non Culturable (VBNC state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to "resuscitate". The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO2-induced VBNC state in S. cerevisiae and that the stress removal allows the "resuscitation" of VBNC cells during the VBNC state.

  12. Entry of Vibrio harveyi and Vibrio fischeri into the viable but nonculturable state.

    Science.gov (United States)

    Ramaiah, N; Ravel, J; Straube, W L; Hill, R T; Colwell, R R

    2002-01-01

    Physiological responses of marine luminous bacteria, Vibrio harveyi (ATCC 14216) and V. fischeri (UM1373) to nutrient-limited normal strength (35 ppt iso-osmolarity) and low (10 ppt hypo-osmolarity) salinity conditions were determined. Plate counts, direct viable counts, actively respiring cell counts, nucleoid-containing cell counts, and total counts were determined. Vibrio harveyi incubated at 22 degrees C in nutrient-limited artificial seawater (ASW) became nonculturable after approximately 62 and 45 d in microcosms of 35 ppt and 10 ppt ASW, respectively. In contrast, V. fischeri became nonculturable at approximately 55 and 31 d in similar microcosms. Recovery of both culturability and luminescence of cells in the viable but nonculturable state was achieved by addition of nutrient broth or nutrient broth supplemented with a carbon source, including luminescence-stimulating compounds. Temperature upshift from 22 degrees C to 30 degrees C or 37 degrees C did not result in recovery from nonculturability. The study confirms entry of V. harveyi and V. fischeri into the viable but nonculturable state under low-nutrient conditions and demonstrates nutrient-dependent resuscitation from this state. This study confirms loss of luminescence of V. harveyi and V. fischeri on entry into the viable but nonculturable state and suggests that enumeration of luminescent cells in water samples may be a rapid method to deduce the nutrient status of a water sample.

  13. Introduce of Viable But Nonculturable Bacteria

    Directory of Open Access Journals (Sweden)

    Mehdi Hassanshahian

    2008-03-01

    Full Text Available Viable-But-Nonculturable-State (VBNC is the condition in which bacteria fail to grow on their routine bacteriological media where they would normally grow and develop into colonies, but are still alive and capable of renewed metabolic activity. VBNC state is useful for evaluating public health and for ascertaining the sterility of drinking water, pharmaceuticals, and foodstuff. A number of bacteria, mostly pathogenic to humans, have been proved to enter into this state in response to natural stresses such as starvation, incubation out of optimum growth temperature, increased osmotic pressure, etc. Once in the VBNC state, they undergo various physiological, structural, and genetic alterations. These alterations result in reduced cell size, conversion from bacilli to coccid, thickened cell walls, and peptidoglycan gaining many cross links. Metabolic changes also occur that include reductions in growth, nutrient transport, and respiratory rate; biosynthesis of new protein, and ATP remaining at a constant level. It has been shown that in the VBNC state, some pathogens conserve their virulence properties. Gene expression continues in the VBNC cell. Nucleic acids remain intact in the early VBNC phase but they gradually undergo degradation with prolonged VBNC. Cytological methods such as direct viable count and reduction of tetrazolium salts, and molecular methods such as reverse transcription polymerase chain reaction and green fluorescent protein have been used for the study of VBNC. Resuscitation from VBNC state starts when the inducing factor(s is/are lifted. Factors that help the resuscitation of VBNC bacteria include addition of certain nutrients and chemicals, introduction of a few culturable cells into the VBNC cell population, and passage through the animal host. As virulence properties are sustained during the VBNC phase, special care must be paid when evaluating sterility of drinking water.

  14. Strategies for improving production performance of probiotic Pediococcus acidilactici viable cell by overcoming lactic acid inhibition.

    Science.gov (United States)

    Othman, Majdiah; Ariff, Arbakariya B; Wasoh, Helmi; Kapri, Mohd Rizal; Halim, Murni

    2017-11-27

    Lactic acid bacteria are industrially important microorganisms recognized for fermentative ability mostly in their probiotic benefits as well as lactic acid production for various applications. Fermentation conditions such as concentration of initial glucose in the culture, concentration of lactic acid accumulated in the culture, types of pH control strategy, types of aeration mode and different agitation speed had influenced the cultivation performance of batch fermentation of Pediococcus acidilactici. The maximum viable cell concentration obtained in constant fed-batch fermentation at a feeding rate of 0.015 L/h was 6.1 times higher with 1.6 times reduction in lactic acid accumulation compared to batch fermentation. Anion exchange resin, IRA 67 was found to have the highest selectivity towards lactic acid compared to other components studied. Fed-batch fermentation of P. acidilactici coupled with lactic acid removal system using IRA 67 resin showed 55.5 and 9.1 times of improvement in maximum viable cell concentration compared to fermentation without resin for batch and fed-batch mode respectively. The improvement of the P. acidilactici growth in the constant fed-batch fermentation indicated the use of minimal and simple process control equipment is an effective approach for reducing by-product inhibition. Further improvement in the cultivation performance of P. acidilactici in fed-bath fermentation with in situ addition of anion-exchange resin significantly helped to enhance the growth of P. acidilactici by reducing the inhibitory effect of lactic acid and thus increasing probiotic production.

  15. Dendritic cells take up and present antigens from viable and apoptotic polymorphonuclear leukocytes.

    Directory of Open Access Journals (Sweden)

    Carlos Alfaro

    Full Text Available Dendritic cells (DC are endowed with the ability to cross-present antigens from other cell types to cognate T cells. DC are poised to meet polymorphonuclear leukocytes (PMNs as a result of being co-attracted by interleukin-8 (IL-8, for instance as produced by tumor cells or infected tissue. Human monocyte-derived and mouse bone marrow-derived DC can readily internalize viable or UV-irradiated PMNs. Such internalization was abrogated at 4°C and partly inhibited by anti-CD18 mAb. In mice, DC which had internalized PMNs containing electroporated ovalbumin (OVA protein, were able to cross-present the antigen to CD8 (OT-1 and CD4 (OT-2 TCR-transgenic T cells. Moreover, in humans, tumor cell debris is internalized by PMNs and the tumor-cell material can be subsequently taken up from the immunomagnetically re-isolated PMNs by DC. Importantly, if human neutrophils had endocytosed bacteria, they were able to trigger the maturation program of the DC. Moreover, when mouse PMNs with E. coli in their interior are co-injected in the foot pad with DC, many DC loaded with fluorescent material from the PMNs reach draining lymph nodes. Using CT26 (H-2(d mouse tumor cells, it was observed that if tumor cells are intracellularly loaded with OVA protein and UV-irradiated, they become phagocytic prey of H-2(d PMNs. If such PMNs, that cannot present antigens to OT-1 T cells, are immunomagnetically re-isolated and phagocytosed by H-2(b DC, such DC productively cross-present OVA antigen determinants to OT-1 T cells. Cross-presentation to adoptively transferred OT-1 lymphocytes at draining lymph nodes also take place when OVA-loaded PMNs (H-2(d are coinjected in the footpad of mice with autologous DC (H-2(b. In summary, our results indicate that antigens phagocytosed by short-lived PMNs can be in turn internalized and productively cross-presented by DC.

  16. Glioma Surgical Aspirate: A Viable Source of Tumor Tissue for Experimental Research

    International Nuclear Information System (INIS)

    Day, Bryan W.; Stringer, Brett W.; Wilson, John; Jeffree, Rosalind L.; Jamieson, Paul R.

    2013-01-01

    Brain cancer research has been hampered by a paucity of viable clinical tissue of sufficient quality and quantity for experimental research. This has driven researchers to rely heavily on long term cultured cells which no longer represent the cancers from which they were derived. Resection of brain tumors, particularly at the interface between normal and tumorigenic tissue, can be carried out using an ultrasonic surgical aspirator (CUSA) that deposits liquid (blood and irrigation fluid) and resected tissue into a sterile bottle for disposal. To determine the utility of CUSA-derived glioma tissue for experimental research, we collected 48 CUSA specimen bottles from glioma patients and analyzed both the solid tissue fragments and dissociated tumor cells suspended in the liquid waste fraction. We investigated if these fractions would be useful for analyzing tumor heterogeneity, using IHC and multi-parameter flow cytometry; we also assessed culture generation and orthotopic xenograft potential. Both cell sources proved to be an abundant, highly viable source of live tumor cells for cytometric analysis, animal studies and in-vitro studies. Our findings demonstrate that CUSA tissue represents an abundant viable source to conduct experimental research and to carry out diagnostic analyses by flow cytometry or other molecular diagnostic procedures

  17. Separation of viable and non-viable tomato (Solanum lycopersicum L.) seeds using single seed near-infrared spectroscopy

    DEFF Research Database (Denmark)

    Shrestha, Santosh; Deleuran, Lise Christina; Gislum, René

    2017-01-01

    Single seed near-infrared (NIR) spectroscopy is a non-destructive technology commonly used for predicting lipids, proteins, carbohydrates and water content of agricultural products. The aim of the current study is to investigate the prospects of NIR spectroscopy in classifying viable and non...... identified as important for classification of viable and non-viable tomato seeds by iPLS-DA. The sensitivity i.e. ability to correctly identify the positive samples and specificity i.e. ability to reject the negative samples of the (iPLS-DA) model on identified spectral regions for prediction of viable......-viable tomato seeds of two cultivars using chemometrics. The data exploration were performed by principal component analysis (PCA). Subsequently, viable and non-viable seeds were classified by partial least squares-discriminant analysis (PLS-DA) and interval PLS-DA (iPLS-DA). The indication of clustering...

  18. Improving carbon dioxide yields and cell efficiencies for ethanol oxidation by potential scanning

    Science.gov (United States)

    Majidi, Pasha; Pickup, Peter G.

    2014-12-01

    An ethanol electrolysis cell with aqueous ethanol supplied to the anode and nitrogen at the cathode has been operated under potential cycling conditions in order to increase the yield of carbon dioxide and thereby increase cell efficiency relative to operation at a fixed potential. At ambient temperature, faradaic yields of CO2 as high as 26% have been achieved, while only transient CO2 production was observed at constant potential. Yields increased substantially at higher temperatures, with maximum values at Pt anodes reaching 45% at constant potential and 65% under potential cycling conditions. Use of a PtRu anode increased the cell efficiency by decreasing the anode potential, but this was offset by decreased CO2 yields. Nonetheless, cycling increased the efficiency relative to constant potential. The maximum yields at PtRu and 80 °C were 13% at constant potential and 32% under potential cycling. The increased yields under cycling conditions have been attributed to periodic oxidative stripping of adsorbed CO, which occurs at lower potentials on PtRu than on Pt. These results will be important in the optimization of operating conditions for direct ethanol fuel cells and for the electrolysis of ethanol to produce clean hydrogen.

  19. Designing primers and evaluation of the efficiency of propidium monoazide – Quantitative polymerase chain reaction for counting the viable cells of Lactobacillus gasseri and Lactobacillus salivarius

    Directory of Open Access Journals (Sweden)

    Chieh-Hsien Lai

    2017-07-01

    Full Text Available The purpose of this study is to evaluate the efficiency of using propidium monoazide (PMA real-time quantitative polymerase chain reaction (qPCR to count the viable cells of Lactobacillus gasseri and Lactobacillus salivarius in probiotic products. Based on the internal transcription spacer and 23S rRNA genes, two primer sets specific for these two Lactobacillus species were designed. For a probiotic product, the total deMan Rogosa Sharpe plate count was 8.65±0.69 log CFU/g, while for qPCR, the cell counts of L. gasseri and L. salivarius were 8.39±0.14 log CFU/g and 8.57±0.24 log CFU/g, respectively. Under the same conditions, for its heat-killed product, qPCR counts for L. gasseri and L. salivarius were 6.70±0.16 log cells/g and 7.67±0.20 log cells/g, while PMA-qPCR counts were 5.33±0.18 log cells/g and 5.05±0.23 log cells/g, respectively. For cell dilutions with a viable cell count of 8.5 log CFU/mL for L. gasseri and L. salivarius, after heat killing, the PMA-qPCR count for both Lactobacillus species was near 5.5 log cells/mL. When the PMA-qPCR counts of these cell dilutions were compared before and after heat killing, although some DNA might be lost during the heat killing, significant qPCR signals from dead cells, i.e., about 4–5 log cells/mL, could not be reduced by PMA treatment. Increasing PMA concentrations from 100 μM to 200 μM or light exposure time from 5 minutes to 15 minutes had no or, if any, only minor effect on the reduction of qPCR signals from their dead cells. Thus, to differentiate viable lactic acid bacterial cells from dead cells using the PMA-qPCR method, the efficiency of PMA to reduce the qPCR signals from dead cells should be notable.

  20. Synthesis of red fluorescent graphene quantum dot-europium complex composites as a viable bio imaging platform

    International Nuclear Information System (INIS)

    Liu, Yanting; Fan, Louzhen; Zhou, Shixin; Fan, Hong

    2016-01-01

    We have prepared graphene quantum dot-europium(III) complex composites by noncovalently connecting chelating ligands dibenzoylmethane (DBM) and 1,10-phenanthroline (Phen) with graphene quantum dots (GQDs) first, followed by coordination to Eu(III). The resulting composites are well water-soluble and display red fluorescence of high color purity. The composites were characterized by transmission electron microscopy, X-ray photoelectron spectroscopy and X-ray diffraction. Aqueous solutions of the composites under 365 nm excitation display fluorescence with a peak at 613 nm and a quantum yield as high as 15.5 %. The good water solubility and stable photoluminescence make the composites very different from other Eu(III)-based coordination complexes. The composites are cell viable and can be used to label both the cell membrane and the cytoplasm of MCF-7 cells. They are also shown to act as bioprobes for in-vivo localization of tumorous tissue. In our perception, such composites are expected to possess wide scope because of the many functionalizations that are possible with GQDs. (author)

  1. Designing primers and evaluation of the efficiency of propidium monoazide - Quantitative polymerase chain reaction for counting the viable cells of Lactobacillus gasseri and Lactobacillus salivarius.

    Science.gov (United States)

    Lai, Chieh-Hsien; Wu, Sih-Rong; Pang, Jen-Chieh; Ramireddy, Latha; Chiang, Yu-Cheng; Lin, Chien-Ku; Tsen, Hau-Yang

    2017-07-01

    The purpose of this study is to evaluate the efficiency of using propidium monoazide (PMA) real-time quantitative polymerase chain reaction (qPCR) to count the viable cells of Lactobacillus gasseri and Lactobacillus salivarius in probiotic products. Based on the internal transcription spacer and 23S rRNA genes, two primer sets specific for these two Lactobacillus species were designed. For a probiotic product, the total deMan Rogosa Sharpe plate count was 8.65±0.69 log CFU/g, while for qPCR, the cell counts of L. gasseri and L. salivarius were 8.39±0.14 log CFU/g and 8.57±0.24 log CFU/g, respectively. Under the same conditions, for its heat-killed product, qPCR counts for L. gasseri and L. salivarius were 6.70±0.16 log cells/g and 7.67±0.20 log cells/g, while PMA-qPCR counts were 5.33±0.18 log cells/g and 5.05±0.23 log cells/g, respectively. For cell dilutions with a viable cell count of 8.5 log CFU/mL for L. gasseri and L. salivarius, after heat killing, the PMA-qPCR count for both Lactobacillus species was near 5.5 log cells/mL. When the PMA-qPCR counts of these cell dilutions were compared before and after heat killing, although some DNA might be lost during the heat killing, significant qPCR signals from dead cells, i.e., about 4-5 log cells/mL, could not be reduced by PMA treatment. Increasing PMA concentrations from 100 μM to 200 μM or light exposure time from 5 minutes to 15 minutes had no or, if any, only minor effect on the reduction of qPCR signals from their dead cells. Thus, to differentiate viable lactic acid bacterial cells from dead cells using the PMA-qPCR method, the efficiency of PMA to reduce the qPCR signals from dead cells should be notable. Copyright © 2016. Published by Elsevier B.V.

  2. Entry of Vibrio harveyi and Vibrio fischeri into the viable but nonculturable state

    Digital Repository Service at National Institute of Oceanography (India)

    Ramaiah, Neelam; Ravel, J.; Straube, W.L.; Hill, R.T.; Colwell, R.R.

    , direct viable counts, actively respiring cell counts, nucleoid-containing cell counts, and total counts were determined. V. harveyi incubated at 22 degrees C nutrient-limited artificial seawater (ASW) became nonculturable after approximately 62 and 45 d...

  3. Construction of a system using a deep learning algorithm to count cell numbers in nanoliter wells for viable single-cell experiments.

    Science.gov (United States)

    Kamatani, Takashi; Fukunaga, Koichi; Miyata, Kaede; Shirasaki, Yoshitaka; Tanaka, Junji; Baba, Rie; Matsusaka, Masako; Kamatani, Naoyuki; Moro, Kazuyo; Betsuyaku, Tomoko; Uemura, Sotaro

    2017-12-04

    For single-cell experiments, it is important to accurately count the number of viable cells in a nanoliter well. We used a deep learning-based convolutional neural network (CNN) on a large amount of digital data obtained as microscopic images. The training set consisted of 103 019 samples, each representing a microscopic grayscale image. After extensive training, the CNN was able to classify the samples into four categories, i.e., 0, 1, 2, and more than 2 cells per well, with an accuracy of 98.3% when compared to determination by two trained technicians. By analyzing the samples for which judgments were discordant, we found that the judgment by technicians was relatively correct although cell counting was often difficult by the images of discordant samples. Based on the results, the system was further enhanced by introducing a new algorithm in which the highest outputs from CNN were used, increasing the accuracy to higher than 99%. Our system was able to classify the data even from wells with a different shape. No other tested machine learning algorithm showed a performance higher than that of our system. The presented CNN system is expected to be useful for various single-cell experiments, and for high-throughput and high-content screening.

  4. The Conceptual Mechanism for Viable Organizational Learning Based on Complex System Theory and the Viable System Model

    Science.gov (United States)

    Sung, Dia; You, Yeongmahn; Song, Ji Hoon

    2008-01-01

    The purpose of this research is to explore the possibility of viable learning organizations based on identifying viable organizational learning mechanisms. Two theoretical foundations, complex system theory and viable system theory, have been integrated to provide the rationale for building the sustainable organizational learning mechanism. The…

  5. Fate of viable but non-culturable Listeria monocytogenes in pig manure microcosms

    Directory of Open Access Journals (Sweden)

    Jeremy eDesneux

    2016-03-01

    Full Text Available The fate of two strains of L. monocytogenes and their ability to become viable but non-culturable (VBNC was investigated in microcosms containing piggery effluents (two raw manures and two biologically treated manures stored for two months at 8°C and 20°C. Levels of L. monocytogenes were estimated using the culture method, qPCR, and propidium monoazide treatment combined with qPCR (qPCRPMA. The chemical composition and the microbial community structure of the manures were also analysed. The strains showed similar decline rates and persisted up to 63 days. At day zero, the percentage of VBNC cells among viable cells was higher in raw manures (81.5-94.8% than in treated manures (67.8-79.2%. The changes in their proportion over time depended on the temperature and on the type of effluent: the biggest increase was observed in treated manures at 20°C and the smallest increase in raw manures at 8°C. The chemical parameters had no influence on the behaviour of the strains, but decrease of the persistence of viable cells was associated with an increase in the microbial richness of the manures. This study demonstrated that storing manure altered the culturability of L. monocytogenes, which rapidly entered the VBNC state, and underlines the importance of including VBNC cells when estimating the persistence of the pathogens in farm effluents.

  6. Polymerase chain reaction-based discrimination of viable from non-viable Mycoplasma gallisepticum

    Directory of Open Access Journals (Sweden)

    Ching Giap Tan

    2014-09-01

    Full Text Available The present study was based on the reverse transcription polymerase chain reaction (RT-PCR of the 16S ribosomal nucleic acid (rRNA of Mycoplasma for detection of viable Mycoplasma gallisepticum. To determine the stability of M. gallisepticum 16S rRNA in vitro, three inactivation methods were used and the suspensions were stored at different temperatures. The 16S rRNA of M. gallisepticum was detected up to approximately 20–25 h at 37 °C, 22–25 h at 16 °C, and 23–27 h at 4 °C. The test, therefore, could detect viable or recently dead M. gallisepticum (< 20 h. The RT-PCR method was applied during an in vivo study of drug efficacy under experimental conditions, where commercial broiler-breeder eggs were inoculated with M. gallisepticum into the yolk. Hatched chicks that had been inoculated in ovo were treated with Macrolide 1. The method was then applied in a flock of day 0 chicks with naturally acquired vertical transmission of M. gallisepticum, treated with Macrolide 2. Swabs of the respiratory tract were obtained for PCR and RT-PCR evaluations to determine the viability of M. gallisepticum. This study proved that the combination of both PCR and RT-PCR enables detection and differentiation of viable from non-viable M. gallisepticum.

  7. Monitoring viable cells of the biological control agent Lactobacillus plantarum PM411 in aerial plant surfaces by means of a strain-specific viability quantitative PCR.

    Science.gov (United States)

    Daranas, Núria; Bonaterra, Anna; Francés, Jesús; Cabrefiga, Jordi; Montesinos, Emilio; Badosa, Esther

    2018-03-09

    A viability qPCR (v-qPCR) assay was developed for the unambiguous detection and quantification of Lactobacillus plantarum PM411 viable cells in aerial plant surfaces. A 972 bp region of a PM411 predicted prophage with mosaic architecture enabled the identification of a PM411 strain-specific molecular marker. Three primer sets, with different amplicon lengths (92, 188, and 317 bp), and one TaqMan probe were designed. All the qPCR assays showed good linearity over a 4-log range and good efficiencies, but differed in sensitivity. The nucleic acid-binding dye PEMAX was used for selectively detecting and enumerating viable bacteria by v-qPCR. The primer set amplifying a 188 bp DNA fragment was selected as the most suitable for v-qPCR. The performance of the method was assessed on apple blossoms, pear, strawberry and kiwifruit leaves in potted plants under controlled environmental conditions, and pear and apple blossoms under field conditions, by comparing v-qPCR population estimation to those obtained by qPCR and specific plate counting on MRS-rifampicin. The population estimation did not differ significantly between methods when conditions were conducive to bacterial survival. However, under stressful conditions, differences between methods were observed due to cell death or viable but non-culturable state induction. While qPCR overestimated the population level, plate counting underestimated this value in comparison to v-qPCR. PM411 attained stable population levels of viable cells on flower environment under high relative humidity. However, the unfavourable conditions onto the leaf surface and the relatively dryness in the field caused an important decrease of viable population. IMPORTANCE The v-qPCR method in combination with plate counting and qPCR is a powerful tool for studies of colonization and survival in field conditions, to improve formulations and delivery strategies of PM411, or to optimize the dose and timing of spray schedules. It is expected that PEMAX

  8. Evaluation of PCR and DNA hybridization protocols for detection of viable enterotoxigenic Clostridium perfringens in irradiated beef

    International Nuclear Information System (INIS)

    Baez, L.A.; Juneja, V.K.; Thayer, D.W.; Sackitey, S.

    1997-01-01

    The sensitivity of DNA hybridization and polymerase chain reaction (PCR), was evaluated in irradiated cooked and raw beef samples. A membrane-based colony hybridization assay and a PCR protocol, both with specificity for the enterotoxin A gene of Clostridium perfringens, were compared with viable plate counts. The results of the colony hybridization procedure were in agreement with viable plate counts for detection and enumeration of enterotoxigenic C. perfringens. The PCR procedure combined a 4 h enrichment followed by a nucleic acid extraction step and assessed the amplification of 183 and 750 base pair enterotoxin gene targets. Detection of C. perfringens by PCR did not show a reliable correlation with viable plate counts or the colony hybridization assay. C. perfringens killed by irradiation were not detected by the plate count or colony hybridization methods; however, killed cells were detected with the PCR technique. By relying on the growth of viable cells for detection and/or enumeration, the colony hybridization and plate count methods provided a direct correlation with the presence of viable bacteria

  9. Interrelationships of somatic cell count, mastitis, and milk yield in a low somatic cell count herd.

    Science.gov (United States)

    Deluyker, H A; Gay, J M; Weaver, L D

    1993-11-01

    In a high yielding low SCC herd, changes in milk yield associated with SCC and occurrence of clinical mastitis and differences in SCC with parity, clinical mastitis, and DIM were investigated. Milk yield data were obtained at every milking, and SCC was measured once every 48 h in 117 cows during the first 119 d postpartum. Effects of SCC and clinical mastitis on cumulative milk yield in the first 119 d postpartum were evaluated with least squares linear regression. Repeated measures ANOVA was used to detect changes in SCC. The SCC was highest at lactation onset, and cows with clinical mastitis had significantly higher SCC. During the 10 d prior to onset of clinical mastitis, SCC was higher in affected cows than in matched unaffected controls and surged just prior to diagnosis. During the 10-d period following a mastitis treatment, SCC differences between treated and control cows remained significant but became smaller with time and returned to the premastitis differences. Occurrence of clinical mastitis was associated with 5% milk yield loss. Cows with mean SCC > 245,000 cells/ml over the 119 d showed 6.2% yield loss compared with cows with SCC 245,000 cells/ml) because a greater percentage of cows (26%) had clinical mastitis than elevated SCC (12.5%).

  10. Spatiotemporal Dynamics of Total Viable Vibrio spp. in a NW Mediterranean Coastal Area.

    Science.gov (United States)

    Girard, Léa; Peuchet, Sébastien; Servais, Pierre; Henry, Annabelle; Charni-Ben-Tabassi, Nadine; Baudart, Julia

    2017-09-27

    A cellular approach combining Direct Viable Counting and Fluorescent In Situ Hybridization using a one-step multiple-probe technique and Solid Phase Cytometry (DVC-FISH-SPC) was developed to monitor total viable vibrios and cover the detection of a large diversity of vibrios. FISH combined three probes in the same assay and targeted sequences located at different positions on the 16S rRNA of Vibrio and Aliivibrio members. We performed a 10-month in situ study to investigate the weekly dynamics of viable vibrios relative to culturable counts at two northwestern Mediterranean coastal sites, and identified the key physicochemical factors for their occurrence in water using a multivariate analysis. Total viable and culturable cell counts showed the same temporal pattern during the warmer season, whereas the ratios between both methods were inverted during the colder seasons (<15°C), indicating that some of the vibrio community had entered into a viable but non-culturable (VBNC) state. We confirmed that Seawater Surface Temperature explained 51-62% of the total variance in culturable counts, and also showed that the occurrence of viable vibrios is controlled by two variables, pheopigment (15%) and phosphate (12%) concentrations, suggesting that other unidentified factors play a role in maintaining viability.

  11. Contrast-Enhanced Ultrasound Improves the Pathological Outcomes of US-Guided Core Needle Biopsy That Targets the Viable Area of Anterior Mediastinal Masses

    Directory of Open Access Journals (Sweden)

    Jian-hua Zhou

    2018-01-01

    Full Text Available Based on the option that ultrasound-guided core needle biopsy (US-CNB of the enhanced portion of anterior mediastinal masses (AMMs identified by contrast-enhanced ultrasound (CEUS would harvest viable tissue and benefit the histological diagnoses, a retrospective study was performed to elucidate the correlation between the prebiopsy CEUS and diagnostic yield of AMMs and found that CEUS potentially improved the diagnostic yield of AMMs compared with conventional US with a significant increase in the cellularity of samples. Furthermore, the marginal blood flow signals and absence of necrosis can predict the diagnostic yield of AMM. It was concluded that US-CNB of the viable part of AMMs, as verified by CEUS, was able to harvest sufficient tissue with more cellularity that could be used for ancillary studies and improve the diagnostic yield. And CEUS was recommended to those patients with AMMs undergoing repeated US-CNB, with the absence of marginal blood signals or presence of necrosis.

  12. Cellular bone matrices: viable stem cell-containing bone graft substitutes.

    Science.gov (United States)

    Skovrlj, Branko; Guzman, Javier Z; Al Maaieh, Motasem; Cho, Samuel K; Iatridis, James C; Qureshi, Sheeraz A

    2014-11-01

    Advances in the field of stem cell technology have stimulated the development and increased use of allogenic bone grafts containing live mesenchymal stem cells (MSCs), also known as cellular bone matrices (CBMs). It is estimated that CBMs comprise greater than 17% of all bone grafts and bone graft substitutes used. To critically evaluate CBMs, specifically their technical specifications, existing published data supporting their use, US Food and Drug Administration (FDA) regulation, cost, potential pitfalls, and other aspects pertaining to their use. Areview of literature. A series of Ovid, Medline, and Pubmed-National Library of Medicine/National Institutes of Health (www.ncbi.nlm.nih.gov) searches were performed. Only articles in English journals or published with English language translations were included. Level of evidence of the selected articles was assessed. Specific technical information on each CBM was obtained by direct communication from the companies marketing the individual products. Five different CBMs are currently available for use in spinal fusion surgery. There is a wide variation between the products with regard to the average donor age at harvest, total cellular concentration, percentage of MSCs, shelf life, and cell viability after defrosting. Three retrospective studies evaluating CBMs and fusion have shown fusion rates ranging from 90.2% to 92.3%, and multiple industry-sponsored trials are underway. No independent studies evaluating spinal fusion rates with the use of CBMs exist. All the commercially available CBMs claim to meet the FDA criteria under Section 361, 21 CFR Part 1271, and are not undergoing FDA premarket review. The CBMs claim to provide viable MSCs and are offered at a premium cost. Numerous challenges exist in regard to MSCs' survival, function, osteoblastic potential, and cytokine production once implanted into the intended host. Cellular bone matrices may be a promising bone augmentation technology in spinal fusion surgery

  13. Quantification of viable bacteria in wastewater treatment plants by using propidium monoazide combined with quantitative PCR (PMA-qPCR).

    Science.gov (United States)

    Li, Dan; Tong, Tiezheng; Zeng, Siyu; Lin, Yiwen; Wu, Shuxu; He, Miao

    2014-02-01

    The detection of viable bacteria in wastewater treatment plants (WWTPs) is very important for public health, as WWTPs are a medium with a high potential for waterborne disease transmission. The aim of this study was to use propidium monoazide (PMA) combined with the quantitative polymerase chain reaction (PMA-qPCR) to selectively detect and quantify viable bacteria cells in full-scale WWTPs in China. PMA was added to the concentrated WWTP samples at a final concentration of 100 micromol/L and the samples were incubated in the dark for 5 min, and then lighted for 4 min prior to DNA extraction and qPCR with specific primers for Escherichia coli and Enterococci, respectively. The results showed that PMA treatment removed more than 99% of DNA from non-viable cells in all the WWTP samples, while matrices in sludge samples markedly reduced the effectiveness of PMA treatment. Compared to qPCR, PMA-qPCR results were similar and highly linearly correlated to those obtained by culture assay, indicating that DNA from non-viable cells present in WWTP samples can be eliminated by PMA treatment, and that PMA-qPCR is a reliable method for detection of viable bacteria in environmental samples. This study demonstrated that PMA-qPCR is a rapid and selective detection method for viable bacteria in WWTP samples, and that WWTPs have an obvious function in removing both viable and non-viable bacteria. The results proved that PMA-qPCR is a promising detection method that has a high potential for application as a complementary method to the standard culture-based method in the future.

  14. Ethanol yields and cell wall properties in divergently bred switchgrass genotypes

    Science.gov (United States)

    Genetic modification of herbaceous plant cell walls to increase biofuels yields from harvested biomass is a primary bioenergy research goal. The focus of much of this research has been on cell wall lignin concentration. Using switchgrass genotypes developed by divergent breeding for ruminant diges...

  15. Supercooling as a viable non-freezing cell preservation method of rat hepatocytes.

    Directory of Open Access Journals (Sweden)

    O Berk Usta

    Full Text Available Supercooling preservation holds the potential to drastically extend the preservation time of organs, tissues and engineered tissue products, and fragile cell types that do not lend themselves well to cryopreservation or vitrification. Here, we investigate the effects of supercooling preservation (SCP at -4(oC on primary rat hepatocytes stored in cryovials and compare its success (high viability and good functional characteristics to that of static cold storage (CS at +4(oC and cryopreservation. We consider two prominent preservation solutions a Hypothermosol (HTS-FRS and b University of Wisconsin solution (UW and a range of preservation temperatures (-4 to -10 (oC. We find that there exists an optimum temperature (-4(oC for SCP of rat hepatocytes which yields the highest viability; at this temperature HTS-FRS significantly outperforms UW solution in terms of viability and functional characteristics (secretions and enzymatic activity in suspension and plate culture. With the HTS-FRS solution we show that the cells can be stored for up to a week with high viability (~56%; moreover we also show that the preservation can be performed in large batches (50 million cells with equal or better viability and no loss of functionality as compared to smaller batches (1.5 million cells performed in cryovials.

  16. Viable group A streptococci in macrophages during acute soft tissue infection.

    Directory of Open Access Journals (Sweden)

    Pontus Thulin

    2006-03-01

    Full Text Available Group A streptococcal severe soft tissue infections, such as necrotizing fasciitis, are rapidly progressive infections associated with high mortality. Group A streptococcus is typically considered an extracellular pathogen, but has been shown to reside intracellularly in host cells.We characterized in vivo interactions between group A streptococci (GAS and cells involved in innate immune responses, using human biopsies (n = 70 collected from 17 patients with soft tissue infections. Immunostaining and in situ image analysis revealed high amounts of bacteria in the biopsies, even in those collected after prolonged antibiotic therapy. Viability of the streptococci was assessed by use of a bacterial viability stain, which demonstrated viable bacteria in 74% of the biopsies. GAS were present both extracellularly and intracellularly within phagocytic cells, primarily within macrophages. Intracellular GAS were predominantly noted in biopsies from newly involved tissue characterized by lower inflammation and bacterial load, whereas purely extracellular GAS or a combination of intra- and extracellular GAS dominated in severely inflamed tissue. The latter tissue was also associated with a significantly increased amount of the cysteine protease streptococcal pyrogenic exotoxin SpeB. In vitro studies confirmed that macrophages serve as reservoirs for viable GAS, and infection with a speB-deletion mutant produced significantly lower frequencies of cells with viable GAS following infection as compared to the wild-type bacteria.This is the first study to demonstrate that GAS survive intracellularly in macrophages during acute invasive infections. This intracellular presence may have evolved as a mechanism to avoid antibiotic eradication, which may explain our finding that high bacterial load is present even in tissue collected after prolonged intravenous antibiotic therapy. This new insight into the pathogenesis of streptococcal soft tissue infections

  17. Viable Group A Streptococci in Macrophages during Acute Soft Tissue Infection.

    Directory of Open Access Journals (Sweden)

    2006-01-01

    Full Text Available BACKGROUND: Group A streptococcal severe soft tissue infections, such as necrotizing fasciitis, are rapidly progressive infections associated with high mortality. Group A streptococcus is typically considered an extracellular pathogen, but has been shown to reside intracellularly in host cells. METHODS AND FINDINGS: We characterized in vivo interactions between group A streptococci (GAS and cells involved in innate immune responses, using human biopsies (n = 70 collected from 17 patients with soft tissue infections. Immunostaining and in situ image analysis revealed high amounts of bacteria in the biopsies, even in those collected after prolonged antibiotic therapy. Viability of the streptococci was assessed by use of a bacterial viability stain, which demonstrated viable bacteria in 74% of the biopsies. GAS were present both extracellularly and intracellularly within phagocytic cells, primarily within macrophages. Intracellular GAS were predominantly noted in biopsies from newly involved tissue characterized by lower inflammation and bacterial load, whereas purely extracellular GAS or a combination of intra- and extracellular GAS dominated in severely inflamed tissue. The latter tissue was also associated with a significantly increased amount of the cysteine protease streptococcal pyrogenic exotoxin SpeB. In vitro studies confirmed that macrophages serve as reservoirs for viable GAS, and infection with a speB-deletion mutant produced significantly lower frequencies of cells with viable GAS following infection as compared to the wild-type bacteria. CONCLUSIONS: This is the first study to demonstrate that GAS survive intracellularly in macrophages during acute invasive infections. This intracellular presence may have evolved as a mechanism to avoid antibiotic eradication, which may explain our finding that high bacterial load is present even in tissue collected after prolonged intravenous antibiotic therapy. This new insight into the pathogenesis

  18. Polyelectrolyte Complex Beads by Novel Two-Step Process for Improved Performance of Viable Whole-Cell Baeyer-Villiger Monoxygenase by Immobilization

    Czech Academy of Sciences Publication Activity Database

    Krajčovič, T.; Bučko, M.; Vikartovská, A.; Lacík, I.; Uhelská, L.; Chorvát, D.; Neděla, Vilém; Tihlaříková, Eva; Gericke, M.; Heinze, T.; Gemeiner, P.

    2017-01-01

    Roč. 7, č. 11 (2017), s. 353-364 ISSN 2073-4344 Institutional support: RVO:68081731 Keywords : polyelectrolyte complex beads * environmental scanning electron microscopy * confocal laser scanning microscopy * Baeyer-Villiger biooxidation * cyclohexanone monoxygenase * immobilization * viable whole-cell biocatalyst Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering OBOR OECD: Bioprocessing technologies (industrial processes relying on biological agents to drive the process) biocatalysis, fermentation Impact factor: 3.082, year: 2016 http://www.mdpi.com/2073-4344/7/11/353

  19. Detection of viable Escherichia coli O157:H7 in ground beef by propidium monoazide real-time PCR.

    Science.gov (United States)

    Liu, Yarui; Mustapha, Azlin

    2014-01-17

    Escherichia coli O157:H7 associated with food has caused many serious public health problems in recent years. However, only viable cells of this pathogen can cause infections, and false-positive detection caused by dead cells can lead to unnecessary product recalls. The objective of this study was to develop and optimize a method that combines propidium monoazide (PMA) staining with real-time PCR to detect only viable cells of E. coli O157:H7 in ground beef. PMA is a DNA intercalating dye that can penetrate compromised membranes of dead cells and bind to cellular DNA, preventing its amplification via a subsequent PCR. Three strains of E. coli O157:H7 (505B, G5310 and C7927) at concentrations of 10(0) to 10(8)CFU/mL were used as live cells. Dead cells were obtained by heating cell suspensions at 85°C for 15 min. Suspensions were treated with PMA and the optimized assay was applied to artificially contaminated ground beef with two different fat contents (10% and 27%). DNA was extracted and amplified by TaqMan® real-time PCR assay targeting the uidA gene for detection of E. coli O157:H7. Plasmid pUC19 was added as an internal amplification control (IAC). A treatment of 25 μM PMA with a 10-min light exposure on ice was sufficient to eliminate DNA from 10(8) dead E. coli O157:H7 cells/mL. The optimized assay could detect as low as 10(2) CFU/mL viable E. coli O157:H7 in pure culture and 10(5) CFU/g in ground beef, in the presence of 10(6)/mL or g of dead cells. With an 8-h enrichment, 1 CFU/g viable E. coli O157:H7 in ground beef was detectable without interference from 10(6) dead cells/g. In conclusion, the PMA real-time PCR could effectively detect viable E. coli O157:H7 without being compromised by dead cells. Copyright © 2013 Elsevier B.V. All rights reserved.

  20. Viable bacteria associated with red blood cells and plasma in freshly drawn blood donations.

    Science.gov (United States)

    Damgaard, Christian; Magnussen, Karin; Enevold, Christian; Nilsson, Martin; Tolker-Nielsen, Tim; Holmstrup, Palle; Nielsen, Claus Henrik

    2015-01-01

    Infection remains a leading cause of post-transfusion mortality and morbidity. Bacterial contamination is, however, detected in less than 0.1% of blood units tested. The aim of the study was to identify viable bacteria in standard blood-pack units, with particular focus on bacteria from the oral cavity, and to determine the distribution of bacteria revealed in plasma and in the red blood cell (RBC)-fraction. Cross-sectional study. Blood were separated into plasma and RBC-suspensions, which were incubated anaerobically or aerobically for 7 days on trypticase soy blood agar (TSA) or blue lactose plates. For identification colony PCR was performed using primers targeting 16S rDNA. Blood donors attending Capital Region Blood Bank, Copenhagen University Hospital, Rigshospitalet, Hvidovre, Denmark, October 29th to December 10th 2013. 60 donors (≥50 years old), self-reported medically healthy. Bacterial growth was observed on plates inoculated with plasma or RBCs from 62% of the blood donations. Growth was evident in 21 (35%) of 60 RBC-fractions and in 32 (53%) of 60 plasma-fractions versus 8 of 60 negative controls (p = 0.005 and p = 2.6x10-6, respectively). Propionibacterium acnes was found in 23% of the donations, and Staphylococcus epidermidis in 38%. The majority of bacteria identified in the present study were either facultative anaerobic (59.5%) or anaerobic (27.8%) species, which are not likely to be detected during current routine screening. Viable bacteria are present in blood from donors self-reported as medically healthy, indicating that conventional test systems employed by blood banks insufficiently detect bacteria in plasma. Further investigation is needed to determine whether routine testing for anaerobic bacteria and testing of RBC-fractions for adherent bacteria should be recommended.

  1. High yield cell-free production of integral membrane proteins without refolding or detergents.

    Science.gov (United States)

    Wuu, Jessica J; Swartz, James R

    2008-05-01

    Integral membrane proteins act as critical cellular components and are important drug targets. However, difficulties in producing membrane proteins have hampered investigations of structure and function. In vivo production systems are often limited by cell toxicity, and previous in vitro approaches have required unnatural folding pathways using detergents or lipid solutions. To overcome these limitations, we present an improved cell-free expression system which produces high yields of integral membrane proteins without the use of detergents or refolding steps. Our cell-free reaction activates an Escherichia coli-derived cell extract for transcription and translation. Purified E. coli inner membrane vesicles supply membrane-bound components and the lipid environment required for insertion and folding. Using this system, we demonstrated successful synthesis of two complex integral membrane transporters, the tetracycline pump (TetA) and mannitol permease (MtlA), in yields of 570+/-50 microg/mL and 130+/-30 microg/mL of vesicle-associated protein, respectively. These yields are up to 400 times typical in vivo concentrations. Insertion and folding of these proteins are verified by sucrose flotation, protease digestion, and activity assays. Whereas TetA incorporates efficiently into vesicle membranes with over two-thirds of the synthesized protein being inserted, MtlA yields appear to be limited by insufficient concentrations of a membrane-associated chaperone.

  2. Three-dimensional spheroid culture of human umbilical cord mesenchymal stem cells promotes cell yield and stemness maintenance.

    Science.gov (United States)

    Li, Yi; Guo, Gang; Li, Li; Chen, Fei; Bao, Ji; Shi, Yu-Jun; Bu, Hong

    2015-05-01

    Mesenchymal stem cell (MSC) transplantation is a promising treatment of many diseases. However, conventional techniques with cells being cultured as a monolayer result in slow cell proliferation and insufficient yield to meet clinical demands. Three-dimensional (3D) culture systems are gaining attention with regard to recreating a complex microenvironment and to understanding the conditions experienced by cells. Our aim is to establish a novel 3D system for the culture of human umbilical cord MSCs (hUC-MSCs) within a real 3D microenvironment but with no digestion or passaging. Primary hUC-MSCs were isolated and grown in serum-free medium (SFM) on a suspension Rocker system. Cell characteristics including proliferation, phenotype and multipotency were recorded. The therapeutic effects of 3D-cultured hUC-MSCs on carbon tetrachloride (CCl4)-induced acute liver failure in mouse models were examined. In the 3D Rocker system, hUC-MSCs formed spheroids in SFM and maintained high viability and active proliferation. Compared with monolayer culture, the 3D-culture system yielded more hUC-MSCs cells within the same volume. The spheroids expressed higher levels of stem cell markers and displayed stronger multipotency. After transplantation into mouse, 3D hUC-MSCs significantly promoted the secretion of interferon-γ and interleukin-6 but inhibited that of tumor necrosis factor-α, thereby alleviating liver necrosis and promoting regeneration following CCl4 injury. The 3D culture of hUC-MSCs thus promotes cell yield and stemness maintenance and represents a promising strategy for hUC-MSCs expansion on an industrial scale with great potential for cell therapy and biotechnology.

  3. Droplet digital PCR improves absolute quantification of viable lactic acid bacteria in faecal samples.

    Science.gov (United States)

    Gobert, Guillaume; Cotillard, Aurélie; Fourmestraux, Candice; Pruvost, Laurence; Miguet, Jean; Boyer, Mickaël

    2018-03-14

    Analysing correlations between the observed health effects of ingested probiotics and their survival in digestive tract allows adapting their preparations for food. Tracking ingested probiotic in faecal samples requires accurate and specific tools to quantify live vs dead cells at strain level. Traditional culture-based methods are simpler to use but they do not allow quantifying viable but non-cultivable (VBNC) cells and they are poorly discriminant below the species level. We have set up a viable PCR (vPCR) assay combining propidium monoazide (PMA) treatment and either real time quantitative PCR (qPCR) or droplet digital PCR (ddPCR) to quantify a Lactobacillus rhamnosus and two Lactobacillus paracasei subsp. paracasei strains in piglet faeces. Adjustments of the PMA treatment conditions and reduction of the faecal sample size were necessary to obtain accurate discrimination between dead and live cells. The study also revealed differences of PMA efficiency among the two L. paracasei strains. Both PCR methods were able to specifically quantify each strain and provided comparable total bacterial counts. However, quantification of lower numbers of viable cells was best achieved with ddPCR, which was characterized by a reduced lower limit of quantification (improvement of up to 1.76 log 10 compared to qPCR). All three strains were able to survive in the piglets' gut with viability losses between 0.78 and 1.59 log 10 /g faeces. This study shows the applicability of PMA-ddPCR to specific quantification of small numbers of viable bacterial cells in the presence of an important background of unwanted microorganisms, and without the need to set up standard curves. It also illustrates the need to adapt PMA protocols according to the final matrix and target strain, even for closely related strains. The PMA-ddPCR approach provides a new tool to quantify bacterial survival in faecal samples from a preclinical and clinical trial. Copyright © 2018 The Authors. Published by

  4. Does enterohemorrhagic Escherichia coli O157:H7 enter the viable but nonculturable state in salted salmon roe?

    Science.gov (United States)

    Makino, S I; Kii, T; Asakura, H; Shirahata, T; Ikeda, T; Takeshi, K; Itoh, K

    2000-12-01

    An outbreak caused by salted salmon roe contaminated with enterohemorrhagic Escherichia coli O157 occurred in Japan in 1998. Since about 0.75 to 1.5 viable cells were estimated to cause infection, we presumed that O157 might enter the viable but nonculturable (VNC) state in salted salmon roe and consequently that viable cell numbers might be underestimated. Although patient-originating O157 cells could not grow on agar plates after 72 h of incubation in 13% NaCl, they were resuscitated in yeast extract broth, and more than 90% of the cells were shown to be viable by fluorescent staining, suggesting that almost all of them could enter the VNC state in NaCl water. Roe-originating O157 was resistant to NaCl because it could grow on agar after 72 h of incubation in NaCl water, but about 20% of cells appeared to enter the VNC state. Therefore, germfree mice were infected with O157 to examine the resuscitation of cells in the VNC state and the retention of pathogenicity. O157 that originated in roe, but not patients, killed mice and was isolated from the intestine. However, these isolates had become sensitive to NaCl. O157 cells of roe origin incubated in normal media also killed mice and were isolated from the intestine, but they also became transiently NaCl sensitive. We therefore propose that bacterial cells might enter the VNC state under conditions of stress, such as those encountered in vivo or in high salt concentrations, and then revive when those conditions have eased. If so, the VNC state in food is potentially dangerous from a public health viewpoint and may have to be considered at the time of food inspection. Finally, the establishment of a simple recovery system for VNC cells should be established.

  5. Does the preference of peripheral versus central venous access in peripheral blood stem cell collection/yield change stem cell kinetics in autologous stem cell transplantation?

    Science.gov (United States)

    Dogu, Mehmet Hilmi; Kaya, Ali Hakan; Berber, Ilhami; Sari, İsmail; Tekgündüz, Emre; Erkurt, Mehmet Ali; Iskender, Dicle; Kayıkçı, Ömur; Kuku, Irfan; Kaya, Emin; Keskin, Ali; Altuntaş, Fevzi

    2016-02-01

    Central venous access is often used during apheresis procedure in stem cell collection. The aim of the present study was to evaluate whether central or peripheral venous access has an effect on stem cell yield and the kinetics of the procedure and the product in patients undergoing ASCT after high dose therapy. A total of 327 patients were retrospectively reviewed. The use of peripheral venous access for stem cell yield was significantly more frequent in males compared to females (p = 0.005). Total volume of the product was significantly lower in central venous access group (p = 0.046). As being a less invasive procedure, peripheral venous access can be used for stem cell yield in eligible selected patients. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Impact of endoscopic stent insertion on detection of viable circulating tumor cells from obstructive colorectal cancer.

    Science.gov (United States)

    Yamashita, Shinya; Tanemura, Masahiro; Sawada, Genta; Moon, Jeongho; Shimizu, Yosuke; Yamaguchi, Toshiki; Kuwai, Toshio; Urata, Yasuo; Kuraoka, Kazuya; Hatanaka, Nobutaka; Yamashita, Yoshinori; Taniyama, Kiyomi

    2018-01-01

    The placement of a self-expanding metallic stent (SEMS) in obstructive colorectal cancer (OCRC) is acknowledged to be a safe and effective procedure for the relief of obstruction. However, there is concern that shear forces acting on the tumor during stent expansion may release cancer cells into the circulation, resulting in a poor prognosis. The aim of the present study was to determine whether colonic stent insertion increases viable circulating tumor cells (v-CTCs). A telomerase-specific replication-selective adenovirus-expressing GFP (TelomeScanF35) detection system was used to detect v-CTCs in 8 OCRC patients with a SEMS before and after stent insertion and after surgical resection. In 7 patients, a SEMS was inserted as a bridge to surgery (BTS), and in one patient, a SEMS was inserted for palliation. Surgical resection (R0) was performed in 7 patients. Four patients had no v-CTCs before SEMS placement, two of four measurable patients had an increased number of v-CTCs after SEMS placement (1-3 v-CTCs), and one of two patients with increased v-CTCs developed distant lymphatic metastasis despite curative resection. Four patients had v-CTCs (1-19 cells) before SEMS placement, and two of these four patients had an increase in the number of v-CTCs (20-21 cells) after SEMS placement, while one of the four patients died early with distant metastasis. The present study demonstrated that endoscopic stent insertion for OCRC may result in tumor cell dissemination into the peripheral circulation and may induce distant metastases.

  7. 14C leucine chloromethylketone interaction with sarcoma 37 cell plasma membrane components

    International Nuclear Information System (INIS)

    Matthews, R.H.; Milo, G.E.; McMichael, T.L.; Lewis, N.J.

    1982-01-01

    Leucine chloromethylketone labelling of viable S37 cells was preferential for the plasma membrane fraction. The pattern of radiolabelling of the plasma membrane proteins was time-dependent. After 5 min the radiolabel was localized with glutamyl transpeptidase, and subsequently, with other physiologically active proteins as a function of time after incubation. Labelling of proteins was temperature-dependent and incubation of viable S37 cells with the radiolabelled substrate at 0 0 C yielded little or no radioactivity localized in the plasma membrane. The molecular weight of one radiolabelled substratemembrane protein complex was estimated on sodium dodecyl sulfate polyacrylamide gel electrophoresis to be between 100,000-200,000. (author)

  8. Microbial Electrolysis Cells for High Yield Hydrogen Gas Production from Organic Matter

    KAUST Repository

    Logan, Bruce E.

    2008-12-01

    The use of electrochemically active bacteria to break down organic matter, combined with the addition of a small voltage (>0.2 V in practice) in specially designed microbial electrolysis cells (MECs), can result in a high yield of hydrogen gas. While microbial electrolysis was invented only a few years ago, rapid developments have led to hydrogen yields approaching 100%, energy yields based on electrical energy input many times greater than that possible by water electrolysis, and increased gas production rates. MECs used to make hydrogen gas are similar in design to microbial fuel cells (MFCs) that produce electricity, but there are important differences in architecture and analytical methods used to evaluate performance. We review here the materials, architectures, performance, and energy efficiencies of these MEC systems that show promise as a method for renewable and sustainable energy production, and wastewater treatment. © 2008 American Chemical Society.

  9. Relative ion yields in mammalian cell components using C60 SIMS

    Science.gov (United States)

    Keskin, Selda; Piwowar, Alan; Hue, Jonathan; Shen, Kan; Winograd, Nicholas

    2013-01-01

    Time of flight secondary ion mass spectrometry has been used to better understand the influence of molecular environment on the relative ion yields of membrane lipid molecules found in high abundance in a model mammalian cell line, RAW264.7. Control lipid mixtures were prepared to simulate lipid–lipid interactions in the inner and outer leaflet of cell membranes. Compared with its pure film, the molecular ion yields of 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine are suppressed when mixed with 2-dipalmitoyl-sn-glycero-3-phosphocholine. In the mixture, proton competition between 1,2-dioleoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, and 2-dipalmitoyl-sn-glycero-3-phosphocholine led to lower ionization efficiency. The possible mechanism for ion suppression was also investigated with 1H and 13C nuclear magnetic resonance spectroscopy. The formation of a hydroxyl bond in lipid mixtures confirms the mechanism involving proton exchange with the surrounding environment. Similar effects were observed for lipid mixtures mimicking the composition of the inner leaflet of cell membranes. The secondary molecular ion yield of 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-L-serine was observed to be enhanced in the presence of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine. PMID:25140069

  10. A propidium monoazide–quantitative PCR method for the detection and quantification of viable Enterococcus faecalis in large-volume samples of marine waters

    KAUST Repository

    Salam, Khaled W.; El-Fadel, Mutasem E.; Barbour, Elie K.; Saikaly, Pascal

    2014-01-01

    The development of rapid detection assays of cell viability is essential for monitoring the microbiological quality of water systems. Coupling propidium monoazide with quantitative PCR (PMA-qPCR) has been successfully applied in different studies for the detection and quantification of viable cells in small-volume samples (0.25-1.00 mL), but it has not been evaluated sufficiently in marine environments or in large-volume samples. In this study, we successfully integrated blue light-emitting diodes for photoactivating PMA and membrane filtration into the PMA-qPCR assay for the rapid detection and quantification of viable Enterococcus faecalis cells in 10-mL samples of marine waters. The assay was optimized in phosphate-buffered saline and seawater, reducing the qPCR signal of heat-killed E. faecalis cells by 4 log10 and 3 log10 units, respectively. Results suggest that high total dissolved solid concentration (32 g/L) in seawater can reduce PMA activity. Optimal PMA-qPCR standard curves with a 6-log dynamic range and detection limit of 102 cells/mL were generated for quantifying viable E. faecalis cells in marine waters. The developed assay was compared with the standard membrane filter (MF) method by quantifying viable E. faecalis cells in seawater samples exposed to solar radiation. The results of the developed PMA-qPCR assay did not match that of the standard MF method. This difference in the results reflects the different physiological states of E. faecalis cells in seawater. In conclusion, the developed assay is a rapid (∼5 h) method for the quantification of viable E. faecalis cells in marine recreational waters, which should be further improved and tested in different seawater settings. © 2014 Springer-Verlag Berlin Heidelberg.

  11. A propidium monoazide–quantitative PCR method for the detection and quantification of viable Enterococcus faecalis in large-volume samples of marine waters

    KAUST Repository

    Salam, Khaled W.

    2014-08-23

    The development of rapid detection assays of cell viability is essential for monitoring the microbiological quality of water systems. Coupling propidium monoazide with quantitative PCR (PMA-qPCR) has been successfully applied in different studies for the detection and quantification of viable cells in small-volume samples (0.25-1.00 mL), but it has not been evaluated sufficiently in marine environments or in large-volume samples. In this study, we successfully integrated blue light-emitting diodes for photoactivating PMA and membrane filtration into the PMA-qPCR assay for the rapid detection and quantification of viable Enterococcus faecalis cells in 10-mL samples of marine waters. The assay was optimized in phosphate-buffered saline and seawater, reducing the qPCR signal of heat-killed E. faecalis cells by 4 log10 and 3 log10 units, respectively. Results suggest that high total dissolved solid concentration (32 g/L) in seawater can reduce PMA activity. Optimal PMA-qPCR standard curves with a 6-log dynamic range and detection limit of 102 cells/mL were generated for quantifying viable E. faecalis cells in marine waters. The developed assay was compared with the standard membrane filter (MF) method by quantifying viable E. faecalis cells in seawater samples exposed to solar radiation. The results of the developed PMA-qPCR assay did not match that of the standard MF method. This difference in the results reflects the different physiological states of E. faecalis cells in seawater. In conclusion, the developed assay is a rapid (∼5 h) method for the quantification of viable E. faecalis cells in marine recreational waters, which should be further improved and tested in different seawater settings. © 2014 Springer-Verlag Berlin Heidelberg.

  12. Defined Combinations of Cryomedia and Thawing Extenders Influence the Viable X-Y Boar Sperm Ratio in Vitro.

    Science.gov (United States)

    Korchunjit, W; Kaeoket, K; Kitiyanant, Y; Taylor, J; Wongtawan, T

    It is believed that plasma membrane X- and Y-chromosome bearing sperm are different; therefore the freezing and thawing process may affect X- and Y-sperm differently. The objective of this study was to investigate the effect of cryomedia and thawing extenders on the survival of X and Y-sperm. Three different cryomedia and thawing extenders were compared. Viable motile sperm were separated using a swim-up technique. Real-time PCR was used to identify the sperm type. Using CryoA for freezing and Beltsville-Thawing-Solution (BTS) as the thawing extender yielded significantly higher numbers of viable motile Y sperm (64 percent) than control (48 percent) (P semen freezing with CryoC and thawing with Androstar Plus gave a significantly lower number of viable motile Y sperm (32 percent) than control (51 percent). Our results revealed that defined combinations of cryomedia and thawing extenders significantly altered the survival ratio of frozen-thawed X-Y sperm in vitro, which has potential implications for artificial insemination.

  13. Bio-electrosprayed multicellular zebrafish embryos are viable and develop normally

    International Nuclear Information System (INIS)

    Clarke, Jonathan D W; Jayasinghe, Suwan N

    2008-01-01

    Bio-electrosprays are rapidly emerging as a viable protocol for directly engineering living cells. This communication reports the bio-electrospraying of multicellular organisms, namely zebrafish embryos. The results demonstrate that the bio-electrospray protocol fails to induce any embryological perturbations. In addition to analysing overall embryo morphology, we use transgenic embryos that express green fluorescent protein in specific brain neurons to determine that neuronal numbers and organization are completely normal. These results demonstrate that the bio-electrospraying protocol does not interfere with the complex gene regulation and cell movements required for the development of a multicellular organism. (communication)

  14. Sinusoidal potential cycling operation of a direct ethanol fuel cell to improving carbon dioxide yields

    Science.gov (United States)

    Majidi, Pasha; Pickup, Peter G.

    2014-12-01

    A direct ethanol fuel cell has been operated under sinusoidal (AC) potential cycling conditions in order to increase the yield of carbon dioxide and thereby increase cell efficiency relative to operation at a fixed potential. At 80 °C, faradaic yields of CO2 as high as 25% have been achieved with a PtRu anode catalyst, while the maximum CO2 production at constant potential was 13%. The increased yields under cycling conditions have been attributed to periodic oxidative stripping of adsorbed CO. These results will be important in the optimization of operating conditions for direct ethanol fuel cells, where the benefits of potential cycling are projected to increase as catalysts that produce CO2 more efficiently are implemented.

  15. A factor converting viable but nonculturable Vibrio cholerae to a culturable state in eukaryotic cells is a human catalase.

    Science.gov (United States)

    Senoh, Mitsutoshi; Hamabata, Takashi; Takeda, Yoshifumi

    2015-08-01

    In our previous work, we demonstrated that viable but nonculturable (VBNC) Vibrio cholerae O1 and O139 were converted to culturable by coculture with eukaryotic cells. Furthermore, we isolated a factor converting VBNC V. cholerae to culturable (FCVC) from a eukaryotic cell line, HT-29. In this study, we purified FCVC by successive column chromatographies comprising UNO Q-6 anion exchange, Bio-Scale CHT2-1 hydroxyapatite, and Superdex 200 10/300 GL. Homogeneity of the purified FCVC was demonstrated by SDS-PAGE. Nano-LC MS/MS analysis showed that the purified FCVC was a human catalase. An experiment of RNAi knockdown of catalase mRNA from HT-29 cells and treatment of the purified FCVC with a catalase inhibitor, 3-amino-1,2,4-triazole confirmed that the FCVC was a catalase. A possible role of the catalase in converting a VBNC V. cholerae to a culturable state in the human intestine is discussed. © 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  16. Sources of Hematopoietic Stem and Progenitor Cells and Methods to Optimize Yields for Clinical Cell Therapy.

    Science.gov (United States)

    Panch, Sandhya R; Szymanski, James; Savani, Bipin N; Stroncek, David F

    2017-08-01

    Bone marrow (BM) aspirates, mobilized peripheral blood, and umbilical cord blood (UCB) have developed as graft sources for hematopoietic stem and progenitor cells (HSPCs) for stem cell transplantation and other cellular therapeutics. Individualized techniques are necessary to enhance graft HSPC yields and cell quality from each graft source. BM aspirates yield adequate CD34 + cells but can result in relative delays in engraftment. Granulocyte colony-stimulating factor (G-CSF)-primed BM HSPCs may facilitate faster engraftment while minimizing graft-versus-host disease in certain patient subsets. The levels of circulating HSPCs are enhanced using mobilizing agents, such as G-CSF and/or plerixafor, which act via the stromal cell-derived factor 1/C-X-C chemokine receptor type 4 axis. Alternate niche pathway mediators, including very late antigen-4/vascular cell adhesion molecule-1, heparan sulfate proteoglycans, parathyroid hormone, and coagulation cascade intermediates, may offer promising alternatives for graft enhancement. UCB grafts have been expanded ex vivo with cytokines, notch-ligand, or mesenchymal stromal cells, and most studies demonstrated greater quantities of CD34 + cells ex vivo and improved short-term engraftment. No significant changes were observed in long-term repopulating potential or in patient survival. Early phase clinical trials using nicotinamide and StemReginin1 may offer improved short- and long-term repopulating ability. Breakthroughs in genome editing and stem cell reprogramming technologies may hasten the generation of pooled, third-party HSPC grafts. This review elucidates past, present, and potential future approaches to HSPC graft optimization. Published by Elsevier Inc.

  17. Viable Bacteria Associated with Red Blood Cells and Plasma in Freshly Drawn Blood Donations

    DEFF Research Database (Denmark)

    Damgaard, Christian; Magnussen, Karin; Enevold, Christian

    2015-01-01

    ) or blue lactose plates. For identification colony PCR was performed using primers targeting 16S rDNA. SETTING: Blood donors attending Capital Region Blood Bank, Copenhagen University Hospital, Rigshospitalet, Hvidovre, Denmark, October 29th to December 10th 2013. PARTICIPANTS: 60 donors (≥50 years old....... CONCLUSIONS: Viable bacteria are present in blood from donors self-reported as medically healthy, indicating that conventional test systems employed by blood banks insufficiently detect bacteria in plasma. Further investigation is needed to determine whether routine testing for anaerobic bacteria and testing......OBJECTIVES: Infection remains a leading cause of post-transfusion mortality and morbidity. Bacterial contamination is, however, detected in less than 0.1% of blood units tested. The aim of the study was to identify viable bacteria in standard blood-pack units, with particular focus on bacteria from...

  18. Fuel cells : a viable fossil fuel alternative

    Energy Technology Data Exchange (ETDEWEB)

    Paduada, M.

    2007-02-15

    This article presented a program initiated by Natural Resources Canada (NRCan) to develop proof-of-concept of underground mining vehicles powered by fuel cells in order to eliminate emissions. Recent studies on American and Canadian underground mines provided the basis for estimating the operational cost savings of switching from diesel to fuel cells. For the Canadian mines evaluated, the estimated ventilation system operating cost reductions ranged from 29 per cent to 75 per cent. In order to demonstrate the viability of a fuel cell-powered vehicle, NRCan has designed a modified Caterpillar R1300 loader with a 160 kW hybrid power plant in which 3 stacks of fuel cells deliver up to 90 kW continuously, and a nickel-metal hydride battery provides up to 70 kW. The battery subsystem transiently boosts output to meet peak power requirements and also accommodates regenerative braking. Traction for the loader is provided by a brushless permanent magnet traction motor. The hydraulic pump motor is capable of a 55 kW load continuously. The loader's hydraulic and traction systems are operated independently. Future fuel cell-powered vehicles designed by the program may include a locomotive and a utility vehicle. Future mines running their operations with hydrogen-fueled equipment may also gain advantages by employing fuel cells in the operation of handheld equipment such as radios, flashlights, and headlamps. However, the proton exchange membrane (PEM) fuel cells used in the project are prohibitively expensive. The catalytic content of a fuel cell can add hundreds of dollars per kW of electric output. Production of catalytic precious metals will be strongly connected to the scale of use and acceptance of fuel cells in vehicles. In addition, the efficiency of hydrogen production and delivery is significantly lower than the well-to-tank efficiency of many conventional fuels. It was concluded that an adequate hydrogen infrastructure will be required for the mining industry

  19. Concerning the role of cell lysis-cryptic growth in anaerobic side-stream reactors: the single-cell analysis of viable, dead and lysed bacteria.

    Science.gov (United States)

    Foladori, P; Velho, V F; Costa, R H R; Bruni, L; Quaranta, A; Andreottola, G

    2015-05-01

    In the Anaerobic Side-Stream Reactor (ASSR), part of the return sludge undergoes alternating aerobic and anaerobic conditions with the aim of reducing sludge production. In this paper, viability, enzymatic activity, death and lysis of bacterial cells exposed to aerobic and anaerobic conditions for 16 d were investigated at single-cell level by flow cytometry, with the objective of contributing to the understanding of the mechanisms of sludge reduction in the ASSR systems. Results indicated that total and viable bacteria did not decrease during the anaerobic phase, indicating that anaerobiosis at ambient temperature does not produce a significant cell lysis. Bacteria decay and lysis occurred principally under aerobic conditions. The aerobic decay rate of total bacteria (bTB) was considered as the rate of generation of lysed bacteria. Values of bTB of 0.07-0.11 d(-1) were measured in anaerobic + aerobic sequence. The enzymatic activity was not particularly affected by the transition from anaerobiosis to aerobiosis. Large solubilisation of COD and NH4(+) was observed only under anaerobic conditions, as a consequence of hydrolysis of organic matter, but not due to cell lysis. The observations supported the proposal of two independent mechanisms contributing equally to sludge reduction: (1) under anaerobic conditions: sludge hydrolysis of non-bacterial material, (2) under aerobic conditions: bacterial cell lysis and oxidation of released biodegradable compounds. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Managing Viable Knowledge

    NARCIS (Netherlands)

    Achterbergh, J.M.I.M.; Vriens, D.J.

    2002-01-01

    In this paper, Beer's Viable System Model (VSM) is applied to knowledge management. Based on the VSM, domains of knowledge are identified that an organization should possess to maintain its viability. The logic of the VSM is also used to support the diagnosis, design and implementation of the

  1. The role of stem cell mobilization regimen on lymphocyte collection yield in patients with multiple myeloma.

    Science.gov (United States)

    Hiwase, D K; Hiwase, S; Bailey, M; Bollard, G; Schwarer, A P

    2008-01-01

    The lymphocyte dose (LY-DO) infused during an autograft influences absolute lymphocyte (ALC) recovery and survival following autologous stem cell transplantation (ASCT) in multiple myeloma (MM) patients. Factors influencing lymphocyte yield (LY-C) during leukapheresis have been poorly studied. Factors that could influence survival, LY-C and CD34(+) cell yield were analyzed in 122 MM patients. Three mobilization regimens were used, granulocyte-colony-stimulating factor (G-CSF) alone (n=13), cyclophosphamide 1-2 g/m(2) plus G-CSF (LD-CY, n=62) and cyclophosphamide 3-4 g/m(2) and G-CSF (ID-CY, n=47). Using multivariate analysis, age, LY-C, ALC on day 30 (ALC-30) and International Staging System stage significantly influenced overall (OS) and progression-free survival (PFS) following ASCT. PFS (56 versus 29 months, P=0.05) and OS (72 versus 49 months; P=0.07) were longer in the LY-C>or=0.12x10(9)/kg group than the LY-Cradiotherapy and number of leukaphereses significantly influenced LY-C. Significantly higher LY-C was obtained with G-CSF alone compared with the LD-CY and ID-CY groups. CD34(+) count on the day of leukapheresis, prior chemotherapy with prednisone, cyclophosphamide, adriamycin and BCNU or melphalan, and stem cell mobilization regimen significantly influenced CD34(+) cell yield. LY-C influenced ALC-15 and survival following ASCT. Factors that influenced CD34(+) cell yield and LY-C during leukapheresis were different. Mobilization should be tailored to maximize the LY-C and CD34(+) cell yield.

  2. The use of flow cytometry to accurately ascertain total and viable counts of Lactobacillus rhamnosus in chocolate.

    Science.gov (United States)

    Raymond, Yves; Champagne, Claude P

    2015-04-01

    The goals of this study were to evaluate the precision and accuracy of flow cytometry (FC) methodologies in the evaluation of populations of probiotic bacteria (Lactobacillus rhamnosus R0011) in two commercial dried forms, and ascertain the challenges in enumerating them in a chocolate matrix. FC analyses of total (FC(T)) and viable (FC(V)) counts in liquid or dried cultures were almost two times more precise (reproducible) than traditional direct microscopic counts (DCM) or colony forming units (CFU). With FC, it was possible to ascertain low levels of dead cells (FC(D)) in fresh cultures, which is not possible with traditional CFU and DMC methodologies. There was no interference of chocolate solids on FC counts of probiotics when inoculation was above 10(7) bacteria per g. Addition of probiotics in chocolate at 40 °C resulted in a 37% loss in viable cells. Blending of the probiotic powder into chocolate was not uniform which raised a concern that the precision of viable counts could suffer. FCT data can serve to identify the correct inoculation level of a sample, and viable counts (FCV or CFU) can subsequently be better interpreted. Crown Copyright © 2014. Published by Elsevier Ltd. All rights reserved.

  3. Rapid cell separation with minimal manipulation for autologous cell therapies

    Science.gov (United States)

    Smith, Alban J.; O'Rorke, Richard D.; Kale, Akshay; Rimsa, Roberts; Tomlinson, Matthew J.; Kirkham, Jennifer; Davies, A. Giles; Wälti, Christoph; Wood, Christopher D.

    2017-02-01

    The ability to isolate specific, viable cell populations from mixed ensembles with minimal manipulation and within intra-operative time would provide significant advantages for autologous, cell-based therapies in regenerative medicine. Current cell-enrichment technologies are either slow, lack specificity and/or require labelling. Thus a rapid, label-free separation technology that does not affect cell functionality, viability or phenotype is highly desirable. Here, we demonstrate separation of viable from non-viable human stromal cells using remote dielectrophoresis, in which an electric field is coupled into a microfluidic channel using shear-horizontal surface acoustic waves, producing an array of virtual electrodes within the channel. This allows high-throughput dielectrophoretic cell separation in high conductivity, physiological-like fluids, overcoming the limitations of conventional dielectrophoresis. We demonstrate viable/non-viable separation efficacy of >98% in pre-purified mesenchymal stromal cells, extracted from human dental pulp, with no adverse effects on cell viability, or on their subsequent osteogenic capabilities.

  4. Detection of viable Salmonella in ice cream by TaqMan real-time polymerase chain reaction assay combining propidium monoazide

    Directory of Open Access Journals (Sweden)

    Yuexia Wang

    2015-09-01

    Full Text Available Real-time polymerase chain reaction (PCR allows rapid detection of Salmonella in frozen dairy products, but it might cause a false positive detection result because it might amplify DNA from dead target cells as well. In this study, Salmonella-free frozen ice cream was initially inoculated with heat-killed Salmonella Typhimurium cells and stored at −18°C. Bacterial DNA extracted from the sample was amplified using TaqMan probe-based real-time PCR targeting the invA gene. Our results indicated that DNA from the dead cells remained stable in frozen ice cream for at least 20 days, and could produce fluorescence signal for real-time PCR as well. To overcome this limitation, propidium monoazide (PMA was combined with real-time PCR. PMA treatment can effectively prevent PCR amplification from heat-killed Salmonella cells in frozen ice cream. The PMA real-time PCR assay can selectively detect viable Salmonella at as low as 103 CFU/mL. Combining 18 hours of pre-enrichment with the assay allows for the detection of viable Salmonella at 100 CFU/mL and avoiding the false-positive result of dead cells. The PMA real-time PCR assay provides an alternative specifically for detection of viable Salmonella in ice cream. However, when the PMA real-time PCR assay was evaluated in ice cream subjected to frozen storage, it obviously underestimated the contamination situation of viable Salmonella, which might lead to a false negative result. According to this result, the use of enrichment prior to PMA real-time PCR analysis remains as the more appropriate approach.

  5. Detection of viable Salmonella in ice cream by TaqMan real-time polymerase chain reaction assay combining propidium monoazide.

    Science.gov (United States)

    Wang, Yuexia; Yang, Ming; Liu, Shuchun; Chen, Wanyi; Suo, Biao

    2015-09-01

    Real-time polymerase chain reaction (PCR) allows rapid detection of Salmonella in frozen dairy products, but it might cause a false positive detection result because it might amplify DNA from dead target cells as well. In this study, Salmonella-free frozen ice cream was initially inoculated with heat-killed Salmonella Typhimurium cells and stored at -18°C. Bacterial DNA extracted from the sample was amplified using TaqMan probe-based real-time PCR targeting the invA gene. Our results indicated that DNA from the dead cells remained stable in frozen ice cream for at least 20 days, and could produce fluorescence signal for real-time PCR as well. To overcome this limitation, propidium monoazide (PMA) was combined with real-time PCR. PMA treatment can effectively prevent PCR amplification from heat-killed Salmonella cells in frozen ice cream. The PMA real-time PCR assay can selectively detect viable Salmonella at as low as 10 3  CFU/mL. Combining 18 hours of pre-enrichment with the assay allows for the detection of viable Salmonella at 10 0  CFU/mL and avoiding the false-positive result of dead cells. The PMA real-time PCR assay provides an alternative specifically for detection of viable Salmonella in ice cream. However, when the PMA real-time PCR assay was evaluated in ice cream subjected to frozen storage, it obviously underestimated the contamination situation of viable Salmonella, which might lead to a false negative result. According to this result, the use of enrichment prior to PMA real-time PCR analysis remains as the more appropriate approach. Copyright © 2015. Published by Elsevier B.V.

  6. Detection and Quantification of Viable and Nonviable Trypanosoma cruzi Parasites by a Propidium Monoazide Real-Time Polymerase Chain Reaction Assay

    Science.gov (United States)

    Cancino-Faure, Beatriz; Fisa, Roser; Alcover, M. Magdalena; Jimenez-Marco, Teresa; Riera, Cristina

    2016-01-01

    Molecular techniques based on real-time polymerase chain reaction (qPCR) allow the detection and quantification of DNA but are unable to distinguish between signals from dead or live cells. Because of the lack of simple techniques to differentiate between viable and nonviable cells, the aim of this study was to optimize and evaluate a straightforward test based on propidium monoazide (PMA) dye action combined with a qPCR assay (PMA-qPCR) for the selective quantification of viable/nonviable epimastigotes of Trypanosoma cruzi. PMA has the ability to penetrate the plasma membrane of dead cells and covalently cross-link to the DNA during exposure to bright visible light, thereby inhibiting PCR amplification. Different concentrations of PMA (50–200 μM) and epimastigotes of the Maracay strain of T. cruzi (1 × 105–10 parasites/mL) were assayed; viable and nonviable parasites were tested and quantified by qPCR with a TaqMan probe specific for T. cruzi. In the PMA-qPCR assay optimized at 100 μM PMA, a significant qPCR signal reduction was observed in the nonviable versus viable epimastigotes treated with PMA, with a mean signal reduction of 2.5 logarithm units and a percentage of signal reduction > 98%, in all concentrations of parasites assayed. This signal reduction was also observed when PMA-qPCR was applied to a mixture of live/dead parasites, which allowed the detection of live cells, except when the concentration of live parasites was low (10 parasites/mL). The PMA-qPCR developed allows differentiation between viable and nonviable epimastigotes of T. cruzi and could thus be a potential method of parasite viability assessment and quantification. PMID:27139452

  7. Enumeration of viable and non-viable larvated Ascaris eggs with quantitative PCR

    Science.gov (United States)

    Aims: The goal of the study was to further develop an incubation-qPCR method for quantifying viable Ascaris eggs. The specific objectives were to characterize the detection limit and number of template copies per egg, determine the specificity of the method, and test the method w...

  8. Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3

    OpenAIRE

    Liu, Z.; Zhang, P.; Zhou, Y.; Qin, H.; Shen, T.

    2010-01-01

    Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithel...

  9. Nut crop yield records show that budbreak-based chilling requirements may not reflect yield decline chill thresholds

    Science.gov (United States)

    Pope, Katherine S.; Dose, Volker; Da Silva, David; Brown, Patrick H.; DeJong, Theodore M.

    2015-06-01

    Warming winters due to climate change may critically affect temperate tree species. Insufficiently cold winters are thought to result in fewer viable flower buds and the subsequent development of fewer fruits or nuts, decreasing the yield of an orchard or fecundity of a species. The best existing approximation for a threshold of sufficient cold accumulation, the "chilling requirement" of a species or variety, has been quantified by manipulating or modeling the conditions that result in dormant bud breaking. However, the physiological processes that affect budbreak are not the same as those that determine yield. This study sought to test whether budbreak-based chilling thresholds can reasonably approximate the thresholds that affect yield, particularly regarding the potential impacts of climate change on temperate tree crop yields. County-wide yield records for almond ( Prunus dulcis), pistachio ( Pistacia vera), and walnut ( Juglans regia) in the Central Valley of California were compared with 50 years of weather records. Bayesian nonparametric function estimation was used to model yield potentials at varying amounts of chill accumulation. In almonds, average yields occurred when chill accumulation was close to the budbreak-based chilling requirement. However, in the other two crops, pistachios and walnuts, the best previous estimate of the budbreak-based chilling requirements was 19-32 % higher than the chilling accumulations associated with average or above average yields. This research indicates that physiological processes beyond requirements for budbreak should be considered when estimating chill accumulation thresholds of yield decline and potential impacts of climate change.

  10. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  11. Successful Isolation of Viable Adipose-Derived Stem Cells from Human Adipose Tissue Subject to Long-Term Cryopreservation: Positive Implications for Adult Stem Cell-Based Therapeutics in Patients of Advanced Age

    Directory of Open Access Journals (Sweden)

    Sean M. Devitt

    2015-01-01

    Full Text Available We examined cell isolation, viability, and growth in adipose-derived stem cells harvested from whole adipose tissue subject to different cryopreservation lengths (2–1159 days from patients of varying ages (26–62 years. Subcutaneous abdominal adipose tissue was excised during abdominoplasties and was cryopreserved. The viability and number of adipose-derived stem cells isolated were measured after initial isolation and after 9, 18, and 28 days of growth. Data were analyzed with respect to cryopreservation duration and patient age. Significantly more viable cells were initially isolated from tissue cryopreserved 2 years, irrespective of patient age. However, this difference did not persist with continued growth and there were no significant differences in cell viability or growth at subsequent time points with respect to cryopreservation duration or patient age. Mesenchymal stem cell markers were maintained in all cohorts tested throughout the duration of the study. Consequently, longer cryopreservation negatively impacts initial live adipose-derived stem cell isolation; however, this effect is neutralized with continued cell growth. Patient age does not significantly impact stem cell isolation, viability, or growth. Cryopreservation of adipose tissue is an effective long-term banking method for isolation of adipose-derived stem cells in patients of varying ages.

  12. Quality of raw cow milk in Republic of Macedonia determined through the testing of somatic cell count and total viable count

    Directory of Open Access Journals (Sweden)

    Angelovski Ljupco

    2008-11-01

    Full Text Available Somatic cells count and total viable count are criteria used to estimate the compliance of raw cow milk with the Book of rules for demands for safety and hygiene and procedures for official controls of milk and milk products, Official Gazette of RM 157/2007. According to the given demands, raw milk operators are obliged to conduct all procedures and to guarantee that milk is in compliance with the criteria laid down in Book of rules. At the same time, Republic of Macedonia have to fulfill EU criteria laid down in Directive 92/46 (Council directive 92/46/EEC laying down the health rules for the production and placing on the market of raw milk, heat-treated milk and milkbased products for quality of raw milk as part of implementation of community legislation and milk production. The independent laboratory for milk quality control at FVM-Skopje, in frame of its activities in the period February- August 2008 has conducted a study for obtaining preliminary results for the situation with raw milk quality produced in R. of Macedonia for somatic cells counts and total viable count. In the study we analyzed 2065 samples for TVC and 1625 samples for SCC of raw milk samples produced in different parts of the country. From the tested samples only 41,8% fulfill criteria for SCC and 41,45% criteria for TVC lay down in Book of rules for 2008. Assessment of the results in light of Council Directive it is obvious that only 42,7% of the samples for SCC and 10,7% for TVC fulfill the criteria of Council Directive having in mind different requirements vs. Book of rules.

  13. Optimizing culture medium composition to improve oligodendrocyte progenitor cell yields in vitro from subventricular zone-derived neural progenitor cell neurospheres.

    Directory of Open Access Journals (Sweden)

    Paula G Franco

    Full Text Available Neural Stem and Progenitor Cells (NSC/NPC are gathering tangible recognition for their uses in cell therapy and cell replacement therapies for human disease, as well as a model system to continue research on overall neural developmental processes in vitro. The Subventricular Zone is one of the largest NSC/NPC niches in the developing mammalian Central Nervous System, and persists through to adulthood. Oligodendrocyte progenitor cell (OPC enriched cultures are usefull tools for in vitro studies as well as for cell replacement therapies for treating demyelination diseases. We used Subventricular Zone-derived NSC/NPC primary cultures from newborn mice and compared the effects of different growth factor combinations on cell proliferation and OPC yield. The Platelet Derived Growth Factor-AA and BB homodimers had a positive and significant impact on OPC generation. Furthermore, heparin addition to the culture media contributed to further increase overall culture yields. The OPC generated by this protocol were able to mature into Myelin Basic Protein-expressing cells and to interact with neurons in an in vitro co-culture system. As a whole, we describe an optimized in vitro method for increasing OPC.

  14. High-yield secretion of recombinant proteins expressed in tobacco cell culture with a designer glycopeptide tag: Process development.

    Science.gov (United States)

    Zhang, Ningning; Gonzalez, Maria; Savary, Brett; Xu, Jianfeng

    2016-03-01

    Low-yield protein production remains the most significant economic hurdle with plant cell culture technology. Fusions of recombinant proteins with hydroxyproline-O-glycosylated designer glycopeptide tags have consistently boosted secreted protein yields. This prompted us to study the process development of this technology aiming to achieve productivity levels necessary for commercial viability. We used a tobacco BY-2 cell culture expressing EGFP as fusion with a glycopeptide tag comprised of 32 repeat of "Ser-Pro" dipeptide, or (SP)32 , to study cell growth and protein secretion, culture scale-up, and establishment of perfusion cultures for continuous production. The BY-2 cells accumulated low levels of cell biomass (~7.5 g DW/L) in Schenk & Hildebrandt medium, but secreted high yields of (SP)32 -tagged EGFP (125 mg/L). Protein productivity of the cell culture has been stable for 6.0 years. The BY-2 cells cultured in a 5-L bioreactor similarly produced high secreted protein yield at 131 mg/L. Successful operation of a cell perfusion culture for 30 days was achieved under the perfusion rate of 0.25 and 0.5 day(-1) , generating a protein volumetric productivity of 17.6 and 28.9 mg/day/L, respectively. This research demonstrates the great potential of the designer glycopeptide technology for use in commercial production of valuable proteins with plant cell cultures. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Generation of high-yield insulin producing cells from human bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Jafarian, Arefeh; Taghikhani, Mohammad; Abroun, Saeid; Pourpak, Zahra; Allahverdi, Amir; Soleimani, Masoud

    2014-07-01

    Allogenic islet transplantation is a most efficient approach for treatment of diabetes mellitus. However, the scarcity of islets and long term need for an immunosuppressant limits its application. Recently, cell replacement therapies that generate of unlimited sources of β cells have been developed to overcome these limitations. In this study we have described a stage specific differentiation protocol for the generation of insulin producing islet-like clusters from human bone marrow mesenchymal stem cells (hBM-MSCs). This specific stepwise protocol induced differentiation of hMSCs into definitive endoderm, pancreatic endoderm and pancreatic endocrine cells that expressed of sox17, foxa2, pdx1, ngn3, nkx2.2, insulin, glucagon, somatostatin, pancreatic polypeptide, and glut2 transcripts respectively. In addition, immunocytochemical analysis confirmed protein expression of the above mentioned genes. Western blot analysis discriminated insulin from proinsulin in the final differentiated cells. In derived insulin producing cells (IPCs), secreted insulin and C-peptide was in a glucose dependent manner. We have developed a protocol that generates effective high-yield human IPCs from hBM-MSCs in vitro. These finding suggest that functional IPCs generated by this procedure can be used as a cell-based approach for insulin dependent diabetes mellitus.

  16. Rapid assessment of viable but non-culturable Bacillus coagulans MTCC 5856 in commercial formulations using Flow cytometry

    Science.gov (United States)

    Majeed, Muhammed; Majeed, Shaheen; Nagabhushanam, Kalyanam; Punnapuzha, Ardra; Philip, Sheena

    2018-01-01

    Accurate enumeration of bacterial count in probiotic formulation is imperative to ensure that the product adheres to regulatory standards and citation in consumer product label. Standard methods like plate count, can enumerate only replicating bacterial population under selected culture conditions. Viable but non culturable bacteria (VBNC) retain characteristics of living cells and can regain cultivability by a process known as resuscitation. This is a protective mechanism adapted by bacteria to evade stressful environmental conditions. B. coagulans MTCC 5856(LactoSpore®) is a probiotic endospore which can survive for decades in hostile environments without dividing. In the present study, we explored the use of flow cytometry to enumerate the viable count of B. coagulans MTCC 5856 under acidic and alkaline conditions, high temperature and in commercial formulations like compressed tablets and capsules. Flow cytometry (FCM) was comparable to plate count method when the spores were counted at physiological conditions. We show that VBNC state is induced in B. coagulans MTCC 5856by high temperature and acidic pH. The cells get resuscitated under physiological conditions and FCM was sensitive to detect the VBNC spores. Flow cytometry showed excellent ability to assess the viable spore count in commercial probiotic formulations of B. coagulans MTCC 5856. The results establish Flow cytometry as a reliable method to count viable bacteria in commercial probiotic preparations. Sporulation as well as existence as VBNC could contribute to the extreme stability of B. coagulans MTCC 5856. PMID:29474436

  17. Rapid assessment of viable but non-culturable Bacillus coagulans MTCC 5856 in commercial formulations using Flow cytometry.

    Directory of Open Access Journals (Sweden)

    Muhammed Majeed

    Full Text Available Accurate enumeration of bacterial count in probiotic formulation is imperative to ensure that the product adheres to regulatory standards and citation in consumer product label. Standard methods like plate count, can enumerate only replicating bacterial population under selected culture conditions. Viable but non culturable bacteria (VBNC retain characteristics of living cells and can regain cultivability by a process known as resuscitation. This is a protective mechanism adapted by bacteria to evade stressful environmental conditions. B. coagulans MTCC 5856(LactoSpore® is a probiotic endospore which can survive for decades in hostile environments without dividing. In the present study, we explored the use of flow cytometry to enumerate the viable count of B. coagulans MTCC 5856 under acidic and alkaline conditions, high temperature and in commercial formulations like compressed tablets and capsules. Flow cytometry (FCM was comparable to plate count method when the spores were counted at physiological conditions. We show that VBNC state is induced in B. coagulans MTCC 5856by high temperature and acidic pH. The cells get resuscitated under physiological conditions and FCM was sensitive to detect the VBNC spores. Flow cytometry showed excellent ability to assess the viable spore count in commercial probiotic formulations of B. coagulans MTCC 5856. The results establish Flow cytometry as a reliable method to count viable bacteria in commercial probiotic preparations. Sporulation as well as existence as VBNC could contribute to the extreme stability of B. coagulans MTCC 5856.

  18. Rapid assessment of viable but non-culturable Bacillus coagulans MTCC 5856 in commercial formulations using Flow cytometry.

    Science.gov (United States)

    Majeed, Muhammed; Majeed, Shaheen; Nagabhushanam, Kalyanam; Punnapuzha, Ardra; Philip, Sheena; Mundkur, Lakshmi

    2018-01-01

    Accurate enumeration of bacterial count in probiotic formulation is imperative to ensure that the product adheres to regulatory standards and citation in consumer product label. Standard methods like plate count, can enumerate only replicating bacterial population under selected culture conditions. Viable but non culturable bacteria (VBNC) retain characteristics of living cells and can regain cultivability by a process known as resuscitation. This is a protective mechanism adapted by bacteria to evade stressful environmental conditions. B. coagulans MTCC 5856(LactoSpore®) is a probiotic endospore which can survive for decades in hostile environments without dividing. In the present study, we explored the use of flow cytometry to enumerate the viable count of B. coagulans MTCC 5856 under acidic and alkaline conditions, high temperature and in commercial formulations like compressed tablets and capsules. Flow cytometry (FCM) was comparable to plate count method when the spores were counted at physiological conditions. We show that VBNC state is induced in B. coagulans MTCC 5856by high temperature and acidic pH. The cells get resuscitated under physiological conditions and FCM was sensitive to detect the VBNC spores. Flow cytometry showed excellent ability to assess the viable spore count in commercial probiotic formulations of B. coagulans MTCC 5856. The results establish Flow cytometry as a reliable method to count viable bacteria in commercial probiotic preparations. Sporulation as well as existence as VBNC could contribute to the extreme stability of B. coagulans MTCC 5856.

  19. Detection of viable but non-culturable Escherichia coli O157:H7 by PCR in combination with propidium monoazide.

    Science.gov (United States)

    Zhong, Junliang; Zhao, Xihong

    2018-01-01

    The aim of this study was to evaluate the applicability of the conventional PCR detection method combined with propidium monoazide (PMA) treatment for the detection of viable but non-culturable (VBNC) state Escherichia coli O157:H7 in ground beef meatballs. Under low temperature, E. coli O157:H7 cells were induced into the VBNC state in ground beef meatballs at - 20 °C after 152 days. The optimal PMA concentration of 5 µg/mL was obtained in beef meatball samples, which could completely inhibit the DNA amplification on dead cells (10 6  cells/mL) but with no inhibition on viable cells. The established PMA-PCR assay revealed that the VBNC counts exceeded 10 7  CFU/mL in artificial contamination beef samples, which could be used for semi-quantitative detection of VBNC cells in beef meatball samples. This study indicated that the PMA-PCR assay might be a potential method for detection of VBNC state E . coli O157:H7 cells in food products.

  20. Estimating milk yield and value losses from increased somatic cell count on US dairy farms.

    Science.gov (United States)

    Hadrich, J C; Wolf, C A; Lombard, J; Dolak, T M

    2018-04-01

    Milk loss due to increased somatic cell counts (SCC) results in economic losses for dairy producers. This research uses 10 mo of consecutive dairy herd improvement data from 2013 and 2014 to estimate milk yield loss using SCC as a proxy for clinical and subclinical mastitis. A fixed effects regression was used to examine factors that affected milk yield while controlling for herd-level management. Breed, milking frequency, days in milk, seasonality, SCC, cumulative months with SCC greater than 100,000 cells/mL, lactation, and herd size were variables included in the regression analysis. The cumulative months with SCC above a threshold was included as a proxy for chronic mastitis. Milk yield loss increased as the number of test days with SCC ≥100,000 cells/mL increased. Results from the regression were used to estimate a monetary value of milk loss related to SCC as a function of cow and operation related explanatory variables for a representative dairy cow. The largest losses occurred from increased cumulative test days with a SCC ≥100,000 cells/mL, with daily losses of $1.20/cow per day in the first month to $2.06/cow per day in mo 10. Results demonstrate the importance of including the duration of months above a threshold SCC when estimating milk yield losses. Cows with chronic mastitis, measured by increased consecutive test days with SCC ≥100,000 cells/mL, resulted in higher milk losses than cows with a new infection. This provides farm managers with a method to evaluate the trade-off between treatment and culling decisions as it relates to mastitis control and early detection. Copyright © 2018 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  1. Clinical validation of an ultra high-throughput spiral microfluidics for the detection and enrichment of viable circulating tumor cells.

    Directory of Open Access Journals (Sweden)

    Bee Luan Khoo

    Full Text Available Circulating tumor cells (CTCs are cancer cells that can be isolated via liquid biopsy from blood and can be phenotypically and genetically characterized to provide critical information for guiding cancer treatment. Current analysis of CTCs is hindered by the throughput, selectivity and specificity of devices or assays used in CTC detection and isolation.Here, we enriched and characterized putative CTCs from blood samples of patients with both advanced stage metastatic breast and lung cancers using a novel multiplexed spiral microfluidic chip. This system detected putative CTCs under high sensitivity (100%, n = 56 (Breast cancer samples: 12-1275 CTCs/ml; Lung cancer samples: 10-1535 CTCs/ml rapidly from clinically relevant blood volumes (7.5 ml under 5 min. Blood samples were completely separated into plasma, CTCs and PBMCs components and each fraction were characterized with immunophenotyping (Pan-cytokeratin/CD45, CD44/CD24, EpCAM, fluorescence in-situ hybridization (FISH (EML4-ALK or targeted somatic mutation analysis. We used an ultra-sensitive mass spectrometry based system to highlight the presence of an EGFR-activating mutation in both isolated CTCs and plasma cell-free DNA (cf-DNA, and demonstrate concordance with the original tumor-biopsy samples.We have clinically validated our multiplexed microfluidic chip for the ultra high-throughput, low-cost and label-free enrichment of CTCs. Retrieved cells were unlabeled and viable, enabling potential propagation and real-time downstream analysis using next generation sequencing (NGS or proteomic analysis.

  2. Pancreatic Beta-Cell Purification by Altering FAD and NAD(PH Metabolism

    Directory of Open Access Journals (Sweden)

    P. de Vos

    2008-07-01

    Full Text Available Isolation of primary beta cells from other cells within in the pancreatic islets is of importance for many fields of islet research. However, up to now, no satisfactory method has been developed that gained high numbers of viable beta cells, without considerable alpha-cell contamination. In this study, we investigated whether rat beta cells can be isolated from nonbeta endocrine cells by manipulating the flavin adenine dinucleotide (FAD and nicotinamide-adenine dinucleotide phosphate (NAD(PH autofluorescence. Beta cells were isolated from dispersed islets by flow cytometry, based on their high FAD and NAD(PH fluorescence. To improve beta cell yield and purity, the cellular FAD and NAD(PH contents were altered by preincubation in culture media containing varying amounts of D-glucose and amino acids. Manipulation of the cellular FAD and NAD(PH fluorescence improves beta cell yield and purity after sorting. This method is also a fast and reliable method to measure beta cell functional viability. A conceivable application is assessing beta cell viability before transplantation.

  3. Viable Mycobacterium avium ssp. paratuberculosis isolated from calf milk replacer.

    Science.gov (United States)

    Grant, Irene R; Foddai, Antonio C G; Tarrant, James C; Kunkel, Brenna; Hartmann, Faye A; McGuirk, Sheila; Hansen, Chungyi; Talaat, Adel M; Collins, Michael T

    2017-12-01

    When advising farmers on how to control Johne's disease in an infected herd, one of the main recommendations is to avoid feeding waste milk to calves and instead feed calf milk replacer (CMR). This advice is based on the assumption that CMR is free of viable Mycobacterium avium ssp. paratuberculosis (MAP) cells, an assumption that has not previously been challenged. We tested commercial CMR products (n = 83) obtained from dairy farms around the United States by the peptide-mediated magnetic separation (PMS)-phage assay, PMS followed by liquid culture (PMS-culture), and direct IS900 quantitative PCR (qPCR). Conventional microbiological analyses for total mesophilic bacterial counts, coliforms, Salmonella, coagulase-negative staphylococci, streptococci, nonhemolytic Corynebacterium spp., and Bacillus spp. were also performed to assess the overall microbiological quality of the CMR. Twenty-six (31.3%) of the 83 CMR samples showed evidence of the presence of MAP. Seventeen (20.5%) tested positive for viable MAP by the PMS-phage assay, with plaque counts ranging from 6 to 1,212 pfu/50 mL of reconstituted CMR (average 248.5 pfu/50 mL). Twelve (14.5%) CMR samples tested positive for viable MAP by PMS-culture; isolates from all 12 of these samples were subsequently confirmed by whole-genome sequencing to be different cattle strains of MAP. Seven (8.4%) CMR samples tested positive for MAP DNA by IS900 qPCR. Four CMR samples tested positive by both PMS-based tests and 5 CMR samples tested positive by IS900 qPCR plus one or other of the PMS-based tests, but only one CMR sample tested positive by all 3 MAP detection tests applied. All conventional microbiology results were within current standards for whole milk powders. A significant association existed between higher total bacterial counts and presence of viable MAP indicated by either of the PMS-based assays. This represents the first published report of the isolation of viable MAP from CMR. Our findings raise concerns

  4. Microbial Electrolysis Cells for High Yield Hydrogen Gas Production from Organic Matter

    NARCIS (Netherlands)

    Logan, B.E.; Call, D.; Cheng, S.; Hamelers, H.V.M.; Sleutels, T.H.J.A.; Jeremiasse, A.W.; Rozendal, R.A.

    2008-01-01

    The use of electrochemically active bacteria to break down organic matter, combined with the addition of a small voltage (>0.2 V in practice) in specially designed microbial electrolysis cells (MECs), can result in a high yield of hydrogen gas. While microbial electrolysis was invented only a few

  5. Effect of Nitrogen and Phosphorus on Yield and Yield Components of Sesame (Sesamumindicum L.)

    OpenAIRE

    Muhammad Ibrahim; Manzoor Hussain; Ahmad Khan; Yousaf Jamal; Muhammad Ali; Muhammad Faisal Anwar Malik

    2014-01-01

    Nitrogen is a structural component of chlorophyll and protein therefore adequate supply of nitrogen is beneficial for both carbohydrates and protein metabolism as it promotes cell division and cell enlargement, resulting in more leaf area and thus ensuring good seed and dry matter yield. Theexperiment entitled effect of nitrogen and phosphorus on yield and yield components of sesame were conducted at New Developmental Farm of the University of Agriculture Peshawar during kharif 2013. Randomiz...

  6. The viable but nonculturable state induction and genomic analyses of Lactobacillus casei BM-LC14617, a beer-spoilage bacterium.

    Science.gov (United States)

    Liu, Junyan; Li, Lin; Peters, Brian M; Li, Bing; Chen, Lequn; Deng, Yang; Xu, Zhenbo; Shirtliff, Mark E

    2017-10-01

    This study aimed to investigate the viable but nonculturable (VBNC) state and genomic features of a beer-spoilage strain, Lactobacillus caseiBM-LC14617. Induction on the VBNC state of L. casei strain BM-LC14617 was conducted by both low-temperature storage and continuous passage in beer, and formation of VBNC state was detected after 196 ± 3.3 days and 32 ± 1.6 subcultures, respectively. Resuscitation of VBNC cells was successfully induced by addition of catalase, and culturable, VBNC, and resuscitated cells shared similar beer-spoilage capability. Whole genome sequencing was performed, and out of a total of 3,964 predicted genes, several potential VBNC and beer-spoilage-associated genes were identified. L. casei is capable of entering into and resuscitating from the VBNC state and possesses beer-spoilage capability. The genomic characterization yield insightful elucidation of VBNC state for L. casei. This study represents the first evidence on VBNC state induction of L. casei and beer-spoilage capability of VBNC and resuscitated cells. Also, this is the first genomic characterization of L. casei as a beer-spoilage bacterium. The current study may aid in further study on L. casei and other beer-spoilage bacteria, and guide the prevention and control of beer spoilage. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  7. Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3

    Directory of Open Access Journals (Sweden)

    Z. Liu

    2010-05-01

    Full Text Available Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithelial cells, while endothelin-3 was added to stimulate their growth. By adding endothelin-3, the achievement ratio (viable cell cultures/total cultures was enhanced to 60% of a total of 10 cultures (initiated from 8 distinct fetal small intestines, allowing the generation of viable epithelial cell cultures. Western blot, real-time PCR and immunofluorescent staining showed that cytokeratins 8, 18 and mouse intestinal mucosa-1/39 had high expression levels in human intestinal epithelial cells. Differentiated markers such as sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV also showed high expression levels in human intestinal epithelial cells. Differentiated human intestinal epithelial cells, with the expression of surface markers (cytokeratins 8, 18 and mouse intestinal mucosa-1/39 and secretion of cytokines (sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV, may be cultured by the thermolysin and endothelin-3 method and maintained for at least 20 passages. This is relatively simple, requiring no sophisticated techniques or instruments, and may have a number of varied applications.

  8. Co-infusion of autologous adipose tissue derived neuronal differentiated mesenchymal stem cells and bone marrow derived hematopoietic stem cells, a viable therapy for post-traumatic brachial plexus injury: A case report

    Directory of Open Access Journals (Sweden)

    Umang G Thakkar

    2014-08-01

    Full Text Available Stem cell therapy is emerging as a viable approach in regenerative medicine. A 31-year-old male with brachial plexus injury had complete sensory-motor loss since 16 years with right pseudo-meningocele at C5-D1 levels and extra-spinal extension up to C7-D1, with avulsion on magnetic resonance imaging and irreversible damage. We generated adipose tissue derived neuronal differentiated mesenchymal stem cells (N-AD-MSC and bone marrow derived hematopoietic stem cells (HSC-BM. Neuronal stem cells expressed β-3 tubulin and glial fibrillary acid protein which was confirmed on immunofluorescence. On day 14, 2.8 ml stem cell inoculum was infused under local anesthesia in right brachial plexus sheath by brachial block technique under ultrasonography guidance with a 1.5-inch-long 23 gauge needle. Nucleated cell count was 2 × 10 4 /μl, CD34+ was 0.06%, and CD45-/90+ and CD45-/73+ were 41.63% and 20.36%, respectively. No untoward effects were noted. He has sustained recovery with re-innervation over a follow-up of 4 years documented on electromyography-nerve conduction velocity study.

  9. High Yield of Adult Oligodendrocyte Lineage Cells Obtained from Meningeal Biopsy

    Directory of Open Access Journals (Sweden)

    Sissi Dolci

    2017-10-01

    Full Text Available Oligodendrocyte loss can lead to cognitive and motor deficits. Current remyelinating therapeutic strategies imply either modulation of endogenous oligodendrocyte precursors or transplantation of in vitro expanded oligodendrocytes. Cell therapy, however, still lacks identification of an adequate source of oligodendrocyte present in adulthood and able to efficiently produce transplantable cells. Recently, a neural stem cell-like population has been identified in meninges. We developed a protocol to obtain high yield of oligodendrocyte lineage cells from one single biopsy of adult rat meningeal tissue. From 1 cm2 of adult rat spinal cord meninges, we efficiently expanded a homogenous culture of 10 millions of meningeal-derived oligodendrocyte lineage cells in a short period of time (approximately 4 weeks. Meningeal-derived oligodendrocyte lineage cells show typical mature oligodendrocyte morphology and express specific oligodendrocyte markers, such as galactosylceramidase and myelin basic protein. Moreover, when transplanted in a chemically demyelinated spinal cord model, meningeal-derived oligodendrocyte lineage cells display in vivo-remyelinating potential. This oligodendrocyte lineage cell population derives from an accessible and adult source, being therefore a promising candidate for autologous cell therapy of demyelinating diseases. In addition, the described method to differentiate meningeal-derived neural stem cells into oligodendrocyte lineage cells may represent a valid in vitro model to dissect oligodendrocyte differentiation and to screen for drugs capable to promote oligodendrocyte regeneration.

  10. Cellular and soluble components decrease the viable pathogen counts in milk from dairy cows with subclinical mastitis.

    Science.gov (United States)

    Koshiishi, Tomoko; Watanabe, Masako; Miyake, Hajime; Hisaeda, Keiichi; Isobe, Naoki

    2017-08-10

    The present study was undertaken to clarify the factors that reduce the viable pathogen count in milk collected from the udders of subclinical mastitic cows during preservation. Milk was centrifuged to divide somatic cells (cellular components, precipitates) and antimicrobial peptides (soluble components, supernatants without fat layer); each fraction was cultured with bacteria, and the number of viable bacteria was assessed prior to and after culture. In 28.8% of milk samples, we noted no viable bacteria immediately after collection; this value increased significantly after a 5-hr incubation of milk with cellular components but not with soluble components (48.1 and 28.8%, respectively). After culture with cellular components, the numbers of bacteria (excluding Staphylococcus aureus and Streptococcus uberis) and yeast decreased dramatically, although the differences were not statistically significant. After cultivation with soluble components, only yeasts showed a tendency toward decreased mean viability, whereas the mean bacterial counts of S. uberis and T. pyogenes tended to increase after 5-hr preservation with soluble components. These results suggest that most pathogens in high somatic cell count (SCC) milk decreased during preservation at 15 to 25°C, due to both the cellular components and antimicrobial components in the milk. Particularly, the cellular components more potently reduced bacterial counts during preservation.

  11. Viable but non-culturable Listeria monocytogenes on parsley leaves and absence of recovery to a culturable state.

    Science.gov (United States)

    Dreux, N; Albagnac, C; Federighi, M; Carlin, F; Morris, C E; Nguyen-the, C

    2007-10-01

    To investigate the presence of viable but non-culturable Listeria monocytogenes during survival on parsley leaves under low relative humidity (RH) and to evaluate the ability of L. monocytogenes to recover from VBNC to culturable state under satured humidity. Under low RH (47-69%) on parsley leaves, the initial number of L. monocytogenes populations counted on non selective media (10(9) L. monocytogenes per leaf on TSA) was reduced by 6 log10 scales in 15 days, whereas number of viable L. monocytogenes counted under the microscope was reduced by 3-4 log10 scales, indicating the presence of VBNC cells. This was demonstrated on three L. monocytogenes strains (EGDe, Bug 1995 and LmP60). Changing from low to 100% RH permitted an increase of the culturable counts of L. monocytogenes and this growth was observed only when residual culturable cells were present. Moreover, VBNC L. monocytogenes inoculated on parsley leaves did not become culturable after incubation under 100% RH. Dry conditions induced VBNC L. monocytogenes on parsley leaves but these VBNC were likely unable to recover culturability after transfer to satured humidity. Enumeration on culture media presumably under-estimates the number of viable L. monocytogenes on fresh produce after exposure to low RH.

  12. Expression of adhesion molecules, chemokines and matrix metallo- proteinases (MMPs) in viable and degenerating stage of Taenia solium metacestode in swine neurocysticercosis.

    Science.gov (United States)

    Singh, Satyendra K; Singh, Aloukick K; Prasad, Kashi N; Singh, Amrita; Singh, Avinash; Rai, Ravi P; Tripathi, Mukesh; Gupta, Rakesh K; Husain, Nuzhat

    2015-11-30

    Neurocysticercosis (NCC) is a parasitic infection of central nervous system (CNS). Expression of adhesion molecules, chemokines and matrix metalloproteinases (MMPs) were investigated on brain tissues surrounding viable (n=15) and degenerating cysticerci (n=15) of Taenia solium in swine by real-time RT-PCR and ELISA. Gelatin gel zymography was performed for MMPs activity. ICAM-1 (intercellular adhesion molecule-1), E-selectin, MIP-1α (macrophage inflammatory protein-1α), Eotaxin-1 and RANTES (regulated on activation, normal T cell expressed and secreted) were associated with degenerating cysticerci (cysts). However, VCAM-1 (vascular cell adhesion molecule-1), MCP-1 (monocyte chemotactic protein-1), MMP-2 and MMP-9 were associated with both viable and degenerating cysts. In conclusion, viable and degenerating cysticerci have different immune molecule profiles and role of these molecules in disease pathogenesis needs to be investigated. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Regulation of Viable and Optimal Cohorts

    Energy Technology Data Exchange (ETDEWEB)

    Aubin, Jean-Pierre, E-mail: aubin.jp@gmail.com [VIMADES (Viabilité, Marchés, Automatique, Décisions) (France)

    2015-10-15

    This study deals with the evolution of (scalar) attributes (resources or income in evolutionary demography or economics, position in traffic management, etc.) of a population of “mobiles” (economic agents, vehicles, etc.). The set of mobiles sharing the same attributes is regarded as an instantaneous cohort described by the number of its elements. The union of instantaneous cohorts during a mobile window between two attributes is a cohort. Given a measure defining the number of instantaneous cohorts, the accumulation of the mobile attributes on a evolving mobile window is the measure of the cohort on this temporal mobile window. Imposing accumulation constraints and departure conditions, this study is devoted to the regulation of the evolutions of the attributes which are1.viable in the sense that the accumulations constraints are satisfied at each instant;2.and, among them, optimal, in the sense that both the duration of the temporal mobile window is maximum and that the accumulation on this temporal mobile window is the largest viable one. This value is the “accumulation valuation” function. Viable and optimal evolutions under accumulation constraints are regulated by an “implicit Volterra integro-differential inclusion” built from the accumulation valuation function, solution to an Hamilton–Jacobi–Bellman partial differential equation under constraints which is constructed for this purpose.

  14. Sorption and precipitation of Mn2+ by viable and autoclaved Shewanella putrefaciens: Effect of contact time

    KAUST Repository

    Chubar, Natalia

    2013-01-01

    The sorption of Mn(II) by viable and inactivated cells of Shewanella putrefaciens, a non-pathogenic, facultative anaerobic, gram-negative bacterium characterised as a Mn(IV) and Fe(III) reducer, was studied under aerobic conditions, as a function of pH, bacterial density and metal loading. During a short contact time (3-24h), the adsorptive behaviour of live and dead bacteria toward Mn(II) was sufficiently similar, an observation that was reflected in the studies on adsorption kinetics at various metal loadings, effects of pH, bacteria density, isotherms and drifting of pH during adsorption. Continuing the experiment for an additional 2-30days demonstrated that the Mn(II) sorption by suspensions of viable and autoclaved cells differed significantly from one another. The sorption to dead cells was characterised by a rapid equilibration and was described by an isotherm. In contrast, the sorption (uptake) to live bacteria exhibited a complex time-dependent uptake. This uptake began as adsorption and ion exchange processes followed by bioprecipitation, and it was accompanied by the formation of polymeric sugars (EPS) and the release of dissolved organic substances. FTIR, EXAFS/XANES and XPS demonstrated that manganese(II) phosphate was the main precipitate formed in 125ml batches, which is the first evidence of the ability of microbes to synthesise manganese phosphates. XPS and XANES spectra did not detect Mn(II) oxidation. Although the release of protein-like compounds by the viable bacteria increased in the presence of Mn2+ (and, by contrast, the release of carbohydrates did not change), electrochemical analyses did not indicate any aqueous complexation of Mn(II) by the organic ligands. © 2012 Elsevier Ltd.

  15. Identification of Viable Helicobacter pylori in Drinking Water Supplies by Cultural and Molecular Techniques.

    Science.gov (United States)

    Santiago, Paula; Moreno, Yolanda; Ferrús, M Antonía

    2015-08-01

    Helicobacter pylori is one of the most common causes of chronic bacterial infection in humans, directly related to peptic ulcer and gastric cancer. It has been suggested that H. pylori can be acquired through different transmission routes, including water. In this study, culture and qPCR were used to detect and identify the presence of H. pylori in drinking water. Furthermore, the combined techniques PMA-qPCR and DVC-FISH were applied for detection of viable cells of H. pylori. Among 24 drinking water samples, 16 samples were positive for the presence of H. pylori, but viable cells were only detected in six samples. Characteristic colonies, covered by a mass of bacterial unspecific growth, were observed on selective agar plates from an only sample, after enrichment. The mixed culture was submitted to DVC-FISH and qPCR analysis, followed by sequencing of the amplicons. Molecular techniques confirmed the growth of H. pylori on the agar plate. Our results demonstrate for the first time that H. pylori can survive and be potentially infective in drinking water, showing that water distribution systems could be a potential route for H. pylori transmission. © 2015 John Wiley & Sons Ltd.

  16. Nutrient regulation by continuous feeding removes limitations on cell yield in the large-scale expansion of Mammalian cell spheroids.

    Directory of Open Access Journals (Sweden)

    Bradley P Weegman

    Full Text Available Cellular therapies are emerging as a standard approach for the treatment of several diseases. However, realizing the promise of cellular therapies across the full range of treatable disorders will require large-scale, controlled, reproducible culture methods. Bioreactor systems offer the scale-up and monitoring needed, but standard stirred bioreactor cultures do not allow for the real-time regulation of key nutrients in the medium. In this study, β-TC6 insulinoma cells were aggregated and cultured for 3 weeks as a model of manufacturing a mammalian cell product. Cell expansion rates and medium nutrient levels were compared in static, stirred suspension bioreactors (SSB, and continuously fed (CF SSB. While SSB cultures facilitated increased culture volumes, no increase in cell yields were observed, partly due to limitations in key nutrients, which were consumed by the cultures between feedings, such as glucose. Even when glucose levels were increased to prevent depletion between feedings, dramatic fluctuations in glucose levels were observed. Continuous feeding eliminated fluctuations and improved cell expansion when compared with both static and SSB culture methods. Further improvements in growth rates were observed after adjusting the feed rate based on calculated nutrient depletion, which maintained physiological glucose levels for the duration of the expansion. Adjusting the feed rate in a continuous medium replacement system can maintain the consistent nutrient levels required for the large-scale application of many cell products. Continuously fed bioreactor systems combined with nutrient regulation can be used to improve the yield and reproducibility of mammalian cells for biological products and cellular therapies and will facilitate the translation of cell culture from the research lab to clinical applications.

  17. Nutrient Regulation by Continuous Feeding Removes Limitations on Cell Yield in the Large-Scale Expansion of Mammalian Cell Spheroids

    Science.gov (United States)

    Weegman, Bradley P.; Nash, Peter; Carlson, Alexandra L.; Voltzke, Kristin J.; Geng, Zhaohui; Jahani, Marjan; Becker, Benjamin B.; Papas, Klearchos K.; Firpo, Meri T.

    2013-01-01

    Cellular therapies are emerging as a standard approach for the treatment of several diseases. However, realizing the promise of cellular therapies across the full range of treatable disorders will require large-scale, controlled, reproducible culture methods. Bioreactor systems offer the scale-up and monitoring needed, but standard stirred bioreactor cultures do not allow for the real-time regulation of key nutrients in the medium. In this study, β-TC6 insulinoma cells were aggregated and cultured for 3 weeks as a model of manufacturing a mammalian cell product. Cell expansion rates and medium nutrient levels were compared in static, stirred suspension bioreactors (SSB), and continuously fed (CF) SSB. While SSB cultures facilitated increased culture volumes, no increase in cell yields were observed, partly due to limitations in key nutrients, which were consumed by the cultures between feedings, such as glucose. Even when glucose levels were increased to prevent depletion between feedings, dramatic fluctuations in glucose levels were observed. Continuous feeding eliminated fluctuations and improved cell expansion when compared with both static and SSB culture methods. Further improvements in growth rates were observed after adjusting the feed rate based on calculated nutrient depletion, which maintained physiological glucose levels for the duration of the expansion. Adjusting the feed rate in a continuous medium replacement system can maintain the consistent nutrient levels required for the large-scale application of many cell products. Continuously fed bioreactor systems combined with nutrient regulation can be used to improve the yield and reproducibility of mammalian cells for biological products and cellular therapies and will facilitate the translation of cell culture from the research lab to clinical applications. PMID:24204645

  18. Viable suspensions of maize (Zea mays L.) pollen with exogenous DNA

    Energy Technology Data Exchange (ETDEWEB)

    Broglia, M. [ENEA, Centro Ricerche Casaccia, Rome (Italy). Dip. Innovazione

    1996-12-01

    A viable suspension of maize pollen in aqueous medium containing exogenous DNA would be a suitable tool in attempting maize genetic transformation via pollen grains by different techniques. In this work the effects of addition of DNA to hypertonic aqueous media able to preserve maize pollen viability were investigated. An almost total loss of viability was found when pollen was incubated with native DNA in water or sucrose medium due to the immediate sticking of DNA on the pollen wall. Calcium in the incubation medium avoided DNA sticking preserving pollen fertilization ability. Pre-washing of pollen in hypertonic sucrose solution was proved to remove DNA binding components from the pollen wall. PEG 20%, that is known to inhibit pollen, and silk nucleases, was also used instead of sucrose, without any reduction in the seed-set yields.

  19. Enhanced spermatogonial stem cell killing and reduced translocation yield from X-irradiated 101/H mice

    Energy Technology Data Exchange (ETDEWEB)

    Cattanach, B M; Kirk, M J

    1987-01-01

    The spermatogonial stem cells of 101/H mice have been found to be more sensitive to killing by acute X-ray doses than those of the 'standard' C3H/HeH x 101/H F/sub 1/ hybrid. Duration of the sterile period was longer throughout the 0.5-8.0-Gy dose range tested and 'recovered' testis weights, taken after recovery of fertility, were more severely reduced. The shapes of the sterile period dose-response curves were similar, but with the 101/H mice the plateau occurred at 3-5 Gy, rather than at 6 Gy. An equivalent observation was made with the testis weight data. The translocation dose-response curve was bell-shaped, as previously found with the hybrid, but yields were lower at all but the lowest doses. Notably, peak yields occurred at 3-5 Gy, rather than at 6 Gy. The altered stem cell killing and genetic responses may be explained either by a higher proportion of radiosensitive cells in the heterogeneous stem cell population or by a higher ratio of cell killing to recoverable chromosome damage which might imply a reduced repair capacity. (Auth.). 43 refs.; 5 figs.; 5 tabs.

  20. Combined evaluation of rest-redistribution thallium-201 tomography and low-dose dobutamine echocardiography enhances the identification of viable myocardium in patients with chronic coronary artery disease

    International Nuclear Information System (INIS)

    Pace, L.; Cuocolo, A.; Salvatore, M.; Perrone-Filardi, P.; Prastaro, M.; Vezzuto, P.; Crisci, T.; Dellegrottaglie, S.; Piscione, F.; Chiariello, M.; Mainenti, P.P.; Varrone, A.

    1998-01-01

    The purpose of this study was to evaluate whether combined evaluation by discriminant analysis of rest-redistribution thallium-201 tomography and low-dose dobutamine echocardiography enhances the accuracy in identifying viable myocardium in patients with chronic coronary artery disease. Rest-redistribution 201 Tl has high sensitivity but low specificity in identifying viable myocardium, while the opposite is true for low-dose dobutamine echocardiography. Forty-six patients underwent low-dose dobutamine echocardiography and rest-redistribution 201 Tl tomography on the same day. Rest echocardiography was repeated at least 30 days (mean 40±20) after myocardial revascularization. Discriminant analysis was applied to the results of 201 Tl tomography and dobutamine echocardiography to classify a/dyskinetic segments as viable or non-viable. In 92 a/dyskinetic segments that were revascularized, rest-redistribution 201 Tl tomography yielded an accuracy of 75%, while the accuracy of dobutamine echocardiography was 70% (P 201 Tl imaging are useful and complementary techniques for identifying viable myocardium in patients with chronic coronary artery disease. Combined evaluation by discriminant analysis significantly improves accuracy, although the cost-effectiveness of such an approach remains to be determined. (orig.)

  1. Promoting resuscitation of viable but nonculturable cells of Vibrio harveyi by a resuscitation-promoting factor-like protein YeaZ.

    Science.gov (United States)

    Li, Y; Chen, J; Zhao, M; Yang, Z; Yue, L; Zhang, X

    2017-02-01

    To demonstrate the resuscitation-promoting activities of recombinant YeaZ from Vibrio harveyi SF-1. The gene of resuscitation-promoting factor YeaZ was cloned from genomic DNA of V. harveyi SF-1. The gene was expressed in Escherichia coli, and the expressed protein was purified by Ni 2+ -affinity chromatography. A yeaZ mutant was constructed by using the suicide plasmid pNQ705 with homologous recombination. Disruption of yeaZ did not affect cell growth significantly in 2216 E broth at 28°C. The wild-type and mutant viable but nonculturable (VBNC) cells could be resuscitated by temperature upshift method. In addition, the recombinant YeaZ increased the culturable counts from 1·27 × 10 4  CFU per ml and 1·99 × 10 4 CFU per ml to 2·88 × 10 5  CFU per ml and 4·59 × 10 5 CFU per ml, respectively. After the VBNC cells of wild-type and mutant cells were maintained at 4°C for 120 days, no resuscitation was obtained by temperature upshift method, but addition of the recombinant YeaZ promoted the resuscitation of the wild-type and mutant cells, with the culturable cell counts of 1·13 × 10 3 and 1·44 × 10 3 CFU per ml, respectively. Disruption of yeaZ decreased the virulence of V. harveyi in zebrafish. The lethal dose 50% of the yeaZ null mutant was more than 10-fold higher than that of the wild-type cells. The recombinant YeaZ could efficiently promote resuscitation of the wild-type and mutant cells of V. harveyi from VBNC to culturable state. The protein also promoted resuscitation of the VBNC wild-type and mutant cells, which were maintained at 4°C for 120 days and not recovered by temperature upshift method. Disruption of yeaZ decreased the virulence of V. harveyi in zebrafish. Here, we show clear evidence of a resuscitation-promoting factor YeaZ of V. harveyi and the roles in resuscitation of the VBNC cells and its pathogenicity. © 2016 The Society for Applied Microbiology.

  2. Computer tomography colonography participation and yield in patients under surveillance for 6-9 mm polyps in a population-based screening trial

    NARCIS (Netherlands)

    Tutein Nolthenius, Charlotte J.; Boellaard, Thierry N.; de Haan, Margriet C.; Nio, C. Yung; Thomeer, Maarten G. J.; Bipat, Shandra; Montauban van Swijndregt, Alexander D.; van de Vijver, Marc J.; Biermann, Katharina; Kuipers, Ernst J.; Dekker, Evelien; Stoker, Jaap

    2016-01-01

    Surveillance CT colonography (CTC) is a viable option for 6-9 mm polyps at CTC screening for colorectal cancer. We established participation and diagnostic yield of surveillance and determined overall yield of CTC screening. In an invitational CTC screening trial 82 of 982 participants harboured 6-9

  3. Particle debonding using different yield criteria

    DEFF Research Database (Denmark)

    Legarth, Brian Nyvang; Kuroda, Mitsutoshi

    2004-01-01

    Effects of plastic anisotropy in relation to debonding of rigid inclusions embedded in an elastic-viscoplastic metal are studied. Full finite strain analyses are carried out for plane cells assuming plane stress or plane strain. The overall stress strain response is calculated, when the cell......-matrix debonding. Keeping all material parameters fixed, the material response of the plane strain cell is considerably affected, due to debonding at a much reduced overall plastic strain compared to the corresponding plane stress cell. (C) 2004 Elsevier SAS. All rights reserved....... extent and shape of the particular yield function considered. The required overall straining of the cell for debonding initiation is related to the extent of the yield surfaces, since a high yield stress promotes debonding. Additionally, the maximum overall stress level for the cell is lower for the Hill...

  4. Relating Nanoscale Accessibility within Plant Cell Walls to Improved Enzyme Hydrolysis Yields in Corn Stover Subjected to Diverse Pretreatments.

    Science.gov (United States)

    Crowe, Jacob D; Zarger, Rachael A; Hodge, David B

    2017-10-04

    Simultaneous chemical modification and physical reorganization of plant cell walls via alkaline hydrogen peroxide or liquid hot water pretreatment can alter cell wall structural properties impacting nanoscale porosity. Nanoscale porosity was characterized using solute exclusion to assess accessible pore volumes, water retention value as a proxy for accessible water-cell walls surface area, and solute-induced cell wall swelling to measure cell wall rigidity. Key findings concluded that delignification by alkaline hydrogen peroxide pretreatment decreased cell wall rigidity and that the subsequent cell wall swelling resulted increased nanoscale porosity and improved enzyme binding and hydrolysis compared to limited swelling and increased accessible surface areas observed in liquid hot water pretreated biomass. The volume accessible to a 90 Å dextran probe within the cell wall was found to be correlated to both enzyme binding and glucose hydrolysis yields, indicating cell wall porosity is a key contributor to effective hydrolysis yields.

  5. Asouzu's Complementary Ontology as a Foundation for a Viable ...

    African Journals Online (AJOL)

    This paper on “Asouzu's Complementary Ontology as a foundation for a viable Ethic of the Environment”, posits that an ethic of the environment can be seen as viable if it considers the whole of reality as ontologically relevant. This point of view would free environmental ethics of anthropocentric bias and its attendant ...

  6. Macrophage conditioned medium induced cellular network formation in MCF-7 cells through enhanced tunneling nanotube formation and tunneling nanotube mediated release of viable cytoplasmic fragments

    International Nuclear Information System (INIS)

    Patheja, Pooja; Sahu, Khageswar

    2017-01-01

    Infiltrating macrophages in tumor microenvironment, through their secreted cytokines and growth factors, regulate several processes of cancer progression such as cancer cell survival, proliferation, invasion, metastasis and angiogenesis. Recently, intercellular cytoplasmic bridges between cancer cells referred as tunneling nanotubes (TNTs) have been recognized as novel mode of intercellular communication between cancer cells. In this study, we investigated the effect of inflammatory mediators present in conditioned medium derived from macrophages on the formation of TNTs in breast adenocarcinoma cells MCF-7. Results show that treatment with macrophage conditioned medium (MφCM) not only enhanced TNT formation between cells but also stimulated the release of independently migrating viable cytoplasmic fragments, referred to as microplasts, from MCF-7 cells. Time lapse microscopy revealed that microplasts were released from parent cancer cells in extracellular space through formation of TNT-like structures. Mitochondria, vesicles and cytoplasm could be transferred from parent cell body to microplasts through connecting TNTs. The microplasts could also be resorbed into the parent cell body by retraction of the connecting TNTs. Microplast formation inhibited in presence cell migration inhibitor, cytochalasin-B. Notably by utilizing migratory machinery within microplasts, distantly located MCF-7 cells formed several TNT based intercellular connections, leading to formation of physically connected network of cells. Together, these results demonstrate novel role of TNTs in microplast formation, novel modes of TNT formation mediated by microplasts and stimulatory effect of MφCM on cellular network formation in MCF-7 cells mediated through enhanced TNT and microplast formation.

  7. Macrophage conditioned medium induced cellular network formation in MCF-7 cells through enhanced tunneling nanotube formation and tunneling nanotube mediated release of viable cytoplasmic fragments

    Energy Technology Data Exchange (ETDEWEB)

    Patheja, Pooja, E-mail: pooja.patheja8@gmail.com [Laser Biomedical Applications Section, Raja Ramanna Centre for Advanced Technology, Indore 452013, Madhya Pradesh (India); Homi Bhabha National Institute, Training School Complex, Anushaktinagar, Mumbai 400094, Maharashtra (India); Sahu, Khageswar [Laser Biomedical Applications Section, Raja Ramanna Centre for Advanced Technology, Indore 452013, Madhya Pradesh (India)

    2017-06-15

    Infiltrating macrophages in tumor microenvironment, through their secreted cytokines and growth factors, regulate several processes of cancer progression such as cancer cell survival, proliferation, invasion, metastasis and angiogenesis. Recently, intercellular cytoplasmic bridges between cancer cells referred as tunneling nanotubes (TNTs) have been recognized as novel mode of intercellular communication between cancer cells. In this study, we investigated the effect of inflammatory mediators present in conditioned medium derived from macrophages on the formation of TNTs in breast adenocarcinoma cells MCF-7. Results show that treatment with macrophage conditioned medium (MφCM) not only enhanced TNT formation between cells but also stimulated the release of independently migrating viable cytoplasmic fragments, referred to as microplasts, from MCF-7 cells. Time lapse microscopy revealed that microplasts were released from parent cancer cells in extracellular space through formation of TNT-like structures. Mitochondria, vesicles and cytoplasm could be transferred from parent cell body to microplasts through connecting TNTs. The microplasts could also be resorbed into the parent cell body by retraction of the connecting TNTs. Microplast formation inhibited in presence cell migration inhibitor, cytochalasin-B. Notably by utilizing migratory machinery within microplasts, distantly located MCF-7 cells formed several TNT based intercellular connections, leading to formation of physically connected network of cells. Together, these results demonstrate novel role of TNTs in microplast formation, novel modes of TNT formation mediated by microplasts and stimulatory effect of MφCM on cellular network formation in MCF-7 cells mediated through enhanced TNT and microplast formation.

  8. Induction of Viable but Nonculturable Salmonella in Exponentially Grown Cells by Exposure to a Low-Humidity Environment and Their Resuscitation by Catalase.

    Science.gov (United States)

    Morishige, Yuta; Koike, Atsushi; Tamura-Ueyama, Ai; Amano, Fumio

    2017-02-01

    Salmonella is a major cause of foodborne disease that sometimes occurs in massive outbreaks around the world. This pathogen is tolerant of low-humidity conditions. We previously described a method for induction of viable but nonculturable (VBNC) Salmonella enterica serovar Enteritidis by treatment with hydrogen peroxide (H 2 O 2 ) and subsequent resuscitation with 0.3 mM sodium pyruvate. Here, we report a new method for the induction of the VBNC state in Salmonella Enteritidis cells, one involving dehydration. Exposure of Salmonella Enteritidis cells to dehydration stress under poor nutritional conditions (0.9% [wt/vol] NaCl) and 10 to 20% relative humidity at room temperature decreased the presence of culturable population to 0.0067%, but respiratory and glucose uptake active populations were maintained at 0.46 and 1.12%, respectively, meaning that approximately 1% may have entered the VBNC state. Furthermore, these VBNC cells could be resuscitated to acquire culturability by incubation with catalase in M9 minimal medium without glucose in a manner dependent on the dose of catalase but not sodium pyruvate. These results suggest that a low-humidity environment could cause Salmonella Enteritidis cells to enter the VBNC state and the cells could then be resuscitated for growth by treatment with catalase, suggesting a potential risk of Salmonella Enteritidis to survive in low water activity foods in the VBNC state and to start regrowth for foodborne illness.

  9. Improved Performance in Mammalian Cell Perfusion Cultures by Growth Inhibition.

    Science.gov (United States)

    Wolf, Moritz K F; Closet, Aurélie; Bzowska, Monika; Bielser, Jean-Marc; Souquet, Jonathan; Broly, Hervé; Morbidelli, Massimo

    2018-05-21

    Mammalian cell perfusion cultures represent a promising alternative to the current fed-batch technology for the production of various biopharmaceuticals. Long-term operation at a fixed viable cell density (VCD) requires a viable culture and a constant removal of excessive cells. Product loss in the cell removing bleed stream deteriorates the process yield. In this study, the authors investigate the use of chemical and environmental growth inhibition on culture performance by either adding valeric acid (VA) to the production media or by reducing the culture temperature (33.0 °C) with respect to control conditions (36.5 °C, no VA). Low temperature significantly reduces cellular growth, thus, resulting in lower bleed rates accompanied by a reduced product loss of 11% compared to 26% under control conditions. Additionally, the cell specific productivity of the target protein improves and maintained stable leading to media savings per mass of product. VA shows initially an inhibitory effect on cellular growth. However, cells seemed to adapt to the presence of the inhibitor resulting in a recovery of the cellular growth. Cell cycle and Western blot analyses support the observed results. This work underlines the role of temperature as a key operating variable for the optimization of perfusion cultures. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Influence of yield surface curvature on the macroscopic yielding and ductile failure of isotropic porous plastic materials

    Science.gov (United States)

    Dæhli, Lars Edvard Bryhni; Morin, David; Børvik, Tore; Hopperstad, Odd Sture

    2017-10-01

    Numerical unit cell models of an approximative representative volume element for a porous ductile solid are utilized to investigate differences in the mechanical response between a quadratic and a non-quadratic matrix yield surface. A Hershey equivalent stress measure with two distinct values of the yield surface exponent is employed as the matrix description. Results from the unit cell calculations are further used to calibrate a heuristic extension of the Gurson model which incorporates effects of the third deviatoric stress invariant. An assessment of the porous plasticity model reveals its ability to describe the unit cell response to some extent, however underestimating the effect of the Lode parameter for the lower triaxiality ratios imposed in this study when compared to unit cell simulations. Ductile failure predictions by means of finite element simulations using a unit cell model that resembles an imperfection band are then conducted to examine how the non-quadratic matrix yield surface influences the failure strain as compared to the quadratic matrix yield surface. Further, strain localization predictions based on bifurcation analyses and imperfection band analyses are undertaken using the calibrated porous plasticity model. These simulations are then compared to the unit cell calculations in order to elucidate the differences between the various modelling strategies. The current study reveals that strain localization analyses using an imperfection band model and a spatially discretized unit cell are in reasonable agreement, while the bifurcation analyses predict higher strain levels at localization. Imperfection band analyses are finally used to calculate failure loci for the quadratic and the non-quadratic matrix yield surface under a wide range of loading conditions. The underlying matrix yield surface is demonstrated to have a pronounced influence on the onset of strain localization.

  11. Honey yield of different commercial apiaries treated with Lactobacillus salivarius A3iob, a new bee-probiotic strain.

    Science.gov (United States)

    Fanciotti, M Novicov; Tejerina, M; Benítez-Ahrendts, M R; Audisio, M C

    2018-02-27

    The main objective of this study was to determine the impact of Lactobacillus salivarius A3iob, a honey bee gut-associated strain (GenBank code access KX198010), on honey yield. Independent assays were conducted from May to September 2014 and 2015, in three commercial apiaries: Tilquiza, El Carmen and Yala, all located in north-western Argentina. Local Apis mellifera L. bees were kept in standard Langstroth hives; treated hives were fed once a month with 1×10 5 cfu/ml viable Lactobacillus cells, administered to the bees through a Doolittle-type feeder in 125 g/l sucrose syrup. Control hives were only given the syrup mixed with MRS sterile broth. The main honey harvest was done in December in all groups and we found that there was an overall increase in honey yield from the treated hives. In 2014, all treated hives produced between 2.3 to 6.5 times more honey than the controls. However, in 2015, higher honey average yields in the treated hives at El Carmen and Yala were obtained, yet not at Tilquiza, because of a slight mishap. They experienced the swarming of several bee colonies due to a higher number of bees without appropriate management, which caused the control group to yield more honey compared to the hives fed with Lactobacillus. Interestingly, at El Carmen, two honey harvests were recorded: one in winter and another in summer (July and December 2015, respectively). This unexpected result arose from the particular flora of the region, mainly Tithonia tubaeformis, which blooms in winter. L. salivarius A3iob cells prove to be a natural alternative that will positively impact the beekeepers' economy by providing a higher honey yield.

  12. Texas Native Plants Yield Compounds with Cytotoxic Activities against Prostate Cancer Cells.

    Science.gov (United States)

    Shaffer, Corena V; Cai, Shengxin; Peng, Jiangnan; Robles, Andrew J; Hartley, Rachel M; Powell, Douglas R; Du, Lin; Cichewicz, Robert H; Mooberry, Susan L

    2016-03-25

    There remains a critical need for more effective therapies for the treatment of late-stage and metastatic prostate cancers. Three Texas native plants yielded three new and three known compounds with antiproliferative and cytotoxic activities against prostate cancer cells with IC50 values in the range of 1.7-35.0 μM. A new sesquiterpene named espadalide (1), isolated from Gochnatia hypoleuca, had low micromolar potency and was highly effective in clonogenic assays. Two known bioactive germacranolides (2 and 3) were additionally isolated from G. hypoleuca. Dalea frutescens yielded two new isoprenylated chalcones, named sanjuanolide (4) and sanjoseolide (5), and the known sesquiterpenediol verbesindiol (6) was isolated from Verbesina virginica. Mechanistic studies showed that 1-4 caused G2/M accumulation and the formation of abnormal mitotic spindles. Tubulin polymerization assays revealed that 4 increased the initial rate of tubulin polymerization, but did not change total tubulin polymer levels, and 1-3 had no effects on tubulin polymerization. Despite its cytotoxic activity, compound 6 did not initiate changes in cell cycle distribution and has a mechanism of action different from the other compounds. This study demonstrates that new compounds with significant biological activities germane to unmet oncological needs can be isolated from Texas native plants.

  13. Introducing a New Experimental Islet Transplantation Model using Biomimetic Hydrogel and a Simple High Yield Islet Isolation Technique.

    Science.gov (United States)

    Mohammadi Ayenehdeh, Jamal; Niknam, Bahareh; Hashemi, Seyed Mahmoud; Rahavi, Hossein; Rezaei, Nima; Soleimani, Masoud; Tajik, Nader

    2017-07-01

    Islet transplantation could be an ideal alternative treatment to insulin therapy for type 1 diabetes Mellitus (T1DM). This clinical and experimental field requires a model that covers problems such as requiring a large number of functional and viable islets, the optimal transplantation site, and the prevention of islet dispersion. Hence, the methods of choice for isolation of functional islets and transplantation are crucial. The present study has introduced an experimental model that overcomes some critical issues in islet transplantation, including in situ pancreas perfusion by digestive enzymes through common bile duct. In comparison with conventional methods, we inflated the pancreas in Petri dishes with only 1 ml collagenase type XI solution, which was followed by hand-picking isolation or Ficoll gradient separation to purify the islets. Then we used a hydrogel composite in which the islets were embedded and transplanted into the peritoneal cavity of the streptozotocin-induced diabetic C57BL/6 mice. As compared to the yield of the classical methods, in our modified technique, the mean yield of isolation was about 130-200 viable islets/mouse pancreas. In vitro glucose-mediated insulin secretion assay indicated an appropriate response in isolated islets. In addition, data from in vivo experiments revealed that the allograft remarkably maintained blood glucose levels under 400 mg/dl and hydrogel composite prevents the passage of immune cells. In the model presented here, the rapid islet isolation technique and the application of biomimetic hydrogel wrapping of islets could facilitate islet transplantation procedures.

  14. Adipose tissue-derived mesenchymal stem cell yield and growth characteristics are affected by the tissue-harvesting procedure

    NARCIS (Netherlands)

    Oedayrajsingh-Varma, M. J.; van Ham, S. M.; Knippenberg, M.; Helder, M. N.; Klein-Nulend, J.; Schouten, T. E.; Ritt, M. J. P. F.; van Milligen, F. J.

    2006-01-01

    Adipose tissue contains a stromal vascular fraction that can be easily isolated and provides a rich source of adipose tissue-derived mesenchymal stem cells (ASC). These ASC are a potential source of cells for tissue engineering. We studied whether the yield and growth characteristics of ASC were

  15. A Novel Application for Low Frequency Electrochemical Impedance Spectroscopy as an Online Process Monitoring Tool for Viable Cell Concentrations

    Directory of Open Access Journals (Sweden)

    Christoph Slouka

    2016-11-01

    Full Text Available New approaches in process monitoring during industrial fermentations are not only limited to classical pH, dO2 and offgas analysis, but use different in situ and online sensors based on different physical principles to determine biomass, product quality, lysis and far more. One of the very important approaches is the in situ accessibility of viable cell concentration (VCC. This knowledge provides increased efficiency in monitoring and controlling strategies during cultivations. Electrochemical impedance spectroscopy—EIS—is used to monitor biomass in a fermentation of E. coli BL21(DE3, producing a recombinant protein using a fed batch-based approach. Increases in the double layer capacitance (Cdl, determined at frequencies below 1 kHz, are proportional to the increase of biomass in the batch and fed batch phase, monitored in offline and online modes for different cultivations. A good correlation of Cdl with cell density is found and in order to get an appropriate verification of this method, different state-of-the-art biomass measurements are performed and compared. Since measurements in this frequency range are largely determined by the double layer region between the electrode and media, rather minor interferences with process parameters (aeration, stirring are to be expected. It is shown that impedance spectroscopy at low frequencies is a powerful tool for cultivation monitoring.

  16. Relation between cell cycle and yield of aberrations observed in irradiated human lymphocytes

    International Nuclear Information System (INIS)

    Leonard, A.; Decat, G.

    1979-01-01

    The bromodeoxyuridine-Giemsa technique has been used to study systematically the incidence of cells in first or subsequent mitoses at differrent fixation times of human lymphocyte control cultures as well as the influence of ionizing radiations on cell kinetics. Second divisions appear (3%) in cultures harvested 48 h after initiation. In 72 h cultures 40% of the dividing cells are in second and 33% in third division. Administration of 200 rads of X-rays before PHA stimulation results in a mitotic delay but does not increase the incidence of SCE. The yield of dicentrics after an exposure to 200 rads was the same for all cells in first mitosis regardless of fixation time. These results demonstrate that there is no evidence for the existence of sensitive subpopulations that could be distinguished by the time of the first mitotic division following stimulation. (author)

  17. Critical review of the molecular design progress in non-fullerene electron acceptors towards commercially viable organic solar cells

    KAUST Repository

    Wadsworth, Andrew; Moser, Maximilian; Marks, Adam; Little, Mark S.; Gasparini, Nicola; Brabec, Christoph J.; Baran, Derya; McCulloch, Iain

    2018-01-01

    Fullerenes have formed an integral part of high performance organic solar cells over the last 20 years, however their inherent limitations in terms of synthetic flexibility, cost and stability have acted as a motivation to develop replacements; the so-called non-fullerene electron acceptors. A rapid evolution of such materials has taken place over the last few years, yielding a number of promising candidates that can exceed the device performance of fullerenes and provide opportunities to improve upon the stability and processability of organic solar cells. In this review we explore the structure-property relationships of a library of non-fullerene acceptors, highlighting the important chemical modifications that have led to progress in the field and provide an outlook for future innovations in electron acceptors for use in organic photovoltaics.

  18. Critical review of the molecular design progress in non-fullerene electron acceptors towards commercially viable organic solar cells

    KAUST Repository

    Wadsworth, Andrew

    2018-04-26

    Fullerenes have formed an integral part of high performance organic solar cells over the last 20 years, however their inherent limitations in terms of synthetic flexibility, cost and stability have acted as a motivation to develop replacements; the so-called non-fullerene electron acceptors. A rapid evolution of such materials has taken place over the last few years, yielding a number of promising candidates that can exceed the device performance of fullerenes and provide opportunities to improve upon the stability and processability of organic solar cells. In this review we explore the structure-property relationships of a library of non-fullerene acceptors, highlighting the important chemical modifications that have led to progress in the field and provide an outlook for future innovations in electron acceptors for use in organic photovoltaics.

  19. Sorting of cells of the same size, shape, and cell cycle stage for a single cell level assay without staining

    Directory of Open Access Journals (Sweden)

    Yomo Tetsuya

    2006-06-01

    cell viability but decreases the proliferative ability of the PC12 cells. Conclusion We demonstrated a pretreatment method to collect well-characterized, viable, single cells without using fluorescent labels and without significant damage to the cells. This method is quantitative, rapid, single-step, and yields cells of high purity, making it applicable for a variety of single-cell level analyses.

  20. Macrophage conditioned medium induced cellular network formation in MCF-7 cells through enhanced tunneling nanotube formation and tunneling nanotube mediated release of viable cytoplasmic fragments.

    Science.gov (United States)

    Patheja, Pooja; Sahu, Khageswar

    2017-06-15

    Infiltrating macrophages in tumor microenvironment, through their secreted cytokines and growth factors, regulate several processes of cancer progression such as cancer cell survival, proliferation, invasion, metastasis and angiogenesis. Recently, intercellular cytoplasmic bridges between cancer cells referred as tunneling nanotubes (TNTs) have been recognized as novel mode of intercellular communication between cancer cells. In this study, we investigated the effect of inflammatory mediators present in conditioned medium derived from macrophages on the formation of TNTs in breast adenocarcinoma cells MCF-7. Results show that treatment with macrophage conditioned medium (MɸCM) not only enhanced TNT formation between cells but also stimulated the release of independently migrating viable cytoplasmic fragments, referred to as microplasts, from MCF-7 cells. Time lapse microscopy revealed that microplasts were released from parent cancer cells in extracellular space through formation of TNT-like structures. Mitochondria, vesicles and cytoplasm could be transferred from parent cell body to microplasts through connecting TNTs. The microplasts could also be resorbed into the parent cell body by retraction of the connecting TNTs. Microplast formation inhibited in presence cell migration inhibitor, cytochalasin-B. Notably by utilizing migratory machinery within microplasts, distantly located MCF-7 cells formed several TNT based intercellular connections, leading to formation of physically connected network of cells. Together, these results demonstrate novel role of TNTs in microplast formation, novel modes of TNT formation mediated by microplasts and stimulatory effect of MɸCM on cellular network formation in MCF-7 cells mediated through enhanced TNT and microplast formation. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. On the use of the serial dilution culture method to enumerate viable phytoplankton in natural communities of plankton subjected to ballast water treatment.

    Science.gov (United States)

    Cullen, John J; MacIntyre, Hugh L

    2016-01-01

    Discharge standards for ballast water treatment (BWT) systems are based on concentrations of living cells, for example, as determined with vital stains. Ultraviolet radiation (UV) stops the reproduction of microorganisms without killing them outright; they are living, but not viable, and ecologically as good as dead. Consequently, UV-treated discharge can be compliant with the intent of regulation while failing a live/dead test. An alternative evaluation of BWT can be proposed based on the assessment of viable, rather than living, cells in discharge water. In principle, the serial dilution culture-most probable number (SDC-MPN) method provides the appropriate measure for phytoplankton. But, the method has been criticized, particularly because it is thought that many phytoplankton species cannot be cultured. A review of the literature shows that although SDC-MPN has been used for more than 50 years-generally to identify and count phytoplankton species that cannot be preserved-its application to enumerate total viable phytoplankton seems to be new, putting past criticisms of the method in a different light. Importantly, viable cells need to grow only enough to be detected, not to be brought into sustained culture, and competition between species in a dilution tube is irrelevant as long as the winner is detectable. Thorough consideration of sources of error leads to recommendations for minimizing and quantifying uncertainties by optimizing growth conditions and conducting systematic comparisons. We conclude that with careful evaluation, SDC-MPN is potentially an effective method for assessing the viability of phytoplankton after BWT.

  2. The impact of silicon solar cell architecture and cell interconnection on energy yield in hot & sunny climates

    KAUST Repository

    Haschke, Jan

    2017-03-23

    Extensive knowledge of the dependence of solar cell and module performance on temperature and irradiance is essential for their optimal application in the field. Here we study such dependencies in the most common high-efficiency silicon solar cell architectures, including so-called Aluminum back-surface-field (BSF), passivated emitter and rear cell (PERC), passivated emitter rear totally diffused (PERT), and silicon heterojunction (SHJ) solar cells. We compare measured temperature coefficients (TC) of the different electrical parameters with values collected from commercial module data sheets. While similar TC values of the open-circuit voltage and the short circuit current density are obtained for cells and modules of a given technology, we systematically find that the TC under maximum power-point (MPP) conditions is lower in the modules. We attribute this discrepancy to additional series resistance in the modules from solar cell interconnections. This detrimental effect can be reduced by using a cell design that exhibits a high characteristic load resistance (defined by its voltage-over-current ratio at MPP), such as the SHJ architecture. We calculate the energy yield for moderate and hot climate conditions for each cell architecture, taking into account ohmic cell-to-module losses caused by cell interconnections. Our calculations allow us to conclude that maximizing energy production in hot and sunny environments requires not only a high open-circuit voltage, but also a minimal series-to-load-resistance ratio.

  3. Reduced infectivity of waterborne viable but nonculturable Helicobacter pylori strain SS1 in mice.

    Science.gov (United States)

    Boehnke, Kevin F; Eaton, Kathryn A; Fontaine, Clinton; Brewster, Rebecca; Wu, Jianfeng; Eisenberg, Joseph N S; Valdivieso, Manuel; Baker, Laurence H; Xi, Chuanwu

    2017-08-01

    Helicobacter pylori infection has been consistently associated with lack of access to clean water and proper sanitation, but no studies have demonstrated that the transmission of viable but nonculturable (VBNC) H. pylori can occur from drinking contaminated water. In this study, we used a laboratory mouse model to test whether waterborne VBNCH. pylori could cause gastric infection. We performed five mouse experiments to assess the infectivity of VBNCH. pylori in various exposure scenarios. VBNC viability was examined using Live/Dead staining and Biolog phenotype metabolism arrays. High doses of VBNCH. pylori in water were chosen to test the "worst-case" scenario for different periods of time. One experiment also investigated the infectious capabilities of VBNC SS1 using gavage. Further, immunocompromised mice were exposed to examine infectivity among potentially vulnerable groups. After exposure, mice were euthanized and their stomachs were examined for H. pylori infection using culture and PCR methodology. VBNC cells were membrane intact and retained metabolic activity. Mice exposed to VBNCH. pylori via drinking water and gavage were not infected, despite the various exposure scenarios (immunocompromised, high doses) that might have permitted infection with VBNCH. pylori. The positive controls exposed to viable, culturable H. pylori did become infected. While other studies that have used viable, culturable SS1 via gavage or drinking water exposures to successfully infect mice, in our study, waterborne VBNC SS1 failed to colonize mice under all test conditions. Future studies could examine different H. pylori strains in similar exposure scenarios to compare the relative infectivity of the VBNC vs the viable, culturable state, which would help inform future risk assessments of H. pylori in water. © 2017 The Authors. Helicobacter Published by John Wiley & Sons Ltd.

  4. Co-infusion of autologous adipose tissue derived insulin-secreting mesenchymal stem cells and bone marrow derived hematopoietic stem cells: Viable therapy for type III.C. a diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Umang G Thakkar

    2014-12-01

    Full Text Available Transition from acute pancreatitis to insulin-dependent diabetes mellitus (IDDM is a rare manifestation of primary hyperparathyroidism caused by parathyroid adenoma because of impaired glucose tolerance and suppresses insulin secretion. We report the case of a 26-year-old male with pancreatic diabetes caused by parathyroid adenoma induced chronic pancreatitis. He had serum C-peptide 0.12 ng/ml, glutamic acid decarboxylase antibody 5.0 IU/ml, and glycosylated hemoglobin (HbA1C 8.9%, and required 72 IU/day of biphasic-isophane insulin injection for uncontrolled hyperglycemia. We treated him with his own adipose tissue derived insulin-secreting mesenchymal stem-cells (IS-ADMSC along with his bone marrow derived hematopoietic stem cells (BM-HSC. Autologous IS-ADMSC + BM-HSC were infused into subcutaneous tissue, portal and thymic circulation without any conditioning. Over a follow-up of 27 months, the patient is maintaining fasting and postprandial blood sugar levels of 132 and 165 mg/dl, respectively, with HbA1C 6.8% and requiring 36 IU/day of biphasic-isophane insulin. Co-infusion of IS-ADMSC + BM-HSC offers a safe and viable therapy for type III.C.a Diabetes Mellitus.

  5. Utilization of Cell-Transfer Technique for Molecular Testing on Hematoxylin-Eosin-Stained Sections: A Viable Option for Small Biopsies That Lack Tumor Tissues in Paraffin Block.

    Science.gov (United States)

    Wu, Howard H; Jovonovich, Stephen M; Randolph, Melissa; Post, Kristin M; Sen, Joyashree D; Curless, Kendra; Cheng, Liang

    2016-12-01

    - In some instances the standard method of doing molecular testing from formalin-fixed, paraffin-embedded block is not possible because of limited tissue. Tumor cell-enriched cell-transfer technique has been proven useful for performing immunocytochemistry and molecular testing on cytologic smears. - To establish the cell-transfer technique as a viable option for isolating tumor cells from hematoxylin-eosin (H&E)-stained slides. - Molecular testing was performed by using the cell-transfer technique on 97 archived H&E-stained slides from a variety of different tumors. Results were compared to the conventional method of molecular testing. - Polymerase chain reaction-based molecular testing via the cell-transfer technique was successfully performed on 82 of 97 samples (85%). This included 39 of 47 cases for EGFR, 10 of 11 cases for BRAF, and 33 of 39 cases for KRAS mutations. Eighty-one of 82 cell-transfer technique samples (99%) showed agreement with previous standard method results, including 4 mutations and 35 wild-type alleles for EGFR, 4 mutations and 6 wild-type alleles for BRAF, and 11 mutations and 21 wild-type alleles for KRAS. There was only 1 discrepancy: a cell-transfer technique with a false-negative >KRAS result (wild type versus G12C). - Molecular testing performed on H&E-stained sections via cell-transfer technique is useful when tissue from cell blocks and small surgical biopsy samples is exhausted and the only available material for testing is on H&E-stained slides.

  6. Differential Effect of Viable Versus Necrotic Neutrophils on Mycobacterium tuberculosis Growth and Cytokine Induction in Whole Blood

    Directory of Open Access Journals (Sweden)

    David M. Lowe

    2018-04-01

    Full Text Available Neutrophils exert both positive and negative influences on the host response to tuberculosis, but the mechanisms by which these differential effects are mediated are unknown. We studied the impact of live and dead neutrophils on the control of Mycobacterium tuberculosis using a whole blood bioluminescence-based assay, and assayed supernatant cytokine concentrations using Luminex™ technology and ELISA. CD15+ granulocyte depletion from blood prior to infection with M. tuberculosis-lux impaired control of mycobacteria by 96 h, with a greater effect than depletion of CD4+, CD8+, or CD14+ cells (p < 0.001. Augmentation of blood with viable granulocytes significantly improved control of mycobacteria by 96 h (p = 0.001, but augmentation with necrotic granulocytes had the opposite effect (p = 0.01. Both augmentations decreased supernatant concentrations of tumor necrosis factor and interleukin (IL-12 p40/p70, but necrotic granulocyte augmentation also increased concentrations of IL-10, G-CSF, GM-CSF, and CCL2. Necrotic neutrophil augmentation reduced phagocytosis of FITC-labeled M. bovis BCG by all phagocytes, whereas viable neutrophil augmentation specifically reduced early uptake by CD14+ cells. The immunosuppressive effect of dead neutrophils required necrotic debris rather than supernatant. We conclude that viable neutrophils enhance control of M. tuberculosis in blood, but necrotic neutrophils have the opposite effect—the latter associated with induction of IL-10, growth factors, and chemoattractants. Our findings suggest a mechanism by which necrotic neutrophils may exert detrimental effects on the host response in active tuberculosis.

  7. Mitochondrial respiration in human viable platelets-Methodology and influence of gender, age and storage

    DEFF Research Database (Denmark)

    Sjövall, Fredrik; Ehinger, Johannes K H; Marelsson, Sigurður E

    2013-01-01

    Studying whole cell preparations with intact mitochondria and respiratory complexes has a clear benefit compared to isolated or disrupted mitochondria due to the dynamic interplay between mitochondria and other cellular compartments. Platelet mitochondria have a potential to serve as a source...... of human viable mitochondria when studying mitochondrial physiology and pathogenic mechanisms, as well as for the diagnostics of mitochondrial diseases. The objective of the present study was to perform a detailed evaluation of platelet mitochondrial respiration using high-resolution respirometry. Further...

  8. Articular Cartilage Repair Using Marrow Stimulation Augmented with a Viable Chondral Allograft: 9-Month Postoperative Histological Evaluation

    Directory of Open Access Journals (Sweden)

    James K. Hoffman

    2015-01-01

    Full Text Available Marrow stimulation is frequently employed to treat focal chondral defects of the knee. However, marrow stimulation typically results in fibrocartilage repair tissue rather than healthy hyaline cartilage, which, over time, predisposes the repair to failure. Recently, a cryopreserved viable chondral allograft was developed to augment marrow stimulation. The chondral allograft is comprised of native viable chondrocytes, chondrogenic growth factors, and extracellular matrix proteins within the superficial, transitional, and radial zones of hyaline cartilage. Therefore, host mesenchymal stem cells that infiltrate the graft from the underlying bone marrow following marrow stimulation are provided with the optimal microenvironment to undergo chondrogenesis. The present report describes treatment of a trochlear defect with marrow stimulation augmented with this novel chondral allograft, along with nine month postoperative histological results. At nine months, the patient demonstrated complete resolution of pain and improvement in function, and the repair tissue consisted of 85% hyaline cartilage. For comparison, a biopsy obtained from a patient 8.2 months after treatment with marrow stimulation alone contained only 5% hyaline cartilage. These outcomes suggest that augmenting marrow stimulation with the viable chondral allograft can eliminate pain and improve outcomes, compared with marrow stimulation alone.

  9. A high-yielding, generic fed-batch process for recombinant antibody production of GS-engineered cell lines

    DEFF Research Database (Denmark)

    Fan, Li; Zhao, Liang; Sun, Yating

    2009-01-01

    An animal component-free and chemically defined fed-batch process for GS-engineered cell lines producing recombinant antibodies has been developed. The fed-batch process relied on supplying sufficient nutrients to match their consumption, simultaneously minimizing the accumulation of byproducts....... This generic and high-yielding fed-batch process would shorten development time, and ensure process stability, thereby facilitating the manufacture of therapeutic antibodies by GS-engineered cell lines....

  10. Coal liquefaction becomes viable

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2005-11-15

    In 2003 the May/June issue of CoalTrans International speculated that coal liquefaction would become viable due to falling coal prices. This has not proved the case but the sustained high oil price is sparking new interest. A survey by Energy Intelligence and Marketing Research during November 2005 revealed a growth in the number of projects under development or at the feasibility stage. The article reports projects in China, the USA, Australia, New Zealand, the Philippines and India. China is commissioning the first wave of large liquefaction plants. The key question is whether other countries, particularly the USA, will follow.

  11. Vicia narbonensis-Avena strigosa mixture, a viable alternative in rainfed cropping systems under Mediterranean conditions

    Energy Technology Data Exchange (ETDEWEB)

    Pedraza, V.; Perea, F.; Saavedra, M.; Fuentes, C.; Alcántara, C.

    2017-07-01

    The demand of vegetable protein for animal feed and the need to diversify the crop rotation in rainfed Mediterranean climates has led to study the viability as new forage crop of the Vicia narbonensis-Avena strigosa mixture. Therefore, a 3-year field trial was conducted at two different and representative locations of the area to evaluate the capacity of both species to form a balanced mixture and to define its potential for high yield and forage quality. Different seeding ratios (65:35, 50:50 and 35:65) were compared with their pure stands and the standard mixture Avena sativa-Vicia sativa (65:35). Forage mixtures establishment and growth varied according to the environmental conditions with a negative influence of a dry year 1 for legumes (<300 mm) and a wet year 2 for oats (>630 mm). However, competition ratio indicated that there were not significant differences between mixed treatments, displaying intermediate ground coverage values, dry matter yield and crude protein regarding pure stands. Higher dry matter yield than control mixture in addition to similar crude protein, acid-detergent fibre, neutral-detergent fibre and digestible dry matter values, were produced at 35:65 in dry years and 65:35 and 50:50 in rainy years with loamy and clay soils, respectively. The appropriate development of both species in the mixture at different soil and rainfall conditions, as well as a good yield often higher than control mixture and a great forage quality, confirm to the narbon bean-black oat mixture as a viable and profitable crop alternative in rainfed cropping systems under Mediterranean conditions.

  12. Primordial germ cell-mediated chimera technology produces viable pure-line Houbara bustard offspring: potential for repopulating an endangered species.

    Directory of Open Access Journals (Sweden)

    Ulrich Wernery

    2010-12-01

    Full Text Available The Houbara bustard (Chlamydotis undulata is a wild seasonal breeding bird populating arid sandy semi-desert habitats in North Africa and the Middle East. Its population has declined drastically during the last two decades and it is classified as vulnerable. Captive breeding programmes have, hitherto, been unsuccessful in reviving population numbers and thus radical technological solutions are essential for the long term survival of this species. The purpose of this study was to investigate the use of primordial germ cell-mediated chimera technology to produce viable Houbara bustard offspring.Embryonic gonadal tissue was dissected from Houbara bustard embryos at eight days post-incubation. Subsequently, Houbara tissue containing gonadal primordial germ cells (gPGCs was injected into White Leghorn chicken (Gallus gallus domesticus embryos, producing 83/138 surviving male chimeric embryos, of which 35 chimeric roosters reached sexual maturity after 5 months. The incorporation and differentiation of Houbara gPGCs in chimeric chicken testis were assessed by PCR with Houbara-specific primers and 31.3% (5/16 gonads collected from the injected chicken embryos showed the presence of donor Houbara cells. A total of 302 semen samples from 34 chimeric roosters were analyzed and eight were confirmed as germline chimeras. Semen samples from these eight roosters were used to artificially inseminate three female Houbara bustards. Subsequently, 45 Houbara eggs were obtained and incubated, two of which were fertile. One egg hatched as a male live born Houbara; the other was female but died before hatching. Genotyping confirmed that the male chick was a pure-line Houbara derived from a chimeric rooster.This study demonstrates for the first time that Houbara gPGCs can migrate, differentiate and eventually give rise to functional sperm in the chimeric chicken testis. This approach may provide a promising tool for propagation and conservation of endangered avian

  13. Strengthening Agricultural Research Capacity for Viable Extension ...

    African Journals Online (AJOL)

    Strengthening Agricultural Research Capacity for Viable Extension Policies in Nigeria: An Exploration of Ricoeur's Hermeneutic Theory for Analysing Extension Research. ... Progressively more, researchers use hermeneutic philosophy to inform the conduct of interpretive research. Analogy between the philosophical ...

  14. Selectable high-yield recombinant protein production in human cells using a GFP/YFP nanobody affinity support.

    Science.gov (United States)

    Schellenberg, Matthew J; Petrovich, Robert M; Malone, Christine C; Williams, R Scott

    2018-03-25

    Recombinant protein expression systems that produce high yields of pure proteins and multi-protein complexes are essential to meet the needs of biologists, biochemists, and structural biologists using X-ray crystallography and cryo-electron microscopy. An ideal expression system for recombinant human proteins is cultured human cells where the correct translation and chaperone machinery are present. However, compared to bacterial expression systems, human cell cultures present several technical challenges to their use as an expression system. We developed a method that utilizes a YFP fusion-tag to generate recombinant proteins using suspension-cultured HEK293F cells. YFP is a dual-function tag that enables direct visualization and fluorescence-based selection of high expressing clones for and rapid purification using a high-stringency, high-affinity anti-GFP/YFP nanobody support. We demonstrate the utility of this system by expressing two large human proteins, TOP2α (340 KDa dimer) and a TOP2β catalytic core (260 KDa dimer). This robustly and reproducibly yields >10 mg/L liter of cell culture using transient expression or 2.5 mg/L using stable expression. Published 2018. This article is a US Government work and is in the public domain in the USA.

  15. Increased cell surface metallopeptidase activity in cells undergoing UV-induced apoptosis

    International Nuclear Information System (INIS)

    Piva, T.J.; Davern, C.M.; Ellem, K.A.O.

    1999-01-01

    Full text: We have previously shown that UVC irradiation activated a range of cell surface peptidases (CSP) in HeLa cell monolayer cultures 20 h post-irradiation (1). In cells undergoing apoptosis there is an increase in CSP activity compared to control viable cells in cultures which have been treated by a wide range of agents including UV-irradiation (2). In order to further understand the mechanism involved in this process, we induced apoptosis in HeLa cells using 500 Jm -2 UVB. The separation of viable, apoptotic and necrotic cells of irradiated HeLa cell cultures was made by FACS analysis and sorting. The three populations were distinguished by their staining with PI and Hoechst 33342 dyes. CSP activity was measured using the P9 assay developed in this laboratory (1-3). The viable fraction of the irradiated cells had a higher level of CSP activity compared to unirradiated controls. The level of CSP activity in the apoptotic fraction was higher than that of the viable fraction, however that of the necrotic fraction was significantly lower. This finding agreed with that seen in UVC-irradiated (50 Jm -2 ) cultures (2). In order to elucidate the mechanism by which CSP activity was increased in UVB-irradiated cells undergoing apoptosis, the cultures were treated with the following agents: bestatin, aminopeptidase inhibitor, DEVD, caspase 3 inhibitor, and 3-aminobenzamide (3AB), PARP activation inhibitor. Bestatin and DEVD did not affect the level of CSP activity in the different cell subpopulations following UVB-irradiation. Treatment with 3AB abolished the increased CSP activity seen in the viable and apoptotic fraction following UVB-irradiation. All treated cells had the same morphology as observed under EM. The degree of phosphatidylserine eversion on the cell membrane was similar as were the cleavage profiles of PARP and actin. Only DEVD-treated cells had reduced caspase 3 activity which confirmed that the activation of CSP activity in apoptotic cells is

  16. Sorghums: viable biomass candidates

    Energy Technology Data Exchange (ETDEWEB)

    McClure, T A; Arthur, M F; Kresovich, S; Scantland, D A

    1980-01-01

    Agronomic studies conducted at Battelle's Columbus Division to evaluate biomass and sugar yields of sweet sorghum are described and the major findings are summarized. Development opportunities for using sorghum cultivars as a large-scale energy crop are discussed. With presently available cultivars, sweet sorghum should produce 3500 to 4000 liters ethanol per hectare from the fermentable sugars alone. Conversion of the stalk fibers into alcohol could increase production by another 1600 to 1900 liters per hectare with existing cultivars. These yields are approximately 30 to 40% greater per hectare than would be obtained from above average yields of grain and stalk fiber with corn. There is reason to believe, that with hybrid sweet sorghum, these yields could be further increased by as much as 30%. Diminishing land availability for agricultural crops necessitates that maximum yields be obtained. Over the next decade, imaginative technological innovations in sorghum harvesting, processing, and crop preservation, coupled with plant breeding research should help this crop realize its full potential as a renewable resource for energy production.

  17. Diploid yeast cells yield homozygous spontaneous mutations

    Science.gov (United States)

    Esposito, M. S.; Bruschi, C. V.; Brushi, C. V. (Principal Investigator)

    1993-01-01

    A leucine-requiring hybrid of Saccharomyces cerevisiae, homoallelic at the LEU1 locus (leu1-12/leu1-12) and heterozygous for three chromosome-VII genetic markers distal to the LEU1 locus, was employed to inquire: (1) whether spontaneous gene mutation and mitotic segregation of heterozygous markers occur in positive nonrandom association and (2) whether homozygous LEU1/LEU1 mutant diploids are generated. The results demonstrate that gene mutation of leu1-12 to LEU1 and mitotic segregation of heterozygous chromosome-VII markers occur in strong positive nonrandom association, suggesting that the stimulatory DNA lesion is both mutagenic and recombinogenic. In addition, genetic analysis of diploid Leu+ revertants revealed that approximately 3% of mutations of leu1-12 to LEU1 result in LEU1/LEU1 homozygotes. Red-white sectored Leu+ colonies exhibit genotypes that implicate post-replicational chromatid breakage and exchange near the site of leu1-12 reversion, chromosome loss, and subsequent restitution of diploidy, in the sequence of events leading to mutational homozygosis. By analogy, diploid cell populations can yield variants homozygous for novel recessive gene mutations at biologically significant rates. Mutational homozygosis may be relevant to both carcinogenesis and the evolution of asexual diploid organisms.

  18. Development of a chemically defined platform fed-batch culture media for monoclonal antibody-producing CHO cell lines with optimized choline content.

    Science.gov (United States)

    Kuwae, Shinobu; Miyakawa, Ichiko; Doi, Tomohiro

    2018-01-11

    A chemically defined platform basal medium and feed media were developed using a single Chinese hamster ovary (CHO) cell line that produces a monoclonal antibody (mAb). Cell line A, which showed a peak viable cell density of 5.9 × 10 6  cells/mL and a final mAb titer of 0.5 g/L in batch culture, was selected for the platform media development. Stoichiometrically balanced feed media were developed using glucose as an indicator of cell metabolism to determine the feed rates of all other nutrients. A fed-batch culture of cell line A using the platform fed-batch medium yielded a 6.4 g/L mAb titer, which was 12-fold higher than that of the batch culture. To examine the applicability of the platform basal medium and feed media, three other cell lines (A16, B, and C) that produce mAbs were cultured using the platform fed-batch medium, and they yielded mAb titers of 8.4, 3.3, and 6.2 g/L, respectively. The peak viable cell densities of the three cell lines ranged from 1.3 × 10 7 to 1.8 × 10 7  cells/mL. These results show that the nutritionally balanced fed-batch medium and feeds worked well for other cell lines. During the medium development, we found that choline limitation caused a lower cell viability, a lower mAb titer, a higher mAb aggregate content, and a higher mannose-5 content. The optimal choline chloride to glucose ratio for the CHO cell fed-batch culture was determined. Our platform basal medium and feed media will shorten the medium-development time for mAb-producing cell lines.

  19. Yield Improvement and Energy Savings Uing Phosphonates as Additives in Kraft pulping

    Energy Technology Data Exchange (ETDEWEB)

    Ulrike W. Tschirner; Timothy Smith

    2007-03-31

    Project Objective: Develop a commercially viable modification to the Kraft process resulting in energy savings, increased yield and improved bleachability. Evaluate the feasibility of this technology across a spectrum of wood species used in North America. Develop detailed fundamental understanding of the mechanism by which phosphonates improve KAPPA number and yield. Evaluate the North American market potential for the use of phosphonates in the Kraft pulping process. Examine determinants of customer perceived value and explore organizational and operational factors influencing attitudes and behaviors. Provide an economic feasibility assessment for the supply chain, both suppliers (chemical supply companies) and buyers (Kraft mills). Provide background to most effectively transfer this new technology to commercial mills.

  20. GROWTH, GAS EXCHANGE AND YIELD OF CORN WHEN FERTIGATED WITH BOVINE BIOFERTILIZER

    OpenAIRE

    THALES VINÍCIUS DE ARAÚJO VIANA; JOÃO GUILHERME ARAÚJO LIMA; GEOCLEBER GOMES DE SOUSA; LUIS GONZAGA PINHEIRO NETO; BENITO MOREIRA DE AZEVEDO

    2014-01-01

    The bovine biofertilizer applied through irrigation water in the soil (bio fertigation), can be a viable organic source to maintain fertility levels in agricultural production systems. So, this work was aimed at evaluating the effects of different concentrations of bovine biofertilizer applied by fertigation on corn growth, gas exchange and yield. The experiment was conducted under full sun exposure, in Fortaleza, Ceara, in 100 liter (100 L) vessels. The experimental design was that of random...

  1. Assessments of Total and Viable Escherichia coli O157:H7 on Field and Laboratory Grown Lettuce

    Science.gov (United States)

    Moyne, Anne-Laure; Harris, Linda J.; Marco, Maria L.

    2013-01-01

    Leafy green produce has been associated with numerous outbreaks of foodborne illness caused by strains of Escherichia coli O157:H7. While the amounts of culturable E. coli O157:H7 rapidly decline after introduction onto lettuce in the field, it remains to be determined whether the reduction in cell numbers is due to losses in cell viability, cell injury and a subsequent inability to be detected by standard laboratory culturing methods, or a lack of adherence and hence rapid removal of the organism from the plants during application. To assess which of these options is most relevant for E. coli O157:H7 on leafy green produce, we developed and applied a propidium monoazide (PMA) real-time PCR assay to quantify viable (with PMA) and total (without PMA) E. coli O157:H7 cells on growth chamber and field-grown lettuce. E. coli O157:H7, suspended in 0.1% peptone, was inoculated onto 4-week-old lettuce plants at a level of approximately 106 CFU/plant. In the growth chamber at low relative humidity (30%), culturable amounts of the nontoxigenic E. coli O157:H7 strain ATCC 700728 and the virulent strain EC4045 declined 100 to 1000-fold in 24 h. Fewer E. coli O157:H7 cells survived when applied onto plants in droplets with a pipette compared with a fine spray inoculation. Total cells for both strains were equivalent to inoculum levels for 7 days after application, and viable cell quantities determined by PMA real-time PCR were approximately 104 greater than found by colony enumeration. Within 2 h after application onto plants in the field, the number of culturable E. coli ATCC 700728 was reduced by up to 1000-fold, whereas PCR-based assessments showed that total cell amounts were equivalent to inoculum levels. These findings show that shortly after inoculation onto plants, the majority of E. coli O157:H7 cells either die or are no longer culturable. PMID:23936235

  2. Establishing a high yielding streptomyces-based cell-free protein synthesis system.

    Science.gov (United States)

    Li, Jian; Wang, He; Kwon, Yong-Chan; Jewett, Michael C

    2017-06-01

    Cell-free protein synthesis (CFPS) has emerged as a powerful platform for applied biotechnology and synthetic biology, with a range of applications in synthesizing proteins, evolving proteins, and prototyping genetic circuits. To expand the current CFPS repertoire, we report here the development and optimization of a Streptomyces-based CFPS system for the expression of GC-rich genes. By developing a streamlined crude extract preparation protocol and optimizing reaction conditions, we were able to achieve active enhanced green fluorescent protein (EGFP) yields of greater than 50 μg/mL with batch reactions lasting up to 3 h. By adopting a semi-continuous reaction format, the EGFP yield could be increased to 282 ± 8 μg/mL and the reaction time was extended to 48 h. Notably, our extract preparation procedures were robust to multiple Streptomyces lividans and Streptomyces coelicolor strains, although expression yields varied. We show that our optimized Streptomyces lividans system provides benefits when compared to an Escherichia coli-based CFPS system for increasing percent soluble protein expression for four Streptomyces-originated high GC-content genes that are involved in biosynthesis of the nonribosomal peptides tambromycin and valinomycin. Looking forward, we believe that our Streptomyces-based CFPS system will contribute significantly towards efforts to express complex natural product gene clusters (e.g., nonribosomal peptides and polyketides), providing a new avenue for obtaining and studying natural product biosynthesis pathways. Biotechnol. Bioeng. 2017;114: 1343-1353. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  3. Investigation of Removal Capacities of Biofilters for Airborne Viable Micro-Organisms

    Science.gov (United States)

    Soret, Rémi; Fanlo, Jean-Louis; Malhautier, Luc; Geiger, Philippe; Bayle, Sandrine

    2018-01-01

    New emerging issues appears regarding the possible aerosolization of micro-organisms from biofilters to the ambient air. Traditional bioaerosol sampling and cultural methods used in literature offer relative efficiencies. In this study, a new method revolving around a particle counter capable of detecting total and viable particles in real time was used. This counter (BioTrak 9510-BD) uses laser-induced fluorescence (LIF) technology to determine the biological nature of the particle. The concentration of viable particles was measured on two semi-industrial pilot scale biofilters in order to estimate the Removal Efficiency in viable particles (REvp) in stable conditions and to examine the influence of pollutant feeding and relative humidification of the gaseous effluent on the REvp. The REvp of biofilters reached near 80% and highlighted both the stability of that removal and the statistical equivalence between two identical biofilters. Pollutant deprivation periods of 12 h, 48 h and 30 days were shown to have no influence on the biofilters’ removal capacity, demonstrating the robustness and adaptation capacities of the flora. In contrast, a 90-day famine period turned the biofilters into emitters of viable particles. Finally, the humidification of the effluent was shown to negatively influence the removal capacity for viable particles, as drying off the air was shown to increase the REvp from 60 to 85%. PMID:29562709

  4. Genomic characterisation of small cell lung cancer patient-derived xenografts generated from endobronchial ultrasound-guided transbronchial needle aspiration specimens.

    Directory of Open Access Journals (Sweden)

    Tracy L Leong

    Full Text Available Patient-derived xenograft (PDX models generated from surgical specimens are gaining popularity as preclinical models of cancer. However, establishment of PDX lines from small cell lung cancer (SCLC patients is difficult due to very limited amount of available biopsy material. We asked whether SCLC cells obtained from endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA could generate PDX lines that maintained the phenotypic and genetic characteristics of the primary tumor. Following successful EBUS-TBNA sampling for diagnostic purposes, we obtained an extra sample for cytologic analysis and implantation into the flanks of immunodeficient mice. Animals were monitored for engraftment for up to 6 months. Histopathologic and immunohistochemical analysis, and targeted next-generation re-sequencing, were then performed in both the primary sample and the derivative PDX line. A total of 12 patients were enrolled in the study. EBUS-TBNA aspirates yielded large numbers of viable tumor cells sufficient to inject between 18,750 and 1,487,000 cells per flank, and to yield microgram quantities of high-quality DNA. Of these, samples from 10 patients generated xenografts (engraftment rate 83% with a mean latency of 104 days (range 63-188. All but one maintained a typical SCLC phenotype that closely matched the original sample. Identical mutations that are characteristic of SCLC were identified in both the primary sample and xenograft line. EBUS-TBNA has the potential to be a powerful tool in the development of new targeting strategies for SCLC patients by providing large numbers of viable tumor cells suitable for both xenografting and complex genomic analysis.

  5. Autolyse the cell in order to save it? Inducing, then blocking, autolysis as a strategy for delaying cell death in the probiotic Lactobacillus reuteri.

    Science.gov (United States)

    Zimmerman, Tahl; Gyawali, Rabin; Ibrahim, Salam

    2017-10-01

    To examine whether choline and its derivatives can be used to preserve viable cells of Lactobacillus reuteri in autolytic models. A phosphate-induced autolytic model in de Man, Rogosa and Sharpe medium (MRS) was used. Viable cell counts were determined by plated on MRS-agar. Choline and hemicholinium-3 (HC-3) significantly blocked autolysis of L. reuteri at 360 mM and 4 mM, respectively. Viable cell counts corroborated these observations. Importantly, autolytically induced cells treated with choline and hemicholinium-3 were significantly more viable then even non-induced cells. Over-production of a known autolytic protein, spirosin, was not attenuated in the presence of choline and hemicholinium-3. Inducing autolysis and then blocking it with choline and its analogs is a promising approach for retaining the viability of L. reuteri cells.

  6. Closing Yield Gaps: How Sustainable Can We Be?

    Directory of Open Access Journals (Sweden)

    Prajal Pradhan

    Full Text Available Global food production needs to be increased by 60-110% between 2005 and 2050 to meet growing food and feed demand. Intensification and/or expansion of agriculture are the two main options available to meet the growing crop demands. Land conversion to expand cultivated land increases GHG emissions and impacts biodiversity and ecosystem services. Closing yield gaps to attain potential yields may be a viable option to increase the global crop production. Traditional methods of agricultural intensification often have negative externalities. Therefore, there is a need to explore location-specific methods of sustainable agricultural intensification. We identified regions where the achievement of potential crop calorie production on currently cultivated land will meet the present and future food demand based on scenario analyses considering population growth and changes in dietary habits. By closing yield gaps in the current irrigated and rain-fed cultivated land, about 24% and 80% more crop calories can respectively be produced compared to 2000. Most countries will reach food self-sufficiency or improve their current food self-sufficiency levels if potential crop production levels are achieved. As a novel approach, we defined specific input and agricultural management strategies required to achieve the potential production by overcoming biophysical and socioeconomic constraints causing yield gaps. The management strategies include: fertilizers, pesticides, advanced soil management, land improvement, management strategies coping with weather induced yield variability, and improving market accessibility. Finally, we estimated the required fertilizers (N, P2O5, and K2O to attain the potential yields. Globally, N-fertilizer application needs to increase by 45-73%, P2O5-fertilizer by 22-46%, and K2O-fertilizer by 2-3 times compared to the year 2010 to attain potential crop production. The sustainability of such agricultural intensification largely depends

  7. Viable-but-Nonculturable Listeria monocytogenes and Salmonella enterica Serovar Thompson Induced by Chlorine Stress Remain Infectious

    Directory of Open Access Journals (Sweden)

    Callum J. Highmore

    2018-04-01

    Full Text Available The microbiological safety of fresh produce is monitored almost exclusively by culture-based detection methods. However, bacterial food-borne pathogens are known to enter a viable-but-nonculturable (VBNC state in response to environmental stresses such as chlorine, which is commonly used for fresh produce decontamination. Here, complete VBNC induction of green fluorescent protein-tagged Listeria monocytogenes and Salmonella enterica serovar Thompson was achieved by exposure to 12 and 3 ppm chlorine, respectively. The pathogens were subjected to chlorine washing following incubation on spinach leaves. Culture data revealed that total viable L. monocytogenes and Salmonella Thompson populations became VBNC by 50 and 100 ppm chlorine, respectively, while enumeration by direct viable counting found that chlorine caused a <1-log reduction in viability. The pathogenicity of chlorine-induced VBNC L. monocytogenes and Salmonella Thompson was assessed by using Caenorhabditis elegans. Ingestion of VBNC pathogens by C. elegans resulted in a significant life span reduction (P = 0.0064 and P < 0.0001, and no significant difference between the life span reductions caused by the VBNC and culturable L. monocytogenes treatments was observed. L. monocytogenes was visualized beyond the nematode intestinal lumen, indicating resuscitation and cell invasion. These data emphasize the risk that VBNC food-borne pathogens could pose to public health should they continue to go undetected.

  8. 9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Detection of viable bacteria and fungi... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.26 Detection of viable bacteria and fungi except... required to be free of viable bacteria and fungi, they shall also be tested as prescribed in this section...

  9. Can Malin's gravitational-field equations be modified to obtain a viable theory of gravity to obtain a viable theory of gravity to obtain a viable theory of gravity

    International Nuclear Information System (INIS)

    Smalley, L.L.; Prestage, J.

    1976-01-01

    Malin's gravitational theory, which was recently shown by Lindblom and Nester to be incorrect, is modified by means of a recently proposed method for obtaining viable gravitational theories. The resulting self-consistent theory, which is in effect a Rastall-type modification of the Einstein theory, exhibits nonconservation of momentum, yet agrees with all experimental limits known to date within the PPN framework

  10. Monotone viable trajectories for functional differential inclusions

    Science.gov (United States)

    Haddad, Georges

    This paper is a study on functional differential inclusions with memory which represent the multivalued version of retarded functional differential equations. The main result gives a necessary and sufficient equations. The main result gives a necessary and sufficient condition ensuring the existence of viable trajectories; that means trajectories remaining in a given nonempty closed convex set defined by given constraints the system must satisfy to be viable. Some motivations for this paper can be found in control theory where F( t, φ) = { f( t, φ, u)} uɛU is the set of possible velocities of the system at time t, depending on the past history represented by the function φ and on a control u ranging over a set U of controls. Other motivations can be found in planning procedures in microeconomics and in biological evolutions where problems with memory do effectively appear in a multivalued version. All these models require viability constraints represented by a closed convex set.

  11. Viable Techniques, Leontief’s Closed Model, and Sraffa’s Subsistence Economies

    Directory of Open Access Journals (Sweden)

    Alberto Benítez

    2014-11-01

    Full Text Available This paper studies the production techniques employed in economies that reproduce themselves. Special attention is paid to the distinction usually made between those that do not produce a surplus and those that do, which are referred to as first and second class economies, respectively. Based on this, we present a new definition of viable economies and show that every viable economy of the second class can be represented as a viable economy of the first class under two different forms, Leontief‘s closed model and Sraffa’s subsistence economies. This allows us to present some remarks concerning the economic interpretation of the two models. On the one hand, we argue that the participation of each good in the production of every good can be considered as a normal characteristic of the first model and, on the other hand, we provide a justification for the same condition to be considered a characteristic of the second model. Furthermore, we discuss three definitions of viable techniques advanced by other authors and show that they differ from ours because they admit economies that do not reproduce themselves completely.

  12. Cell and protein compatible 3D bioprinting of mechanically strong constructs for bone repair

    International Nuclear Information System (INIS)

    Sawkins, M J; Mistry, P; Shakesheff, K M; Yang, J; Brown, B N; Bonassar, L J

    2015-01-01

    Rapid prototyping of bone tissue engineering constructs often utilizes elevated temperatures, organic solvents and/or UV light for materials processing. These harsh conditions may prevent the incorporation of cells and therapeutic proteins in the fabrication processes. Here we developed a method for using bioprinting to produce constructs from a thermoresponsive microparticulate material based on poly(lactic-co-glycolic acid) at ambient conditions. These constructs could be engineered with yield stresses of up to 1.22 MPa and Young’s moduli of up to 57.3 MPa which are within the range of properties of human cancellous bone. Further study showed that protein-releasing microspheres could be incorporated into the bioprinted constructs. The release of the model protein lysozyme from bioprinted constructs was sustainted for a period of 15 days and a high degree of protein activity could be measured up to day 9. This work suggests that bioprinting is a viable route to the production of mechanically strong constructs for bone repair under mild conditions which allow the inclusion of viable cells and active proteins. (paper)

  13. [Milk yield and environmental factors: Multiple regression analysis of the association between milk yield and udder health, fertility data and replacement rate].

    Science.gov (United States)

    Fölsche, C; Staufenbiel, R

    2014-01-01

    The relationship between milk yield and both fertility and general animal health in dairy herds is discussed from opposing viewpoints. The hypothesis (1) that raising the herd milk yield would decrease fertility results, the number of milk cells as an indicator for udder health and the replacement rate as a global indicator for animal health as well as increasing the occurrence of specific diseases as a herd problem was compared to the opposing hypotheses that there is no relationship (2) or that there is a differentiated and changing relationship (3). A total of 743 herd examinations, considered independent, were performed in 489 herds between 1995 and 2010. The milk yield, fertility rate, milk cell count, replacement rate, categorized herd problems and management information were recorded. The relationship between the milk yield and both the fertility data and animal health was evaluated using simple and multiple regression analyses. The period between calving and the first service displayed no significant relationship to the herd milk yield. Simple regression analysis showed that the period between calving and gestation, the calving interval and the insemination number were significantly positively associated with the herd milk yield. This positive correlation was lost in multiple regression analysis. The milk cell count and replacement rate using both the simple and multiple regression analyses displayed a significant negative relationship to the milk yield. The alternative hypothesis (3) was confirmed. A higher milk yield has no negative influence on the milk cell count and the replacement rate in terms of the udder and general health. When parameterizing the fertility, the herd milk yield should be considered. Extending the resting time may increase the milk yield while preventing a decline in the insemination index.

  14. Development of a propidium monoazide-polymerase chain reaction assay for detection of viable Lactobacillus brevis in beer.

    Science.gov (United States)

    Ma, Yanlin; Deng, Yang; Xu, Zhenbo; Liu, Junyan; Dong, Jianjun; Yin, Hua; Yu, Junhong; Chang, Zongming; Wang, Dongfeng

    The spoilage of beer by bacteria is of great concern to the brewer as this can lead to turbidity and abnormal flavors. The polymerase chain reaction (PCR) method for detection of beer-spoilage bacteria is highly specific and provides results much faster than traditional microbiology techniques. However, one of the drawbacks is the inability to differentiate between live and dead cells. In this paper, the combination of propidium monoazide (PMA) pretreatment and conventional PCR had been described. The established PMA-PCR identified beer spoilage Lactobacillus brevis based not on their identity, but on the presence of horA gene which we show to be highly correlated with the ability of beer spoilage LAB to grow in beer. The results suggested that the use of 30μg/mL or less of PMA did not inhibit the PCR amplification of DNA derived from viable L. brevis cells. The minimum amount of PMA to completely inhibit the PCR amplification of DNA derived from dead L. brevis cells was 2.0μg/mL. The detection limit of PMA-PCR assay described here was found to be 10 colony forming units (CFU)/reaction for the horA gene. Moreover, the horA-specific PMA-PCR assays were subjected to 18 reference isolates, representing 100% specificity with no false positive amplification observed. Overall the use of horA-specific PMA-PCR allows for a substantial reduction in the time required for detection of potential beer spoilage L. brevis and efficiently differentiates between viable and nonviable cells. Copyright © 2017 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  15. Timing of Peripheral Blood Stem Cell Yield: Comparison of Alternative Methods with the Classic Method for CD34+ Cell Determination

    Directory of Open Access Journals (Sweden)

    I. Fatorova

    2014-01-01

    Full Text Available Hematopoietic stem cells (HSCs, still represent a certain mystery in biology, have a unique property of dividing into equal cells and repopulating the hematopoietic tissue. This potential enables their use in transplantation treatments. The quality of the HSC grafts for transplantation is evaluated by flow cytometric determination of the CD34+ cells, which enables optimal timing of the first apheresis and the acquisition of maximal yield of the peripheral blood stem cells (PBSCs. To identify a more efficient method for evaluating CD34+ cells, we compared the following alternative methods with the reference method: hematopoietic progenitor cells (HPC enumeration (using the Sysmex XE-2100 analyser, detection of CD133+ cells, and quantification of aldehyde dehydrogenase activity in the PBSCs. 266 aphereses (84 patients were evaluated. In the preapheretic blood, the new methods produced data that were in agreement with the reference method. The ROC curves have shown that for the first-day apheresis target, the optimal predictive cut-off value was 0.032 cells/mL for the HPC method (sensitivity 73.4%, specificity 69.3%. HPC method exhibited a definite practical superiority as compared to other methods tested. HPC enumeration could serve as a supplementary method for the optimal timing of the first apheresis; it is simple, rapid, and cheap.

  16. Primordial Germ Cell-Mediated Chimera Technology Produces Viable Pure-Line Houbara Bustard Offspring: Potential for Repopulating an Endangered Species

    Science.gov (United States)

    Wernery, Ulrich; Liu, Chunhai; Baskar, Vijay; Guerineche, Zhor; Khazanehdari, Kamal A.; Saleem, Shazia; Kinne, Jörg; Wernery, Renate

    2010-01-01

    Background The Houbara bustard (Chlamydotis undulata) is a wild seasonal breeding bird populating arid sandy semi-desert habitats in North Africa and the Middle East. Its population has declined drastically during the last two decades and it is classified as vulnerable. Captive breeding programmes have, hitherto, been unsuccessful in reviving population numbers and thus radical technological solutions are essential for the long term survival of this species. The purpose of this study was to investigate the use of primordial germ cell-mediated chimera technology to produce viable Houbara bustard offspring. Methodology/Principal Findings Embryonic gonadal tissue was dissected from Houbara bustard embryos at eight days post-incubation. Subsequently, Houbara tissue containing gonadal primordial germ cells (gPGCs) was injected into White Leghorn chicken (Gallus gallus domesticus) embryos, producing 83/138 surviving male chimeric embryos, of which 35 chimeric roosters reached sexual maturity after 5 months. The incorporation and differentiation of Houbara gPGCs in chimeric chicken testis were assessed by PCR with Houbara-specific primers and 31.3% (5/16) gonads collected from the injected chicken embryos showed the presence of donor Houbara cells. A total of 302 semen samples from 34 chimeric roosters were analyzed and eight were confirmed as germline chimeras. Semen samples from these eight roosters were used to artificially inseminate three female Houbara bustards. Subsequently, 45 Houbara eggs were obtained and incubated, two of which were fertile. One egg hatched as a male live born Houbara; the other was female but died before hatching. Genotyping confirmed that the male chick was a pure-line Houbara derived from a chimeric rooster. Conclusion This study demonstrates for the first time that Houbara gPGCs can migrate, differentiate and eventually give rise to functional sperm in the chimeric chicken testis. This approach may provide a promising tool for propagation

  17. Increasing Hematopoietic Stem Cell Yield to Develop Mice with Human Immune Systems

    Directory of Open Access Journals (Sweden)

    Juan-Carlos Biancotti

    2013-01-01

    Full Text Available Hematopoietic stem cells (HSCs are unique in their capacity to give rise to all mature cells of the immune system. For years, HSC transplantation has been used for treatment of genetic and neoplastic diseases of the hematopoietic and immune systems. The sourcing of HSCs from human umbilical cord blood has salient advantages over isolation from mobilized peripheral blood. However, poor sample yield has prompted development of methodologies to expand HSCs ex vivo. Cytokines, trophic factors, and small molecules have been variously used to promote survival and proliferation of HSCs in culture, whilst strategies to lower the concentration of inhibitors in the culture media have recently been applied to promote HSC expansion. In this paper, we outline strategies to expand HSCs in vitro, and to improve engraftment and reconstitution of human immune systems in immunocompromised mice. To the extent that these “humanized” mice are representative of the endogenous human immune system, they will be invaluable tools for both basic science and translational medicine.

  18. Culture of somatic cells isolated from frozen-thawed equine semen using fluorescence-assisted cell sorting.

    Science.gov (United States)

    Brom-de-Luna, Joao Gatto; Canesin, Heloísa Siqueira; Wright, Gus; Hinrichs, Katrin

    2018-03-01

    Nuclear transfer using somatic cells from frozen semen (FzSC) would allow cloning of animals for which no other genetic material is available. Horses are one of the few species for which cloning is commercially feasible; despite this, there is no information available on the culture of equine FzSC. After preliminary trials on equine FzSC, recovered by density-gradient centrifugation, resulted in no growth, we hypothesized that sperm in the culture system negatively affected cell proliferation. Therefore, we evaluated culture of FzSC isolated using fluorescence-assisted cell sorting. In Exp. 1, sperm were labeled using antibodies to a sperm-specific antigen, SP17, and unlabeled cells were collected. This resulted in high sperm contamination. In Exp. 2, FzSC were labeled using an anti-MHC class I antibody. This resulted in an essentially pure population of FzSC, 13-25% of which were nucleated. Culture yielded no proliferation in any of nine replicates. In Exp. 3, 5 × 10 3 viable fresh, cultured horse fibroblasts were added to the frozen-thawed, washed semen, then this suspension was labeled and sorted as for Exp. 2. The enriched population had a mean of five sperm per recovered somatic cell; culture yielded formation of monolayers. In conclusion, an essentially pure population of equine FzSC could be obtained using sorting for presence of MHC class I antigens. No equine FzSC grew in culture; however, the proliferation of fibroblasts subjected to the same processing demonstrated that the labeling and sorting methods, and the presence of few sperm in culture, were compatible with cell viability. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. The viable but non-culturable state in pathogenic Escherichia coli: A general review

    Directory of Open Access Journals (Sweden)

    Jennifer A. Pienaar

    2016-05-01

    Objectives: This review discusses various general aspects of the VBNC state, the mechanisms and possible public health impact of indicator and pathogenic E. coli entering into the VBNC state. Method: A literature review was conducted to ascertain the possibleimpact of E. coli entering into the VBNC state. Results: Escherichia coli enter into the VBNC state by means of several induction mechanisms. Various authors have found that E. coli can be resuscitated post-VBNC. Certain strains of pathogenic E. coli are still able to produce toxins in the VBNC state, whilst others are avirulent during the VBNC state but are able to regain virulence after resuscitation. Conclusion: Pathogenic and indicator E. coli entering into the VBNC state could have an adverse effect on public health if conventional detection methods are used, where the number of viable cells could be underestimated and the VBNC cells still produce toxins or could, at anytime, be resuscitated and become virulent again.

  20. Tradeoffs between water requirements and yield stability in annual vs. perennial crops

    Science.gov (United States)

    Vico, Giulia; Brunsell, Nathaniel A.

    2018-02-01

    Population growth and changes in climate and diets will likely further increase the pressure on agriculture and water resources globally. Currently, staple crops are obtained from annuals plants. A shift towards perennial crops may enhance many ecosystem services, but at the cost of higher water requirements and lower yields. It is still unclear when the advantages of perennial crops overcome their disadvantages and perennial crops are thus a sustainable solution. Here we combine a probabilistic description of the soil water balance and crop development with an extensive dataset of traits of congeneric annuals and perennials to identify the conditions for which perennial crops are more viable than annual ones with reference to yield, yield stability, and effective use of water. We show that the larger and more developed roots of perennial crops allow a better exploitation of soil water resources and a reduction of yield variability with respect to annual species, but their yields remain lower when considering grain crops. Furthermore, perennial crops have higher and more variable irrigation requirements and lower water productivity. These results are important to understand the potential consequences for yield, its stability, and water resource use of a shift from annual to perennial crops and, more generally, if perennial crops may be more resilient than annual crops in the face of climatic fluctuations.

  1. Cryopreservation of human insulin expressing cells macro-encapsulated in a durable therapeutic immunoisolating device theracyte.

    Science.gov (United States)

    Yakhnenko, Ilya; Wong, Wallace K; Katkov, Igor I; Itkin-Ansari, Pamela

    2012-01-01

    Encapsulating insulin producing cells (INPCs) in an immunoisolation device have been shown to cure diabetes in rodents without the need for immunosuppression. However, micro-encapsulation in semi-solid gels raises longevity and safety concerns for future use of stem cell derived INPCs. We have focused on a durable and retrievable macro-encapsulation (> 10(6) cells) device (TheraCyte). Cryopreservation (CP) of cells preloaded into the device is highly desirable but may require prolonged exposure to cryoprotectants during loading and post-thaw manipulations. Here, we are reporting survival and function of a human islet cell line frozen as single cells or as islet-like cell clusters. The non-clusterized cells exhibited high cryosurvival after prolonged pre-freeze or post-thaw exposure to 10 percent DMSO. However, both clusterization and especially loading INPCs into the device reduced viable yield even without CP. The survived cryopreserved macro-encapsulated INPCs remained fully functional suggesting that CP of macro-encapsulated cells is a promising tool for cell based therapies.

  2. Growth Phase, Oxygen, Temperature and Starvation Affect the Development of Viable but Non-Culturable State of Vibrio cholerae

    Directory of Open Access Journals (Sweden)

    Bin eWu

    2016-03-01

    Full Text Available AbstractVibrio cholerae can enter into a viable but non-culturable (VBNC state in order to survive in unfavourable environments. In this study, we studied the roles of five physicochemical and microbiological factors or states, namely, different strains, growth phases, oxygen, temperature, and starvation, on the development of VBNC of V. cholerae in artificial sea water (ASW. Different strains of the organism, the growth phase, and oxygen levels affected the progress of VBNC development. It was found that the VBNC state was induced faster in V. cholerae serogroup O1 classical biotype strain O395 than in O1 El Tor biotype strains C6706 and N16961. When cells in different growth phases were used for VBNC induction, stationary-phase cells lost their culturability more quickly than exponential-phase cells, while induction of a totally non-culturable state took longer to achieve for stationary-phase cells in all three strains, suggesting that heterogeneity of cells should be considered. Aeration strongly accelerated the loss of culturability. During the development of the VBNC state, the culturable cell count under aeration conditions was almost 106-fold lower than under oxygen-limited conditions for all three strains. The other two factors, temperature and nutrients-rich environment, may prevent the induction of VBNC cells. At 22°C or 37°C in ASW, most of the cells rapidly died and the culturable cell count reduced from about 108 CFU/mL to 106–105 CFU/mL. The total cell counts showed that cells that lost viability were decomposed, and the viable cell counts were the same as culturable cell counts, indicating that the cells did not reach the VBNC state. VBNC state development was blocked when ASW was supplied with Luria-Bertani broth (LB, but it was not affected in ASW with M9, suggesting that specific nutrients in LB may prevent the development of VBNC state. These results revealed that the five factors evaluated in this study had different

  3. Association of viable Mycobacterium leprae with Type 1 reaction in leprosy.

    Science.gov (United States)

    Save, Mrudula Prakash; Dighe, Anju Rajaram; Natrajan, Mohan; Shetty, Vanaja Prabhakaran

    2016-03-01

    The working hypothesis is that, viable Mycobacterium leprae (M. leprae) play a crucial role in the precipitation of Type 1 reaction (T1R) in leprosy. A total of 165 new multibacillary patients were studied. To demonstrate presence of viable M. leprae in reactional lesion (T1R+), three tests were used concurrently viz. growth in the mouse foot pad (MFP), immunohistochemical detection of M. leprae secretory protein Ag85, and 16s rRNA--using in situ RT-PCR. Mirror biopsies and non reactional lesions served as controls (T1R-). A significantly higher proportion of lesion biopsy homogenates obtained at onset, from T1R(+) cases have shown unequivocal growth in MFP, proving the presence of viable bacteria, as compared to T1R(-) (P leprae is a component/prerequisite and the secretory protein Ag 85, might be the trigger for precipitation of T1R.

  4. Closing Yield Gaps: How Sustainable Can We Be?

    Science.gov (United States)

    Pradhan, Prajal; Fischer, Günther; van Velthuizen, Harrij; Reusser, Dominik E.; Kropp, Juergen P.

    2015-01-01

    Global food production needs to be increased by 60–110% between 2005 and 2050 to meet growing food and feed demand. Intensification and/or expansion of agriculture are the two main options available to meet the growing crop demands. Land conversion to expand cultivated land increases GHG emissions and impacts biodiversity and ecosystem services. Closing yield gaps to attain potential yields may be a viable option to increase the global crop production. Traditional methods of agricultural intensification often have negative externalities. Therefore, there is a need to explore location-specific methods of sustainable agricultural intensification. We identified regions where the achievement of potential crop calorie production on currently cultivated land will meet the present and future food demand based on scenario analyses considering population growth and changes in dietary habits. By closing yield gaps in the current irrigated and rain-fed cultivated land, about 24% and 80% more crop calories can respectively be produced compared to 2000. Most countries will reach food self-sufficiency or improve their current food self-sufficiency levels if potential crop production levels are achieved. As a novel approach, we defined specific input and agricultural management strategies required to achieve the potential production by overcoming biophysical and socioeconomic constraints causing yield gaps. The management strategies include: fertilizers, pesticides, advanced soil management, land improvement, management strategies coping with weather induced yield variability, and improving market accessibility. Finally, we estimated the required fertilizers (N, P2O5, and K2O) to attain the potential yields. Globally, N-fertilizer application needs to increase by 45–73%, P2O5-fertilizer by 22–46%, and K2O-fertilizer by 2–3 times compared to the year 2010 to attain potential crop production. The sustainability of such agricultural intensification largely depends on the way

  5. The yield of genome mutations in cells of intact and regenerating rat liver in normal conditions and after γ-irradiation

    International Nuclear Information System (INIS)

    Gil'yano, N.Ya.; Malinovskij, O.V.

    1984-01-01

    A comparative study was made of spontaneus and induced polyploidy in cells of resting and regenerating rat liver. Polyploidy was shown to play a major role in the ontogenesis and during regeneration after partial hepatectomy. An essential difference was revealed in the radiation response of cells of intact and regenerating liver with respect to the yield of polyploid cells. This distinction was referped to different effectiveness of the processes of repair and fixation of radiation damages in the actively proliferating and resting cells

  6. Single cell adhesion force measurement for cell viability identification using an AFM cantilever-based micro putter

    Science.gov (United States)

    Shen, Yajing; Nakajima, Masahiro; Kojima, Seiji; Homma, Michio; Kojima, Masaru; Fukuda, Toshio

    2011-11-01

    Fast and sensitive cell viability identification is a key point for single cell analysis. To address this issue, this paper reports a novel single cell viability identification method based on the measurement of single cell shear adhesion force using an atomic force microscopy (AFM) cantilever-based micro putter. Viable and nonviable yeast cells are prepared and put onto three kinds of substrate surfaces, i.e. tungsten probe, gold and ITO substrate surfaces. A micro putter is fabricated from the AFM cantilever by focused ion beam etching technique. The spring constant of the micro putter is calibrated using the nanomanipulation approach. The shear adhesion force between the single viable or nonviable cell and each substrate is measured using the micro putter based on the nanorobotic manipulation system inside an environmental scanning electron microscope. The adhesion force is calculated based on the deflection of the micro putter beam. The results show that the adhesion force of the viable cell to the substrate is much larger than that of the nonviable cell. This identification method is label free, fast, sensitive and can give quantitative results at the single cell level.

  7. Sycamore produces viable seed after six years

    Science.gov (United States)

    A. F. Ike

    1966-01-01

    In the early stages of any tree improvement program it is desirable to know how soon progenies of selected parents can themselves be included in a breeding program. How soon will they produce viable pollen and seed? In the case of sycamore (Platanus occidentalis L.), the information is meager: the Woody- Plant Seed Manual lists the minimum commercial seedbearing age...

  8. Optimisation of the Factor VIII yield in mammalian cell cultures by reducing the membrane bound fraction

    DEFF Research Database (Denmark)

    Kolind, Mille Petersen; Nørby, Peder Lisby; Berchtold, Martin Werner

    2011-01-01

    and forms the tenase complex together with clotting Factor IX. In vitro, during serum free production of recombinant FVIII (rFVIII), production cells also expose PS, and since vWF is not present to hinder interaction of secreted rFVIII with PS, rFVIII is partly associated with the cell membrane...... of active membrane bound rFVIII to the culture medium. Moreover, the attachment of rFVIII to cell membranes of un-transfected HEK293 cells was studied in the presence of compounds that competes for interactions between rFVIII and PS. Competitive assays between iodinated rFVIII (¹²5I-rFVIII) and annexin V...... or ortho-phospho-L-serine (OPLS) demonstrated that annexin V and OPLS were able to reduce the membrane bound fraction of rFVIII by 70% and 30%, respectively. Finally, adding OPLS to CHO cells stably expressing FVIII increased the yield by 50%. Using this new knowledge, the recovery of rFVIII could...

  9. Susceptibility of various cell lines to Neospora caninum tachyzoites cultivation

    Directory of Open Access Journals (Sweden)

    Khordadmehr, M.,

    2014-05-01

    Full Text Available Neospora caninum is a coccidian protozoan parasite which is a major cause of bovine abortions and neonatal mortality in cattle, sheep, goat and horse. Occasionally, cultured cells are used for isolation and multiplication of the agent in vitro with several purposes. In this study the tachyzoite yields of N. caninum were compared in various cell cultures as the host cell lines. Among the cell cultures tested, two presented good susceptibility to the agent: cell lines Vero and MA-104. SW742 and TLI (in vitro suspension culture of lymphoid cells infected with Theileria lestoquardi showed moderate sensitivity. No viable tachyzoite were detected in the culture of MDCK and McCoy cell lines. These results demonstrate that MA-104 and SW742 cells present adequate susceptibility to N. caninum compared to Vero cells, which have been largely used to multiply the parasite in vitro. Moreover, these have easy manipulation, fast multiplication and relatively low nutritional requirements. In addition, the result of this study showed that TLI cell line as a suspension cell culture is susceptible to Nc-1 tachyzoites infection and could be used as an alternative host cell line for tachyzoites culture in vitro studies.

  10. Issues of organizational cybernetics and viability beyond Beer's viable systems model

    Science.gov (United States)

    Nechansky, Helmut

    2013-11-01

    The paper starts summarizing the claims of Beer's viable systems model to identify five issues any viable organizations has to deal with in an unequivocal hierarchical structure of five interrelated systems. Then the evidence is introduced for additional issues and related viable structures of organizations, which deviate from Beer's model. These issues are: (1) the establishment and (2) evolution of an organization; (3) systems for independent top-down control (like "Six Sigma"); (4) systems for independent bottom-up correction of performance problems (like "Kaizen"), both working outside a hierarchical structure; (5) pull production systems ("Just in Time") and (6) systems for checks and balances of top-level power (like boards and shareholder meetings). Based on that an evolutionary approach to organizational cybernetics is outlined, addressing the establishment of organizations and possible courses of developments, including recent developments in quality and production engineering, as well as problems of setting and changing goal values determining organizational policies.

  11. Formas cocoides de Helicobacter pylori: viables o degenerativas

    Directory of Open Access Journals (Sweden)

    Felipe Cava

    2003-06-01

    Full Text Available De los trabajos presentados acerca de las formas cocoides de Helicobacter pylori se deduce una controversia mucho mayor que la resultante del mero estudio clínico de este microorganismo. Parece claro que existe una conversión tanto in vivo como in vitro de las formas espirales a las formas cocoides inducida por varios motivos, como cultivos prolongados, estrés físico y químico, y agentes antimicrobianos. En esta revisión repasamos los puntos de vista que han dividido a investigadores de esta área en dos grupos bien definidos: Los que consideran a estas formas cocoides como un producto no viable de degeneración celular y los que piensan que estas formas son estructuras viables,durmientes o de resistencia frente a condiciones ambientales adversas. Esta discrepancia conlleva a que interrogantes sobre la relación entre la transmisión de la enfermedad y estas formas cocoides permanezcan sin respuesta todavía.

  12. Casticin impairs cell growth and induces cell apoptosis via cell cycle arrest in human oral cancer SCC-4 cells.

    Science.gov (United States)

    Chou, Guan-Ling; Peng, Shu-Fen; Liao, Ching-Lung; Ho, Heng-Chien; Lu, Kung-Wen; Lien, Jin-Cherng; Fan, Ming-Jen; La, Kuang-Chi; Chung, Jing-Gung

    2018-02-01

    Casticin, a polymethoxyflavone, present in natural plants, has been shown to have biological activities including anti-cancer activities. Herein, we investigated the anti-oral cancer activity of casticin on SCC-4 cells in vitro. Viable cells, cell cycle distribution, apoptotic cell death, reactive oxygen species (ROS) production, and Ca 2+ production, levels of ΔΨ m and caspase activity were measured by flow cytometric assay. Cell apoptosis associated protein expressions were examined by Western blotting and confocal laser microscopy. Results indicated that casticin induced cell morphological changes, DNA condensation and damage, decreased the total viable cells, induced G 2 /M phase arrest in SCC-4 cells. Casticin promoted ROS and Ca 2+ productions, decreases the levels of ΔΨ m , promoted caspase-3, -8, and -9 activities in SCC-4 cells. Western blotting assay demonstrated that casticin affect protein level associated with G2/M phase arrest and apoptosis. Confocal laser microscopy also confirmed that casticin increased the translocation of AIF and cytochrome c in SCC-4 cells. In conclusion, casticin decreased cell number through G 2 /M phase arrest and the induction of cell apoptosis through caspase- and mitochondria-dependent pathways in SCC-4 cells. © 2017 Wiley Periodicals, Inc.

  13. Photo-oxidation of cells generates long-lived intracellular protein peroxides

    DEFF Research Database (Denmark)

    Wright, Adam; Hawkins, Clare Louise; Davies, Michael Jonathan

    2003-01-01

    Singlet oxygen is generated by several cellular, enzymatic, and chemical reactions as well as by exposure to UV or visible light in the presence of a sensitizer. Consequently, this oxidant has been proposed to be a damaging agent many pathologies. Proteins are major targets for singlet oxygen...... as a result of their abundance and high rate constants for reaction. In this study, we show that illumination of viable rose bengal-loaded THP-1 (human monocyte-like) cells with visible light gives rise to intracellular protein-derived peroxides. The peroxide yield increases with illumination time, requires....../2) about 4 h at 37 degrees C. Decomposition of protein peroxides formed within cells, or on isolated cellular proteins, by metal ions gives rise to radicals as detected by EPR spin trapping. These studies demonstrate that exposure of intact cells to visible light in the presence of a sensitizer leads...

  14. Mouse embryonic stem cells efficiently lipofected with nuclear localization peptide result in a high yield of chimeric mice and retain germline transmission potency.

    Science.gov (United States)

    Ma, Haiching; Liu, Qin; Diamond, Scott L; Pierce, Eric A

    2004-06-01

    Embryonic stem (ES) cells are an important tool in developmental biology, genomics, and transgenic methods, as well as in potential clinical applications such as gene therapy or tissue engineering. Electroporation is the standard transfection method for mouse ES (mES) cells because lipofection is quite inefficient. It is also unclear if mES cells treated with cationic lipids maintain pluripotency. We have developed a simple lipofection method for high efficiency transfection and stable transgene expression by employing the nonclassical nuclear localization signal M9 derived from the heterogeneous nuclear ribonucleoprotein A1. In contrast to using 20 microg DNA for 10 x 10(6) cells via electroporation which resulted in 10-20 positive cells/mm2, M9-assisted lipofection of 2 x 10(5) cells with 2 microg DNA resulted in > 150 positive cells/mm2. Electroporation produced only 0.16% EGFP positive cells with fluorescence intensity (FI) > 1000 by FACS assay, while M9-lipofection produced 36-fold more highly EGFP positive cells (5.75%) with FI > 1000. Using 2.5 x 10(6) ES cells and 6 microg linearized DNA followed by selection with G418, electroporation yielded 17 EGFP expressing colonies, while M9-assisted lipofection yielded 72 EGFP expressing colonies. The mES cells that stably expressed EGFP following M9-assisted lipofection yielded > 66% chimeric mice (8 of 12) and contributed efficiently to the germline. In an example of gene targeting, a knock-in mouse was produced from an ES clone screened from 200 G418-resistant colonies generated via M9-assisted lipofection. To our knowledge, this is the first report of generation of transgenic or knock-in mice obtained from lipofected mES cells and this method may facilitate large scale genomic studies of ES developmental biology or large scale generation of mouse models of human disease. Copyright 2003 Elsevier Inc.

  15. Extracellular delivery induced by ultrasound and microbubbles in cells

    Science.gov (United States)

    Hussein, Farah; Antonescu, Costin; Karshafian, Raffi

    2017-03-01

    Ultrasound and microbubble treatment (USMB) can enhance the intracellular uptake of molecules, which otherwise would be excluded from the cell, through USMB-mediated transient membrane disruption and through enhanced endocytosis. However, the effect of USMB on the outward movement of molecules from cells is not well understood. This study investigates the effects of USMB on the release of molecules from various cellular compartments including cytoplasm, lysosomes, and recycling endosomes. In vitro ARPE-19 (RPE henceforth) cells were loaded with Alexa fluor-labeled transferrin as a marker for recycling endosomes, LAMP-1 antibody was used to detect the fusion of lysosomes with the plasma membrane, GFP-transfected RPE cells were used to examine the release of GFP from the cytoplasm, and 7-AAD was used to assess cell viability. Subsequently, cells were exposed to USMB (106 cells/mL, 300 kPa peak negative pressure, 1 min treatment duration, and 20 µL/mL Definity microbubbles). Following USMB, the release of the fluorescent markers was examined at 1.5, 11.5, and 21.5 minutes from the start of USMB. The mean fluorescent intensity (MFI) of untreated and USMB treated samples were measured using flow cytometry. USMB increased the extracellular delivery of GFP molecules from the cytoplasm; the MFI in USMB treated GFP-transfected RPE cells decreased by 17% in viable cells and this MFI decreased by 70% in non-viable cells. This could be due to diffusion of GFP through the membrane disruptions induced by USMB. Additionally, the MFI of viable cells stained with LAMP-1 antibody increased by 50% and this increase was 15 folds in the non-viable cells indicating lysosome exocytosis as a mechanism for membrane repair. Furthermore, the MFI of cells loaded with fluorescent transferrin decreased by 22% after USMB treatment in viable cells, indicating a significant increase in transferrin recycling to the cell membrane. However, the increased recycling was not statistically significant

  16. First study on the formation and resuscitation of viable but nonculturable state and beer spoilage capability of Lactobacillus lindneri.

    Science.gov (United States)

    Liu, Junyan; Li, Lin; Li, Bing; Peters, Brian M; Deng, Yang; Xu, Zhenbo; Shirtliff, Mark E

    2017-06-01

    This study aimed to investigate the spoilage capability of Lactobacillus lindneri during the induction and resuscitation of viable but nonculturable (VBNC) state. L. lindneri strain was identified by sequencing the PCR product (amplifying 16S rRNA gene) using ABI Prism 377 DNA Sequencer. During the VBNC state induction by low temperature storage and beer adaption, total, culturable, and viable cells were assessed by acridine orange direct counting, plate counting, and Live/Dead BacLight bacterial viability kit, respectively. Organic acids and diacetyl concentration were measured by reversed-phase high performance liquid chromatography and head dpace gas chromatography, respectively. VBNC state of L. lindneri was successfully induced by both beer adaption and low temperature storage, and glycerol frozen stock was the optimal way to maintain the VBNC state. Addition of catalase was found to be an effective method for the resuscitation of VBNC L. lindneri cells. Furthermore, spoilage capability remained similar during the induction and resuscitation of VBNC L. lindneri. This is the first report of induction by low temperature storage and resuscitation of VBNC L. lindneri strain, as well as the first identification of spoilage capability of VBNC and resuscitated L. lindneri cells. This study indicated that the potential colonization of L. lindneri strain in brewery environment, formation and resuscitation of VBNC state, as well as maintenance in beer spoilage capability, may be an important risk factor for brewery environment. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. A BioDesign Approach to Obtain High Yields of Biosimilars by Anti-apoptotic Cell Engineering: a Case Study to Increase the Production Yield of Anti-TNF Alpha Producing Recombinant CHO Cells.

    Science.gov (United States)

    Gulce Iz, Sultan; Inevi, Muge Anil; Metiner, Pelin Saglam; Tamis, Duygu Ayyildiz; Kisbet, Nazli

    2018-01-01

    Recent developments in medical biotechnology have facilitated to enhance the production of monoclonal antibodies (mAbs) and recombinant proteins in mammalian cells. Human mAbs for clinical applications have focused on three areas, particularly cancer, immunological disorders, and infectious diseases. Tumor necrosis factor alpha (TNF-α), which has both proinflammatory and immunoregulatory functions, is an important target in biopharmaceutical industry. In this study, a humanized anti-TNF-α mAb producing stable CHO cell line which produces a biosimilar of Humira (adalimumab) was used. Adalimumab is a fully human anti-TNF mAb among the top-selling mAb products in recent years as a biosimilar. Products from mammalian cell bioprocesses are a derivative of cell viability and metabolism, which is mainly disrupted by cell death in bioreactors. Thus, different strategies are used to increase the product yield. Suppression of apoptosis, also called anti-apoptotic cell engineering, is the most remarkable strategy to enhance lifetime of cells for a longer production period. In fact, using anti-apoptotic cell engineering as a BioDesign approach was inspired by nature; nature gives prolonged life span to some cells like stem cells, tumor cells, and memory B and T cells, and researchers have been using this strategy for different purposes. In this study, as a biomimicry approach, anti-apoptotic cell engineering was used to increase the anti-TNF-α mAb production from the humanized anti-TNF-α mAb producing stable CHO cell line by Bcl-xL anti-apoptotic protein. It was shown that transient transfection of CHO cells by the Bcl-xL anti-apoptotic protein expressing plasmid prolonged the cell survival rate and protected cells from apoptosis. The transient expression of Bcl-xL using CHO cells enhanced the anti-TNF-α production. The production of anti-TNF-α in CHO cells was increased up to 215 mg/L with an increase of 160% after cells were transfected with Bcl-xL expressing plasmid

  18. Membrane Lipids as Indicators for Viable Bacterial Communities Inhabiting Petroleum Systems.

    Science.gov (United States)

    Gruner, Andrea; Mangelsdorf, Kai; Vieth-Hillebrand, Andrea; Horsfield, Brian; van der Kraan, Geert M; Köhler, Thomas; Janka, Christoph; Morris, Brandon E L; Wilkes, Heinz

    2017-08-01

    Microbial activity in petroleum reservoirs has been implicated in a suite of detrimental effects including deterioration of petroleum quality, increases in oil sulfur content, biofouling of steel pipelines and other infrastructures, and well plugging. Here, we present a biogeochemical approach, using phospholipid fatty acids (PLFAs), for detecting viable bacteria in petroleum systems. Variations within the bacterial community along water flow paths (producing well, topside facilities, and injection well) can be elucidated in the field using the same technique, as shown here within oil production plants in the Molasse Basin of Upper Austria. The abundance of PLFAs is compared to total cellular numbers, as detected by qPCR of the 16S rDNA gene, to give an overall comparison between the resolutions of both methods in a true field setting. Additionally, the influence of biocide applications on lipid- and DNA-based quantification was investigated. The first oil field, Trattnach, showed significant PLFA abundances and cell numbers within the reservoir and topside facilities. In contrast, the second field (Engenfeld) showed very low PLFA levels overall, likely due to continuous treatment of the topside facilities with a glutaraldehyde-based antimicrobial. In comparison, Trattnach is dosed once per week in a batch fashion. Changes within PLFA compositions across the flow path, throughout the petroleum production plants, point to cellular adaptation within the system and may be linked to shifts in the dominance of certain bacterial types in oil reservoirs versus topside facilities. Overall, PLFA-based monitoring provides a useful tool to assess the abundance and high-level taxonomic diversity of viable microbial populations in oil production wells, topside infrastructure, pipelines, and other related facilities.

  19. High-yield production of a stable Vero cell-based vaccine candidate against the highly pathogenic avian influenza virus H5N1

    International Nuclear Information System (INIS)

    Zhou, Fangye; Zhou, Jian; Ma, Lei; Song, Shaohui; Zhang, Xinwen; Li, Weidong; Jiang, Shude; Wang, Yue; Liao, Guoyang

    2012-01-01

    Highlights: ► Vero cell-based HPAI H5N1 vaccine with stable high yield. ► Stable high yield derived from the YNVa H3N2 backbone. ► H5N1/YNVa has a similar safety and immunogenicity to H5N1delta. -- Abstract: Highly pathogenic avian influenza (HPAI) viruses pose a global pandemic threat, for which rapid large-scale vaccine production technology is critical for prevention and control. Because chickens are highly susceptible to HPAI viruses, the supply of chicken embryos for vaccine production might be depleted during a virus outbreak. Therefore, developing HPAI virus vaccines using other technologies is critical. Meeting vaccine demand using the Vero cell-based fermentation process has been hindered by low stability and yield. In this study, a Vero cell-based HPAI H5N1 vaccine candidate (H5N1/YNVa) with stable high yield was achieved by reassortment of the Vero-adapted (Va) high growth A/Yunnan/1/2005(H3N2) (YNVa) virus with the A/Anhui/1/2005(H5N1) attenuated influenza vaccine strain (H5N1delta) using the 6/2 method. The reassorted H5N1/YNVa vaccine maintained a high hemagglutination (HA) titer of 1024. Furthermore, H5N1/YNVa displayed low pathogenicity and uniform immunogenicity compared to that of the parent virus.

  20. Preliminary Study on Testicular Germ Cell Transplantation of Endemic Species Oryzias celebensis

    Science.gov (United States)

    Andriani, I.; Agustiani, F.; Hassan, M.; Parenrengi, A.; Inoue, K.

    2018-03-01

    The research has been conducted to study some technical steps for male germ-plasm from endemic fish species such as some species of Oryzias fish in Indonesia to preserve and propagate through germ cell transplantation technology. For preliminary research, the study was started with germ cell characterization of testes, cryopreservation of TGC and the transplantation of Oryzias celebensis as candidates for surrogate broodstock of Oryzias fish male germ plasm. The data analized included the potential number of TGC as donor, the viability of cryopreserved TGC in two types of cryoprotectans and the survival rate of O.celebensis larvae as recipient after transplantation. The result showed that the average amount of TGC yielded after dissociation was 131000 ± 31349 with 74.2 % viability of TGC each. Cryoprotectan10% DMSO +glucose yielded higher viable of TGC. More than 80 % of O.celebensis larvae survived after transplantation. In conclusion, these preliminary data of O.celebensis as surrogate broodstock candidate will support the application of TGC transplantation technology in Oryzias endemic species.

  1. A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Shima P Damodaran

    Full Text Available To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers and a significant subpopulation of slowly dividing cells (slow-growers. These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

  2. Efficacy of zinc with nitrogen as foliar feeding on growth, yield and quality of tomato grown under poly tunnel

    International Nuclear Information System (INIS)

    Awan, A.A.; Abbas, S.J.; Ullah, E.

    2012-01-01

    Zinc (Zn) deficiency is considered one of nutritional constraints for crop yield worldwide. In recent past, the deficiency of Zn has heavily declined the quality of vegetables, especially tomato. Thus, a study was planned to enhance the growth, yield and quality of tomato plant using Zn and N alone and in combination. ZnSO/sub 4/ was used as a source of Zn (10% and 12%) and urea as source of N (1% and 2%). The results showed that application of either Zn or N alone at both concentrations enhanced growth, yield and quality of tomato plants under poly tunnel. However, the combined use of both Zn and N further enhanced the growth, yield and fruit quality with application of Zn (12%) plus N (2%). Hence the combined use of Zn and N can be a viable strategy for improving yield and quality of tomato. (author)

  3. Modeling the yield potential of dryland canola under current and future climates in California

    Science.gov (United States)

    George, N.; Kaffka, S.; Beeck, C.; Bucaram, S.; Zhang, J.

    2012-12-01

    Models predict that the climate of California will become hotter, drier and more variable under future climate change scenarios. This will lead to both increased irrigation demand and reduced irrigation water availability. In addition, it is predicted that most common Californian crops will suffer a concomitant decline in productivity. To remain productive and economically viable, future agricultural systems will need to have greater water use efficiency, tolerance of high temperatures, and tolerance of more erratic temperature and rainfall patterns. Canola (Brassica napus) is the third most important oilseed globally, supporting large and well-established agricultural industries in Canada, Europe and Australia. It is an agronomically useful and economically valuable crop, with multiple end markets, that can be grown in California as a dryland winter rotation with little to no irrigation demand. This gives canola great potential as a new crop for Californian farmers both now and as the climate changes. Given practical and financial limitations it is not always possible to immediately or widely evaluate a crop in a new region. Crop production models are therefore valuable tools for assessing the potential of new crops, better targeting further field research, and refining research questions. APSIM is a modular modeling framework developed by the Agricultural Production Systems Research Unit in Australia, it combines biophysical and management modules to simulate cropping systems. This study was undertaken to examine the yield potential of Australian canola varieties having different water requirements and maturity classes in California using APSIM. The objective of the work was to identify the agricultural regions of California most ideally suited to the production of Australian cultivars of canola and to simulate the production of canola in these regions to estimate yield-potential. This will establish whether the introduction and in-field evaluation of better

  4. Higher biomolecules yield in phytoplankton under copper exposure.

    Science.gov (United States)

    Silva, Jaqueline Carmo; Echeveste, Pedro; Lombardi, Ana Teresa

    2018-05-30

    Copper is an important metal for industry, and its toxic threshold in natural ecosystems has increased since the industrial revolution. As an essential nutrient, it is required in minute amounts, being toxic in slightly increased concentrations, causing great biochemical transformation in microalgae. This study aimed at investigating the physiology of Scenedesmus quadricauda, a cosmopolitan species, exposed to copper concentrations including those that trigger intracellular biochemical modifications. The Cu exposure concentrations tested ranged from 0.1 to 25 µM, thus including environmentally important levels. Microalgae cultures were kept under controlled environmental conditions and monitored daily for cell density, in vivo chlorophyll a, and photosynthetic quantum yield (Φ M ). After 24 h growth, free Cu 2+ ions were determined, and after 96 h, cellular Cu concentration, total carbohydrates, proteins, lipids, and cell volume were determined. The results showed that both free Cu 2+ ions and cellular Cu increased with Cu increase in culture medium. Microalgae cell abundance and in vivo chlorophyll a were mostly affected at 2.5 µM Cu exposure (3.8 pg Cu cell -1 ) and above. Approximately 31% decrease of photosynthetic quantum yield was obtained at the highest Cu exposure concentration (25 µM; 25 pg Cu cell -1 ) in comparison with the control. However, at environmentally relevant copper concentrations (0.5 µM Cu; 0.4 pg Cu cell -1 ) cell volume increased in comparison with the control. Considering biomolecules accumulation per unit cell volume, the highest carbohydrates and proteins yield was obtained at 1.0 µM Cu (1.1 pg Cu cell -1 ), while for lipids higher Cu was necessary (2.5 µM Cu; 3.8 pg Cu cell -1 ). This study is a contribution to the understanding of the effects of environmentally significant copper concentrations in the physiology of S. quadricauda, as well as to biotechnological approach to increase biomolecule yield in

  5. Effect of plastic mulch on growth and yield of chilli (Capsicum annuum L.

    Directory of Open Access Journals (Sweden)

    M Ashrafuzzaman

    2011-04-01

    Full Text Available In this work a field study was conducted to evaluate the effect of coloured plastic mulch on growth and yield of chilli from October 2005 to April 2006. The plastic mulches were transparent, blue, and black and bare soil was the control. Different mulches generated higher soil temperature and soil moisture under mulch over the control. Transparent and blue plastic mulches encouraged weed population which were suppressed under black plastic. Plant height, number of primary branches, stem base diameter, number of leaves and yield were better for the plants on plastic. At the mature green stage, fruits had the highest vitamin-C content on the black plastic. Mulching produced the fruits with the highest chlorophyll-a, chlorophyll-b and total chlorophyll contents and also increased the number of fruits per plant and yield. However, mulching did not affect the length and diameter of the fruits and number of seeds per fruit. Plants on black plastic mulch had the maximum number of fruits and highest yield. Thus, mulching appears to be a viable tool to increase the chilli production under tropical conditions.

  6. Detection of viable Helicobacter pylori inside free-living amoebae in wastewater and drinking water samples from Eastern Spain.

    Science.gov (United States)

    Moreno-Mesonero, Laura; Moreno, Yolanda; Alonso, José Luis; Ferrús, M Antonia

    2017-10-01

    Helicobacter pylori is one of the most concerning emerging waterborne pathogens. It has been suggested that it could survive in water inside free-living amoebae (FLA), but nobody has studied this relationship in the environment yet. Thus, we aimed to detect viable H. pylori cells from inside FLA in water samples. Sixty-nine wastewater and 31 drinking water samples were collected. FLA were purified and identified by PCR and sequencing. For exclusively detecting H. pylori inside FLA, samples were exposed to sodium hypochlorite and assayed by specific PMA-qPCR, DVC-FISH and culture. FLA were detected in 38.7% of drinking water and 79.7% of wastewater samples, even after disinfection. In wastewater, Acanthamoeba spp. and members of the family Vahlkampfiidae were identified. In drinking water, Acanthamoeba spp. and Echinamoeba and/or Vermamoeba were present. In 39 (58.2%) FLA-positive samples, H. pylori was detected by PMA-qPCR. After DVC-FISH, 21 (31.3%) samples harboured viable H. pylori internalized cells. H. pylori was cultured from 10 wastewater samples. To our knowledge, this is the first report that demonstrates that H. pylori can survive inside FLA in drinking water and wastewater, strongly supporting the hypothesis that FLA could play an important role in the transmission of H. pylori to humans. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  7. Production process reproducibility and product quality consistency of transient gene expression in HEK293 cells with anti-PD1 antibody as the model protein.

    Science.gov (United States)

    Ding, Kai; Han, Lei; Zong, Huifang; Chen, Junsheng; Zhang, Baohong; Zhu, Jianwei

    2017-03-01

    Demonstration of reproducibility and consistency of process and product quality is one of the most crucial issues in using transient gene expression (TGE) technology for biopharmaceutical development. In this study, we challenged the production consistency of TGE by expressing nine batches of recombinant IgG antibody in human embryonic kidney 293 cells to evaluate reproducibility including viable cell density, viability, apoptotic status, and antibody yield in cell culture supernatant. Product quality including isoelectric point, binding affinity, secondary structure, and thermal stability was assessed as well. In addition, major glycan forms of antibody from different batches of production were compared to demonstrate glycosylation consistency. Glycan compositions of the antibody harvested at different time periods were also measured to illustrate N-glycan distribution over the culture time. From the results, it has been demonstrated that different TGE batches are reproducible from lot to lot in overall cell growth, product yield, and product qualities including isoelectric point, binding affinity, secondary structure, and thermal stability. Furthermore, major N-glycan compositions are consistent among different TGE batches and conserved during cell culture time.

  8. Production of viable male unreduced gametes in Brassica interspecific hybrids is genotype specific and stimulated by cold temperatures

    Directory of Open Access Journals (Sweden)

    Cowling Wallace A

    2011-06-01

    Full Text Available Abstract Background Unreduced gametes (gametes with the somatic chromosome number may provide a pathway for evolutionary speciation via allopolyploid formation. We evaluated the effect of genotype and temperature on male unreduced gamete formation in Brassica allotetraploids and their interspecific hybrids. The frequency of unreduced gametes post-meiosis was estimated in sporads from the frequency of dyads or giant tetrads, and in pollen from the frequency of viable giant pollen compared with viable normal pollen. Giant tetrads were twice the volume of normal tetrads, and presumably resulted from pre-meiotic doubling of chromosome number. Giant pollen was defined as pollen with more than 1.5 × normal diameter, under the assumption that the doubling of DNA content in unreduced gametes would approximately double the pollen cell volume. The effect of genotype was assessed in five B. napus, two B. carinata and one B. juncea parents and in 13 interspecific hybrid combinations. The effect of temperature was assessed in a subset of genotypes in hot (day/night 30°C/20°C, warm (25°C/15°C, cool (18°C/13°C and cold (10°C/5°C treatments. Results Based on estimates at the sporad stage, some interspecific hybrid genotypes produced unreduced gametes (range 0.06 to 3.29% at more than an order of magnitude higher frequency than in the parents (range 0.00% to 0.11%. In nine hybrids that produced viable mature pollen, the frequency of viable giant pollen (range 0.2% to 33.5% was much greater than in the parents (range 0.0% to 0.4%. Giant pollen, most likely formed from unreduced gametes, was more viable than normal pollen in hybrids. Two B. napus × B. carinata hybrids produced 9% and 23% unreduced gametes based on post-meiotic sporad observations in the cold temperature treatment, which was more than two orders of magnitude higher than in the parents. Conclusions These results demonstrate that sources of unreduced gametes, required for the triploid

  9. Expression of proposed implantation marker genes CDX2 and HOXB7 in the blastocyst does not distinguish viable from non-viable human embryos

    DEFF Research Database (Denmark)

    Kirkegaard, Kirstine; Hindkjær, Johnny Juhl; Ingerslev, Hans Jakob

    2012-01-01

    expression differs between viable and non-viable embryos in both human and non-humans, suggesting transcriptome analysis of trophectoderm (TE) as a novel method of improving embryo selection. Potential candidate marker genes have been identified with array studies on animal blastocysts. The aim of this study...... was to investigate the expression of selected genes in human blastocysts in relation to the outcome of implantation. Materials and methods: Embryos from 10 oatients undergoing in vitro fertilization treatment were included in the project. A single blastocyst was chosen for biopsy on the morning of day 5 after oocyte...... of 15 key genes associated with developmental competence in animals were evaluated in high quality human embryos with monogenic or chromosomal disorders from a pre-implantation genetic disorder program. Triplicate cDNA amplifications for quantitative (q) RT-PCR were performed using pre-designed gene...

  10. High-yield production of a stable Vero cell-based vaccine candidate against the highly pathogenic avian influenza virus H5N1

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Fangye; Zhou, Jian; Ma, Lei; Song, Shaohui; Zhang, Xinwen; Li, Weidong; Jiang, Shude [No. 5, Department of Bioproducts, Institute of Medical Biology, Chinese Academy of Medical Science and Pecking Union Medical College, Jiaoling Avenue 935, Kunming, Yunnan Province 650102, People' s Republic of China (China); Wang, Yue [National Institute for Viral Disease Control and Prevention, China Center for Disease Control and Prevention, Yingxin Lane 100, Xicheng District, Beijing 100052, People' s Republic of China (China); Liao, Guoyang [No. 5, Department of Bioproducts, Institute of Medical Biology, Chinese Academy of Medical Science and Pecking Union Medical College, Jiaoling Avenue 935, Kunming, Yunnan Province 650102, People' s Republic of China (China)

    2012-05-18

    Highlights: Black-Right-Pointing-Pointer Vero cell-based HPAI H5N1 vaccine with stable high yield. Black-Right-Pointing-Pointer Stable high yield derived from the YNVa H3N2 backbone. Black-Right-Pointing-Pointer H5N1/YNVa has a similar safety and immunogenicity to H5N1delta. -- Abstract: Highly pathogenic avian influenza (HPAI) viruses pose a global pandemic threat, for which rapid large-scale vaccine production technology is critical for prevention and control. Because chickens are highly susceptible to HPAI viruses, the supply of chicken embryos for vaccine production might be depleted during a virus outbreak. Therefore, developing HPAI virus vaccines using other technologies is critical. Meeting vaccine demand using the Vero cell-based fermentation process has been hindered by low stability and yield. In this study, a Vero cell-based HPAI H5N1 vaccine candidate (H5N1/YNVa) with stable high yield was achieved by reassortment of the Vero-adapted (Va) high growth A/Yunnan/1/2005(H3N2) (YNVa) virus with the A/Anhui/1/2005(H5N1) attenuated influenza vaccine strain (H5N1delta) using the 6/2 method. The reassorted H5N1/YNVa vaccine maintained a high hemagglutination (HA) titer of 1024. Furthermore, H5N1/YNVa displayed low pathogenicity and uniform immunogenicity compared to that of the parent virus.

  11. Quantitative real-time PCR combined with propidium monoazide for the selective quantification of viable periodontal pathogens in an in vitro subgingival biofilm model.

    Science.gov (United States)

    Sánchez, M C; Marín, M J; Figuero, E; Llama-Palacios, A; León, R; Blanc, V; Herrera, D; Sanz, M

    2014-02-01

    Differentiation of live and dead cells is an important challenge when using molecular diagnosis for microbial identification. This is particularly relevant when bacteria have been exposed to antimicrobial agents. The objective of this study was to test a method using quantitative real-time polymerase chain reaction (qPCR) combined with propidium monoazide (PMA), developed for the selective quantification of viable P. gingivalis, A. actinomycetemcomitans, F. nucleatum and total bacteria in an in vitro biofilm model after antimicrobial treatment. PMA-qPCR method was tested in an in vitro biofilm model, using isopropyl alcohol as the antimicrobial agent. Matured biofilms were exposed for 1, 5, 10 and 30 min to isopropyl alcohol by immersion. Biofilms were disrupted and PMA added (final concentration of 100 μm). After DNA isolation, qPCR was carried out using specific primers and probes for the target bacteria. The differentiation of live and dead cells was tested by analysis of variance. When PMA was used in the presence of viable target bacterial cells, no statistically significant inhibition of qPCR amplification was detected (p > 0.05 in all cases). Conversely, after immersion in isopropyl alcohol of the biofilm, PMA resulted in a significant total reduction of qPCR amplification of about 4 log10 . P. gingivalis showed a vitality reduction in the biofilm of 3 log10 , while A. actinomycetemcomitans and F. nucleatum showed a 2 log10 reduction. These results demonstrate the efficiency of PMA for differentiating viable and dead P. gingivalis, A. actinomycetemcomitans and F. nucleatum cells, as well as total bacteria, in an in vitro biofilm model, after being exposed to an antimicrobial agent. Hence, this PMA-qPCR method may be useful for studying the effect of antimicrobial agents aimed at oral biofilms. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  12. Genetic relationships among linear type traits, milk yield, body weight, fertility and somatic cell count in primiparous dairy cows

    NARCIS (Netherlands)

    Berry, D.P.; Buckley, F.; Dillon, P.P.; Evans, R.D.; Veerkamp, R.F.

    2004-01-01

    Phenotypic and genetic (co)variances among type traits, milk yield, body weight, fertility and somatic cell count were estimated. The data analysed included 3,058 primiparous spring-calving Holstein-Friesian cows from 80 farms throughout the south of Ireland. Heritability estimates for the type

  13. Genetic relationship of lactation persistency with milk yield, somatic cell score, reproductive traits, and longevity in Slovak Holstein cattle

    OpenAIRE

    Strapáková, Eva; Candrák, Juraj; Strapák, Peter

    2016-01-01

    The objective of this study was to estimate the breeding values (BVs) of lactation persistency, the test day of milk yield, the somatic cell score, reproductive traits (calving interval, days open), longevity in Slovak Holstein dairy cattle. BVs were used for the detection of relationships among the persistency of lactation and other selected traits. Data for the estimation of BVs of milk production and somatic cell score were collected from 855 240 cows. BVs for reproductive t...

  14. Cell wall metabolism and hexose allocation contribute to biomass accumulation in high yielding extreme segregants of a Saccharum interspecific F2 population.

    Science.gov (United States)

    Wai, Ching Man; Zhang, Jisen; Jones, Tyler C; Nagai, Chifumi; Ming, Ray

    2017-10-11

    Sugarcane is an emerging dual-purpose biofuel crop for energy and sugar production, owing to its rapid growth rate, high sucrose storage in the stems, and high lignocellulosic yield. It has the highest biomass production reaching 1.9 billion tonnes in 2014 worldwide. To improve sugarcane biomass accumulation, we developed an interspecific cross between Saccharum officinarum 'LA Purple' and Saccharum robustum 'MOL5829'. Selected F1 individuals were self-pollinated to generate a transgressive F2 population with a wide range of biomass yield. Leaf and stem internodes of fourteen high biomass and eight low biomass F2 extreme segregants were used for RNA-seq to decipher the molecular mechanism of rapid plant growth and dry weight accumulation. Gene Ontology terms involved in cell wall metabolism and carbohydrate catabolism were enriched among 3274 differentially expressed genes between high and low biomass groups. Up-regulation of cellulose metabolism, pectin degradation and lignin biosynthesis genes were observed in the high biomass group, in conjunction with higher transcript levels of callose metabolic genes and the cell wall loosening enzyme expansin. Furthermore, UDP-glucose biosynthesis and sucrose conversion genes were differentially expressed between the two groups. A positive correlation between stem glucose, but not sucrose, levels and dry weight was detected. We thus postulated that the high biomass sugarcane plants rapidly convert sucrose to UDP-glucose, which is the building block of cell wall polymers and callose, in order to maintain the rapid plant growth. The gene interaction of cell wall metabolism, hexose allocation and cell division contributes to biomass yield.

  15. Sedimentation properties of DNA-membrane complexes and yield of DNA breaks at irradiation of mammalian cells

    International Nuclear Information System (INIS)

    Erzgraber, G.; Kozubek, S.; Lapidus, I.L.

    1985-01-01

    The dependence of the relative sedimentation velocity of DNA-membrane complexes on the dose of irradiation and time of incubation of Chinese Hamster cells is analysed. It is concluded that the initial part of the curve provides the information on the occurrence of single strand breaks in DNA; the position of the local maximum allows us to calculate the yield of DNA double strand breaks. The reparation decay constant can be estimated as well

  16. Immunogenicity of ascites tumor cells following in vitro hyperthermia

    International Nuclear Information System (INIS)

    Dickson, J.A.; Jasiewicz, M.L.; Simpson, A.C.

    1982-01-01

    The concept that host immunization may be achieved by heat-induced antigenic modifications of cancer cells and/or the release of immunogenic products by dead or dying tumor cells following in vitro heating was examined. Ehrlich ascites cells were used, inasmuch as it was claimed that in vitro hyperthermia increased the immunogenicity of these cells. Tumor cell populations of different viability were obtained by heating Ehrlich cells at 42.5 degrees, 45 degrees, or 60 degrees C. Viable and nonviable cells were separated by Ficoll-Hypaque density centrifugation; viable nonreplicating cells were obtained by treatment with mitomycin C. Cell populations of different viability after heating were left to die slowly over 3 days at 37 degrees C. Swiss TO mice were then given injections of the treated cells and/or medium. No survival benefit occurred in mice inoculated with any of these different components and then challenged with viable tumor cells. Injection of irradiated cells, however, did produce host immunity. Similarly, D23 rat hepatoma ascites cells produced host immunity after 15,000 rad but not after heating. The claim that in vitro hyperthermia increases the immunogenicity of tumor cells was not confirmed

  17. Skills training workshops as a viable strategy for improving ...

    African Journals Online (AJOL)

    Skills training workshops as a viable strategy for improving smallholder and cooperative agribusiness management: A case study of Vhembe District, Limpopo Province, South Africa. ... South African Journal of Agricultural Extension ... Empirical evidence from this study shows that six months after attending the workshops, ...

  18. Examination of an indicative tool for rapidly estimating viable organism abundance in ballast water

    Science.gov (United States)

    Vanden Byllaardt, Julie; Adams, Jennifer K.; Casas-Monroy, Oscar; Bailey, Sarah A.

    2018-03-01

    Regulatory discharge standards stipulating a maximum allowable number of viable organisms in ballast water have led to a need for rapid, easy and accurate compliance assessment tools and protocols. Some potential tools presume that organisms present in ballast water samples display the same characteristics of life as the native community (e.g. rates of fluorescence). This presumption may not prove true, particularly when ships' ballast tanks present a harsh environment and long transit times, negatively impacting organism health. Here, we test the accuracy of a handheld pulse amplitude modulated (PAM) fluorometer, the Hach BW680, for detecting photosynthetic protists at concentrations above or below the discharge standard (< 10 cells·ml- 1) in comparison to microscopic counts using fluorescein diacetate as a viability probe. Testing was conducted on serial dilutions of freshwater harbour samples in the lab and in situ untreated ballast water samples originating from marine, freshwater and brackish sources utilizing three preprocessing techniques to target organisms in the size range of ≥ 10 and < 50 μm. The BW680 numeric estimates were in agreement with microscopic counts when analyzing freshly collected harbour water at all but the lowest concentrations (< 38 cells·ml- 1). Chi-square tests determined that error is not independent of preprocessing methods: using the filtrate method or unfiltered water, in addition to refining the conversion factor of raw fluorescence to cell size, can decrease the grey area where exceedance of the discharge standard cannot be measured with certainty (at least for the studied populations). When examining in situ ballast water, the BW680 detected significantly fewer viable organisms than microscopy, possibly due to factors such as organism size or ballast water age. Assuming both the BW680 and microscopy with FDA stain were measuring fluorescence and enzymatic activity/membrane integrity correctly, the observed discrepancy

  19. Talons and beaks are viable but underutilized samples for detecting ...

    African Journals Online (AJOL)

    Talons and beaks are viable but underutilized samples for detecting organophosphorus and carbamate pesticide poisoning in raptors. Ngaio Richards, Irene Zorrilla, Joseph Lalah, Peter Otieno, Isabel Fernandez, Monica Calvino, Joaquin Garcia ...

  20. The yield of DNA double strand breaks determined after exclusion of those forming from heat-labile lesions predicts tumor cell radiosensitivity to killing.

    Science.gov (United States)

    Cheng, Yanlei; Li, Fanghua; Mladenov, Emil; Iliakis, George

    2015-09-01

    The radiosensitivity to killing of tumor cells and in-field normal tissue are key determinants of radiotherapy response. In vitro radiosensitivity of tumor- and normal-tissue-derived cells often predicts radiation response, but high determination cost in time and resources compromise utility as routine response-predictor. Efforts to use induction or repair of DNA double-strand-breaks (DSBs) as surrogate-predictors of cell radiosensitivity to killing have met with limited success. Here, we re-visit this issue encouraged by our recent observations that ionizing radiation (IR) induces not only promptly-forming DSBs (prDSBs), but also DSBs developing after irradiation from the conversion to breaks of thermally-labile sugar-lesions (tlDSBs). We employ pulsed-field gel-electrophoresis and flow-cytometry protocols to measure total DSBs (tDSB=prDSB+tlDSBs) and prDSBs, as well as γH2AX and parameters of chromatin structure. We report a fully unexpected and in many ways unprecedented correlation between yield of prDSBs and radiosensitivity to killing in a battery of ten tumor cell lines that is not matched by yields of tDSBs or γH2AX, and cannot be explained by simple parameters of chromatin structure. We propose the introduction of prDSBs-yield as a novel and powerful surrogate-predictor of cell radiosensitivity to killing with potential for clinical application. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  1. Quantitative assessment of viable Cryptosporidium parvum load in commercial oysters (Crassostrea virginica) in the Chesapeake Bay.

    Science.gov (United States)

    Graczyk, Thaddeus K; Lewis, Earl J; Glass, Gregory; Dasilva, Alexandre J; Tamang, Leena; Girouard, Autumn S; Curriero, Frank C

    2007-01-01

    The epidemiological importance of increasing reports worldwide on Cryptosporidium contamination of oysters remains unknown in relation to foodborne cryptosporidiosis. Thirty market-size oysters (Crassostrea virginica), collected from each of 53 commercial harvesting sites in Chesapeake Bay, MD, were quantitatively tested in groups of six for Cryptosporidium sp. oocysts by immunofluorescent antibody (IFA). After IFA analysis, the samples were retrospectively retested for viable Cryptosporidium parvum oocysts by combined fluorescent in situ hybridization (FISH) and IFA. The mean cumulative numbers of Cryptosporidium sp. oocysts in six oysters (overall, 42.1+/-4.1) were significantly higher than in the numbers of viable C. parvum oocysts (overall, 28.0+/-2.9). Of 265 oyster groups, 221 (83.4%) contained viable C. parvum oocysts, and overall, from 10-32% (mean, 23%) of the total viable oocysts were identified in the hemolymph as distinct from gill washings. The amount of viable C. parvum oocysts was not related to oyster size or to the level of fecal coliforms at the sampling site. This study demonstrated that, although oysters are frequently contaminated with oocysts, the levels of viable oocysts may be too low to cause infection in healthy individuals. FISH assay for identification can be retrospectively applied to properly stored samples.

  2. An invention of thermo-responsive polymer surface, yielding cell sheet based regenerative therapies in cardiology and ophthalmology

    Directory of Open Access Journals (Sweden)

    Sawa Y

    2015-12-01

    Full Text Available The Invention: In vitro cell culture methodologies provide a conducive environment for the cells taken out of their native environment to grow and proliferate in a non-physiological environment, the culture dish. Research experiments have been focusing on various criteria for assessing how far it is possible to recapitulate the native extra-cellular environment in vitro. Scaffolds, culture media, growth factors and cell surface modified culture dishes are some of the components that provide a conducive environment for in vitro cell culture. Cells that are grown in culture dishes using conventional methodologies are usually detached using enzymatic treatment with Trypsin, Collagenase etc [1], to be transplanted when it comes to a clinical or experimental application. Such enzymes used in separating the cells may have some damaging effects to the cell membranes which might impair the cell function [1]. However, cells if can be grown as a monolayer and be harvested as a contiguous cell sheet, it is considered suitable for transplantation in certain specific applications. In addition to that, if enzymatic digestion which has some detrimental effects on the detached cells could be avoided, that is an added advantage. The work by Prof. Okano and team from the Tokyo Women's Medical University, Japan, on thermo-responsive polymer surfaces has yielded a solution which has both the advantages viz., detachability of cells grown as a monolayer in the form of a cell sheet, that too without the use of enzymes. Their research into biomaterials for more than two decades has yielded a thermo-responsive polymer, the poly(N-isopropylacrylamide (PIPAAm [1] coated culture dish for cell sheet engineering. In their technology, PIPAAm is polymerized and grafted to tissue culture polystyrene (TCPS dishes. Cells have been found to grow confluent on PIPAAm-TCPS at 37 °C. Once confluent as a monolayer, by merely reducing the temperature of the PIPAAm-TCPS to 20 °C, it

  3. Molecular Characterization of Gastric Epithelial Cells Using Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Kevin A. Bockerstett

    2018-04-01

    Full Text Available The ability to analyze individual epithelial cells in the gastric mucosa would provide important insight into gastric disease, including chronic gastritis and progression to gastric cancer. However, the successful isolation of viable gastric epithelial cells (parietal cells, neck cells, chief cells, and foveolar cells from gastric glands has been limited due to difficulties in tissue processing. Furthermore, analysis and interpretation of gastric epithelial cell flow cytometry data has been difficult due to the varying sizes and light scatter properties of the different epithelial cells, high levels of autofluorescence, and poor cell viability. These studies were designed to develop a reliable method for isolating viable single cells from the corpus of stomachs and to optimize analyses examining epithelial cells from healthy and diseased stomach tissue by flow cytometry. We performed a two stage enzymatic digestion in which collagenase released individual gastric glands from the stromal tissue of the corpus, followed by a Dispase II digestion that dispersed these glands into greater than 1 × 106 viable single cells per gastric corpus. Single cell suspensions were comprised of all major cell lineages found in the normal gastric glands. A method describing light scatter, size exclusion, doublet discrimination, viability staining, and fluorescently-conjugated antibodies and lectins was used to analyze individual epithelial cells and immune cells. This technique was capable of identifying parietal cells and revealed that gastric epithelial cells in the chronically inflamed mucosa significantly upregulated major histocompatibility complexes (MHC I and II but not CD80 or CD86, which are costimulatory molecules involved in T cell activation. These studies describe a method for isolating viable single cells and a detailed description of flow cytometric analysis of cells from healthy and diseased stomachs. These studies begin to identify effects of

  4. Potential limitations for potato yields in raised soil field systems near Lake Titicaca

    Directory of Open Access Journals (Sweden)

    Lozada Diego Sánchez de

    2006-01-01

    Full Text Available During the last two decades, various non-governmental organizations have strongly encouraged Bolivian farmers in the Altiplano region near Lake Titicaca to resume the ancestral agricultural practice of constructing raised fields. In addition to improved drainage and possibilities this practice affords for sub-irrigation, advocated benefits of this system traditionally include frost mitigation and high crop yields. Until recently, reliable data to assess the extent of these benefits were unfortunately lacking. In this context, field experiments on raised fields were designed and carried out at two locations in the Bolivian Altiplano to obtain reliable potato yield and temperature data. Observed yields ranged from 2.73 to 10.80 t ha-1 at the first site, where salinity caused significant yield variability (R² = 0.79. At the second site, yields per raised platform varied between 8.25 and 33.45 t ha-1. However, comparable yields were obtained in flat control plots in spite of a mid-season frost, and the minimum temperatures differed only by 1ºC in the conventional plots relative to the raised fields. These results suggest that, under the experimental conditions, the potential benefits of raised fields in terms of frost mitigation or increased yields might only be observable in exceptionally bad years, when extreme frosts wipe out entire potato crops on conventional fields. Nevertheless, it is argued that in spite of these marginally supportive observations, raised-field agriculture may still be a viable option for farmers to consider if the water-filled channels between the raised fields are managed for fish and fertilizer production.

  5. Economics and yields of energy plantations: Status and potential, 1992-1993 update

    International Nuclear Information System (INIS)

    Gambles, R.L.; Kenney, W.A.

    1994-01-01

    An update is presented of a study carried out to: determine the factors affecting the cost of energy conversion feedstocks in short rotation intensive culture plantations of trees; determine the factors influencing biomass yield; identify interrelationships between the previous two objectives; present estimates of potential biomass yields and associated economics; and to identify gaps in the knowledge of the economics and yields of biomass production and their interrelationships. Developments in economics and yields in short rotation intensive silviculture for the production of biomass energy since 1991 are documented. The most substantial changes have been: the introduction of new clones in Sweden with a 20% increase in yield; illustrating the potential genetic gains achievable through selection and breeding; and halving of harvesting costs with new machinery. Harvesting costs with chipping incorporated have fallen to $51.21/dry tonne. The twin row ESM and Frobbester harvesters have lower estimated costs of $36.62 and $ 33.69 respectively. Agricultural based machines have further reduced costs to $19.42 and $26.12/dry tonne. Using these new data, three new scenarios were developed for cost of production analysis, using contract labour, farm labour or farm labour plus a subsidy. A contracted operation is now viable with an annual equivalent net value (AENV) of $35/ha. With the use of farm labour for most operations and omitting land rent, profitability increased to $127/ha. With a subsidy of $75/ha, the AENV increases to $205/ha. 25 refs., 1 fig., 3 tabs

  6. Comparison of egg and high yielding MDCK cell-derived live attenuated influenza virus for commercial production of trivalent influenza vaccine: in vitro cell susceptibility and influenza virus replication kinetics in permissive and semi-permissive cells.

    Science.gov (United States)

    Hussain, Althaf I; Cordeiro, Melissa; Sevilla, Elizabeth; Liu, Jonathan

    2010-05-14

    Currently MedImmune manufactures cold-adapted (ca) live, attenuated influenza vaccine (LAIV) from specific-pathogen free (SPF) chicken eggs. Difficulties in production scale-up and potential exposure of chicken flocks to avian influenza viruses especially in the event of a pandemic influenza outbreak have prompted evaluation and development of alternative non-egg based influenza vaccine manufacturing technologies. As part of MedImmune's effort to develop the live attenuated influenza vaccine (LAIV) using cell culture production technologies we have investigated the use of high yielding, cloned MDCK cells as a substrate for vaccine production by assessing host range and virus replication of influenza virus produced from both SPF egg and MDCK cell production technologies. In addition to cloned MDCK cells the indicator cell lines used to evaluate the impact of producing LAIV in cells on host range and replication included two human cell lines: human lung carcinoma (A549) cells and human muco-epidermoid bronchiolar carcinoma (NCI H292) cells. The influenza viruses used to infect the indicators cell lines represented both the egg and cell culture manufacturing processes and included virus strains that composed the 2006-2007 influenza seasonal trivalent vaccine (A/New Caledonia/20/99 (H1N1), A/Wisconsin/67/05 (H3N2) and B/Malaysia/2506/04). Results from this study demonstrate remarkable similarity between influenza viruses representing the current commercial egg produced and developmental MDCK cell produced vaccine production platforms. MedImmune's high yielding cloned MDCK cells used for the cell culture based vaccine production were highly permissive to both egg and cell produced ca attenuated influenza viruses. Both the A549 and NCI H292 cells regardless of production system were less permissive to influenza A and B viruses than the MDCK cells. Irrespective of the indicator cell line used the replication properties were similar between egg and the cell produced

  7. How Close We Are to Achieving Commercially Viable Large-Scale Photobiological Hydrogen Production by Cyanobacteria: A Review of the Biological Aspects

    Science.gov (United States)

    Sakurai, Hidehiro; Masukawa, Hajime; Kitashima, Masaharu; Inoue, Kazuhito

    2015-01-01

    Photobiological production of H2 by cyanobacteria is considered to be an ideal source of renewable energy because the inputs, water and sunlight, are abundant. The products of photobiological systems are H2 and O2; the H2 can be used as the energy source of fuel cells, etc., which generate electricity at high efficiencies and minimal pollution, as the waste product is H2O. Overall, production of commercially viable algal fuels in any form, including biomass and biodiesel, is challenging, and the very few systems that are operational have yet to be evaluated. In this paper we will: briefly review some of the necessary conditions for economical production, summarize the reports of photobiological H2 production by cyanobacteria, present our schemes for future production, and discuss the necessity for further progress in the research needed to achieve commercially viable large-scale H2 production. PMID:25793279

  8. Solar System constraints on a cosmologically viable f(R) theory

    Energy Technology Data Exchange (ETDEWEB)

    Bisabr, Yousef, E-mail: y-bisabr@srttu.ed [Department of Physics, Shahid Rajaee Teacher Training University, Lavizan, Tehran 16788 (Iran, Islamic Republic of)

    2010-01-18

    Recently, a model f(R) theory is proposed (Miranda et al. (2009)) which is cosmologically viable and distinguishable from LAMBDACDM. We use chameleon mechanism to investigate viability of the model in terms of Solar System experiments.

  9. Solar System constraints on a cosmologically viable f(R) theory

    International Nuclear Information System (INIS)

    Bisabr, Yousef

    2010-01-01

    Recently, a model f(R) theory is proposed (Miranda et al. (2009)) which is cosmologically viable and distinguishable from ΛCDM. We use chameleon mechanism to investigate viability of the model in terms of Solar System experiments.

  10. POOR HEMOPOIETIC STEM CELL MOBILIZERS IN MULTIPLE MYELOMA : A SINGLE INSTITUTION EXPERIENCE

    Directory of Open Access Journals (Sweden)

    Guillermo Jose Ruiz-Delgado

    2010-06-01

    Full Text Available In a single institution, in a group of 28 myeloma patients deemed eligible for autologous transplant, stem cell mobilization was attempted using filgrastim: 26 individuals were given 31 autografts employing 1-4 (median three apheresis sessions, to obtain a target stem cell dose of 1 x 106 CD34 viable cells / Kg of the recipient. The median number of grafted CD34 cells was 7.56 x 106  / Kg of the recipient; the range being 0.92 to 14.8.  By defining as poor mobilizers individuals in which a cell collection of 1 x 106 CD34 viable cells / Kg was better (80% at 80 months than those grafted with < 1 x 106 CD34 viable cells / Kg (67% at 76 months. Methods to improve stem cell mobilization are needed and may result in obtaining better results when autografting multiple myeloma patients.

  11. Comparison of diagnostic yield of endoscopic ultrasound-guided fine-needle aspiration cytology and cell block in solid lesions

    Directory of Open Access Journals (Sweden)

    Avinash Bhat Balekuduru

    2017-01-01

    Full Text Available Background and Aim: Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA is a procedure of choice for the diagnostic evaluation of submucosal and periluminal lesions. Tissue sample can be obtained by EUS-FNA cytology (FNAC or cell block (CB. The aim of the present study is to compare diagnostic yield of EUS-FNA CB and cytology in the absence of onsite pathologist following a protocol-based EUS-FNA approach in solid lesions. Patients and Methods: Participants who underwent EUS-FNA at our center for solid submucosal or periluminal lesions (pancreas, lymph node, and liver between 2014 and 2016 were included, retrospectively. The indication for the procedure along with the clinical and other investigation details and the final etiological diagnosis were recorded on uniform structured data forms. The diagnostic yield of cytology and CB were compared using McNemar's test. The P < 0.05 was considered statistically significant. Results: EUS-FNA for solid lesion was performed in 130 lesions in 101 patients during the study period. Their mean age was 52.5 ± 12 years and 42.5% were female. Pancreatic masses were the most common lesions (37.7% followed by lymph nodes (36.9%. Submucosal lesions (17.7% and liver lesions (7.7% accounted for rest of the cases. The overall diagnostic yield for EUS-FNAC (70% and CB (74.6% was not significantly different (P = 0.3 and their combined yield was 85.3%. For the 23 patients with submucosal lesion, diagnostic yield of CB (82.6% was significantly better than cytology (47.8%, P = 0.04. Conclusions: EUS-guided CB has better yield compared to cytology in gastrointestinal submucosal lesions. The combination of CB with cytology improves the overall yield of the procedure; and hence, they should be considered complimentary rather than alternatives.

  12. Evaluation of the transforming growth factor-beta activity in normal and dry eye human tears by CCL-185 cell bioassay.

    Science.gov (United States)

    Zheng, Xiaofen; De Paiva, Cintia S; Rao, Kavita; Li, De-Quan; Farley, William J; Stern, Michael; Pflugfelder, Stephen C

    2010-09-01

    To develop a new bioassay method using human lung epithelial cells (CCL-185) to assess activity of transforming growth factor beta (TGF-beta) in human tear fluid from normal subjects and patients with dry eye. Two epithelial cell lines, mink lung cells (CCL-64) and human lung cells (CCL-185), were compared to detect the active form of TGF-beta by BrdU incorporation (quantitation of cell DNA synthesis) and WST assay (metabolic activity of viable cells). The effect of TGF-beta on the growth of CCL-185 cells was observed microscopically. Human tears from normal control subjects and patients with dry eye (DE) with and without Sjögren syndrome were evaluated for TGF-beta concentration by Luminex microbead assay, and TGF-beta activity by the CCL-185 cell growth inhibition bioassay. The metabolic activity of viable CCL-185 cells, measured by WST, was shown to be proportional to the TGF-beta1 concentration (R = 0.919) and confirmed by BrdU assay (R = 0.969). Compared with CCL-185, metabolic activity of viable cells and DNA synthesis, measured by WST and BrdU incorporation assays, were shown to be less proportional to the TGF-beta1 concentration in the CCL-64 line (R = 0.42 and 0.17, respectively). Coincubation with human anti-TGF-beta1 antibody (MAB-240) yielded a dose-dependent inhibition of TGF-beta1 (0.3 ng/mL) activity. CCL-185 cell growth observed microscopically was noted to decrease in response to increasing TGF-beta1 concentrations. Levels of immuodetectable TGF-beta1 and TGF-beta2 were similar in normal and DE tears. TGF-beta bioactivity in DE human tears measured by the CCL-185 cells assay was found to be higher (9777.5 +/- 10481.9 pg/mL) than those in normal controls (4129.3 +/- 1342.9 pg/mL) (P tears and 37.6% TGF-beta in normal tears were found to be biologically active. The CCL-185 cell assay was found to be a suitable tool for assessing TGF-beta activity in human tears. Tear TGF-beta bioactivity increases in DE, particularly in Sjögren syndrome, where

  13. A viable real estate economy with disruption and blockchain

    NARCIS (Netherlands)

    Veuger, Jan

    2017-01-01

    Two titles in one cover. On page 56-112 there's the English version of the book: 'A viable real estate economy with disruption and blockchain. Does real estate still have the value that it had, or is the valuation of real estate going to change due to surprising products and services, innovative

  14. Promoting Women Participation in Aquaculture as a Viable Tool for ...

    African Journals Online (AJOL)

    Promoting Women Participation in Aquaculture as a Viable Tool for Poverty Alleviation in the Rural Areas of Nigeria. ... Open Access DOWNLOAD FULL TEXT ... a source of income, also the paper focus on the roles of women in aquaculture, ...

  15. A carbon dioxide stripping model for mammalian cell culture in manufacturing scale bioreactors.

    Science.gov (United States)

    Xing, Zizhuo; Lewis, Amanda M; Borys, Michael C; Li, Zheng Jian

    2017-06-01

    Control of carbon dioxide within the optimum range is important in mammalian bioprocesses at the manufacturing scale in order to ensure robust cell growth, high protein yields, and consistent quality attributes. The majority of bioprocess development work is done in laboratory bioreactors, in which carbon dioxide levels are more easily controlled. Some challenges in carbon dioxide control can present themselves when cell culture processes are scaled up, because carbon dioxide accumulation is a common feature due to longer gas-residence time of mammalian cell culture in large scale bioreactors. A carbon dioxide stripping model can be used to better understand and optimize parameters that are critical to cell culture processes at the manufacturing scale. The prevailing carbon dioxide stripping models in literature depend on mass transfer coefficients and were applicable to cell culture processes with low cell density or at stationary/cell death phase. However, it was reported that gas bubbles are saturated with carbon dioxide before leaving the culture, which makes carbon dioxide stripping no longer depend on a mass transfer coefficient in the new generation cell culture processes characterized by longer exponential growth phase, higher peak viable cell densities, and higher specific production rate. Here, we present a new carbon dioxide stripping model for manufacturing scale bioreactors, which is independent of carbon dioxide mass transfer coefficient, but takes into account the gas-residence time and gas CO 2 saturation time. The model was verified by CHO cell culture processes with different peak viable cell densities (7 to 12 × 10 6  cells mL -1 ) for two products in 5,000-L and 25,000-L bioreactors. The model was also applied to a next generation cell culture process to optimize cell culture conditions and reduce carbon dioxide levels at manufacturing scale. The model provides a useful tool to understand and better control cell culture carbon dioxide

  16. Cybernetically sound organizational structures II: Relating de Sitter's design theory to Beer's viable system model

    NARCIS (Netherlands)

    Achterbergh, J.M.I.M.; Vriens, D.J.

    2011-01-01

    - Purpose – The purpose of this paper is to show how the viable system model (VSM) and de Sitter's design theory can complement each other in the context of the diagnosis and design of viable organizations. - Design/methodology/approach – Key concepts from Beer's model and de Sitter's design theory

  17. Infra-red laser ablative micromachining of parylene-C on SiO2 substrates for rapid prototyping, high yield, human neuronal cell patterning

    International Nuclear Information System (INIS)

    Raos, B J; Unsworth, C P; Costa, J L; Rohde, C A; Simpson, M C; Doyle, C S; Dickinson, M E; Bunting, A S; Murray, A F; Delivopoulos, E; Graham, E S

    2013-01-01

    Cell patterning commonly employs photolithographic methods for the micro fabrication of structures on silicon chips. These require expensive photo-mask development and complex photolithographic processing. Laser based patterning of cells has been studied in vitro and laser ablation of polymers is an active area of research promising high aspect ratios. This paper disseminates how 800 nm femtosecond infrared (IR) laser radiation can be successfully used to perform laser ablative micromachining of parylene-C on SiO 2 substrates for the patterning of human hNT astrocytes (derived from the human teratocarcinoma cell line (hNT)) whilst 248 nm nanosecond ultra-violet laser radiation produces photo-oxidization of the parylene-C and destroys cell patterning. In this work, we report the laser ablation methods used and the ablation characteristics of parylene-C for IR pulse fluences. Results follow that support the validity of using IR laser ablative micromachining for patterning human hNT astrocytes cells. We disseminate the variation in yield of patterned hNT astrocytes on parylene-C with laser pulse spacing, pulse number, pulse fluence and parylene-C strip width. The findings demonstrate how laser ablative micromachining of parylene-C on SiO 2 substrates can offer an accessible alternative for rapid prototyping, high yield cell patterning with broad application to multi-electrode arrays, cellular micro-arrays and microfluidics. (paper)

  18. Response surface optimization for the transesterification of karanja oil using immobilized whole cells of Rhizopus oryzae in n-hexane system

    Energy Technology Data Exchange (ETDEWEB)

    Ganesan, Devanesan; Rajendran, Aravindan; Thangavelu, Viruthagiri [Annamalai University, Department of Chemical Engineering, Faculty of Engineering and Technology, Biochemical Engineering Laboratory, Annamalai Nagar, Tamil Nadu (India)

    2012-03-15

    Non-edible oils represent one of the most viable alternative feed stocks for the production of large volumes of biodiesel at cheaper cost in tropical countries. The objective of the present study is to investigate the ability of the immobilized whole cells of Rhizopus oryzae MTCC 262 to catalyze the biodiesel production from karanja oil in n-hexane system. Response surface methodology was employed to evaluate the effects of synthesis parameters, such as molar ratio of oil to alcohol, reaction temperature and reaction time on percentage biodiesel (methyl esters) yield. Transesterification was performed in shake flasks containing immobilized cells in the reaction mixture with 10% oil weight of n-hexane. The quadratic effects of molar ratio of oil to alcohol and reaction time proved to be the significant at 1% and 5% levels, respectively. The optimum synthesis conditions were found to be: molar ratio of oil to alcohol 1:2.73, reaction temperature 41.39 C and reaction time 73.97 h. Biodiesel yield (methyl ester) was 75.98 (wt.%) under the optimal conditions and the subsequent verification experiments with biodiesel yield of 78.0 (wt.%) confirmed the validity of the proposed model. (orig.)

  19. Preliminary applied study of assessment ischemic/viable myocardium by 99Tcm-HL91

    International Nuclear Information System (INIS)

    Liu Gang; Wu Hua

    2004-01-01

    Objective: To investigate the representation of 99 Tc m -HL91 in the ischemic myocardium, evaluate the diagnosis value of 99 Tc m -HL91 on hypoxic but viable myocardium. Methods: Six patients with cardiac infarction all underwent 99 Tc m -MIBI SPECT and 99 Tc m -HL91 SPECT. Average radioactivity of ischemic area and normal area were respectively obtained by ROI (2 x 2 pixels) on heart minor axis of images, And the radioactivity ratios of target (ischemic area)-to-non target(normal area)were calculated. Results: In image of 99 Tc m -HL91 SPECT, two patients who's radioactivity coloboma of 99 Tc m -MIBI image could be filled with 99 Tc m -HL91, four patients were not caught sight of obvious filling up. Conclusion 99 Tc m -HL91 can be selectively uptaken by ischemic and hypoxic but viable myocardium. it combination of 99 Tc m -MIBI SPECT may be good for accurate diagnosis and differentiation of viable myocardium. (authors)

  20. High-Yield Lithium-Injection Fusion-Energy (HYLIFE) reactor

    International Nuclear Information System (INIS)

    Blink, J.A.; Hogam, W.J.; Hovingh, J.; Meier, E.R.; Pitts, J.H.

    1985-01-01

    The High-Yield Lithium-Injection Fusion Energy (HYLIFE) concept to convent inertial confinement fusion energy into electric power has undergone intensive research and refinement at LLNL since 1978. This paper reports on the final HYLIFE design, focusing on five major areas: the HYLIFE reaction chamber (which includes neutronics, liquid-metal jet-array hydrocynamics, and structural design), supporting systems, primary steam system and balance of plant, safety and environmental protection, and costs. An annotated bibliography of reports applicable to HYLIFE is also provided. We conclude that HYLIFE is a particularly viable concept for the safe, clean production of electrical energy. The liquid-metal jet array, HYLIFE's key design feature, protects the surrounding structural components from x-rays, fusion fuel-pellet debris, neutron damage and activation, and high temperatures and stresses, allowing the structure to last for the plant's entire 30-year lifetime without being replaced. 127 refs., 18 figs

  1. Large-scale Isolation of Highly Pure "Untouched" Regulatory T Cells in a GMP Environment for Adoptive Cell Therapy.

    Science.gov (United States)

    Haase, Doreen; Puan, Kia Joo; Starke, Mireille; Lai, Tuck Siong; Soh, Melissa Yan Ling; Karunanithi, Iyswariya; San Luis, Boris; Poh, Tuang Yeow; Yusof, Nurhashikin; Yeap, Chun Hsien; Phang, Chew Yen; Chye, Willis Soon Yuan; Chan, Marieta; Koh, Mickey Boon Chai; Goh, Yeow Tee; Bertin-Maghit, Sebastien; Nardin, Alessandra; Ho, Liam Pock; Rotzschke, Olaf

    2015-01-01

    Adoptive cell therapy is an emerging treatment strategy for a number of serious diseases. Regulatory T (Treg) cells represent 1 cell type of particular interest for therapy of inflammatory conditions, as they are responsible for controlling unwanted immune responses. Initial clinical trials of adoptive transfer of Treg cells in patients with graft-versus-host disease were shown to be safe. However, obtaining sufficient numbers of highly pure and functional Treg cells with minimal contamination remains a challenge. We developed a novel approach to isolate "untouched" human Treg cells from healthy donors on the basis of negative selection using the surface markers CD49d and CD127. This procedure, which uses an antibody cocktail and magnetic beads for separation in an automated system (RoboSep), was scaled up and adapted to be compatible with good manufacturing practice conditions. With this setup we performed 9 Treg isolations from large-scale leukapheresis samples in a good manufacturing practice facility. These runs yielded sufficient numbers of "untouched" Treg cells for immediate use in clinical applications. The cell preparations consisted of viable highly pure FoxP3-positive Treg cells that were functional in suppressing the proliferation of effector T cells. Contamination with CD4 effector T cells was cell types did not exceed 2% in the final product. Remaining isolation reagents were reduced to levels that are considered safe. Treg cells isolated with this procedure will be used in a phase I clinical trial of adoptive transfer into leukemia patients developing graft-versus-host disease after stem cell transplantation.

  2. Modulation and modeling of monoclonal antibody N-linked glycosylation in mammalian cell perfusion reactors.

    Science.gov (United States)

    Karst, Daniel J; Scibona, Ernesto; Serra, Elisa; Bielser, Jean-Marc; Souquet, Jonathan; Stettler, Matthieu; Broly, Hervé; Soos, Miroslav; Morbidelli, Massimo; Villiger, Thomas K

    2017-09-01

    Mammalian cell perfusion cultures are gaining renewed interest as an alternative to traditional fed-batch processes for the production of therapeutic proteins, such as monoclonal antibodies (mAb). The steady state operation at high viable cell density allows the continuous delivery of antibody product with increased space-time yield and reduced in-process variability of critical product quality attributes (CQA). In particular, the production of a confined mAb N-linked glycosylation pattern has the potential to increase therapeutic efficacy and bioactivity. In this study, we show that accurate control of flow rates, media composition and cell density of a Chinese hamster ovary (CHO) cell perfusion bioreactor allowed the production of a constant glycosylation profile for over 20 days. Steady state was reached after an initial transition phase of 6 days required for the stabilization of extra- and intracellular processes. The possibility to modulate the glycosylation profile was further investigated in a Design of Experiment (DoE), at different viable cell density and media supplement concentrations. This strategy was implemented in a sequential screening approach, where various steady states were achieved sequentially during one culture. It was found that, whereas high ammonia levels reached at high viable cell densities (VCD) values inhibited the processing to complex glycan structures, the supplementation of either galactose, or manganese as well as their synergy significantly increased the proportion of complex forms. The obtained experimental data set was used to compare the reliability of a statistical response surface model (RSM) to a mechanistic model of N-linked glycosylation. The latter outperformed the response surface predictions with respect to its capability and reliability in predicting the system behavior (i.e., glycosylation pattern) outside the experimental space covered by the DoE design used for the model parameter estimation. Therefore, we can

  3. Studies on chlorophyll and viable mutations in green gram (Vigna radiata L. Wilczek) II: Response to mutagen

    Energy Technology Data Exchange (ETDEWEB)

    Krishnaswami, S; Rathinam, M [Tamil Nadu Agricultural Univ., Coimbatore (India). Dept. of Agricultural Botany

    1980-09-01

    The frequency and spectrum of chlorophyll and viable mutations in relation to type and dose of mutagen and cluster progenies were studied in four green gram cultivars viz., Kopergaon, Pusa Baisakhi, L. 24/2 and Sel. 122 subjected to two levels of EMS and gamma irradiation, severally and in conjunction. While chlorophyll mutations did not vary with the mutagen dose, viable mutations exhibited a direct relationship. Combinations of the mutagens were more effective in mutation induction. While no difference was manifested between the cluster families in respect of chlorophyll mutations, progenies of the second cluster recorded less viable mutations than either the first or the third. Viridis and xanthoviridis among chlorophyll mutations, and those affecting plant duration and stature among viable were more predominant.

  4. Canola Oil Fuel Cell Demonstration: Volume 2 - Market Availability of Agricultural Crops for Fuel Cell Applications

    National Research Council Canada - National Science Library

    Adams, John W; Cassarino, Craig; Spangler, Lee; Johnson, Duane; Lindstrom, Joel; Binder, Michael J; Holcomb, Franklin H; Lux, Scott M

    2006-01-01

    .... The reformation of vegetable oil crops for fuel cell uses is not well known; yet vegetable oils such as canola oil represent a viable alternative and complement to traditional fuel cell feedstocks...

  5. Parameters for MDBK cell growth on microcarriers and BoHV-1 virus production

    Directory of Open Access Journals (Sweden)

    Ethel C. Freitas

    2017-09-01

    Full Text Available Bovine herpes virus 1 (BoHV-1 is an important veterinary agent , which causes infectious bovine rhinotra-cheitis. This disease affects the respiratory tract or genitals, causing weight loss, reduced milk production and abor-tion. Several vaccines against BoHV-1 have been developed. In this paper, we study the parameters for MDBK growth on microcarriers (Cytodex 1 and for BoHV-1 virus production. The cell culture attached to microcarriers is an effi-cient method to enlarge the surface of cell growth and for large-scale cell production. Our studies reveal that MDBK adhered to MCs in 30 minutes and that initial agitation of culture did not influence on the efficiency of adhesion or cell growth. In our experiments, we detected no relevant influence of agitation on initial cell adhesion of MDBK to MCs. The maximum cell yield was similar to all initial conditions of agitation studied. The maximum yield obtained in culture started with 15, 20 and 30 cells / MC, was respectively.8.7 x105, 9.3 x105 and 9.8 x105 cells / ml . The cellular distribution on the MCs at the beginning of the culture was more heterogeneous in higher initial densities. After three medium exchanges during MDBK cell culture, the increase in the final yield was 100% higher than that from culture performed without medium change (0.93 x 106 cells / mL. Replacing 50% of the culture medium with fresh medium after 24 hours of growth, the concentration of glucose (5 mM and glutamine (1.8 mM were almost completely res-tored. In these studies, BoHV-1 infections of MDBK were performed after 48, 72 and 86 hours with daily exchanges of 50% of the medium. The increase in viral titer was proportional to the number of viable cells present at the time of infection. The best result of BoHV-1 production was achieved when the infection was performed from 86 hours of cell culture, reaching about 3.7 x108 (TCID50/ml after 24-48 hours of infection, being on average four times higher when compared to the

  6. Improvements to the solar cell efficiency and production yields of low-lifetime wafers with effective phosphorus gettering

    International Nuclear Information System (INIS)

    Lu, Jiunn-Chenn; Chen, Ping-Nan; Chen, Chih-Min; Wu, Chung-Han

    2013-01-01

    Highlights: • Variable-temperature gettering improves efficiencies when the wafer quality is poor. • High-quality wafers need not be used for variable-temperature gettering. • The proposed gettering method is based on an existing diffusion process. • It has a potential interest for hot-spot prevention. -- Abstract: This research focuses on the improvement of solar cell efficiencies in low-lifetime wafers by implementing an appropriate gettering method of the diffusion process. The study also considers a reduction in the value of the reverse current at −12 V, an important electrical parameter related to the hot-spot heating of solar cells and modules, to improve the product's quality during commercial mass production. A practical solar cell production case study is examined to illustrate the use of the proposed method. The results of this case study indicate that variable-temperature gettering significantly improves solar cell efficiencies by 0.14% compared to constant-temperature methods when the wafer quality is poor. Moreover, this study finds that variable-temperature gettering raises production yields of low quality wafers by more than 30% by restraining the measurement value of the reverse current at −12 V during solar cell manufacturing

  7. In vitro study of tumor seeking radiopharmaceutical uptake by human breast cancer cell line MCF-7 after paclitaxel treatment

    International Nuclear Information System (INIS)

    Choi, Joon Young; Choi, Yong; Choe, Yearn Seong; Lee, Kyung Han; Kim, Byung Tae

    2007-01-01

    This study was designed to investigate the cellular uptake of various tumor imaging radiopharmaceuticals in human breast cancer cells before and after paclitaxel exposure considering viable cell number. F-18-fluorodeoxyglucose, C-11-methionine. TI-201, Tc-99m-MIBI, and Tc-99m-tetrofosmin were used to evaluate the cellular uptake in MCF-7 cells. MCF-7 cells were cultured in multi-well plates. Wells were divided into DMSO exposure control group, and paclitaxel exposure group. The exposure durations of paclitaxel with 10 nM or 100 nM were 2 h, 6 h, 12 h, 24 h, and 48 h. Viable cell fraction was reduced as the concentration and exposure time of paclitaxel increased. After 10 nM paclitaxel exposure, the cellular uptake of all 5 radiopharmaceuticals was not reduced significantly, irrespective of exposure time and viable cell fraction. After 100 nM paclitaxel exposure, the cellular uptake of all 5 radiopharmaceuticals was enhanced significantly irrespective of viable cell fraction. The peak uptake was observed in experimental groups with paclitaxel exposure for 6 to 48 h according the type of radiopharmaceutical. When the cellular uptake was adjusted for the viable cell fraction and cell count, the peak cellular uptake was observed in experimental groups with paclitaxel exposure for 48 h, irrespective of the type of radiopharmaceutical. The cellular uptake of F-18-fluorodeoxyglucose, C-11-methionine, TI-201, Tc-99m-MIBI, and Tc-99m-tetrofosmin did not reflect viable cell number in MCF-7 cells after paclitaxel exposure for up to 48 h

  8. Whey utilization for single-cell protein production

    Energy Technology Data Exchange (ETDEWEB)

    Barraquio, V; Silverio, L G; Revilleza, R P; Fernadez, W L

    1980-01-01

    The production of single-cell protein by yeast assimilation of lactose in soft cheese whey was studied using Candida pseudotropicalis as a test organism. Under shake-flask cultivation conditions with deproteinized whey as the medium, lactose (initially 4.20%) was completely assimilated in 48h; cell mass was 5.56 mg/mL after 72h; and average protein content of the dried mass was approximately 11.8%. Batch cultivation using undeproteinized whey resulted in a faster lactose utilization rate from an initial 3.93% to a residual 0.56% in 12 h; cell mass was 8.41 mg/mL in 10 h; and average protein was approximately 37.7%. In a semicontinuous culture with 10 to the power of 7 viable cells/mL as initial cell concentration, 15.69 mg/mL cell mass with a mean protein content of approximately 21.4% could be produced and lactose could be considerably consumed (from an initial 4.75% to a residual 0.42%) within 13-14 h. Supplementation with (NH/sub 4/)/sub 2/S0/sub 4/ and KH/sub 2/P0/sub 4/ did not increase cell mass (12.47 mg/mL in 12 h) and hasten lactose assimulation (from initial 4.49% to residual 0.3% in 12 h). Average protein content was approximately 31%. Cell mass yield was established as 0.29 mg yeast cell/mg lactose consumed. Factors that might have affected protein content are also discussed.

  9. Acupuntura un tratamiento viable para las adicciones en Colombia

    Directory of Open Access Journals (Sweden)

    Hernán López Seuscún

    2013-07-01

    Los tratamientos con auriculoterapia, como el protocolo NADA (National Acupuncture Detoxification Association, son los métodos más usados para las adicciones en el mundo, y aunque no se ha logrado evidenciar su efectividad, por su costo, facilidad y el poco riesgo de efectos adversos se hace viable en un país con pocos recursos económicos como Colombia.

  10. Are GM Crops for Yield and Resilience Possible?

    Science.gov (United States)

    Paul, Matthew J; Nuccio, Michael L; Basu, Shib Sankar

    2018-01-01

    Crop yield improvements need to accelerate to avoid future food insecurity. Outside Europe, genetically modified (GM) crops for herbicide- and insect-resistance have been transformative in agriculture; other traits have also come to market. However, GM of yield potential and stress resilience has yet to impact on food security. Genes have been identified for yield such as grain number, size, leaf growth, resource allocation, and signaling for drought tolerance, but there is only one commercialized drought-tolerant GM variety. For GM and genome editing to impact on yield and resilience there is a need to understand yield-determining processes in a cell and developmental context combined with evaluation in the grower environment. We highlight a sugar signaling mechanism as a paradigm for this approach. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Wavelet-SVM classification and automatic recognition of unstained viable cells in phase-contrast microscopy

    International Nuclear Information System (INIS)

    Skoczylas, M.; Rakowski, W.; Cherubini, R.; Gerardi, S.

    2011-01-01

    Irradiation of individual cultured mammalian cells with a pre-selected number of ions down to one ion per single cell is a useful experimental approach to investigating the low-dose ionising radiation exposure effects and thus contributing to a more realistic human cancer risk assessment. One of the crucial tasks of all the microbeam apparatuses is the visualisation, recognition and positioning of every individual cell of the cell culture to be irradiated. Before irradiations, mammalian cells (specifically, Chinese hamster V79 cells) are seeded and grown as a monolayer on a mylar surface used as the bottom of a specially designed holder. Manual recognition of unstained cells in a bright-field microscope is a time-consuming procedure; therefore, a parallel algorithm has been conceived and developed in order to speed up this irradiation protocol step. Many technical problems have been faced to overcome the complexity of the images to be analysed: cell discrimination in an inhomogeneous background, among many disturbing bodies mainly due to the mylar surface roughness and culture medium bodies; cell shapes, depending on how they attach to the surface, which phase of the cell cycle they are in and on cell density. Preliminary results of the recognition and classification based on a method of wavelet kernels for the support vector machine classifier will be presented. (authors)

  12. Flexible Nanosomes (SECosomes) Enable Efficient siRNA Delivery in Cultured Primary Skin Cells and in the Viable Epidermis of Ex Vivo Human Skin

    NARCIS (Netherlands)

    Geusens, Barbara; Van Gele, Mireille; Braat, Sien; De Smedt, Stefaan C.; Stuart, Marc C. A.; Prow, Tarl W.; Sanchez, Washington; Roberts, Michael S.; Sanders, Niek N.; Lambert, Jo

    2010-01-01

    The extent to which nanoscale-engineered systems cross intact human skin and can exert pharmacological effects in viable epidermis is controversial. This research seeks to develop a new lipid-based nanosome that enables the effective delivery of siRNA into human skin. The major finding is that an

  13. Fluorescently labeled dengue viruses as probes to identify antigen-specific memory B cells by multiparametric flow cytometry.

    Science.gov (United States)

    Woda, Marcia; Mathew, Anuja

    2015-01-01

    Low frequencies of memory B cells in the peripheral blood make it challenging to measure the functional and phenotypic characteristics of this antigen experienced subset of B cells without in vitro culture. To date, reagents are lacking to measure ex vivo frequencies of dengue virus (DENV)-specific memory B cells. We wanted to explore the possibility of using fluorescently labeled DENV as probes to detect antigen-specific memory B cells in the peripheral blood of DENV immune individuals. Alexa Fluor dye-labeled DENV yielded viable virus that could be stored at -80°C for long periods of time. Using a careful gating strategy and methods to decrease non-specific binding, we were able to identify a small frequency of B cells from dengue immune individuals that bound labeled DENV. Sorted DENV(+) B cells from immune, but not naïve donors secreted antibodies that bound DENV after in vitro stimulation. Overall, Alexa Fluor dye-labeled DENVs are useful reagents to enable the detection and characterization of memory B cells in DENV immune individuals. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. A cell transportation solution that preserves live circulating tumor cells in patient blood samples

    International Nuclear Information System (INIS)

    Stefansson, Steingrimur; Adams, Daniel L.; Ershler, William B.; Le, Huyen; Ho, David H.

    2016-01-01

    Circulating tumor cells (CTCs) are typically collected into CellSave fixative tubes, which kills the cells, but preserves their morphology. Currently, the clinical utility of CTCs is mostly limited to their enumeration. More detailed investigation of CTC biology can be performed on live cells, but obtaining live CTCs is technically challenging, requiring blood collection into biocompatible solutions and rapid isolation which limits transportation options. To overcome the instability of CTCs, we formulated a sugar based cell transportation solution (SBTS) that stabilizes cell viability at ambient temperature. In this study we examined the long term viability of human cancer cell lines, primary cells and CTCs in human blood samples in the SBTS for transportation purposes. Four cell lines, 5 primary human cells and purified human PBMCs were tested to determine the viability of cells stored in the transportation solution at ambient temperature for up to 7 days. We then demonstrated viability of MCF-7 cells spiked into normal blood with SBTS and stored for up to 7 days. A pilot study was then run on blood samples from 3 patients with metastatic malignancies stored with or without SBTS for 6 days. CTCs were then purified by Ficoll separation/microfilter isolation and identified using CTC markers. Cell viability was assessed using trypan blue or CellTracker™ live cell stain. Our results suggest that primary/immortalized cell lines stored in SBTS remain ~90 % viable for > 72 h. Further, MCF-7 cells spiked into whole blood remain viable when stored with SBTS for up to 7 days. Finally, live CTCs were isolated from cancer patient blood samples kept in SBTS at ambient temperature for 6 days. No CTCs were isolated from blood samples stored without SBTS. In this proof of principle pilot study we show that viability of cell lines is preserved for days using SBTS. Further, this solution can be used to store patient derived blood samples for eventual isolation of viable CTCs

  15. A cell transportation solution that preserves live circulating tumor cells in patient blood samples.

    Science.gov (United States)

    Stefansson, Steingrimur; Adams, Daniel L; Ershler, William B; Le, Huyen; Ho, David H

    2016-05-06

    Circulating tumor cells (CTCs) are typically collected into CellSave fixative tubes, which kills the cells, but preserves their morphology. Currently, the clinical utility of CTCs is mostly limited to their enumeration. More detailed investigation of CTC biology can be performed on live cells, but obtaining live CTCs is technically challenging, requiring blood collection into biocompatible solutions and rapid isolation which limits transportation options. To overcome the instability of CTCs, we formulated a sugar based cell transportation solution (SBTS) that stabilizes cell viability at ambient temperature. In this study we examined the long term viability of human cancer cell lines, primary cells and CTCs in human blood samples in the SBTS for transportation purposes. Four cell lines, 5 primary human cells and purified human PBMCs were tested to determine the viability of cells stored in the transportation solution at ambient temperature for up to 7 days. We then demonstrated viability of MCF-7 cells spiked into normal blood with SBTS and stored for up to 7 days. A pilot study was then run on blood samples from 3 patients with metastatic malignancies stored with or without SBTS for 6 days. CTCs were then purified by Ficoll separation/microfilter isolation and identified using CTC markers. Cell viability was assessed using trypan blue or CellTracker™ live cell stain. Our results suggest that primary/immortalized cell lines stored in SBTS remain ~90% viable for > 72 h. Further, MCF-7 cells spiked into whole blood remain viable when stored with SBTS for up to 7 days. Finally, live CTCs were isolated from cancer patient blood samples kept in SBTS at ambient temperature for 6 days. No CTCs were isolated from blood samples stored without SBTS. In this proof of principle pilot study we show that viability of cell lines is preserved for days using SBTS. Further, this solution can be used to store patient derived blood samples for eventual isolation of viable CTCs after

  16. The amount of viable and dyssynchronous myocardium is associated with response to cardiac resynchronization therapy: initial clinical results using multiparametric ECG-gated [{sup 18}F]FDG PET

    Energy Technology Data Exchange (ETDEWEB)

    Lehner, Sebastian; Uebleis, Christopher; Haug, Alexander; Bartenstein, Peter [University of Munich, Department of Nuclear Medicine, Munich (Germany); Schuessler, Franziska; Kaeaeb, Stefan; Estner, Heidi [University of Munich, Medical Department I, Munich (Germany); Van Kriekinge, Serge D.; Germano, Guido [UCLA, Cedars-Sinai Medical Center, Los Angeles and David Geffen School of Medicine, Los Angeles, CA (United States); Hacker, Marcus [Medical University of Vienna, Division of Nuclear Medicine, Department of Biomedical Imaging and Image-guided Theraphy, Vienna (Austria)

    2013-12-15

    There is still a significant amount of patients who do not sufficiently respond to cardiac resynchronization therapy (CRT). Previous studies demonstrated that the amount of dyssynchronous myocardium was predictive of response to CRT. Otherwise, non-response is frequently associated with high amounts of scar tissue. The combination of these parameters might yield a more accurate prediction of response. We hypothesized that the probability of a CRT response increases with the presence of high amounts of ''viable and dyssynchronous'' myocardium. A total of 19 patients (17 male, 61 {+-} 10 years) underwent ECG-gated [{sup 18}F]fluorodeoxyglucose (FDG) myocardial positron emission tomography (PET) before CRT device implantation and were followed for 6 months. Response to CRT was defined as clinical improvement of at least one New York Heart Association (NYHA) class in combination with left ventricular (LV) ejection fraction (EF) improvement of >5 %. Twelve responders (71 %) and seven non-responders (29 %) were identified. For each patient bullseye maps of FDG uptake and phase analysis were calculated (QPS/QGS 2012, Cedars-Sinai, Los Angeles, CA, USA) and fused. Amounts of myocardium representing ''viable and synchronous'', ''scar and synchronous'', viable and dyssynchronous or ''scar and dyssynchronous'' myocardium were quantified by planimetric measurements of the fused bullseye maps. Responders by definition showed significant decrease in NYHA class and significant increase of LVEF. Furthermore, a significantly higher amount of viable and dyssynchronous myocardium was found as compared to non-responders (21 {+-} 13 % vs 6 {+-} 5 %; p < 0.05). Combined assessment of myocardial viability and LV dyssynchrony is feasible using multiparametric [{sup 18}F]FDG PET and could improve conventional response prediction criteria for CRT. (orig.)

  17. Human Ocular Epithelial Cells Endogenously Expressing SOX2 and OCT4 Yield High Efficiency of Pluripotency Reprogramming.

    Directory of Open Access Journals (Sweden)

    Ming-Wai Poon

    Full Text Available A variety of pluripotency reprogramming frequencies from different somatic cells has been observed, indicating cell origin is a critical contributor for efficiency of pluripotency reprogramming. Identifying the cell sources for efficient induced pluripotent stem cells (iPSCs generation, and defining its advantages or disadvantages on reprogramming, is therefore important. Human ocular tissue-derived conjunctival epithelial cells (OECs exhibited endogenous expression of reprogramming factors OCT4A (the specific OCT 4 isoform on pluripotency reprogramming and SOX2. We therefore determined whether OECs could be used for high efficiency of iPSCs generation. We compared the endogenous expression levels of four pluripotency factors and the pluripotency reprograming efficiency of human OECs with that of ocular stromal cells (OSCs. Real-time PCR, microarray analysis, Western blotting and immunostaining assays were employed to compare OECiPSCs with OSCiPSCs on molecular bases of reprogramming efficiency and preferred lineage-differentiation potential. Using the traditional KMOS (KLF4, C-MYC, OCT4 and SOX2 reprogramming protocol, we confirmed that OECs, endogenously expressing reprogramming factors OCT4A and SOX2, yield very high efficiency of iPSCs generation (~1.5%. Furthermore, higher efficiency of retinal pigmented epithelial differentiation (RPE cells was observed in OECiPSCs compared to OSCiPSCs or skin fibroblast iMR90iPSCs. The findings in this study suggest that conjunctival-derived epithelial (OECs cells can be easier converted to iPSCs than conjunctival-derived stromal cells (OSCs. This cell type may also have advantages in retinal pigmented epithelial differentiation.

  18. Human Ocular Epithelial Cells Endogenously Expressing SOX2 and OCT4 Yield High Efficiency of Pluripotency Reprogramming.

    Science.gov (United States)

    Poon, Ming-Wai; He, Jia; Fang, Xiaowei; Zhang, Zhao; Wang, Weixin; Wang, Junwen; Qiu, Fangfang; Tse, Hung-Fat; Li, Wei; Liu, Zuguo; Lian, Qizhou

    2015-01-01

    A variety of pluripotency reprogramming frequencies from different somatic cells has been observed, indicating cell origin is a critical contributor for efficiency of pluripotency reprogramming. Identifying the cell sources for efficient induced pluripotent stem cells (iPSCs) generation, and defining its advantages or disadvantages on reprogramming, is therefore important. Human ocular tissue-derived conjunctival epithelial cells (OECs) exhibited endogenous expression of reprogramming factors OCT4A (the specific OCT 4 isoform on pluripotency reprogramming) and SOX2. We therefore determined whether OECs could be used for high efficiency of iPSCs generation. We compared the endogenous expression levels of four pluripotency factors and the pluripotency reprograming efficiency of human OECs with that of ocular stromal cells (OSCs). Real-time PCR, microarray analysis, Western blotting and immunostaining assays were employed to compare OECiPSCs with OSCiPSCs on molecular bases of reprogramming efficiency and preferred lineage-differentiation potential. Using the traditional KMOS (KLF4, C-MYC, OCT4 and SOX2) reprogramming protocol, we confirmed that OECs, endogenously expressing reprogramming factors OCT4A and SOX2, yield very high efficiency of iPSCs generation (~1.5%). Furthermore, higher efficiency of retinal pigmented epithelial differentiation (RPE cells) was observed in OECiPSCs compared to OSCiPSCs or skin fibroblast iMR90iPSCs. The findings in this study suggest that conjunctival-derived epithelial (OECs) cells can be easier converted to iPSCs than conjunctival-derived stromal cells (OSCs). This cell type may also have advantages in retinal pigmented epithelial differentiation.

  19. The search for viable local government system in Nigeria: an ...

    African Journals Online (AJOL)

    The history of the Nigerian local government system has been one long episode of trails and errors aimed at achieving viable local government institution without much success. Local government in the country began its long series of reforms from the colonial period when the colonial government attempted to ...

  20. Viable Syntax: Rethinking Minimalist Architecture

    Directory of Open Access Journals (Sweden)

    Ken Safir

    2010-03-01

    Full Text Available Hauser et al. (2002 suggest that the human language faculty emerged as a genetic innovation in the form of what is called here a ‘keystone factor’—a single, simple, formal mental capability that, interacting with the pre-existing faculties of hominid ancestors, caused a cascade of effects resulting in the language faculty in modern humans. They take Merge to be the keystone factor, but instead it is posited here that Merge is the pre-existing mechanism of thought made viable by a principle that permits relations interpretable at the interfaces to be mapped onto c-command. The simplified minimalist architecture proposed here respects the keystone factor as closely as possible, but is justified on the basis of linguistic analyses it makes available, including a relativized intervention theory applicable across Case, scope, agreement, selection and linearization, a derivation of the A/A’-distinction from Case theory, and predictions such as why in situ wh-interpretation is island-insensitive, but susceptible to intervention effects.

  1. Kinetics of killing Listeria monocytogenes by macrophages: correlation of 3H-DNA release from labeled bacteria and changes in numbers of viable organisms by mathematical model

    International Nuclear Information System (INIS)

    Davies, W.A.

    1982-01-01

    Conventional methods of assessing antibacterial activities of macrophages by viable counting are limited by the precision of the statistics and are difficult to interpret quantitatively because of unrestrained extracellular growth of bacteria. An alternative technique based on the release of radioactive DNA from labeled bacteria has been offered as overcoming these drawbacks. To assess it for use with macrophages I have made a correlation with the conventional viable counting method using a mathematical model. Opsonized Listeria monocytogenes labeled with 3 H-thymidine were exposed to rat macrophages for periods up to 4 hr. Numbers of viable bacteria determined after sonication increased exponentially in the absence of live cells and this growth rate was progressively inhibited by increasing numbers of macrophages. After a lag period of 30-60 min soluble 3 H appeared in the supernatant, the amount increasing with time and numbers of macrophages. To correlate these data I developed a mathematical model that considered that changes in numbers of viable organisms were due to the difference between rates of 1) growth of extracellular bacteria and 2) killing within the macrophage. On the basis of this model curves of best fit to the viable counts data were used to predict the release of radioactivity, assuming that death of a bacterium led to the total release of its label. These predictions and the experimental data agreed well, the lag period of 30-60 min between death of the bacterium and release of radioactivity being consistent with intracellular digestion. Release of soluble radioactivity appears to be an accurate reflection of the number of bacteria killed within the macrophage

  2. Carbon Monoxide Releasing Molecule-A1 (CORM-A1) Improves Neurogenesis: Increase of Neuronal Differentiation Yield by Preventing Cell Death.

    Science.gov (United States)

    Almeida, Ana S; Soares, Nuno L; Vieira, Melissa; Gramsbergen, Jan Bert; Vieira, Helena L A

    2016-01-01

    Cerebral ischemia and neurodegenerative diseases lead to impairment or death of neurons in the central nervous system. Stem cell based therapies are promising strategies currently under investigation. Carbon monoxide (CO) is an endogenous product of heme degradation by heme oxygenase (HO) activity. Administration of CO at low concentrations produces several beneficial effects in distinct tissues, namely anti-apoptotic and anti-inflammatory. Herein the CO role on modulation of neuronal differentiation was assessed. Three different models with increasing complexity were used: human neuroblastoma SH-S5Y5 cell line, human teratocarcinoma NT2 cell line and organotypic hippocampal slice cultures (OHSC). Cell lines were differentiated into post-mitotic neurons by treatment with retinoic acid (RA) supplemented with CO-releasing molecule A1 (CORM-A1). CORM-A1 positively modulated neuronal differentiation, since it increased final neuronal production and enhanced the expression of specific neuronal genes: Nestin, Tuj1 and MAP2. Furthermore, during neuronal differentiation process, there was an increase in proliferative cell number (ki67 mRNA expressing cells) and a decrease in cell death (lower propidium iodide (PI) uptake, limitation of caspase-3 activation and higher Bcl-2 expressing cells). CO supplementation did not increase the expression of RA receptors. In the case of SH-S5Y5 model, small amounts of reactive oxygen species (ROS) generation emerges as important signaling molecules during CO-promoted neuronal differentiation. CO's improvement of neuronal differentiation yield was validated using OHSC as ex vivo model. CORM-A1 treatment of OHSC promoted higher levels of cells expressing the neuronal marker Tuj1. Still, CORM-A1 increased cell proliferation assessed by ki67 expression and also prevented cell death, which was followed by increased Bcl-2 expression, decreased levels of active caspase-3 and PI uptake. Likewise, ROS signaling emerged as key factors in CO

  3. Development of a robust, versatile, and scalable inoculum train for the production of a DNA vaccine.

    Science.gov (United States)

    Okonkowski, J; Kizer-Bentley, L; Listner, K; Robinson, D; Chartrain, M

    2005-01-01

    For many microbial fermentation processes, the inoculum train can have a substantial impact on process performance in terms of productivity, profitability, and process control. In general, it is understood that a well-characterized and flexible inoculum train is essential for future scale-up and implementation of the process in a pilot plant or manufacturing setting. A fermentation process utilizing E. coli DH5 for the production of plasmid DNA carrying the HIV gag gene for use as a vaccine is currently under development in our laboratory. As part of the development effort, we evaluated inoculum train schemes that incorporate one, two, or three stages. In addition, we investigated the effect of inoculum viable-cell concentrations, either thawed or actively growing, over a wide range (from 2.5 x 10(4) to 1.0 x 10(8) viable cells/mL or approximately 0.001% to 4% of final working volume). The various inoculum trains were evaluated in terms of final plasmid yield, process time, reproducibility, robustness, and feasibility at large scale. The results of these studies show that final plasmid yield remained in the desired range, despite the number of stages or inoculation viable-cell concentrations comprising the inoculum train. On the basis of these observations and because it established a large database, the first part of these investigations supports an exceptional flexibility in the design of scalable inoculum trains for this DNA vaccine process. This work also highlighted that a slightly higher level of process reproducibility, as measured by the time for the culture to reach mid-exponential growth, was observed when using actively growing versus frozen cells. It also demonstrated the existence of a viable-cell concentration threshold for the one-stage process, since we observed that inoculation of the production stage with very low amounts of viable cells from a frozen source could lead to increased process sensitivity to external factors such as variation in the

  4. Development of image analysis software for quantification of viable cells in microchips.

    Science.gov (United States)

    Georg, Maximilian; Fernández-Cabada, Tamara; Bourguignon, Natalia; Karp, Paola; Peñaherrera, Ana B; Helguera, Gustavo; Lerner, Betiana; Pérez, Maximiliano S; Mertelsmann, Roland

    2018-01-01

    Over the past few years, image analysis has emerged as a powerful tool for analyzing various cell biology parameters in an unprecedented and highly specific manner. The amount of data that is generated requires automated methods for the processing and analysis of all the resulting information. The software available so far are suitable for the processing of fluorescence and phase contrast images, but often do not provide good results from transmission light microscopy images, due to the intrinsic variation of the acquisition of images technique itself (adjustment of brightness / contrast, for instance) and the variability between image acquisition introduced by operators / equipment. In this contribution, it has been presented an image processing software, Python based image analysis for cell growth (PIACG), that is able to calculate the total area of the well occupied by cells with fusiform and rounded morphology in response to different concentrations of fetal bovine serum in microfluidic chips, from microscopy images in transmission light, in a highly efficient way.

  5. AMMONOX-Ammonia for enhancing biogas yield & reducing NOx

    DEFF Research Database (Denmark)

    Gavala, Hariklia N.; Kristensen, P.G.; Paamand, K.

    2013-01-01

    The continuously increasing demand for renewable energy sources renders anaerobic digestion to one of the most promising technologies for renewable energy production. Due to the animal production intensification, manure is being used as the primary feedstock for most of the biogas plants. However...... of innovative ammonia recovery technology and c) the coupling of the excess ammonia obtained from manure with the catalytic elimination of NOx emissions when the biogas is used for subsequent electricity generation with gas engines.......The continuously increasing demand for renewable energy sources renders anaerobic digestion to one of the most promising technologies for renewable energy production. Due to the animal production intensification, manure is being used as the primary feedstock for most of the biogas plants. However......, biogas plants digesting liquid manure alone are not economically viable due to the relatively low organic content of the manure, usually 3-5%.Thus, their economical profitable operation relies partly on increasing the methane yield from manure, and especially of its solid fraction, usually called...

  6. Quantitative measures of mutagenicity and multability based on mutant yield data

    International Nuclear Information System (INIS)

    Eckhardt, F.; Haynes, R.H.

    1980-01-01

    We describe, how mutant yield data (mutants per cell treated) can be used both to compare the mutagenenicity of different mutagens, and to characterize the mutability of different cell types. Yield curves reveal the net effect of the lethal and genetic actions of mutagens on cells. Normally, yields are the quantities measured in assays for mutagenesis, and rectilinear plots of such data baldly reveal the amount of experimental error and the extent of actual mutant induction above the background level. Plots of yield versus lethal hits can be used to quantify the relative mutagenenic efficiency (RME) of agents whose physical exposure doses otherwise would be incommensurable, as well as the relative mutability (Rmt) of different strains to the same mutagen. Plots of yield versus log dose provide an unambiguous way of assessing the relative mutational sensitivities (Rms) and mutational resolutions (Rmr) of different strains against a given mutagen. Such analysis is important for evaluation of the relative merits of excision-proficient and excision-deficient strains of the same organism as mutagen-testing systems. The mathematical approach outlined here is applied, by way of example, to measurements of UV and 4-NQO induced mutagenesis in both repair-deficient and repair-proficient haploid strains of the yeast Sacccharomyces cerevsiae. (orig.)

  7. El modelo de sistema viable: un instrumento para la organización efectiva

    Directory of Open Access Journals (Sweden)

    Norlando Sánchez Rueda

    2015-05-01

    Full Text Available RESUMEN En este ensayo se presenta una interpretación teórica del denominado Modelo de Sistema Viable (MSV, de Stafford Beer y su Potencial Aplicación en Tareas de Diagnóstico  y diseño empresarial, al igual que para Mejorar las capacidades Organizacionales de Auto- Regulación  y Auto- Organización. Se explica como el Modelo del Sistema Viable permite conocer e interpretar  los mecanismos de estabilidad y adaptabilidad de las organizaciones, pilares para el crecimiento de una verdadera organización Efectiva.

  8. Nitrogen Fertilizer Sources and Application Timing Affects Wheat and Inter-Seeded Red Clover Yields on Claypan Soils

    Directory of Open Access Journals (Sweden)

    Kelly A. Nelson

    2014-11-01

    Full Text Available Controlled-release N fertilizer, such as polymer-coated urea (PCU, may be a fall N management option for wheat (Triticum aestivum L. grown in poorly-drained claypan soils. Field research evaluated (1 urea release from fall-applied PCU in 2006 and 2007; (2 broadcast fall-spring split (25%:75% of N sources; and (3 a single fall (100% application of PCU, urea, urea plus NBPT (N-(n-butyl thiophosphoric triamide] (U + NBPT, ammonium nitrate (AN, or urea ammonium nitrate (UAN at 0, 56, 84, and 112 kg·N·ha−1 on wheat yield, wheat biomass, N uptake by wheat, and frost-seeded red clover (FSC (Trifolium pratense L. forage yield (2004–2007. PCU applied in fall released less than 30% urea by February. Urea released from PCU by harvest was 60% and 85% in 2006 and 2007, respectively. In poorly-drained soils, wheat yields ranked PCU > AN > U + NBPT > urea ≥ UAN over the rates evaluated for fall-only application. PCU was a viable fall-applied N source, with yields similar to or greater than urea or U + NBPT split-applied. Split-N applications of AN, urea, UAN, and U + NBPT generally resulted in greater wheat yields than a fall application. Enhanced efficiency fertilizers provide farmers with flexible options for maintaining high yielding production systems.

  9. Axenic isolation of viable Giardia muris trophozoites.

    Science.gov (United States)

    Tillotson, K D; Buret, A; Olson, M E

    1991-06-01

    Large numbers of viable Giardia muris trophozoites were isolated from the duodenum of experimentally infected mice 6 days after inoculation with 1,000 G. muris cysts. A series of shaking, incubation, and washing steps in the presence of the broad-spectrum antibiotic piperacillin readily provided 4.9 +/- 1.5 x 10(5) G. muris trophozoites per mouse, free of detectable contaminant organisms. Anaerobic and microaerophilic culturing and scanning electron microscopy demonstrated axenic status and high purity of the isolates. The viability of trophozoites was 98 +/- 2%. Application of this technique should permit novel immunological and epidemiological analyses of G. muris infection and biochemical investigations of this protozoan parasite.

  10. Comparison of culture-based, vital stain and PMA-qPCR methods for the quantitative detection of viable hookworm ova.

    Science.gov (United States)

    Gyawali, P; Sidhu, J P S; Ahmed, W; Jagals, P; Toze, S

    2017-06-01

    Accurate quantitative measurement of viable hookworm ova from environmental samples is the key to controlling hookworm re-infections in the endemic regions. In this study, the accuracy of three quantitative detection methods [culture-based, vital stain and propidium monoazide-quantitative polymerase chain reaction (PMA-qPCR)] was evaluated by enumerating 1,000 ± 50 Ancylostoma caninum ova in the laboratory. The culture-based method was able to quantify an average of 397 ± 59 viable hookworm ova. Similarly, vital stain and PMA-qPCR methods quantified 644 ± 87 and 587 ± 91 viable ova, respectively. The numbers of viable ova estimated by the culture-based method were significantly (P methods. Therefore, both PMA-qPCR and vital stain methods appear to be suitable for the quantitative detection of viable hookworm ova. However, PMA-qPCR would be preferable over the vital stain method in scenarios where ova speciation is needed.

  11. Characterization and functional properties of gastric tissue-resident memory T cells from children, adults and the elderly

    Directory of Open Access Journals (Sweden)

    Jayaum S. Booth

    2014-06-01

    Full Text Available T cells are the main orchestrators of protective immunity in the stomach; however, limited information on the presence and function of the gastric T subsets is available mainly due to the difficulty in recovering high numbers of viable cells from human gastric biopsies. To overcome this shortcoming we optimized a cell isolation method that yielded high numbers of viable lamina propria mononuclear cells (LPMC from gastric biopsies. Classic memory T (TM subsets were identified in gastric LPMC and compared to peripheral blood mononuclear cells (PBMC obtained from children, adults and the elderly using an optimized 14 color flow cytometry panel. A dominant effector memory (TEM phenotype was observed in gastric LPMC CD4+ and CD8+ T cells in all age groups. We then evaluated whether these cells represented a population of gastric tissue-resident memory T (TRM cells by assessing expression of CD103 and CD69. The vast majority of gastric LPMC CD8+ T cells either co-expressed CD103/CD69 (>70% or expressed CD103 alone (~20%. Gastric LPMC CD4+ T cells also either co-expressed CD103/CD69 (>35% or expressed at least one of these markers. Thus, gastric LPMC CD8+ and CD4+ T cells had the characteristics of TRM cells. Gastric CD8+ and CD4+ TRM cells produced multiple cytokines (IFN-γ, IL-2, TNF-α, IL-17A, MIP-1β and up-regulated CD107a upon stimulation. However, marked differences were observed in their cytokine and multi-cytokine profiles when compared to their PBMC TEM counterparts. Furthermore, gastric CD8+ TRM and CD4+ TRM cells demonstrated differences in the frequency, susceptibility to activation and cytokine/multi-cytokine production profiles among the age groups. Most notably, children’s gastric TRM cells responded differently to stimuli than gastric TRM cells from adults or the elderly. In conclusion, we demonstrate the presence of gastric TRM which exhibit diverse functional characteristics in children, adults and the elderly.

  12. Routes to a commercially viable PEM fuel cell stack

    Energy Technology Data Exchange (ETDEWEB)

    Newton, J.; Foster, S.E.; Hodgson, D.; Marrett, A.

    2002-07-01

    This report describes the results of a project to design and build a 10 kW{sub e} proton exchange membrane fuel cell (PEMFC) stack, including membrane electrode assemblies (MEAs), bipolar plates and stack hardware. The aim was to prove the design concept and to demonstrate functionality by operating the stack at >1 kW{sub e}/L and 500 W/kg for 200 hours operation. The project was extended to include the assembly and testing of two additional 1 kW{sub e} PEMFC stacks based on coated metal components. Low equivalent weight perfluorinated ionomer ion exchange membranes were prepared and were found to give a superior electrochemical performance to commercial materials. A technique to etch various stainless steel grades and control processes was successfully developed and optimised. Coatings for stainless steel and titanium were successfully developed and met the required performance criteria. All PEMFC stack components were selected and designed to enable subsequent commercial manufacture.

  13. Teaching the relation between solar cell efficiency and annual energy yield

    International Nuclear Information System (INIS)

    Sark, Wilfried G J H M van

    2007-01-01

    To reach a sustainable world the use of renewable energy sources is imperative. Photovoltaics (PV) is but one of the technologies that use the power of the sun and its deployment is growing very fast. Several master programs have been developed over the world, including Utrecht University, that teach these technologies. Within the framework of a course on energy conversion technologies, we have developed a classroom problem that focuses on the difference between PV efficiency and annual yield for the two locations: the Utrecht University campus and the African Sahara desert. In spreadsheet format, students calculate annual yield, and they find a best method to do so. The exercise can be done in about three hours, and students will learn that the annual yield in the Sahara is only twice that at Utrecht University,

  14. A Rapid Method for Quantifying Viable Mycobacterium avium subsp. paratuberculosis in Cellular Infection Assays

    Science.gov (United States)

    Pooley, Hannah B.; de Silva, Kumudika; Purdie, Auriol C.; Begg, Douglas J.; Whittington, Richard J.

    2016-01-01

    ABSTRACT Determining the viability of bacteria is a key outcome of in vitro cellular infection assays. Currently, this is done by culture, which is problematic for fastidious slow-growing bacteria such as Mycobacterium avium subsp. paratuberculosis, where it can take up to 4 months to confirm growth. This study aimed to identify an assay that can rapidly quantify the number of viable M. avium subsp. paratuberculosis cells in a cellular sample. Three commercially available bacterial viability assays along with a modified liquid culture method coupled with high-throughput quantitative PCR growth detection were assessed. Criteria for assessment included the ability of each assay to differentiate live and dead M. avium subsp. paratuberculosis organisms and their accuracy at low bacterial concentrations. Using the culture-based method, M. avium subsp. paratuberculosis growth was reliably detected and quantified within 2 weeks. There was a strong linear association between the 2-week growth rate and the initial inoculum concentration. The number of viable M. avium subsp. paratuberculosis cells in an unknown sample was quantified based on the growth rate, by using growth standards. In contrast, none of the commercially available viability assays were suitable for use with samples from in vitro cellular infection assays. IMPORTANCE Rapid quantification of the viability of Mycobacterium avium subsp. paratuberculosis in samples from in vitro cellular infection assays is important, as it allows these assays to be carried out on a large scale. In vitro cellular infection assays can function as a preliminary screening tool, for vaccine development or antimicrobial screening, and also to extend findings derived from experimental animal trials. Currently, by using culture, it takes up to 4 months to obtain quantifiable results regarding M. avium subsp. paratuberculosis viability after an in vitro infection assay; however, with the quantitative PCR and liquid culture method

  15. Endocochlear potential generation is associated with intercellular communication in the stria vascularis: structural analysis in the viable dominant spotting mouse mutant.

    Science.gov (United States)

    Carlisle, L; Steel, K; Forge, A

    1990-11-01

    Deafness in the viable dominant spotting mouse mutant is due to a primary defect of the stria vascularis which results in absence of the positive endocochlear potential in scala media. Endocochlear potentials were measured and the structure of stria vascularis of mutants with potentials close to zero was compared with that in normal littermate controls by use of morphometric methods. The stria vascularis was significantly thinner in mutants. Marginal cells were not significantly different from controls in terms of volume density or intramembrane particle density but the network density of tight junctions was significantly reduced in the mutants. A virtual absence of gap junctions between basal cells and marginal or intermediate cells was observed, but intramembrane particle density and junctional complexes between adjacent basal cells were not different from controls. The volume density of basal cells was significantly greater in mutants. Intermediate cells accounted for a significantly smaller volume density of the stria vascularis in mutants and had a lower density of intramembrane particles than controls. Melanocytes were not identified in the stria vascularis of mutants. These results suggest that communication between marginal, intermediate and basal cells might be important to the normal function of the stria vascularis.

  16. Enrichment of unlabeled human Langerhans cells from epidermal cell suspensions by discontinuous density gradient centrifugation

    NARCIS (Netherlands)

    Teunissen, M. B.; Wormmeester, J.; Kapsenberg, M. L.; Bos, J. D.

    1988-01-01

    In this report we introduce an alternative procedure for enrichment of human epidermal Langerhans cells (LC) from epidermal cell suspensions of normal skin. By means of discontinuous Ficoll-Metrizoate density gradient centrifugation, a fraction containing high numbers of viable, more than 80% pure

  17. Principles of root water uptake, soil salinity and crop yield for optimizing irrigation management

    International Nuclear Information System (INIS)

    Dirksen, C.

    1983-01-01

    The paper reviews the principles of water and salt transport, root water uptake, crop salt tolerance, water quality, and irrigation methods which should be considered in optimizing irrigation management for sustained, viable agriculture with protection of the quality of land and water resources. In particular, the advantages of high-frequency irrigation at small leaching fractions with closed systems are discussed, for which uptake-weighted mean salinity is expected to correlate best with crop yields. Optimization of irrigation management depends on the scale considered. Non-technical problems which are often much harder to solve than technical problems, may well be most favourable for new projects in developing countries. (author)

  18. Experimental human-like model to assess the part of viable Legionella reaching the thoracic region after nebulization.

    Directory of Open Access Journals (Sweden)

    Jérémie Pourchez

    Full Text Available The incidence of Legionnaires' disease (LD in European countries and the USA has been constantly increasing since 1998. Infection of humans occurs through aerosol inhalation. To bridge the existing gap between the concentration of Legionella in a water network and the deposition of bacteria within the thoracic region (assessment of the number of viable Legionella, we validated a model mimicking realistic exposure through the use of (i recent technology for aerosol generation and (ii a 3D replicate of the human upper respiratory tract. The model's sensitivity was determined by monitoring the deposition of (i aerosolized water and Tc99m radio-aerosol as controls, and (ii bioaerosols generated from both Escherichia coli and Legionella pneumophila sg 1 suspensions. The numbers of viable Legionella prior to and after nebulization were provided by culture, flow cytometry and qPCR. This study was designed to obtain more realistic data on aerosol inhalation (vs. animal experimentation and deposition at the thoracic region in the context of LD. Upon nebulization, 40% and 48% of the initial Legionella inoculum was made of cultivable and non-cultivable cells, respectively; 0.7% of both populations reached the filter holder mimicking the thoracic region in this setup. These results are in agreement with experimental data based on quantitative microbial risk assessment methods and bring new methods that may be useful for preventing LD.

  19. Enumeration of Vibrio parahaemolyticus in the viable but nonculturable state using direct plate counts and recognition of individual gene fluorescence in situ hybridization.

    Science.gov (United States)

    Griffitt, Kimberly J; Noriea, Nicholas F; Johnson, Crystal N; Grimes, D Jay

    2011-05-01

    Vibrio parahaemolyticus is a gram-negative, halophilic bacterium indigenous to marine and estuarine environments and it is capable of causing food and water-borne illness in humans. It can also cause disease in marine animals, including cultured species. Currently, culture-based techniques are used for quantification of V. parahaemolyticus in environmental samples; however, these can be misleading as they fail to detect V. parahaemolyticus in a viable but nonculturable (VBNC) state which leads to an underestimation of the population density. In this study, we used a novel fluorescence visualization technique, called recognition of individual gene fluorescence in situ hybridization (RING-FISH), which targets chromosomal DNA for enumeration. A polynucleotide probe labeled with Cyanine 3 (Cy3) was created corresponding to the ubiquitous V. parahaemolyticus gene that codes for thermolabile hemolysin (tlh). When coupled with the Kogure method to distinguish viable from dead cells, RING-FISH probes reliably enumerated total, viable V. parahaemolyticus. The probe was tested for sensitivity and specificity against a pure culture of tlh(+), tdh(-), trh(-)V. parahaemolyticus, pure cultures of Vibrio vulnificus, Vibrio harveyi, Vibrio alginolyticus and Vibrio fischeri, and a mixed environmental sample. This research will provide additional tools for a better understanding of the risk these environmental organisms pose to human health. Copyright © 2011 Elsevier B.V. All rights reserved.

  20. Contracting of energy services: often a viable alternative

    International Nuclear Information System (INIS)

    Milic, M.; Bruendler, M.

    2001-01-01

    This article discusses the outsourcing of energy services as a viable alternative to the operation of own energy facilities. The advantages of contracting for enterprises wanting to focus on their core competencies and have their energy infrastructure financed, built, maintained and operated by a third party are discussed. Financial aspects are looked at and examples in connection with the calculation of actual energy costs are given. The article is concluded with tips on the evaluation of offers for contracting services and on the definition of ownership aspects and property boundaries

  1. Evaluation of the BioVigilant IMD-A, a novel optical spectroscopy technology for the continuous and real-time environmental monitoring of viable and nonviable particles. Part II. Case studies in environmental monitoring during aseptic filling, intervention assessments, and glove integrity testing in manufacturing isolators.

    Science.gov (United States)

    Miller, Michael J; Walsh, Michael R; Shrake, Jerry L; Dukes, Randall E; Hill, Daniel B

    2009-01-01

    This paper describes the use of the BioVigilant IMD-A, a real-time and continuous monitoring technology based on optical spectroscopy, to simultaneously and instantaneously detect, size, and enumerate both viable and nonviable particles in a variety of filling and transfer isolator environments during an aseptic fill, transfer of sterilized components, and filling interventions. Continuous monitoring of three separate isolators for more than 16 h and representing more than 28 m3 of air per isolator (under static conditions) yielded a mean viable particle count of zero (0) per cubic meter. Although the mean count per cubic meter was zero, the detection of very low levels of single viable particles was randomly observed in each of these sampling runs. No viable particles were detected during the manual transfer of sterilized components from transfer isolators into a filling isolator, and similar results were observed during an aseptic fill, a filling needle change-out procedure, and during disassembly, movement, and reassembly of a vibrating stopper bowl. During the continuous monitoring of a sample transfer port and a simulated mousehole, no viable particles were detected; however, when the sampling probe was inserted beyond the isolator-room interface, the IMD-A instantaneously detected and enumerated both viable and nonviable particles originating from the surrounding room. Data from glove pinhole studies showed no viable particles being observed, although significant viable particles were immediately detected when the gloves were removed and a bare hand was allowed to introduce microorganisms into the isolator. The IMD-A technology offers the industry an unprecedented advantage over growth-based bioaerosol samplers for monitoring the state of microbiological control in pharmaceutical manufacturing environments, and represents significant progress toward the acceptance of microbiology process analytical technology solutions for the industry.

  2. A multicenter study of viable PCR using propidium monoazide to detect Legionella in water samples.

    Science.gov (United States)

    Scaturro, Maria; Fontana, Stefano; Dell'eva, Italo; Helfer, Fabrizia; Marchio, Michele; Stefanetti, Maria Vittoria; Cavallaro, Mario; Miglietta, Marilena; Montagna, Maria Teresa; De Giglio, Osvalda; Cuna, Teresa; Chetti, Leonarda; Sabattini, Maria Antonietta Bucci; Carlotti, Michela; Viggiani, Mariagabriella; Stenico, Alberta; Romanin, Elisa; Bonanni, Emma; Ottaviano, Claudio; Franzin, Laura; Avanzini, Claudio; Demarie, Valerio; Corbella, Marta; Cambieri, Patrizia; Marone, Piero; Rota, Maria Cristina; Bella, Antonino; Ricci, Maria Luisa

    2016-07-01

    Legionella quantification in environmental samples is overestimated by qPCR. Combination with a viable dye, such as Propidium monoazide (PMA), could make qPCR (named then vPCR) very reliable. In this multicentre study 717 artificial water samples, spiked with fixed concentrations of Legionella and interfering bacterial flora, were analysed by qPCR, vPCR and culture and data were compared by statistical analysis. A heat-treatment at 55 °C for 10 minutes was also performed to obtain viable and not-viable bacteria. When data of vPCR were compared with those of culture and qPCR, statistical analysis showed significant differences (P 0.05). Overall this study provided a good experimental reproducibility of vPCR but also highlighted limits of PMA in the discriminating capability of dead and live bacteria, making vPCR not completely reliable. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Three methods for isolating viable anthozoan endoderm cells with their intracellular symbiotic dinoflagellates

    Science.gov (United States)

    Gates, R. D.; Muscatine, L.

    1992-09-01

    Three maceration methods are described for the isolation of single endoderm cells from marine cnidarians. Two are enzymatic treatments suitable for fleshy anthozoans such as sea anemones and zoanthids. The third employs calcium free sea water and is suitable for stony corals. The viability and morphology of the endoderm cells is described using fluorogenic dyes and scanning and transmission electron microscopy.

  4. Symbiotic Role of the Viable but Nonculturable State of Vibrio fischeri in Hawaiian Coastal Seawater.

    Science.gov (United States)

    Lee, K; Ruby, E G

    1995-01-01

    To achieve functional bioluminescence, the developing light organ of newly hatched juveniles of the Hawaiian squid Euprymna scolopes must become colonized by luminous, symbiosis-competent Vibrio fischeri present in the ambient seawater. This benign infection occurs rapidly in animals placed in seawater from the host's natural habitat. Therefore, it was surprising that colony hybridization studies with a V. fischeri-specific luxA gene probe indicated the presence of only about 2 CFU of V. fischeri per ml of this infective seawater. To examine this paradox, we estimated the total concentration of V. fischeri cells present in seawater from the host's habitat in two additional ways. In the first approach, the total bacterial assemblage in samples of seawater was collected on polycarbonate membrane filters and used as a source of both a crude cell lysate and purified DNA. These preparations were then assayed by quantitative DNA-DNA hybridization with the luxA gene probe. The results suggested the presence of between 200 and 400 cells of V. fischeri per ml of natural seawater, a concentration more than 100 times that revealed by colony hybridization. In the second approach, we amplified V. fischeri-specific luxA sequences from microliter volumes of natural seawater by PCR. Most-probable-number analyses of the frequency of positive PCR results from cell lysates in these small volumes gave an estimate of the concentration of V. fischeri luxA gene targets of between 130 and 1,680 copies per ml. From these measurements, we conclude that in their natural seawater environment, the majority of V. fischeri cells become nonculturable while remaining viable and symbiotically infective. Experimental studies indicated that V. fischeri cells suspended in natural Hawaiian seawater enter such a state within a few days.

  5. Brazilian Soybean Yields and Yield Gaps Vary with Farm Size

    Science.gov (United States)

    Jeffries, G. R.; Cohn, A.; Griffin, T. S.; Bragança, A.

    2017-12-01

    Understanding the farm size-specific characteristics of crop yields and yield gaps may help to improve yields by enabling better targeting of technical assistance and agricultural development programs. Linking remote sensing-based yield estimates with property boundaries provides a novel view of the relationship between farm size and yield structure (yield magnitude, gaps, and stability over time). A growing literature documents variations in yield gaps, but largely ignores the role of farm size as a factor shaping yield structure. Research on the inverse farm size-productivity relationship (IR) theory - that small farms are more productive than large ones all else equal - has documented that yield magnitude may vary by farm size, but has not considered other yield structure characteristics. We examined farm size - yield structure relationships for soybeans in Brazil for years 2001-2015. Using out-of-sample soybean yield predictions from a statistical model, we documented 1) gaps between the 95th percentile of attained yields and mean yields within counties and individual fields, and 2) yield stability defined as the standard deviation of time-detrended yields at given locations. We found a direct relationship between soy yields and farm size at the national level, while the strength and the sign of the relationship varied by region. Soybean yield gaps were found to be inversely related to farm size metrics, even when yields were only compared to farms of similar size. The relationship between farm size and yield stability was nonlinear, with mid-sized farms having the most stable yields. The work suggests that farm size is an important factor in understanding yield structure and that opportunities for improving soy yields in Brazil are greatest among smaller farms.

  6. Yields, photosynthetic efficiencies, and proximate chemical composition of dense cultures of marine microalgae. A subcontract report

    Energy Technology Data Exchange (ETDEWEB)

    Thomas, W.H.; Seibert, D.L.R.; Alden, M.; Eldridge, P.; Neori, A.

    1983-07-01

    The yields, photosynthetic efficiencies, and proximate composition of several microalgae were compared in dense cultures grown at light intensities up to 70% sunlight. Yields ranged from 3.4 to 21.7 g dry weight/m/sup 2/ day. The highest yield was obtained with Phaeodactylum; the lowest in Botryococcus cultures. The same species had the highest and lowest efficiencies of utilization of photosynthetically active radiation. In nitrogen-sufficient cells of all but one species, most of the dry weight consisted of protein. Lipid content of all species was 20 to 29%, and carbohydrate content 11 to 23%. Lipid content increased somewhat in N-deficient Phaeodactylum and Isochrysis cells, but decreased in deficient Monallanthus cells. Because the overall dry weight yield was reduced by deficiency, lipid yields did not increase. However, since the carbohydrate content increased to about 65% in N-deficient Dunaliella and Tetraselmis cells, the carbohydrate yield increased. In Phaeodactylum the optimum light intensity was about 40% of full sunlight. Most experimets with this alga included a CUSO/sub 4/ filter to decrease infrared irradiance. When this filter was removed, the yield increased because more red light in the photosynthetically active spectral range was included. These results should prove useful to workers attempting to maximize yields and efficiencies, but additional studies are needed. 69 references, 27 figures, 18 tables.

  7. Fuel cells are a commercially viable alternative for the production of "clean" energy.

    Science.gov (United States)

    Niakolas, Dimitris K; Daletou, Maria; Neophytides, Stylianos G; Vayenas, Constantinos G

    2016-01-01

    Fuel cells present a highly efficient and environmentally friendly alternative technology for decentralized energy production. The scope of the present study is to provide an overview of the technological and commercialization readiness level of fuel cells. Specifically, there is a brief description of their general advantages and weaknesses in correlation with various technological actions and political strategies, which are adopted towards their proper positioning in the global market. Some of the most important key performance indicators are also discussed, alongside with a few examples of broad commercialization. It is concluded that the increasing number of companies which utilize and invest on this technology, in combination with the supply chain improvements and the concomitant technological maturity and recognition, reinforce the fuel cell industry so as to become well-aligned for global success.

  8. Simultaneous treatment (cell disruption and lipid extraction) of wet microalgae using hydrodynamic cavitation for enhancing the lipid yield.

    Science.gov (United States)

    Lee, Ilgyu; Han, Jong-In

    2015-06-01

    Simultaneous treatment (combining with cell disruption and lipid extraction) using hydrodynamic cavitation (HC) was applied to Nannochloropsis salina to demonstrate a simple and integrated way to produce oil from wet microalgae. A high lipid yield from the HC (25.9-99.0%) was observed compared with autoclave (16.2-66.5%) and ultrasonication (5.4-26.9%) in terms of the specific energy input (500-10,000 kJ/kg). The optimal conditions for the simultaneous treatment were established using a statistical approach. The efficiency of the simultaneous method was also demonstrated by comparing each separate treatment. The maximum lipid yield (predicted: 45.9% and experimental: 45.5%) was obtained using 0.89% sulfuric acid with a cavitation number of 1.17 for a reaction time of 25.05 min via response surface methodology. Considering its comparable extractability, energy-efficiency, and potential for scale-up, HC may be a promising method to achieve industrial-scale microalgae operation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. The immunomodulatory properties of viable Lactobacillus salivarius ssp. salivarius CECT5713 are not restricted to the large intestine.

    Science.gov (United States)

    Arribas, Belén; Garrido-Mesa, Natividad; Perán, Laura; Camuesco, Desirée; Comalada, Mònica; Bailón, Elvira; Olivares, Mónica; Xaus, Jordi; Kruidenier, Laurens; Sanderson, Ian R; Zarzuelo, Antonio; Rodríguez-Cabezas, Maria Elena; Gálvez, Julio

    2012-04-01

    The aim of this study was to better characterise the biological effects of Lactobacillus salivarius ssp. salivarius CECT5713, a probiotic with immunomodulatory properties. Live or dead probiotic was assayed in the TNBS model of rat colitis to determine whether viability was a requisite to exert the beneficial effects. In vitro studies were also performed in Caco-2 cells to evaluate its effects on epithelial cell recovery and IL-8 production. Finally, the probiotic was assayed in the LPS model of septic shock in mice to establish its effects when there is an altered systemic immune response. The viability of the probiotic was required for its anti-inflammatory activity. The probiotic inhibited IL-8 production in stimulated Caco-2 cells and facilitated the recovery of damaged intestinal epithelium. In LPS-treated mice, the probiotic inhibited the production of TNFα in plasma and lungs and increased the hepatic glutathione content. These effects were associated with an improvement in the altered production of the T-cell cytokines in splenocytes, by reducing IL-2 and IL-5 and by increasing IL-10. Finally, it reduced the increased plasma IgG production in LPS-treated mice. The anti-inflammatory effects of viable L. salivarius ssp. salivarius CECT5713 are not restricted to the gastrointestinal tract.

  10. Collateral circulation as a marker of the presence of viable myocardium in patients with recent myocardial infarction

    International Nuclear Information System (INIS)

    Fujita, M.; Ohno, A.; Wada, O.; Miwa, K.; Nozawa, T.; Yamanishi, K.; Sasayama, S.

    1991-01-01

    The relationship between the presence of viable myocardium and the extent of coronary collateral circulation to the infarct area was evaluated in 20 patients with a recent anterior myocardial infarction who had complete obstruction of the left anterior descending coronary artery. The viability of myocardial tissue was assessed by exercise thallium-201 myocardial scintigraphy, and the collateral circulation was angiographically evaluated by means of a collateral index ranging from 0 to 3. Patients were divided into two groups according to the presence (group 1, n = 10) or absence (group 2, n = 10) of viable myocardium in the perfusion territory of the infarct-related artery. The collateral index in group 1 was 2.5 ± 0.5 (SD), which was significantly higher than the 0.7 ± 0.8 in group 2. These findings indicate that the presence of ischemic but viable myocardium is intimately related to the development of collateral circulation in patients with myocardial infarction, and the existence of well-developed collateral channels predicts the presence of viable myocardium in the infarct area

  11. Engineering tolerance to industrially relevant stress factors in yeast cell factories.

    Science.gov (United States)

    Deparis, Quinten; Claes, Arne; Foulquié-Moreno, Maria R; Thevelein, Johan M

    2017-06-01

    The main focus in development of yeast cell factories has generally been on establishing optimal activity of heterologous pathways and further metabolic engineering of the host strain to maximize product yield and titer. Adequate stress tolerance of the host strain has turned out to be another major challenge for obtaining economically viable performance in industrial production. Although general robustness is a universal requirement for industrial microorganisms, production of novel compounds using artificial metabolic pathways presents additional challenges. Many of the bio-based compounds desirable for production by cell factories are highly toxic to the host cells in the titers required for economic viability. Artificial metabolic pathways also turn out to be much more sensitive to stress factors than endogenous pathways, likely because regulation of the latter has been optimized in evolution in myriads of environmental conditions. We discuss different environmental and metabolic stress factors with high relevance for industrial utilization of yeast cell factories and the experimental approaches used to engineer higher stress tolerance. Improving stress tolerance in a predictable manner in yeast cell factories should facilitate their widespread utilization in the bio-based economy and extend the range of products successfully produced in large scale in a sustainable and economically profitable way. © FEMS 2017.

  12. Engineering tolerance to industrially relevant stress factors in yeast cell factories

    Science.gov (United States)

    Deparis, Quinten; Claes, Arne; Foulquié-Moreno, Maria R.

    2017-01-01

    Abstract The main focus in development of yeast cell factories has generally been on establishing optimal activity of heterologous pathways and further metabolic engineering of the host strain to maximize product yield and titer. Adequate stress tolerance of the host strain has turned out to be another major challenge for obtaining economically viable performance in industrial production. Although general robustness is a universal requirement for industrial microorganisms, production of novel compounds using artificial metabolic pathways presents additional challenges. Many of the bio-based compounds desirable for production by cell factories are highly toxic to the host cells in the titers required for economic viability. Artificial metabolic pathways also turn out to be much more sensitive to stress factors than endogenous pathways, likely because regulation of the latter has been optimized in evolution in myriads of environmental conditions. We discuss different environmental and metabolic stress factors with high relevance for industrial utilization of yeast cell factories and the experimental approaches used to engineer higher stress tolerance. Improving stress tolerance in a predictable manner in yeast cell factories should facilitate their widespread utilization in the bio-based economy and extend the range of products successfully produced in large scale in a sustainable and economically profitable way. PMID:28586408

  13. Monitoring of viable airborne SARS virus in ambient air

    Science.gov (United States)

    Agranovski, Igor E.; Safatov, Alexander S.; Pyankov, Oleg V.; Sergeev, Alexander N.; Agafonov, Alexander P.; Ignatiev, Georgy M.; Ryabchikova, Elena I.; Borodulin, Alexander I.; Sergeev, Artemii A.; Doerr, Hans W.; Rabenau, Holger F.; Agranovski, Victoria

    Due to recent SARS related issues (Science 300 (5624) 1394; Nature 423 (2003) 240; Science 300 (5627) 1966), the development of reliable airborne virus monitoring procedures has become galvanized by an exceptional sense of urgency and is presently in a high demand (In: Cox, C.S., Wathers, C.M. (Eds.), Bioaerosols Handbook, Lewis Publishers, Boca Raton, FL, 1995, pp. 247-267). Based on engineering control method (Aerosol Science and Technology 31 (1999) 249; 35 (2001) 852), which was previously applied to the removal of particles from gas carriers, a new personal bioaerosol sampler has been developed. Contaminated air is bubbled through porous medium submerged into liquid and subsequently split into multitude of very small bubbles. The particulates are scavenged by these bubbles, and, thus, effectively removed. The current study explores its feasibility for monitoring of viable airborne SARS virus. It was found that the natural decay of such virus in the collection fluid was around 0.75 and 1.76 lg during 2 and 4 h of continuous operation, respectively. Theoretical microbial recovery rates of higher than 55 and 19% were calculated for 1 and 2 h of operation, respectively. Thus, the new sampling method of direct non-violent collection of viable airborne SARS virus into the appropriate liquid environment was found suitable for monitoring of such stress sensitive virus.

  14. In vitro inhibition of Eimeria tenella sporozoite invasion into host cells by probiotics.

    Science.gov (United States)

    Hessenberger, S; Schatzmayr, G; Teichmann, K

    2016-10-15

    The aim was to study the effects of probiotics isolated from the intestinal tract of livestock animals on Eimeria tenella invasion into Madin-Darby bovine kidney (MDBK) cells in vitro. E. tenella sporozoites were purified and labeled with 5(6)-carboxyfluorescein diacetate N-succinimidyl ester before seeding on cell cultures, and invasion was evaluated by fluorescence microscopy. Two protocols (A and B) were used. In protocol A, Enterococcus faecium # 589 or Lactobacillus salivarius subsp. salivarius # 505 were added together with sporozoites to MDBK cell cultures and invasion was evaluated after incubation for approximately 20h. Viable, dead, or spent culture supernatants of probiotics were tested. In protocol B, viable probiotics were incubated with MDBK cells for one hour before sporozoites were added and invasion was evaluated after two more hours of incubation. Parasite invasion of viable, dead, or spent culture supernatant of E. faecium # 589 was assessed. Using protocol A, it was shown that parasite invasion was inhibited by viable (80%) or dead (75%) E. faecium # 589. While inhibition by viable L. salivarius subsp. salivarius # 505 was not valid at the highest concentration and not significant at the other test concentrations, dead cells inhibited parasite invasion up to 45%. Spent culture supernatants of both probiotics had no influence on parasite invasion. Using protocol B, it was shown that viable Bifidobacterium animalis subsp. animalis # 503, E. faecium # 497, E. faecium # 589, L. reuteri # 514, L. salivarius subsp. salivarius # 505, and Bacillus subtilis # 588 inhibited parasite invasion into MDBK cells up to 80%. Anticoccidial activity was strain-specific for E. faecium strains, and the strongest effect was shown by E. faecium # 589. Anticoccidial effects of some of the tested probiotics have already been shown in vivo, which makes them candidates to prevent coccidiosis. These findings have now been confirmed in vitro. The used parasite invasion

  15. Resveratrol production in bioreactor: Assessment of cell physiological states and plasmid segregational stability

    Directory of Open Access Journals (Sweden)

    Margarida S. Afonso

    2015-03-01

    Full Text Available Resveratrol is a plant secondary metabolite commonly found in peanuts and grapevines with significant health benefits. Recombinant organisms can produce large amounts of resveratrol and, in this work, Escherichia coli BW27784 was used to produce resveratrol in bioreactors while monitoring cell physiology and plasmid stability through flow cytometry and real-time qPCR, respectively. Initially, the influence of culture conditions and precursor addition was evaluated in screening assays and the data gathered was used to perform the bioreactor assays, allowing the production of 160 μg/mL of resveratrol. Cellular physiology and plasmid instability affected the final resveratrol production, with lower viability and plasmid copy numbers associated with lower yields. In sum, this study describes new tools to monitor the bioprocess, evaluating the effect of culture conditions, and its correlation with cell physiology and plasmid segregational stability, in order to define a viable and scalable bioprocess to fulfill the need for larger quantities of resveratrol.

  16. Roots of success: cultivating viable community forestry

    Energy Technology Data Exchange (ETDEWEB)

    MacQueen, Duncan

    2009-05-15

    Is community forestry emerging from the shadows? The evidence shows that locally controlled enterprises can be economically viable, and often build on stronger social and environmental foundations than the big private-sector players. Certainly this is an industry in need of a shakeup. Many forests have become flashpoints where agro-industry, large-scale logging concerns and conservation interests clash, while forest-dependent communities are left out in the cold. Meanwhile, governments – driven by concerns over the climate impacts of deforestation – are having to gear up for legal, sustainable forestry production. Community forestry could be crucial to solving many of these challenges. By building on local core capabilities and developing strategic partnerships, they are forging key new business models that could transform the sector.

  17. Rapid Quantification of Viable Campylobacter Bacteria on Chicken Carcasses, Using Real-Time PCR and Propidium Monoazide Treatment, as a Tool for Quantitative Risk Assessment

    DEFF Research Database (Denmark)

    Josefsen, Mathilde Hartmann; Löfström, Charlotta; Hansen, Tina Beck

    2010-01-01

    A number of intervention strategies against Campylobacter contaminated poultry focus on post-slaughter reduction of the number of cells, emphasizing the need for rapid and reliable quantitative detection of only viable Campylobacter. We present a new and rapid quantitative approach for enumeration...... method does not detect DNA from dead Campylobacter, but recognises the infectious potential of the VBNC state, and is thereby able to assess the effect of control strategies, and provide trustworthy data for risk assessment....

  18. Design and validation of a consistent and reproducible manufacture process for the production of clinical-grade bone marrow-derived multipotent mesenchymal stromal cells.

    Science.gov (United States)

    Codinach, Margarita; Blanco, Margarita; Ortega, Isabel; Lloret, Mireia; Reales, Laura; Coca, Maria Isabel; Torrents, Sílvia; Doral, Manel; Oliver-Vila, Irene; Requena-Montero, Miriam; Vives, Joaquim; Garcia-López, Joan

    2016-09-01

    Multipotent mesenchymal stromal cells (MSC) have achieved a notable prominence in the field of regenerative medicine, despite the lack of common standards in the production processes and suitable quality controls compatible with Good Manufacturing Practice (GMP). Herein we describe the design of a bioprocess for bone marrow (BM)-derived MSC isolation and expansion, its validation and production of 48 consecutive batches for clinical use. BM samples were collected from the iliac crest of patients for autologous therapy. Manufacturing procedures included: (i) isolation of nucleated cells (NC) by automated density-gradient centrifugation and plating; (ii) trypsinization and expansion of secondary cultures; and (iii) harvest and formulation of a suspension containing 40 ± 10 × 10(6) viable cells. Quality controls were defined as: (i) cell count and viability assessment; (ii) immunophenotype; and (iii) sterility tests, Mycoplasma detection, endotoxin test and Gram staining. A 3-week manufacturing bioprocess was first designed and then validated in 3 consecutive mock productions, prior to producing 48 batches of BM-MSC for clinical use. Validation included the assessment of MSC identity and genetic stability. Regarding production, 139.0 ± 17.8 mL of BM containing 2.53 ± 0.92 × 10(9) viable NC were used as starting material, yielding 38.8 ± 5.3 × 10(6) viable cells in the final product. Surface antigen expression was consistent with the expected phenotype for MSC, displaying high levels of CD73, CD90 and CD105, lack of expression of CD31 and CD45 and low levels of HLA-DR. Tests for sterility, Mycoplasma, Gram staining and endotoxin had negative results in all cases. Herein we demonstrated the establishment of a feasible, consistent and reproducible bioprocess for the production of safe BM-derived MSC for clinical use. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  19. Effect of Induced Pluripotent Stem Cell Technology in Blood Banking

    Science.gov (United States)

    Focosi, Daniele

    2016-01-01

    Summary Population aging has imposed cost-effective alternatives to blood donations. Artificial blood is still at the preliminary stages of development, and the need for viable cells seems unsurmountable. Because large numbers of viable cells must be promptly available for clinical use, stem cell technologies, expansion, and banking represent ideal tools to ensure a regular supply. Provided key donors can be identified, induced pluripotent stem cell (iPSC) technology could pave the way to a new era in transfusion medicine, just as it is already doing in many other fields of medicine. The present review summarizes the current state of research on iPSC technology in the field of blood banking, highlighting hurdles, and promises. Significance The aging population in Western countries is causing a progressive reduction of blood donors and a constant increase of blood recipients. Because blood is the main therapeutic option to treat acute hemorrhage, cost-effective alternatives to blood donations are being actively investigated. The enormous replication capability of induced pluripotent stem cells and their promising results in many other fields of medicine could be an apt solution to produce the large numbers of viable cells required in transfusion and usher in a new era in transfusion medicine. The present report describes the potentiality, technological hurdles, and promises of induced pluripotent stem cells to generate red blood cells by redifferentiation. PMID:26819256

  20. Electron-deficient N-alkyloyl derivatives of thieno[3,4-c]pyrrole-4,6-dione yield efficient polymer solar cells with open-circuit voltages > 1 v

    KAUST Repository

    Warnan, Julien

    2014-05-13

    Poly(benzo[1,2-b:4,5-b′]dithiophene-thieno[3,4-c]pyrrole-4,6-dione) (PBDTTPD) polymer donors yield some of the highest open-circuit voltages (V OC, ca. 0.9 V) and fill factors (FF, ca. 70%) in conventional bulk-heterojunction (BHJ) solar cells with PCBM acceptors. Recent work has shown that the incorporation of ring substituents into the side chains of the BDT motifs in PBDTTPD can induce subtle variations in material properties, resulting in an increase of the BHJ device VOC to ∼1 V. In this contribution, we report on the synthesis of N-alkyloyl-substituted TPD motifs (TPD(CO)) and show that the electron-deficient motifs can further lower both the polymer LUMO and HOMO levels, yielding device VOC > 1 V (up to ca. 1.1 V) in BHJ solar cells with PCBM. Despite the high VOC achieved (i.e., low polymer HOMO), BHJ devices cast from TPD(CO)-based polymer donors can reach power conversion efficiencies (PCEs) of up to 6.7%, making these promising systems for use in the high-band-gap cell of tandem solar cells. © 2014 American Chemical Society.

  1. Mapping In Vivo Tumor Oxygenation within Viable Tumor by 19F-MRI and Multispectral Analysis

    Directory of Open Access Journals (Sweden)

    Yunzhou Shi

    2013-11-01

    Full Text Available Quantifying oxygenation in viable tumor remains a major obstacle toward a better understanding of the tumor microenvironment and improving treatment strategies. Current techniques are often complicated by tumor heterogeneity. Herein, a novel in vivo approach that combines 19F magnetic resonance imaging (19F-MRIR1 mapping with diffusionbased multispectral (MS analysis is introduced. This approach restricts the partial pressure of oxygen (pO2 measurements to viable tumor, the tissue of therapeutic interest. The technique exhibited sufficient sensitivity to detect a breathing gas challenge in a xenograft tumor model, and the hypoxic region measured by MS 19F-MRI was strongly correlated with histologic estimates of hypoxia. This approach was then applied to address the effects of antivascular agents on tumor oxygenation, which is a research question that is still under debate. The technique was used to monitor longitudinal pO2 changes in response to an antibody to vascular endothelial growth factor (B20.4.1.1 and a selective dual phosphoinositide 3-kinase/mammalian target of rapamycin inhibitor (GDC-0980. GDC-0980 reduced viable tumor pO2 during a 3-day treatment period, and a significant reduction was also produced by B20.4.1.1. Overall, this method provides an unprecedented view of viable tumor pO2 and contributes to a greater understanding of the effects of antivascular therapies on the tumor's microenvironment.

  2. Models to estimate lactation curves of milk yield and somatic cell count in dairy cows at the herd level for the use in simulations and predictive models

    Directory of Open Access Journals (Sweden)

    Kaare Græsbøll

    2016-12-01

    Full Text Available Typically, central milk recording data from dairy herds are recorded less than monthly. Over-fitting early in lactation periods is a challenge, which we explored in different ways by reducing the number of parameters needed to describe the milk yield and somatic cell count of individual cows. Furthermore, we investigated how the parameters of lactation models correlate between parities and from dam to offspring. The aim of the study was to provide simple and robust models for cow level milk yield and somatic cell count (SCC for fitting to sparse data to parameterise herd- and cow-specific simulation of dairy herds.Data from 610 Danish Holstein herds were used to determine parity traits in milk production regarding milk yield and SCC of individual cows. Parity was stratified in first, second and third and higher for milk, and first to sixth and higher for SCC. Fitting of herd level parameters allowed for cow level lactation curves with three, two or one-parameters per lactation. Correlations of milk yield and SCC were estimated between lactations and between dam and offspring.The shape of the lactation curves varied markedly between farms. The correlation between lactations for milk yield and SCC were 0.2-0.6 and significant on more than 95% of farms. The variation in the daily milk yield was observed to be a source of variation to the SCC, and the total SCC was less correlated with the milk production than somatic cells per ml. A positive correlation was found between relative levels of the total SCC and the milk yield.The variation of lactation and SCC curves between farms highlights the importance of a herd level approach. The one-parameter per cow model using a herd level curve allows for estimating the cow production level from first the recording in the parity, while a two-parameter model requires more recordings for a credible estimate, but may more precisely predict persistence, and given the independence of parameters, these can be

  3. Effects of sowing date on the growth and yield of maize cultivars ...

    African Journals Online (AJOL)

    a

    Full Length Research Paper. Effects of sowing date on the ... days (GDD) and was the highest for seed yield and all yield components. The plant sown on 5th ... volume and weight), while plant development refers to processes related to cell ...

  4. Comparison of perimeter trap crop varieties: effects on herbivory, pollination, and yield in butternut squash.

    Science.gov (United States)

    Adler, L S; Hazzard, R V

    2009-02-01

    Perimeter trap cropping (PTC) is a method of integrated pest management (IPM) in which the main crop is surrounded with a perimeter trap crop that is more attractive to pests. Blue Hubbard (Cucurbita maxima Duch.) is a highly effective trap crop for butternut squash (C. moschata Duch. ex Poir) attacked by striped cucumber beetles (Acalymma vittatum Fabricius), but its limited marketability may reduce adoption of PTC by growers. Research comparing border crop varieties is necessary to provide options for growers. Furthermore, pollinators are critical for cucurbit yield, and the effect of PTC on pollination to main crops is unknown. We examined the effect of five border treatments on herbivory, pollination, and yield in butternut squash and manipulated herbivory and pollination to compare their importance for main crop yield. Blue Hubbard, buttercup squash (C. maxima Duch.), and zucchini (C. pepo L.) were equally attractive to cucumber beetles. Border treatments did not affect butternut leaf damage, but butternut flowers had the fewest beetles when surrounded by Blue Hubbard or buttercup squash. Yield was highest in the Blue Hubbard and buttercup treatments, but this effect was not statistically significant. Native bees accounted for 87% of pollinator visits, and pollination did not limit yield. There was no evidence that border crops competed with the main crop for pollinators. Our results suggest that both buttercup squash and zucchini may be viable alternatives to Blue Hubbard as borders for the main crop of butternut squash. Thus, growers may have multiple border options that reduce pesticide use, effectively manage pests, and do not disturb mutualist interactions with pollinators.

  5. Redox balance is key to explaining full vs. partial switching to low-yield metabolism

    Directory of Open Access Journals (Sweden)

    van Hoek Milan JA

    2012-03-01

    Full Text Available Abstract Background Low-yield metabolism is a puzzling phenomenon in many unicellular and multicellular organisms. In abundance of glucose, many cells use a highly wasteful fermentation pathway despite the availability of a high-yield pathway, producing many ATP molecules per glucose, e.g., oxidative phosphorylation. Some of these organisms, including the lactic acid bacterium Lactococcus lactis, downregulate their high-yield pathway in favor of the low-yield pathway. Other organisms, including Escherichia coli do not reduce the flux through the high-yield pathway, employing the low-yield pathway in parallel with a fully active high-yield pathway. For what reasons do some species use the high-yield and low-yield pathways concurrently and what makes others downregulate the high-yield pathway? A classic rationale for metabolic fermentation is overflow metabolism. Because the throughput of metabolic pathways is limited, influx of glucose exceeding the pathway's throughput capacity is thought to be redirected into an alternative, low-yield pathway. This overflow metabolism rationale suggests that cells would only use fermentation once the high-yield pathway runs at maximum rate, but it cannot explain why cells would decrease the flux through the high-yield pathway. Results Using flux balance analysis with molecular crowding (FBAwMC, a recent extension to flux balance analysis (FBA that assumes that the total flux through the metabolic network is limited, we investigate the differences between Saccharomyces cerevisiae and L. lactis that downregulate the high-yield pathway at increasing glucose concentrations, and E. coli, which keeps the high-yield pathway functioning at maximal rate. FBAwMC correctly predicts the metabolic switching mode in these three organisms, suggesting that metabolic network architecture is responsible for differences in metabolic switching mode. Based on our analysis, we expect gradual, "overflow-like" switching behavior in

  6. A method for the isolation and culture of adult rat retinal pigment epithelial (RPE cells to study retinal diseases

    Directory of Open Access Journals (Sweden)

    Janosch Peter Heller

    2015-11-01

    Full Text Available Diseases such as age-related macular degeneration (AMD affect the retinal pigment epithelium (RPE and lead to the death of the epithelial cells and ultimately blindness. RPE transplantation is currently a major focus of eye research and clinical trials using human stem cell-derived RPE cells are ongoing. However, it remains to be established to which extent the source of RPE cells for transplantation affects their therapeutic efficacy and this needs to be explored in animal models. Autotransplantation of RPE cells has attractions as a therapy, but existing protocols to isolate adult RPE cells from rodents are technically difficult, time-consuming, have a low yield and are not optimized for long-term cell culturing. Here, we report a newly devised protocol which facilitates reliable and simple isolation and culture of RPE cells from adult rats. Incubation of a whole rat eyeball in 20 U/ml papain solution for 50 minutes yielded 4 x 104 viable RPE cells. These cells were hexagonal and pigmented upon culture. Using immunostaining, we demonstrated that the cells expressed RPE cell-specific marker proteins including cytokeratin 18 and RPE65, similar to RPE cells in vivo. Additionally, the cells were able to produce and secrete Bruch’s membrane matrix components similar to in vivo situation. Similarly, the cultured RPE cells adhered to isolated Bruch’s membrane as has previously been reported. Therefore, the protocol described in this article provides an efficient method for the rapid and easy isolation of high quantities of adult rat RPE cells. This provides a reliable platform for studying the therapeutic targets, testing the effects of drugs in a preclinical setup and to perform in vitro and in vivo transplantation experiments to study retinal diseases.

  7. Role of chromosome stability and telomere length in the production of viable cell lines for somatic cell nuclear transfer

    Directory of Open Access Journals (Sweden)

    Betts Dean H

    2006-08-01

    Full Text Available Abstract Background Somatic cell nuclear transfer (SCNT provides an appealing alternative for the preservation of genetic material in non-domestic and endangered species. An important prerequisite for successful SCNT is the availability of good quality donor cells, as normal embryo development is dependent upon proper reprogramming of the donor genome so that embryonic genes can be appropriately expressed. The characteristics of donor cell lines and their ability to produce embryos by SCNT were evaluated by testing the effects of tissue sample collection (DART biopsy, PUNCH biopsy, post-mortem EAR sample and culture initiation (explant, collagenase digestion techniques. Results Differences in initial sample size based on sample collection technique had an effect on the amount of time necessary for achieving primary confluence and the number of population doublings (PDL produced. Thus, DART and PUNCH biopsies resulted in cultures with decreased lifespans (50 PDL and chromosomally stable (>70% normal cells at 20 PDL cultures produced by post-mortem EAR samples. Chromosome stability was influenced by sample collection technique and was dependent upon the culture's initial telomere length and its rate of shortening over cell passages. Following SCNT, short-lived cultures resulted in significantly lower blastocyst development (≤ 0.9% compared to highly proliferative cultures (11.8%. Chromosome stability and sample collection technique were significant factors in determining blastocyst development outcome. Conclusion These data demonstrate the influence of culture establishment techniques on cell culture characteristics, including the viability, longevity and normality of cells. The identification of a quantifiable marker associated with SCNT embryo developmental potential, chromosome stability, provides a means by which cell culture conditions can be monitored and improved.

  8. High Harvest Yield, High Expansion, and Phenotype Stability of CD146 Mesenchymal Stromal Cells from Whole Primitive Human Umbilical Cord Tissue

    Directory of Open Access Journals (Sweden)

    Rebecca C. Schugar

    2009-01-01

    Full Text Available Human umbilical cord blood is an excellent primitive source of noncontroversial stem cells for treatment of hematologic disorders; meanwhile, new stem cell candidates in the umbilical cord (UC tissue could provide therapeutic cells for nonhematologic disorders. We show novel in situ characterization to identify and localize a panel of some markers expressed by mesenchymal stromal cells (MSCs; CD44, CD105, CD73, CD90 and CD146 in the UC. We describe enzymatic isolation and purification methods of different UC cell populations that do not require manual separation of the vessels and stroma of the coiled, helical-like UC tissue. Unique quantitation of in situ cell frequency and stromal cell counts upon harvest illustrate the potential to obtain high numerical yields with these methods. UC stromal cells can differentiate to the osteogenic and chondrogenic lineages and, under specific culturing conditions, they exhibit high expandability with unique long-term stability of their phenotype. The remarkable stability of the phenotype represents a novel finding for human MSCs, from any source, and supports the use of these cells as highly accessible stromal cells for both basic studies and potentially therapeutic applications such as allogeneic clinical use for musculoskeletal disorders.

  9. Pyramiding genes and alleles for improving energy cane biomass yield

    Energy Technology Data Exchange (ETDEWEB)

    Ming, Ray [University of Illinois at Urbana-Champaign; Nagai, Chifumi [Hawaii Agriculture Research Center; Yu, Qingyi [Texas A & M AgriLife Research

    2018-03-23

    The overall goal of this project is to identify genes and gene interaction networks contributed to the extreme segregants with 30 folds biomass yield difference in sugarcane F2 populations. Towards achieving this goal, yield trials of 108 F2 extreme segregants from S. officinarum LA Purple and S. robustum MOL5829 (LM population) were carried out in two locations in three years. A yield trial of the second F2 population from S. officinarum LA Purple and S. spontaneum US56-14-4 (LU population) was installed in the summer of 2014 and the first set of yield component data was collected. For genotyping, transcriptomes from leaves and stalks of 70 extreme segregants of the LM F2 population and 119 individuals of the LU F2 populations were sequenced. The genomes of 91 F1 individuals from the LM populations are being sequenced to construct ultra-high density genetic maps for each of the two parents for both assisting the LA Purple genome assembling and for testing a hypothesis of female restitution. The genomes of 110 F2 individuals from single F1 in the LU population, a different set from the 119 F2 individuals used for transcriptome sequencing, are being sequenced for mapping genes and QTLs affecting biomass yield and for testing a hypothesis of female restitution. Gene expression analysis between extreme segregants of high and low biomass yield showed up-regulation of cellulose synthase, cellulose, and xylan synthase in high biomass yield segregants among 3,274 genes differentially expressed between the two extremes. Our transcriptome results revealed not only the increment of cell wall biosynthesis pathway is essential, but the rapid turnover of certain cell wall polymers as well as carbohydrate partitioning are also important for recycling and energy conservation during rapid cell growth in high biomass sugarcane. Seventeen differentially expressed genes in auxin, one in ethylene and one in gibberellin related signaling and biosynthesis pathways were identified, which

  10. Comparing milk yield, chemical properties and somatic cell count from organic and conventional mountain farming systems

    Directory of Open Access Journals (Sweden)

    Marcello Bianchi

    2010-01-01

    Full Text Available A study was undertaken to investigate the effects of farming systems (organic vs. conventional, diet (hay/concentrate vs. pasture and their interaction on milk yield, gross composition and fatty acid (FA profile of dairy cows bred in mountainous areas. For this purpose four dairy farms (two organic and two conventional were chosen in the alpine territory of Aosta Valley (NW Italy; individual milk yield was recorded daily and bulk milk samples were collected monthly from February to September 2007 to cover dietary variations. Higher levels of milk production (P<0.05 and lower milk protein amounts (P<0.01 were observed in the organic farms with respect to the conventional ones, while no significant differences were noticed in milk fat and lactose contents and in somatic cell count. Concerning fatty acids, only small differences were detected between organic and conventional milk and such differences seemed to be related mainly to the stabled period. Diet affected almost all variables studied: pasture feeding provided a significant improvement in the fatty acid composition in both organic and conventional systems leading to lower hypercholesterolemic saturated fatty acids, higher mono- and polyunsaturated fatty acids and conjugated linoleic acid amounts (P<0.001.

  11. Effect of DNA sequence of Fab fragment on yield characteristics and cell growth of E. coli.

    Science.gov (United States)

    Kulmala, Antti; Huovinen, Tuomas; Lamminmäki, Urpo

    2017-06-19

    Codon usage is one of the factors influencing recombinant protein expression. We were interested in the codon usage of an antibody Fab fragment gene exhibiting extreme toxicity in the E. coli host. The toxic synthetic human Fab gene contained domains optimized by the "one amino acid-one codon" method. We redesigned five segments of the Fab gene with a "codon harmonization" method described by Angov et al. and studied the effects of these changes on cell viability, Fab yield and display on filamentous phage using different vectors and bacterial strains. The harmonization considerably reduced toxicity, increased Fab expression from negligible levels to 10 mg/l, and restored the display on phage. Testing the impact of the individual redesigned segments revealed that the most significant effects were conferred by changes in the constant domain of the light chain. For some of the Fab gene variants, we also observed striking differences in protein yields when cloned from a chloramphenicol resistant vector into an identical vector, except with ampicillin resistance. In conclusion, our results show that the expression of a heterodimeric secretory protein can be improved by harmonizing selected DNA segments by synonymous codons and reveal additional complexity involved in heterologous protein expression.

  12. Tumour T1 changes in vivo are highly predictive of response to chemotherapy and reflect the number of viable tumour cells – a preclinical MR study in mice

    International Nuclear Information System (INIS)

    Weidensteiner, Claudia; Allegrini, Peter R; Sticker-Jantscheff, Melanie; Romanet, Vincent; Ferretti, Stephane; McSheehy, Paul MJ

    2014-01-01

    measure of cell density but reflects the decreased number of remaining viable and proliferating tumour cells due to perhaps cell and tissue destruction releasing proteins and/or metals that cause T 1 relaxation. ΔT 1 is a highly sensitive and specific predictor of response. This MRI method provides the opportunity to stratify a patient population during tumour therapy in the clinic

  13. Cell killing and radiosensitization by caffeic acid phenethyl ester (CAPE) in lung cancer cells

    International Nuclear Information System (INIS)

    Chen, Miao-Fen; Chen, Wen-Cheng; Wu, Chun-Te; King, P.C.

    2004-01-01

    Caffeic acid phenethyl ester (CAPE) is a biologically active ingredient of honeybee propoplis. The cytotoxicity and radiation sensitization effects of CAPE were evaluated in human lung cancer A549 cells and normal lung fibroblast WI-38 cells. A549 cells treated with 6 μg/ml CAPE showed marked growth inhibition (60%) at 48 hr after treatments. During the same time, the number of viable cells decreased to 46% of the control value. In contrast, WI-38 cells showed 20% growth inhibition with no change in the number of viable cells under the same treatment conditions. At 72 hr after CAPE treatment (6 μg/ml), the percentage of apoptotic cells in A549 cultures increased significantly to 67% and an S/G2 arrest was also detected in the culture. Furthermore, there was a significant decrease in the level of intracellular glutathione and hydrogen peroxide contents within one hr after CAPE treatment, and the expression of cyclin B 1 was reduced 6 hr after treatment. The radiation sensitization effect of CAPE on A549 cells was determined from the clonogenic survival curves, and the results showed a small but significant difference in radiation survival between cells treated with or without CAPE. Taken together, our results suggest that the effects of CAPE on differential cytotoxicity, apoptosis, and radiosensitization are associated with glutathione depletion that occurred shortly after treatments. (author)

  14. Effect of NPK and Poultry Manure on Growth, Yield, and Proximate Composition of Three Amaranths

    Directory of Open Access Journals (Sweden)

    Stephen Oyedeji

    2014-01-01

    Full Text Available The study compares the growth, yield, and proximate composition of Amaranthus hybridus, Amaranthus cruentus, and Amaranthus deflexus, grown with poultry manure and NPK in relation to the unfertilized soil of Ilorin, Nigeria. Viable seeds of the Amaranths raised in nursery for two weeks were transplanted (one plant per pot into unfertilized soil (control and soils fertilized with either NPK or poultry manure (PM at 30 Kg ha−1 rate arranged in randomized complete block design with four replicates. Data were collected on plant height, stem girth, number of leaves, leaf area, and number of branches from 1 week after transplanting (1 WAT. Fresh weight, dry weight, and proximate composition were determined at 6 WAT. Except for the length, breadth, and number of leaves, the order of growth parameters and yield in the three Amaranthus species was NPK > PM > control. NPK grown Amaranthus species had the highest protein while PM-grown vegetables had the highest ash content. Crude fibre in A. cruentus grown with PM was significantly higher than NPK and the control. The NPK treatment of A. hybridus and A. deflexus had the highest crude fibre content. NPK and PM favoured growth and yield of the Amaranthus species but influenced proximate composition differently.

  15. New approach for dry formulation techniques for rhizobacteria

    Science.gov (United States)

    Elchin, A. A.; Mashinistova, A. V.; Gorbunova, N. V.; Muratov, V. S.; Kydralieva, K. A.; Jorobekova, Sh. J.

    2009-04-01

    Two beneficial Pseudomonas isolates selected from rhizosphere of abundant weed - couch-grass Elytrigia repens L. Nevski have been found to have biocontrol activity. An adequate biocontrol effect requires high yield and long stability of the bacterial preparation [1], which could be achieved by an effective and stable formulation. This study was aimed to test various approaches to dry formulation techniques for Pseudomonas- based preparations. To reach this goal, two drying formulation techniques have been tested: the first one, spray drying and the second, low-temperature contact-convective drying in fluidized bed. The optimal temperature parameters for each technique were estimated. Main merits of the selected approach to dry technique are high yield, moderate specific energy expenditures per 1 kg of evaporated moisture, minimal time of contact of the drying product with drying agent. The technological process for dry formulation included the following stages: the obtaining of cell liquids, the low-temperature concentrating and the subsequent drying of a concentrate. The preliminary technological stages consist in cultivation of the rhizobacteria cultures and concentrating the cell liquids. The following requirements for cultivation regime in laboratory conditions were proposed: optimal temperatures are 26-28°С in 3 days, concentration of viable cells in cell liquid makes 1010-1011 cell/g of absolutely dry substance (ADS). For concentrating the cell liquids the method of a vacuum evaporation, which preserves both rhizobacteria cells and the secondary metabolites of cell liquid, has been used. The process of concentrating was conducted at the minimum possible temperature, i.e. not above 30-33°С. In this case the concentration of viable cells has decreased up to 109-1010 cell/g of ADS. For spray drying the laboratory up-dated drier BUCHI 190, intended for the drying of thermolabile products, was used. The temperatures of an in- and outcoming air did not exceed

  16. Inosine Released from Dying or Dead Cells Stimulates Cell Proliferation via Adenosine Receptors

    Directory of Open Access Journals (Sweden)

    Yi Zhao

    2017-04-01

    Full Text Available IntroductionMany antitumor therapies induce apoptotic cell death in order to cause tumor regression. Paradoxically, apoptotic cells are also known to promote wound healing, cell proliferation, and tumor cell repopulation in multicellular organisms. We aimed to characterize the nature of the regenerative signals concentrated in the micromilieu of dead and dying cells.MethodsCultures of viable melanoma B16F10 cells, mouse fibroblasts, and primary human fibroblast-like synoviocytes (FLS in the presence of dead and dying cells, their supernatants (SNs, or purified agonists and antagonists were used to evaluate the stimulation of proliferation. Viable cell quantification was performed by either flow cytometry of harvested cells or by crystal violet staining of adherent cells. High-performance liquid chromatography and liquid chromatography coupled with mass spectrometry of cell SNs were deployed to identify the nature of growth-promoting factors. Coimplantation of living cells in the presence of SNs collected from dead and dying cells and specific agonists was used to evaluate tumor growth in vivo.ResultsThe stimulation of proliferation of few surviving cells by bystander dead cells was confirmed for melanoma cells, mouse fibroblasts, and primary FLS. We found that small soluble molecules present in the protein-free fraction of SNs of dead and dying cells were responsible for the promotion of proliferation. The nucleoside inosine released by dead and dying cells acting via adenosine receptors was identified as putative inducer of proliferation of surviving tumor cells after irradiation and heat treatment.ConclusionInosine released by dead and dying cells mediates tumor cell proliferation via purinergic receptors. Therapeutic strategies surmounting this pathway may help to reduce the rate of recurrence after radio- and chemotherapy.

  17. Viability in holder of irradiated cells: distinguish between repair and cell multiplication

    International Nuclear Information System (INIS)

    Araujo, A.C. de.

    1980-01-01

    In experiments in which liquid holding recovery (LHR) was measured, the majority of cellular population is formed by non-viable cells and cell multiplication may be important for LHR expression. In order to distinguish between recuperation of viability (true LHR) and cell multiplication, it was necessary to employ improved plating techniques and a fluctuation test based on Poisson distribution. Our results are an indication that this fluctuation test, used together with the traditional method, is a good tool to distinguish repair from cell multiplication. (author)

  18. Toward interplay between substructure evolution, dislocation configuration, and yield strength in a microalloyed steel

    International Nuclear Information System (INIS)

    Venkatsurya, P.K.C.; Misra, R.D.K.; Mulholland, M.D.; Manohar, M.; Hartmann, J.E.

    2014-01-01

    We focus our attention here on the directional dependence of yield strength in high strength microalloyed steel using transmission electron microscopy and x-ray diffraction. The primary objective is to study the interplay between substructural evolution, notably cell size, dense dislocation walls (DDWs), dislocation tangle zones (DTZs), lamellar boundaries, crystallographic texture, and yield strength. The study elucidates for the first time the strong impact of thermo-mechanical deformation-induced dislocation and lamellar structures, which are likely to modify the slip pattern, leading to directional dependence of yield strength. Majority of the dislocations tend to pile along the {110} slip planes as dense dislocation walls. At low strains, grains are first divided into cell blocks that are nearly dislocation-free. At higher strains and with progress in thermo-mechanical processing dislocation tangled zones and lamellar boundaries develop. It is hypothesized that the differences in dislocation configurations, dislocations cells and cell blocks, and lamellar boundaries synergistically contribute to directional dependence of the yield strength in the high strength ferrous alloy. The presumption is envisaged on the basis of observations that the microstructural constituents were similar in the entire plane of the hot rolled strip and the crystallographic texture was weak

  19. Effectiveness of rabbit manure biofertilizer in barley crop yield.

    Science.gov (United States)

    Islas-Valdez, Samira; Lucho-Constantino, Carlos A; Beltrán-Hernández, Rosa I; Gómez-Mercado, René; Vázquez-Rodríguez, Gabriela A; Herrera, Juan M; Jiménez-González, Angélica

    2017-11-01

    The quality of biofertilizers is usually assessed only in terms of the amount of nutrients that they supply to the crops and their lack of viable pathogens and phytotoxicity. The goal of this study was to determine the effectiveness of a liquid biofertilizer obtained from rabbit manure in terms of presence of pathogens, phytotoxicity, and its effect on the grain yield and other agronomic traits of barley (Hordeum vulgare L.). Environmental effects of the biofertilizer were also evaluated by following its influence on selected soil parameters. We applied the biofertilizer at five combinations of doses and timings each and in two application modes (foliar or direct soil application) within a randomized complete block design with three replicates and using a chemical fertilizer as control. The agronomic traits evaluated were plant height, root length, dry weight, and number of leaves and stems at three growth stages: tillering, jointing, and flowering. The effectiveness of the biofertilizer was significantly modified by the mode of application, the growth stage of the crop, and the dose of biofertilizer applied. The results showed that the foliar application of the biofertilizer at the tillering stage produced the highest increase in grain yield (59.7 %, p biofertilizer caused significant changes in soil, particularly concerning pH, EC, Ca, Zn, Mg, and Mn. It is our view that the production and use of biofertilizers are a reliable alternative to deal with a solid waste problem while food security is increased.

  20. Oxidative Damage and Mutagenic Potency of Fast Neutron and UV-B Radiation in Pollen Mother Cells and Seed Yield of Vicia faba L.

    Directory of Open Access Journals (Sweden)

    Ekram Abdel Haliem

    2013-01-01

    Full Text Available In recent years, there has been a great deal of attention toward free radicals, reactive oxygen species (ROS generated by exposure of crop plant cells to physical radiations. Henceforth, the current study was planned to compare oxidative stress and mutagenic potential of different irradiation doses of fast neutron (FN and UV-B on meiotic-pollen mother cells (PMCs, pollen grains (PGs and seeds yielded from irradiated faba beans seedlings. On the cytogenetic level, each irradiation type had special interference with DNA of PMC and exhibited wide range of mutagenic action on the frequency and type of chromosomal anomalies, fertility of PGs and seed yield productivity based on the irradiation exposure dose and radiation sensitivity of faba bean plants compared with un-irradiated ones. On the molecular level, SDS-PAGE and RPAD-PCR analyses of seeds yielded from irradiated seedlings exhibited distinctive polymorphisms based on size, intensity, appearance, and disappearance of polypeptides bands compared with un-irradiated ones. The total values of protein and DNA polymorphisms reached 88% and 90.80% respectively. The neutron fluency (2.3 × 106 n/cm2 and UV-B dose for 1 hr were recorded as bio-positive effects. The present study proved that genetic variations revealed by cytogenetic test could be supported by gene expression (alterations in RAPD and protein profiles.

  1. Silicon Solar Cell Turns 50

    Energy Technology Data Exchange (ETDEWEB)

    Perlin, J.

    2004-08-01

    This short brochure describes a milestone in solar (or photovoltaic, PV) research-namely, the 50th anniversary of the invention of the first viable silicon solar cell by three researchers at Bell Laboratories.

  2. Advanced Analysis to Distinguish between Physical Decrease and Inactivation of Viable Phages in Aerosol by Quantitating Phage-Specific Particles.

    Science.gov (United States)

    Shimasaki, Noriko; Nojima, Yasuhiro; Sakakibara, Masaya; Kikuno, Ritsuko; Iizuka, Chiori; Okaue, Akira; Okuda, Shunji; Shinohara, Katsuaki

    2018-01-01

     Recent studies have investigated the efficacy of air-cleaning products against pathogens in the air. A standard method to evaluate the reduction in airborne viruses caused by an air cleaner has been established using a safe bacteriophage instead of pathogenic viruses; the reduction in airborne viruses is determined by counting the number of viable airborne phages by culture, after operating the air cleaner. The reduction in the number of viable airborne phages could be because of "physical decrease" or "inactivation". Therefore, to understand the mechanism of reduction correctly, an analysis is required to distinguish between physical decrease and inactivation. The purpose of this study was to design an analysis to distinguish between the physical decrease and inactivation of viable phi-X174 phages in aerosols. We established a suitable polymerase chain reaction (PCR) system by selecting an appropriate primer-probe set for PCR and validating the sensitivity, linearity, and specificity of the primer-probe set to robustly quantify phi-X174-specific airborne particles. Using this quantitative PCR system and culture assay, we performed a behavior analysis of the phage aerosol in a small chamber (1 m 3 ) at different levels of humidity, as humidity is known to affect the number of viable airborne phages. The results revealed that the reduction in the number of viable airborne phages was caused not only by physical decrease but also by inactivation under particular levels of humidity. Our study could provide an advanced analysis to differentiate between the physical decrease and inactivation of viable airborne phages.

  3. Amniotic Mesenchymal Stromal Cells Exhibit Preferential Osteogenic and Chondrogenic Differentiation and Enhanced Matrix Production Compared With Adipose Mesenchymal Stromal Cells.

    Science.gov (United States)

    Topoluk, Natasha; Hawkins, Richard; Tokish, John; Mercuri, Jeremy

    2017-09-01

    Therapeutic efficacy of various mesenchymal stromal cell (MSC) types for orthopaedic applications is currently being investigated. While the concept of MSC therapy is well grounded in the basic science of healing and regeneration, little is known about individual MSC populations in terms of their propensity to promote the repair and/or regeneration of specific musculoskeletal tissues. Two promising MSC sources, adipose and amnion, have each demonstrated differentiation and extracellular matrix (ECM) production in the setting of musculoskeletal tissue regeneration. However, no study to date has directly compared the differentiation potential of these 2 MSC populations. To compare the ability of human adipose- and amnion-derived MSCs to undergo osteogenic and chondrogenic differentiation. Controlled laboratory study. MSC populations from the human term amnion were quantified and characterized via cell counting, histologic assessment, and flow cytometry. Differentiation of these cells in comparison to commercially purchased human adipose-derived mesenchymal stromal cells (hADSCs) in the presence and absence of differentiation media was evaluated via reverse transcription polymerase chain reaction (PCR) for bone and cartilage gene transcript markers and histology/immunohistochemistry to examine ECM production. Analysis of variance and paired t tests were performed to compare results across all cell groups investigated. The authors confirmed that the human term amnion contains 2 primary cell types demonstrating MSC characteristics-(1) human amniotic epithelial cells (hAECs) and (2) human amniotic mesenchymal stromal cells (hAMSCs)-and each exhibited more than 90% staining for MSC surface markers (CD90, CD105, CD73). Average viable hAEC and hAMSC yields at harvest were 2.3 × 10 6 ± 3.7 × 10 5 and 1.6 × 10 6 ± 4.7 × 10 5 per milliliter of amnion, respectively. As well, hAECs and hAMSCs demonstrated significantly greater osteocalcin ( P = .025), aggrecan ( P

  4. Metabolic and Kinetic analyses of influenza production in perfusion HEK293 cell culture

    Directory of Open Access Journals (Sweden)

    Lohr Verena

    2011-09-01

    Full Text Available Abstract Background Cell culture-based production of influenza vaccine remains an attractive alternative to egg-based production. Short response time and high production yields are the key success factors for the broader adoption of cell culture technology for industrial manufacturing of pandemic and seasonal influenza vaccines. Recently, HEK293SF cells have been successfully used to produce influenza viruses, achieving hemagglutinin (HA and infectious viral particle (IVP titers in the highest ranges reported to date. In the same study, it was suggested that beyond 4 × 106 cells/mL, viral production was limited by a lack of nutrients or an accumulation of toxic products. Results To further improve viral titers at high cell densities, perfusion culture mode was evaluated. Productivities of both perfusion and batch culture modes were compared at an infection cell density of 6 × 106 cells/mL. The metabolism, including glycolysis, glutaminolysis and amino acids utilization as well as physiological indicators such as viability and apoptosis were extensively documented for the two modes of culture before and after viral infection to identify potential metabolic limitations. A 3 L bioreactor with a perfusion rate of 0.5 vol/day allowed us to reach maximal titers of 3.3 × 1011 IVP/mL and 4.0 logHA units/mL, corresponding to a total production of 1.0 × 1015 IVP and 7.8 logHA units after 3 days post-infection. Overall, perfusion mode titers were higher by almost one order of magnitude over the batch culture mode of production. This improvement was associated with an activation of the cell metabolism as seen by a 1.5-fold and 4-fold higher consumption rates of glucose and glutamine respectively. A shift in the viral production kinetics was also observed leading to an accumulation of more viable cells with a higher specific production and causing an increase in the total volumetric production of infectious influenza particles. Conclusions These results

  5. Biogas generation in landfills. Equilibria, rates and yields

    Energy Technology Data Exchange (ETDEWEB)

    Aakesson, M

    1997-05-01

    Landfilling in `cells` has become more common in recent years. Different waste streams are guided to different cells, among which the biocell is a landfill designed for biogas production. In this thesis, the dependence of biogas generation on waste composition was investigated. Six 8,000 m{sup 3} test cells, with contents ranging from mainly commercial waste to pure domestic waste and equipped with gas extraction systems and bottom plastic liners, were monitored for seven years. Great emphasis was given to the characterization of conversion processes and governing mechanism in the topics of bio-energetics, kinetics and capacities. A thermodynamic model, in which the oxidations of volatile fatty acids (VFA) (2yields (in this study found to be 150-200 and 40-70 Nm{sup 3}/dry tonne, respectively), the possible nutritional limitation was investigated. Pools and emissions of chemical oxygen demand, N, P and K were quantified. Biomass pools were estimated from methane yields, growth yield coefficients, and bacterial mineral contents. However, results from commercial waste test cells showed that the assimilation of P exceeded the refuse content, which suggests the turnover of microbial biomass and questions the notion of nutritional limitation. In sum, the results showed that the advantages of a reduced content of readily biodegradable material, achieved by guidance or pretreatment, encompass several aspects of the performance. 84 refs, 6 figs, 1 tab

  6. Electricity Generation in Microbial Fuel Cell (MFC) by Bacterium Isolated from Rice Paddy Field Soil

    Science.gov (United States)

    Fakhirruddin, Fakhriah; Amid, Azura; Salim, Wan Wardatul Amani Wan; Suhaida Azmi, Azlin

    2018-03-01

    Microbial fuel cell (MFC) is an alternative approach in generating renewable energy by utilising bacteria that will oxidize organic or inorganic substrates, producing electrons yielded as electrical energy. Different species of exoelectrogenic bacteria capable of generating significant amount of electricity in MFC has been identified, using various organic compounds for fuel. Soil sample taken from rice paddy field is proven to contain exoelectrogenic bacteria, thus electricity generation using mixed culture originally found in the soil, and pure culture isolated from the soil is studied. This research will isolate the exoelectrogenic bacterial species in the rice paddy field soil responsible for energy generation. Growth of bacteria isolated from the MFC is observed by measuring the optical density (OD), cell density weight (CDW) and viable cell count. Mixed bacterial species found in paddy field soil generates maximum power of 77.62 μW and 0.70 mA of current. In addition, the research also shows that the pure bacterium in rice paddy field soil can produce maximum power and current at 51.32 μW and 0.28 mA respectively.

  7. Vertical farming increases lettuce yield per unit area compared to conventional horizontal hydroponics.

    Science.gov (United States)

    Touliatos, Dionysios; Dodd, Ian C; McAinsh, Martin

    2016-08-01

    Vertical farming systems (VFS) have been proposed as an engineering solution to increase productivity per unit area of cultivated land by extending crop production into the vertical dimension. To test whether this approach presents a viable alternative to horizontal crop production systems, a VFS (where plants were grown in upright cylindrical columns) was compared against a conventional horizontal hydroponic system (HHS) using lettuce ( Lactuca sativa L . cv. "Little Gem") as a model crop. Both systems had similar root zone volume and planting density. Half-strength Hoagland's solution was applied to plants grown in perlite in an indoor controlled environment room, with metal halide lamps providing artificial lighting. Light distribution (photosynthetic photon flux density, PPFD) and yield (shoot fresh weight) within each system were assessed. Although PPFD and shoot fresh weight decreased significantly in the VFS from top to base, the VFS produced more crop per unit of growing floor area when compared with the HHS. Our results clearly demonstrate that VFS presents an attractive alternative to horizontal hydroponic growth systems and suggest that further increases in yield could be achieved by incorporating artificial lighting in the VFS.

  8. Spatial Rice Yield Estimation Based on MODIS and Sentinel-1 SAR Data and ORYZA Crop Growth Model

    Directory of Open Access Journals (Sweden)

    Tri D. Setiyono

    2018-02-01

    Full Text Available Crop insurance is a viable solution to reduce the vulnerability of smallholder farmers to risks from pest and disease outbreaks, extreme weather events, and market shocks that threaten their household food and income security. In developing and emerging countries, the implementation of area yield-based insurance, the form of crop insurance preferred by clients and industry, is constrained by the limited availability of detailed historical yield records. Remote-sensing technology can help to fill this gap by providing an unbiased and replicable source of the needed data. This study is dedicated to demonstrating and validating the methodology of remote sensing and crop growth model-based rice yield estimation with the intention of historical yield data generation for application in crop insurance. The developed system combines MODIS and SAR-based remote-sensing data to generate spatially explicit inputs for rice using a crop growth model. MODIS reflectance data were used to generate multitemporal LAI maps using the inverted Radiative Transfer Model (RTM. SAR data were used to generate rice area maps using MAPScape-RICE to mask LAI map products for further processing, including smoothing with logistic function and running yield simulation using the ORYZA crop growth model facilitated by the Rice Yield Estimation System (Rice-YES. Results from this study indicate that the approach of assimilating MODIS and SAR data into a crop growth model can generate well-adjusted yield estimates that adequately describe spatial yield distribution in the study area while reliably replicating official yield data with root mean square error, RMSE, of 0.30 and 0.46 t ha−1 (normalized root mean square error, NRMSE of 5% and 8% for the 2016 spring and summer seasons, respectively, in the Red River Delta of Vietnam, as evaluated at district level aggregation. The information from remote-sensing technology was also useful for identifying geographic locations with

  9. Good manufacturing practice-compliant expansion of marrow-derived stem and progenitor cells for cell therapy.

    Science.gov (United States)

    Gastens, Martin H; Goltry, Kristin; Prohaska, Wolfgang; Tschöpe, Diethelm; Stratmann, Bernd; Lammers, Dirk; Kirana, Stanley; Götting, Christian; Kleesiek, Knut

    2007-01-01

    Ex vivo expansion is being used to increase the number of stem and progenitor cells for autologous cell therapy. Initiation of pivotal clinical trials testing the efficacy of these cells for tissue repair has been hampered by the challenge of assuring safe and high-quality cell production. A strategy is described here for clinical-scale expansion of bone marrow (BM)-derived stem cells within a mixed cell population in a completely closed process from cell collection through postculture processing using sterile connectable devices. Human BM mononuclear cells (BMMNC) were isolated, cultured for 12 days, and washed postharvest using either standard open procedures in laminar flow hoods or using automated closed systems. Conditions for these studies were similar to long-term BM cultures in which hematopoietic and stromal components are cultured together. Expansion of marrow-derived stem and progenitor cells was then assessed. Cell yield, number of colony forming units (CFU), phenotype, stability, and multilineage differentiation capacity were compared from the single pass perfusion bioreactor and standard flask cultures. Purification of BMMNC using a closed Ficoll gradient process led to depletion of 98% erythrocytes and 87% granulocytes, compared to 100% and 70%, respectively, for manual processing. After closed system culture, mesenchymal progenitors, measured as CD105+CD166+CD14-CD45- and fibroblastic CFU, expanded 317- and 364-fold, respectively, while CD34+ hematopoietic progenitors were depleted 10-fold compared to starting BMMNC. Cultured cells exhibited multilineage differentiation by displaying adipogenic, osteogenic, and endothelial characteristics in vitro. No significant difference was observed between manual and bioreactor cultures. Automated culture and washing of the cell product resulted in 181 x 10(6) total cells that were viable and contained fibroblastic CFU for at least 24 h of storage. A combination of closed, automated technologies enabled

  10. Improved identification of viable myocardium using second harmonic imaging during dobutamine stress echocardiography

    NARCIS (Netherlands)

    F. Sozzi (Fabiola); D. Poldermans (Don); J.J. Bax (Jeroen); A. Elhendy (Abdou); E.C. Vourvouri (Eleni); R. Valkema (Roelf); J. de Sutter; A.F.L. Schinkel (Arend); A. Borghetti; J.R.T.C. Roelandt (Jos)

    2001-01-01

    textabstractOBJECTIVE: To determine whether, compared with fundamental imaging, second harmonic imaging can improve the accuracy of dobutamine stress echocardiography for identifying viable myocardium, using nuclear imaging as a reference. PATIENTS: 30 patients with chronic left

  11. Changes in total viable count and TVB-N content in marinated chicken breast fillets during storage

    Science.gov (United States)

    Baltić, T.; Ćirić, J.; Velebit, B.; Petronijević, R.; Lakićević, B.; Đorđević, V.; Janković, V.

    2017-09-01

    Marination is a popular technique for enhancing meat properties. Depending on the marinade type and ingredients added, marination can improve sensory, chemical and microbiological quality of meat products. In this study, the total viable count and total volatile basic nitrogen (TVB-N) content in marinated chicken breast fillets were investigated. The possible correlation between bacterial growth and formation of TVB-N was also tested. Chicken breast fillets were immersed in a solution of table salt (as a control) orthree different marinades,which consisted of table salt, sodium tripolyphosphate and/or sodium citrate, and stored in air for nine days at 4±1°C. Analyses of the total viable count and TVB-N were performed on days0, 3, 6 and 9 day of storage. The total viable count gradually increased in all examined groups, and statistically significant differences (pchicken were significantly higher (pchicken marinated with sodium citrate was established (pchicken marinated with sodium tripolyphosphate.

  12. Drug and radiation sensitivity measurements of successful primary monolayer culturing of human tumor cells using cell-adhesive matrix and supplemented medium

    International Nuclear Information System (INIS)

    Baker, F.L.; Spitzer, G.; Ajani, J.A.

    1986-01-01

    The limitations of the agar suspension culture method for primary culturing of human tumor cells prompted development of a monolayer system optimized for cell adhesion and growth. This method grew 83% of fresh human tumor cell biopsy specimens, cultured and not contaminated, from a heterogeneous group of 396 tumors including lung cancer (93 of 114, 82%); melanoma (54 of 72, 75%); sarcoma (46 of 59, 78%); breast cancer (35 of 39, 90%); ovarian cancer (16 of 21, 76%); and a miscellaneous group consisting of gastrointestinal, genitourinary, mesothelioma, and unknown primaries (78 of 91, 86%). Cell growth was characterized morphologically with Papanicolaoustained coverslip cultures and cytogenetically with Giemsastained metaphase spreads. Morphological features such as nuclear pleomorphism, chromatin condensation, basophilic cytoplasm, and melanin pigmentation were routinely seen. Aneuploid metaphases were seen in 90% of evaluable cultures, with 15 of 28 showing 70% or more aneuploid metaphases. Colony-forming efficiency ranged between 0.01 and 1% of viable tumor cells, with a median efficiency of 0.2%. This culture system uses a low inoculum of 25,000 viable cells per well which permitted chemosensitivity testing of nine drugs at four doses in duplicate from 2.2 X 10(6) viable tumor cells and radiation sensitivity testing at five doses in quadruplicate from 0.6 X 10(6) cells. Cultures were analyzed for survival by computerized image analysis of crystal violet-stained cells. Drug sensitivity studies showed variability in sensitivity and in survival curve shape with exponential cell killing for cisplatin, Adriamycin, and etoposide, and shouldered survival curves for 5-fluorouracil frequently seen. Radiation sensitivity studies also showed variability in both sensitivity and survival curve shape. Many cultures showed exponential cell killing, although others had shouldered survival curves

  13. Rapid quantification of viable Legionella in nuclear cooling tower waters using filter cultivation, fluorescent in situ hybridization and solid-phase cytometry.

    Science.gov (United States)

    Baudart, J; Guillaume, C; Mercier, A; Lebaron, P; Binet, M

    2015-05-01

    To develop a rapid and sensitive method to quantify viable Legionella spp. in cooling tower water samples. A rapid, culture-based method capable of quantifying as few as 600 Legionella microcolonies per litre within 2 days in industrial waters was developed. The method combines a short cultivation step of microcolonies on GVPC agar plate, specific detection of Legionella cells by a fluorescent in situ hybridization (FISH) approach, and a sensitive enumeration using a solid-phase cytometer. Following optimization of the cultivation conditions, the qualitative and quantitative performance of the method was assessed and the method was applied to 262 nuclear power plant cooling water samples. The performance of this method was in accordance with the culture method (NF-T 90-431) for Legionella enumeration. The rapid detection of viable Legionella in water is a major concern to the effective monitoring of this pathogenic bacterium in the main water sources involved in the transmission of legionellosis infection (Legionnaires' disease). The new method proposed here appears to be a robust, efficient and innovative means for rapidly quantifying cultivable Legionella in cooling tower water samples within 48 h. © 2015 The Society for Applied Microbiology.

  14. Optimizing rice yields while minimizing yield-scaled global warming potential.

    Science.gov (United States)

    Pittelkow, Cameron M; Adviento-Borbe, Maria A; van Kessel, Chris; Hill, James E; Linquist, Bruce A

    2014-05-01

    To meet growing global food demand with limited land and reduced environmental impact, agricultural greenhouse gas (GHG) emissions are increasingly evaluated with respect to crop productivity, i.e., on a yield-scaled as opposed to area basis. Here, we compiled available field data on CH4 and N2 O emissions from rice production systems to test the hypothesis that in response to fertilizer nitrogen (N) addition, yield-scaled global warming potential (GWP) will be minimized at N rates that maximize yields. Within each study, yield N surplus was calculated to estimate deficit or excess N application rates with respect to the optimal N rate (defined as the N rate at which maximum yield was achieved). Relationships between yield N surplus and GHG emissions were assessed using linear and nonlinear mixed-effects models. Results indicate that yields increased in response to increasing N surplus when moving from deficit to optimal N rates. At N rates contributing to a yield N surplus, N2 O and yield-scaled N2 O emissions increased exponentially. In contrast, CH4 emissions were not impacted by N inputs. Accordingly, yield-scaled CH4 emissions decreased with N addition. Overall, yield-scaled GWP was minimized at optimal N rates, decreasing by 21% compared to treatments without N addition. These results are unique compared to aerobic cropping systems in which N2 O emissions are the primary contributor to GWP, meaning yield-scaled GWP may not necessarily decrease for aerobic crops when yields are optimized by N fertilizer addition. Balancing gains in agricultural productivity with climate change concerns, this work supports the concept that high rice yields can be achieved with minimal yield-scaled GWP through optimal N application rates. Moreover, additional improvements in N use efficiency may further reduce yield-scaled GWP, thereby strengthening the economic and environmental sustainability of rice systems. © 2013 John Wiley & Sons Ltd.

  15. The potential of satellite-observed crop phenology to enhance yield gap assessments in smallholder landscapes

    Directory of Open Access Journals (Sweden)

    John M A Duncan

    2015-08-01

    Full Text Available Many of the undernourished people on the planet obtain their entitlements to food via agricultural-based livelihood strategies, often on underperforming croplands and smallholdings. In this context, expanding cropland extent is not a viable strategy for smallholders to meet their food needs. Therefore, attention must shift to increasing productivity on existing plots and ensuring yield gaps do not widen. Thus, supporting smallholder farmers to sustainably increase the productivity of their lands is one part of a complex solution to realising universal food security. However, the information (e.g. location and causes of cropland underperformance required to support measures to close yield gaps in smallholder landscapes are often not available. This paper reviews the potential of crop phenology, observed from satellites carrying remote sensing sensors, to fill this information gap. It is suggested that on a theoretical level phenological approaches can reveal greater intra-cropland thematic detail, and increase the accuracy of crop extent maps and crop yield estimates. However, on a practical level the spatial mismatch between the resolution at which crop phenology can be estimated from satellite remote sensing data and the scale of yield variability in smallholder croplands inhibits its use in this context. Similarly, the spatial coverage of remote sensing-derived phenology offers potential for integration with ancillary spatial datasets to identify causes of yield gaps. To reflect the complexity of smallholder cropping systems requires ancillary datasets at fine spatial resolutions which, often, are not available. This further precludes the use of crop phenology in attempts to unpick the causes of yield gaps. Research agendas should focus on generating fine spatial resolution crop phenology, either via data fusion or through new sensors (e.g. Sentinel-2 in smallholder croplands. This has potential to transform the applied use of remote sensing

  16. Natural environmental water sources in endemic regions of northeastern Brazil are potential reservoirs of viable Mycobacterium leprae.

    Science.gov (United States)

    Arraes, Maria Luisa Bezerra de Macedo; Holanda, Maísa Viana de; Lima, Luana Nepomuceno Gondim Costa; Sabadia, José Antônio Beltrão; Duarte, Cynthia Romariz; Almeida, Rosa Livia Freitas; Kendall, Carl; Kerr, Ligia Regina Sansigolo; Frota, Cristiane Cunha

    2017-12-01

    The detection of live Mycobacterium leprae in soil and animals other than humans suggests that the environment plays a role in the transmission of leprosy. The objective of this study was to investigate the presence of viable M. leprae in natural water sources used by the local population in five municipalities in the state of Ceará, northeastern Brazil. Samples were collected from 30 different sources. Viable bacilli were identified by reverse transcriptase polymerase chain reaction (PCR) of the M. leprae gyrA gene and sequencing of the PCR products. Physicochemical properties of each water source were also assessed. M. leprae gyrA mRNA was found in 23 (76.7%) of the water sources. No association was found between depth of the water and sample positivity, nor was there any association between the type of water used by the population and sample positivity. An association between viable M. leprae and temperature and pH was found. Georeferencing showed a relation between the residences of leprosy cases and water source containing the bacterium. The finding of viable M. leprae in natural water sources associated with human contact suggests that the environment plays an important role in maintaining endemic leprosy in the study region.

  17. Metabolic enzyme cost explains variable trade-offs between microbial growth rate and yield.

    Directory of Open Access Journals (Sweden)

    Meike T Wortel

    2018-02-01

    Full Text Available Microbes may maximize the number of daughter cells per time or per amount of nutrients consumed. These two strategies correspond, respectively, to the use of enzyme-efficient or substrate-efficient metabolic pathways. In reality, fast growth is often associated with wasteful, yield-inefficient metabolism, and a general thermodynamic trade-off between growth rate and biomass yield has been proposed to explain this. We studied growth rate/yield trade-offs by using a novel modeling framework, Enzyme-Flux Cost Minimization (EFCM and by assuming that the growth rate depends directly on the enzyme investment per rate of biomass production. In a comprehensive mathematical model of core metabolism in E. coli, we screened all elementary flux modes leading to cell synthesis, characterized them by the growth rates and yields they provide, and studied the shape of the resulting rate/yield Pareto front. By varying the model parameters, we found that the rate/yield trade-off is not universal, but depends on metabolic kinetics and environmental conditions. A prominent trade-off emerges under oxygen-limited growth, where yield-inefficient pathways support a 2-to-3 times higher growth rate than yield-efficient pathways. EFCM can be widely used to predict optimal metabolic states and growth rates under varying nutrient levels, perturbations of enzyme parameters, and single or multiple gene knockouts.

  18. Delayed uptake and washout of contrast in non-viable infarcted myocardium shown with dynamic computed tomography

    DEFF Research Database (Denmark)

    Udholm, Sebastian; Laugesen, Sofie; Agger, Peter

    2014-01-01

    BACKGROUND: Assessment of ischemic but potentially viable myocardium plays an important role in the planning of coronary revascularization. Until now SPECT, PET, and MRI have been used to identify viable myocardium. Computed tomography (CT) is increasingly used to diagnose coronary atherosclerosis...... descending artery, followed by removal of the balloon and reperfusion. Four pigs died due to refractory ventricular fibrillation. After 6 weeks, dynamic cardiac CT was performed assessing both wall motion and contrast attenuation. Measurements of attenuation values in Hounsfield units (HU) in the infarct...

  19. Overcoming the response plateau in multiple myeloma: a novel bortezomib-based strategy for secondary induction and high-yield CD34+ stem cell mobilization.

    Science.gov (United States)

    Niesvizky, Ruben; Mark, Tomer M; Ward, Maureen; Jayabalan, David S; Pearse, Roger N; Manco, Megan; Stern, Jessica; Christos, Paul J; Mathews, Lena; Shore, Tsiporah B; Zafar, Faiza; Pekle, Karen; Xiang, Zhaoying; Ely, Scott; Skerret, Donna; Chen-Kiang, Selina; Coleman, Morton; Lane, Maureen E

    2013-03-15

    This phase II study evaluated bortezomib-based secondary induction and stem cell mobilization in 38 transplant-eligible patients with myeloma who had an incomplete and stalled response to, or had relapsed after, previous immunomodulatory drug-based induction. Patients received up to six 21-day cycles of bortezomib plus dexamethasone, with added liposomal doxorubicin for patients not achieving partial response or better by cycle 2 or very good partial response or better (≥VGPR) by cycle 4 (DoVeD), followed by bortezomib, high-dose cyclophosphamide, and filgrastim mobilization. Gene expression/signaling pathway analyses were conducted in purified CD34+ cells after bortezomib-based mobilization and compared against patients who received only filgrastim ± cyclophosphamide. Plasma samples were similarly analyzed for quantification of associated protein markers. The response rate to DoVeD relative to the pre-DoVeD baseline was 61%, including 39% ≥ VGPR. Deeper responses were achieved in 10 of 27 patients who received bortezomib-based mobilization; postmobilization response rate was 96%, including 48% ≥ VGPR, relative to the pre-DoVeD baseline. Median CD34+ cell yield was 23.2 × 10(6) cells/kg (median of 1 apheresis session). After a median follow-up of 46.6 months, median progression-free survival was 47.1 months from DoVeD initiation; 5-year overall survival rate was 76.4%. Grade ≥ 3 adverse events included thrombocytopenia (13%), hand-foot syndrome (11%), peripheral neuropathy (8%), and neutropenia (5%). Bortezomib-based mobilization was associated with modulated expression of genes involved in stem cell migration. Bortezomib-based secondary induction and mobilization could represent an alternative strategy for elimination of tumor burden in immunomodulatory drug-resistant patients that does not impact stem cell yield.

  20. An Effective Approach for Immunotherapy Using Irradiated Tumor Cells

    International Nuclear Information System (INIS)

    Mostafa, D.M.B.

    2011-01-01

    This study has been aimed to investigate the effect of injection of Irradiated Ehrlich tumor cells alone or concurrent with immunomodulator in mice before and after challenge with viable Ehrlich tumor cells for enhancement of immune system. This study includes the estimation of survival, tumor size, lymphocyte count, LDH, MTT, granzyme B, and DNA fragmentation. In order to fulfill the target of this study, a total of 120 female swiss albino mice were used. They were divided into two classes vaccinated (injection of vaccine before challenge) and therapeutic class (injection of vaccine after challenge). Each class was divided into four groups, group (1) mice injected with viable Ehrlich tumor cells (G1), group (2) mice injected with irradiated tumor cells (G2), group (3) mice injected with immunomodulator (G3), and group (4) mice injected with irradiated tumor cells + immunomodulator (G4). Results obtained from this study demonstrated that, the lymphocyte count and granzyme B activity were increased in both the vaccinated and therapeutic classes compared with control group. LDH activity was decreased in all groups of vaccinated class and also in G2 and G4 groups of therapeutic class compared with control group. There was a significant increase in percent apoptosis of tumor cells cultured with spleenocytes of the groups of vaccinated class as compared with control group. Cellular DNA from Ehrlich tumor cell line cultured with spleenocytes of immunized groups was fragmented into discrete bands of approximate multiples of 200 bp. Revealing significant apoptosis in tumor cells due to vaccination. It is concluded that, vaccination with irradiated tumor cells is an effective approach in stimulation of immune system against viable tumor cells.

  1. Monitoring of yeast cell concentration using a micromachined impedance sensor

    NARCIS (Netherlands)

    Krommenhoek, E.E.; Gardeniers, Johannes G.E.; Bomer, Johan G.; van den Berg, Albert; Li, X.; Ottens, M.; van der Wielen, L.A.M.; van Dedem, G.W.K.; van Leeuwen, M.; van Gulik, W.M.; Heijnen, J.J.

    2005-01-01

    The paper describes the design, modelling and experimental characterization of a micromachined impedance sensor for on-line monitoring of the viable yeast cell concentration (biomass) in a miniaturized cell assay. Measurements in a Saccharomyces cerevisiae cell culture show that the permittivity of

  2. Senior health clinics: are they financially viable?

    Science.gov (United States)

    McAtee, Robin E; Crandall, Debra; Wright, Larry D; Beverly, Claudia J

    2009-07-01

    Are hospital-based outpatient interdisciplinary clinics a financially viable alternative for caring for our burgeoning population of older adults in America? Although highly popular, with high patient satisfaction rates among older adults and their families, senior health clinics (SHCs) can be expensive to operate, with limited quantifiable health outcomes. This study analyzed three geriatric hospital-based interdisciplinary clinics in rural Arkansas by examining their patient profiles, revenues, and expenses. It closely examined the effects of the downstream revenue using the multiplier effect and acknowledged other factors that weigh heavily on the success of SHCs and the care of older adults. The findings highlight the similarities and differences in the three clinics' operating and financial structures in addition to the clinics' and providers' productivity. The analysis presents an evidence-based illustration that SHCs can break even or lose large amounts of money.

  3. Quorum sensing alters the microbial community of electrode-respiring bacteria and hydrogen scavengers toward improving hydrogen yield in microbial electrolysis cells

    International Nuclear Information System (INIS)

    Cai, Weiwei; Zhang, Zhaojing; Ren, Ge; Shen, Qiuxuan; Hou, Yanan; Ma, Anzhou; Deng, Ye; Wang, Aijie; Liu, Wenzong

    2016-01-01

    Highlights: • Enhanced hydrogen yield has been achieved with addition of AHL. • AHL regulated exoelectrogens resulting in electrochemical activity enhancement. • Microbial community shift in cathodic biofilm inhibited hydrogen loss. - Abstract: Quorum sensing has been widely applied to enhance the energy recovery of bioelectrochemical system as a sustainable pathway to enhance communication between cells and electrodes. However, how signalling molecules (acyl-homoserine lactones, AHLs) regulate the microbial community to improve hydrogen generation in microbial electrolysis cells (MECs) is not well understood, especially the subsequent influence on interspecies relationships among not only electrode-respiring bacteria but also hydrogen scavengers. Understanding AHL regulation in a complicated and actual biofilm system will be valuable for future applications of microbial electrochemical technology. Herein, we added short-chain AHLs (3OC6) to regulate the biofilm community on bio-electrodes in MECs. As a result, hydrogen yields were enhanced with AHL addition, increasing by 5.57%, 38.68%, and 81.82% with varied external voltages (0.8 V, 0.6 V, and 0.4 V, respectively). Accordingly, overall reactor performance was enhanced, including coulombic efficiency, electron recovery efficiency, and energy efficiency. Based on an electrochemical impedance spectra analysis, the structured biofilm under simple nutrient conditions (acetate) showed a lower internal resistance with AHL addition, indicating that the microbial communities were altered to enhance electron transfer between the biofilm and electrode. The change in the cathodic microbial structure with more electrochemically active bacteria and fewer hydrogen scavengers could contribute to a higher electron recovery and hydrogen yield with AHL addition. The regulation of the microbial community structure by AHLs represents a potential strategy to enhance electron transfer and hydrogen generation in

  4. Progress in emerging techniques for characterization of immobilized viable whole-cell biocatalysts

    Czech Academy of Sciences Publication Activity Database

    Bučko, M.; Vikartovská, A.; Schenkmayerová, A.; Tkáč, J.; Filip, J.; Chorvát Jr., D.; Neděla, Vilém; Ansorge-Schumacher, M.B.; Gemeiner, P.

    2017-01-01

    Roč. 71, č. 11 (2017), s. 2309-2324 ISSN 0366-6352 Institutional support: RVO:68081731 Keywords : bioelectrocatalysis * imaging techniques * immobilized whole-cell biocatalyst * multienzyme cascade reactions * online kinetics Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering OBOR OECD: Bioprocessing technologies (industrial processes relying on biological agents to drive the process) biocatalysis, fermentation Impact factor: 1.258, year: 2016

  5. Physiological basis of barley yield under near optimal and stress conditions

    Directory of Open Access Journals (Sweden)

    Pržulj Novo

    2004-01-01

    Full Text Available Average barley yield fall below its potential due to incidence of stresses. Water stress is the main environmental factor limiting yield. The component a priori more sensitive to most stresses is the amount of radiation absorbed. The effect of stresses influence on the total amount of radiation absorbed by barley crop during its vegetation and the photosynthetic efficiency of radiation conversion. Growth inhibition is accompanied by reductions in leaf and cell wall extensibility. Grain yield under drought conditions is source limited. Supply of assimilates to the developing inflorescence plays a critical role in establishing final grain number and grain size. Grain weight is negatively affected by drought, high temperature, and any other factors that may reduce grain filling duration and grain filling rate. Awns and glaucousness confer better performance of barley under drought stress conditions. Barley responds with an increased accumulation of a number of proteins when subjected to different stress inducing cell dehydration. Screening techniques that are able to identify desirable genotypes based on the evaluation of physiological traits related to stress evasion and stress resistance maybe useful in breeding barley for resistance to stress, particularly drought stress. Crop management and breeding can reduce the incidence of stress on yield. The effect of these practices is sustained by an understanding of their physiology. In this paper the physiological basis of the processes determining barley yield and the incidence of stresses on photosynthetic metabolism that determine grain yield of barley is discussed. .

  6. Estimated yield of double-strand breaks from internal exposure to tritium.

    Science.gov (United States)

    Chen, Jing

    2012-08-01

    Internal exposure to tritium may result in DNA lesions. Of those, DNA double-strand breaks (DSBs) are believed to be important. However, experimental and computational data of DSBs induction by tritium are very limited. In this study, microdosimetric characteristics of uniformly distributed tritium were determined in dimensions of critical significance in DNA DSBs. Those characteristics were used to identify other particles comparable to tritium in terms of microscopic energy deposition. The yield of DSBs could be strongly dependent on biological systems and cellular environments. After reviewing theoretically predicted and experimentally determined DSB yields available in the literature for low-energy electrons and high-energy protons of comparable microdosimetric characteristics to tritium in the dimensions relevant to DSBs, it is estimated that the average DSB yields of 2.7 × 10(-11), 0.93 × 10(-11), 2.4 × 10(-11) and 1.6 × 10(-11) DSBs Gy(-1) Da(-1) could be reasonable estimates for tritium in plasmid DNAs, yeast cells, Chinese hamster V79 cells and human fibroblasts, respectively. If a biological system is not specified, the DSB yield from tritium exposure can be estimated as (2.3 ± 0.7) × 10(-11) DSBs Gy(-1) Da(-1), which is a simple average over experimentally determined yields of DSBs for low-energy electrons in various biological systems without considerations of variations caused by different techniques used and obvious differences among different biological systems where the DSB yield was measured.

  7. Predictors of mother and child DNA yields in buccal cell samples collected in pediatric cancer epidemiologic studies: a report from the Children's Oncology group.

    Science.gov (United States)

    Poynter, Jenny N; Ross, Julie A; Hooten, Anthony J; Langer, Erica; Blommer, Crystal; Spector, Logan G

    2013-08-12

    Collection of high-quality DNA is essential for molecular epidemiology studies. Methods have been evaluated for optimal DNA collection in studies of adults; however, DNA collection in young children poses additional challenges. Here, we have evaluated predictors of DNA quantity in buccal cells collected for population-based studies of infant leukemia (N = 489 mothers and 392 children) and hepatoblastoma (HB; N = 446 mothers and 412 children) conducted through the Children's Oncology Group. DNA samples were collected by mail using mouthwash (for mothers and some children) and buccal brush (for children) collection kits and quantified using quantitative real-time PCR. Multivariable linear regression models were used to identify predictors of DNA yield. Median DNA yield was higher for mothers in both studies compared with their children (14 μg vs. mothers or children in this analysis. The association with seasonality suggests that conditions during transport may influence DNA yield. The low yields observed in most children in these studies highlight the importance of developing alternative methods for DNA collection in younger age groups.

  8. Economically viable large-scale hydrogen liquefaction

    Science.gov (United States)

    Cardella, U.; Decker, L.; Klein, H.

    2017-02-01

    The liquid hydrogen demand, particularly driven by clean energy applications, will rise in the near future. As industrial large scale liquefiers will play a major role within the hydrogen supply chain, production capacity will have to increase by a multiple of today’s typical sizes. The main goal is to reduce the total cost of ownership for these plants by increasing energy efficiency with innovative and simple process designs, optimized in capital expenditure. New concepts must ensure a manageable plant complexity and flexible operability. In the phase of process development and selection, a dimensioning of key equipment for large scale liquefiers, such as turbines and compressors as well as heat exchangers, must be performed iteratively to ensure technological feasibility and maturity. Further critical aspects related to hydrogen liquefaction, e.g. fluid properties, ortho-para hydrogen conversion, and coldbox configuration, must be analysed in detail. This paper provides an overview on the approach, challenges and preliminary results in the development of efficient as well as economically viable concepts for large-scale hydrogen liquefaction.

  9. Economically viable biochemical processes for the advanced rural biorefinery and downstream recovery operations

    Science.gov (United States)

    Rural biorefineries offer an alternative to traditional ethanol production by providing the opportunity to produce fuel on site to reduce costs associated with biomass transportation thus making the fuel economically viable. Widespread installation of rural biorefineries could lead to increased upt...

  10. Effect of Integrated Nutrient Management on Yield and Yield ...

    African Journals Online (AJOL)

    Declining soil fertility is one of the major problems causing yield reduction of barley ... (VC) with inorganic NP on growth, yield and yield components of food barley. ... The experiments were laid out in a randomized complete block design with ...

  11. Oxidative Stress Induces Senescence in Cultured RPE Cells.

    Science.gov (United States)

    Aryan, Nona; Betts-Obregon, Brandi S; Perry, George; Tsin, Andrew T

    2016-01-01

    The aim of this research is to determine whether oxidative stress induces cellular senescence in human retinal pigment epithelial cells. Cultured ARPE19 cells were subjected to different concentrations of hydrogen peroxide to induce oxidative stress. Cells were seeded into 24-well plates with hydrogen peroxide added to cell medium and incubated at 37°C + 5% CO2 for a 90-minute period [at 0, 300, 400 and 800 micromolar (MCM) hydrogen peroxide]. The number of viable ARPE19 cells were recorded using the Trypan Blue Dye Exclusion Method and cell senescence was measured by positive staining for senescence-associated beta-galactosidase (SA-beta-Gal) protein. Without hydrogen peroxide treatment, the number of viable ARPE19 cells increased significantly from 50,000 cells/well to 197,000 within 72 hours. Treatment with hydrogen peroxide reduced this level of cell proliferation significantly (to 52,167 cells at 400 MCM; to 49,263 cells at 800 MCM). Meanwhile, cells with a high level of positive senescence-indicator SA-Beta-Gal-positive staining was induced by hydrogen peroxide treatment (from a baseline level of 12% to 80% at 400 MCM and at 800 MCM). Our data suggests that oxidative stress from hydrogen peroxide treatment inhibited ARPE19 cell proliferation and induced cellular senescence.

  12. Breaking down the barriers to commercialization of fuel cells in transportation through Government - industry R&D programs

    Energy Technology Data Exchange (ETDEWEB)

    Chalk, S.G. [Dept. of Energy, Washington, DC (United States); Venkateswaran, S.R. [Energetics, Inc., Columbia, MD (United States)

    1996-12-31

    PEM fuel cell technology is rapidly emerging as a viable propulsion alternative to the internal combustion engine. Fuel cells offer the advantages of low emissions, high efficiency, fuel flexibility, quiet and continuous operation, and modularity. Over the last decade, dramatic advances have been achieved in the performance and cost of PEM fuel cell technologies for automotive applications. However, significant technical barriers remain to making fuel cell propulsion systems viable alternatives to the internal combustion engine. This paper focuses on the progress achieved and remaining technical barriers while highlighting Government-industry R&D efforts that are accelerating fuel cell technology toward commercialization.

  13. Immunization against chlamydial genital infection in guinea pigs with UV-inactivated and viable chlamydiae administered by different routes

    International Nuclear Information System (INIS)

    Rank, R.G.; Batteiger, B.E.; Soderberg, L.S.

    1990-01-01

    Female guinea pigs were immunized with viable or UV light-inactivated chlamydiae, belonging to the species Chlamydia psittaci, by intravenous, subcutaneous, oral, or ocular routes. All animals were then inoculated vaginally with viable chlamydiae to determine the extent of protection against challenge infection induced by the various regimens. The course of genital infection was significantly reduced in intensity in all groups of animals except the unimmunized controls and those animals immunized orally with inactivated antigen. Guinea pigs immunized with viable antigen were more likely to develop resistance to challenge infection and, in general, had a significantly greater degree of protection than animals immunized with inactivated antigen. No one route seemed superior in producing a protective response. Animals in all groups demonstrating protection developed serum and secretion immunoglobulin G antibody responses to chlamydiae. Lymphocyte proliferative reactions to chlamydial antigen were variable among groups. Immunoblot analysis of serum and secretions indicated a wide range of antibody specificities, but most protected animals produced antibodies to the major outer membrane protein, lipopolysaccharide, and the 61-kilodalton protein. No definitive associations could be made between the increased ability of immunization with viable organisms to produce resistance to challenge infection and a particular immune parameter. These data indicate that viable chlamydiae given by various routes are able to induce a strong immune response which can provide resistance against reinfection in some cases or at least reduce the degree of infection to a greater degree than inactivated antigen. However, complete resistance to genital tract infection may be difficult to obtain and alternate immunizations strategies may have to be developed

  14. Flexible inverted polymer solar cells with an indium-free tri-layer cathode

    International Nuclear Information System (INIS)

    El Hajj, Ahmad; Lucas, Bruno; Schirr-Bonnans, Martin; Ratier, Bernard; Kraft, Thomas M.; Torchio, Philippe

    2014-01-01

    Indium tin oxide (ITO)-free inverted polymer solar cells (PSCs) have been fabricated without the need of an additional electron transport layer. The indium-free transparent electrode consists of a tri-layer stack ZnO (30 nm)/Ag (14 nm)/ZnO (30 nm) deposited on glass and plastic substrates via ion-beam sputtering. The tri-layer electrodes exhibit similar physical properties to its ITO counterpart, specifically yielding high transmittance and low resistivity (76.5% T at 550 nm, R sq of 8 Ω/◻) on plastic substrates. The novel tri-layer electrode allows for the fabrication of inverted PSCs without the additional ZnO interfacial layer commonly deposited between ITO and the photoactive layer. This allows for the preparation of thinner plastic solar cells using less material than conventional architectures. Initial studies involving the newly realized architecture (tri-layer electrode/P3HT:PCBM/PEDOT:PSS/Ag) have shown great promise for the transition from ITO to other viable electrodes in organic electronics

  15. Cannabidiol Reduces Leukemic Cell Size - But Is It Important?

    Science.gov (United States)

    Kalenderoglou, Nikoletta; Macpherson, Tara; Wright, Karen L

    2017-01-01

    The anti-cancer effect of the plant-derived cannabinoid, cannabidiol, has been widely demonstrated both in vivo and in vitro . However, this body of preclinical work has not been translated into clinical use. Key issues around this failure can be related to narrow dose effects, the cell model used and incomplete efficacy. A model of acute lymphoblastic disease, the Jurkat T cell line, has been used extensively to study the cannabinoid system in the immune system and cannabinoid-induced apoptosis. Using these cells, this study sought to investigate the outcome of those remaining viable cells post-treatment with cannabidiol, both in terms of cell size and tracking any subsequent recovery. The phosphorylation status of the mammalian Target of Rapamycin (mTOR) signaling pathway and the downstream target ribosomal protein S6, were measured. The ability of cannabidiol to exert its effect on cell viability was also evaluated in physiological oxygen conditions. Cannabidiol reduced cell viability incompletely, and slowed the cell cycle with fewer cells in the G2/M phase of the cell cycle. Cannabidiol reduced phosphorylation of mTOR, PKB and S6 pathways related to survival and cell size. The remaining population of viable cells that were cultured in nutrient rich conditions post-treatment were able to proliferate, but did not recover to control cell numbers. However, the proportion of viable cells that were gated as small, increased in response to cannabidiol and normally sized cells decreased. This proportion of small cells persisted in the recovery period and did not return to basal levels. Finally, cells grown in 12% oxygen (physiological normoxia) were more resistant to cannabidiol. In conclusion, these results indicate that cannabidiol causes a reduction in cell size, which persists post-treatment. However, resistance to cannabidiol under physiological normoxia for these cells would imply that cannabidiol may not be useful in the clinic as an anti-leukemic agent.

  16. Stereotaxic implantation of dispersed cell suspensions into brain. A systematic appraisal of cell placement and survival

    International Nuclear Information System (INIS)

    Plunkett, R.J.; Weber, R.J.; Oldfield, E.H.

    1988-01-01

    The application of several recent advances in cell biology, brain implantation, and cell-mediated tumor immunotherapy requires successful and reproducible placement of viable cell suspensions into brain. Stereotaxic implantation is being used to inject cytotoxic lymphocytes into gliomas and to replace dopaminergic cells in parkinsonian models. Systematic assessment of the factors that influence success in implantation of cell suspensions into solid tissues is needed. A model was developed for investigation of stereotaxic implantation using radiolabeled rat lymphokine-activated killer (LAK) cells. Anesthetized rats received microliter injections of cell suspension into the right caudate nucleus. The injection volume, cell concentration, infusion rate, and needle size were varied systematically. The animals were sacrificed 1 hour after injection; the brain was removed and sectioned, and the radioactivity was counted. Three aliquots of the suspension were injected into counting tubes for control analysis. Recovery of radioactivity was expressed as the percent of mean counts per minute (cpm) in the right frontal lobe/mean cpm in the three control tubes. To assess the viability of implanted cells, the right frontal region was mechanically dissociated in media and centrifuged, and the pellet and supernatant were counted. By using small needles and slow infusion of volumes of 10 microliters or less, 85% to 90% of the radioactivity was recovered in the caudate nucleus. At least half of the implanted cells were viable. Consistent, accurate implantation of dispersed cells into brain over a range of volumes, cell concentrations, infusion rates, and needle sizes was achieved

  17. Yield trends and yield gap analysis of major crops in the world

    OpenAIRE

    Hengsdijk, H.; Langeveld, J.W.A.

    2009-01-01

    This study aims to quantify the gap between current and potential yields of major crops in the world, and the production constraints that contribute to this yield gap. Using an expert-based evaluation of yield gaps and the literature, global and regional yields and yield trends of major crops are quantified, yield gaps evaluated by crop experts, current yield progress by breeding estimated, and different yield projections compared. Results show decreasing yield growth for wheat and rice, but ...

  18. In search of viable business models for development: sustainable energy in developing countries

    NARCIS (Netherlands)

    Kolk, A.; van den Buuse, D.

    2012-01-01

    Purpose - Although the crucial role of business, and of business-based approaches, in development is increasingly emphasised by academics and practitioners, insight is lacking into the "whether and how" of viable business models, in environmental, social and economical terms. This article analyses

  19. Equity yields

    NARCIS (Netherlands)

    Vrugt, E.; van Binsbergen, J.H.; Koijen, R.S.J.; Hueskes, W.

    2013-01-01

    We study a new data set of dividend futures with maturities up to ten years across three world regions: the US, Europe, and Japan. We use these asset prices to construct equity yields, analogous to bond yields. We decompose the equity yields to obtain a term structure of expected dividend growth

  20. Enterocolic lymphocytic phlebitis: statistical analysis of histology features in viable and ischemic bowel.

    Science.gov (United States)

    Medlicott, Shaun A C; Guggisberg, Kelly A; DesCôteaux, Jean-Gaston; Beck, Paul

    2006-07-01

    Enterocolic lymphocytic phlebitis is a rare cause of segmental ischemic enterocolitis. This artery-sparing transmural vasculitis is classically a circumferential phlebitis with perivenular lymphocyte cuffing and thrombi in the absence of systemic manifestations. Myointimal hyperplasia may represent a chronic phase of enterocolic lymphocytic phlebitis. Subclinical or early stage enterocolic lymphocytic phlebitis is not well delineated. We analyzed 600 submucosal and subserosal veins from both ischemic and intact bowel segments to discern if vascular morphology varied between sites. Crescentic and circumferential lymphocytic phlebitis is more common in viable bowel than in the ischemic segment. A nonsignificant trend was found for increased crescentic morphology between intact bowel remote from the ischemic focus compared with that adjacent to the ischemic focus. Hallmarks of ischemic bowel are necrotizing phlebitis and thrombi formation. Thrombophlebitis morphology is distinctly different in viable and ischemic bowel, changing from the classic lymphocytic to necrotizing lesions respectively.

  1. Blood flow, flow reserve, and glucose utilization in viable and nonviable myocardium in patients with ischemic cardiomyopathy.

    Science.gov (United States)

    Zhang, Xiaoli; Schindler, Thomas H; Prior, John O; Sayre, James; Dahlbom, Magnus; Huang, Sung-Cheng; Schelbert, Heinrich R

    2013-04-01

    The aim of the study was to determine whether glucose uptake in viable myocardium of ischemic cardiomyopathy patients depends on rest myocardial blood flow (MBF) and the residual myocardial flow reserve (MFR). Thirty-six patients with ischemic cardiomyopathy (left ventricular ejection fraction 25 ± 10 %) were studied with (13)N-ammonia and (18)F-fluorodeoxyglucose (FDG) positron emission tomography (PET). Twenty age-matched normals served as controls. Regional MBF was determined at rest and during dipyridamole hyperemia and regional FDG extraction was estimated from regional FDG to (13)N-ammonia activity ratios. Rest MBF was reduced in viable (0.42 ± 0.18 ml/min per g) and nonviable regions (0.32 ± 0.09 ml/min per g) relative to remote regions (0.68 ± 0.23 ml/min per g, p MFRs did not differ significantly (p > 0.05). Compared to MFR in remote myocardium, MFRs in viable regions were similar (1.39 ± 0.56 vs 1.70 ± 0.45, p > 0.05) but were significantly lower in nonviable regions (1.23 ± 0.43, p MFRs (r =-0.424, p MFRs in viable myocardium are associated with increasing glucose extraction that likely reflects a metabolic adaptation of remodeling hibernating myocytes.

  2. Maize Residue as a Viable Substrate for Farm Scale Cultivation of Oyster Mushroom (Pleurotus ostreatus

    Directory of Open Access Journals (Sweden)

    Abena O. Adjapong

    2015-01-01

    Full Text Available In the search for alternatives to sawdust as growing media in commercial mushroom cultivation, three organic substrates obtainable as crop residue, maize husk, maize cob, and maize stalk, with each being supplemented with rice bran, were evaluated as growth media for the oyster mushroom, Pleurotus ostreatus (Kummer. For the tested alternatives to sawdust, the harvested weight of fruiting bodies that sprouted on a kilogram maize husk media per crop (32.99 g was the highest. Sawdust media supported significantly (P<0.001 heavier fruiting bodies (42.18 than the maize residues. The peak mushroom harvests for the various substrates were obtained between the first and seventh fruiting body flushes. The biological efficiency of the substrates, which measured usable nutrients indicated that maize stalk supplemented with rice bran, was 39% compared to that of the sawdust media (60%. The maize husk media and the maize cob media had biological efficiencies of 32% and 9.5%, respectively. These results indicate that two of the tested growing media (maize stalk or husk produced mushrooms with yield characteristics that were comparable to the well-used sawdust in the cultivation of oyster mushrooms. The environmental and economic parameters involved in the use and carting of sawdust make these on-farm crop residues a viable alternative for mushroom cultivation in especially nonforest zones of Ghana.

  3. Finding viable models in SUSY parameter spaces with signal specific discovery potential

    Science.gov (United States)

    Burgess, Thomas; Lindroos, Jan Øye; Lipniacka, Anna; Sandaker, Heidi

    2013-08-01

    Recent results from ATLAS giving a Higgs mass of 125.5 GeV, further constrain already highly constrained supersymmetric models such as pMSSM or CMSSM/mSUGRA. As a consequence, finding potentially discoverable and non-excluded regions of model parameter space is becoming increasingly difficult. Several groups have invested large effort in studying the consequences of Higgs mass bounds, upper limits on rare B-meson decays, and limits on relic dark matter density on constrained models, aiming at predicting superpartner masses, and establishing likelihood of SUSY models compared to that of the Standard Model vis-á-vis experimental data. In this paper a framework for efficient search for discoverable, non-excluded regions of different SUSY spaces giving specific experimental signature of interest is presented. The method employs an improved Markov Chain Monte Carlo (MCMC) scheme exploiting an iteratively updated likelihood function to guide search for viable models. Existing experimental and theoretical bounds as well as the LHC discovery potential are taken into account. This includes recent bounds on relic dark matter density, the Higgs sector and rare B-mesons decays. A clustering algorithm is applied to classify selected models according to expected phenomenology enabling automated choice of experimental benchmarks and regions to be used for optimizing searches. The aim is to provide experimentalist with a viable tool helping to target experimental signatures to search for, once a class of models of interest is established. As an example a search for viable CMSSM models with τ-lepton signatures observable with the 2012 LHC data set is presented. In the search 105209 unique models were probed. From these, ten reference benchmark points covering different ranges of phenomenological observables at the LHC were selected.

  4. Foodservice yield and fabrication times for beef as influenced by purchasing options and merchandising styles.

    Science.gov (United States)

    Weatherly, B H; Griffin, D B; Johnson, H K; Walter, J P; De La Zerda, M J; Tipton, N C; Savell, J W

    2001-12-01

    steaks in terms of yield and labor cost, foodservice operators will be better equipped to decide what option is more viable for their operation.

  5. Electron-deficient N-alkyloyl derivatives of thieno[3,4-c]pyrrole-4,6-dione yield efficient polymer solar cells with open-circuit voltages > 1 v

    KAUST Repository

    Warnan, Julien; Cabanetos, Clement; Bude, Romain; El Labban, Abdulrahman; LI, LIANG; Beaujuge, Pierre

    2014-01-01

    Poly(benzo[1,2-b:4,5-b′]dithiophene-thieno[3,4-c]pyrrole-4,6-dione) (PBDTTPD) polymer donors yield some of the highest open-circuit voltages (V OC, ca. 0.9 V) and fill factors (FF, ca. 70%) in conventional bulk-heterojunction (BHJ) solar cells

  6. Evolution of β-Cell Replacement Therapy in Diabetes Mellitus: Islet Cell Transplantation

    Science.gov (United States)

    Jahansouz, Cyrus; Jahansouz, Cameron; Kumer, Sean C.; Brayman, Kenneth L.

    2011-01-01

    Diabetes mellitus remains one of the leading causes of morbidity and mortality worldwide. According to the Centers for Disease Control and Prevention, approximately 23.6 million people in the United States are affected. Of these individuals, 5 to 10% have been diagnosed with Type 1 diabetes mellitus (T1DM), an autoimmune disease. Although it often appears in childhood, T1DM may manifest at any age, leading to significant morbidity and decreased quality of life. Since the 1960s, the surgical treatment for diabetes mellitus has evolved to become a viable alternative to insulin administration, beginning with pancreatic transplantation. While islet cell transplantation has emerged as another potential alternative, its role in the treatment of T1DM remains to be solidified as research continues to establish it as a truly viable alternative for achieving insulin independence. In this paper, the historical evolution, procurement, current status, benefits, risks, and ongoing research of islet cell transplantation are explored. PMID:22013505

  7. Survival or revival: long-term preservation induces a reversible viable but non-culturable state in methane-oxidizing bacteria.

    Directory of Open Access Journals (Sweden)

    Sven Hoefman

    Full Text Available Knowledge on long-term preservation of micro-organisms is limited and research in the field is scarce despite its importance for microbial biodiversity and biotechnological innovation. Preservation of fastidious organisms such as methane-oxidizing bacteria (MOB has proven difficult. Most MOB do not survive lyophilization and only some can be cryopreserved successfully for short periods. A large-scale study was designed for a diverse set of MOB applying fifteen cryopreservation or lyophilization conditions. After three, six and twelve months of preservation, the viability (via live-dead flow cytometry and culturability (via most-probable number analysis and plating of the cells were assessed. All strains could be cryopreserved without a significant loss in culturability using 1% trehalose in 10-fold diluted TSB (TT as preservation medium and 5% DMSO as cryoprotectant. Several other cryopreservation and lyophilization conditions, all of which involved the use of TT medium, also allowed successful preservation but showed a considerable loss in culturability. We demonstrate here that most of these non-culturables survived preservation according to viability assessment indicating that preservation induces a viable but non-culturable (VBNC state in a significant fraction of cells. Since this state is reversible, these findings have major implications shifting the emphasis from survival to revival of cells in a preservation protocol. We showed that MOB cells could be significantly resuscitated from the VBNC state using the TT preservation medium.

  8. The ghost of extinction: Preservation values and minimum viable population in wildlife models

    NARCIS (Netherlands)

    Eiswerth, M.E.; Kooten, van G.C.

    2009-01-01

    The inclusion of a minimum viable population in bioeconomic modeling creates at least two complications that are not resolved by using a modified logistic growth function. The first complication can be dealt with by choosing a different depensational growth function. The second complication relates

  9. Effect of elevated postirradiation pH on the yield of double-strand breaks in DNA from irradiated bacterial cells

    International Nuclear Information System (INIS)

    Tilby, M.J.; Loverock, P.S.; Fielden, E.M.

    1984-01-01

    Exposure of DNA isolated from irradiated cells of Escherichia coli to a pH of 9.6 caused a marked increase in the yield of double-strand breaks (dsb). After incubation for 4 hr at 37 0 C and pH 9.6 the dsb yields were 95% and 71% higher than when incubation was at pH 7.0 for irradiation under oxic and anoxic conditions, respectively. This effect was not apparent when dsb were induced enzymatically and it was linearly related to radiation dose. After oxic irradiation, the increase in dsb at pH 9.6 was consistent with first-order kinetics over >2 half-lives (t/sub 1/2/ = 1.6 hr at 37 0 C). It is propsoed that the effects of elevated pH revealed the presence in intracellularly irradiated DNA of previously unidentified sites where both strands of the DNA were damaged as a result of single radiation events. The possible nature of the proposed sites and the relevance of these findings to the ''neutral'' elution technique are discussed

  10. The Effects of Royal Jelly on In-Vitro Cytotoxicity of K562 Cells and Peripheral Blood Mononuclear Cells

    Directory of Open Access Journals (Sweden)

    SE Hosseini

    2014-02-01

    Full Text Available Abstract Background & aim: Royal jelly, secreted by worker bees, has different biological activities on cells and tissues. The aim of this study was to evaluate the effects of royal jelly on peripheral blood mononuclear cells and on the tumor category of K562 cell line. Methods: In the present experimental study, three subjects were selected separately with three repetitions. K562 (104 cells and PBMC (105 cells with different concentrations of royal jelly (5, 10, 25, 50 and 100 mg/ml were cultured under standard conditions for 48 and 72 h separately. The fatality rate on PBMC cells and K562 cancer cells was evaluated by using MTT (Tetrazolium Dye-Reduction Assay. The number of viable cells in PBMC that were exposed for 48 hours with Royal Jelly was evaluated by trypan blue staining. Data were analyzed by ANOVA. Results: The royal jelly had no cytotoxicity effect on PBMC cells but at concentration of 50 and 100 mg/mL the cytotoxicity effect were observed on k562 cells whereas, at 10 and 25 mg/ml the number of PBMC viable cells increased. Conclusion: Due to the lack of lethality of royal jelly on PBMC cells and PBMC cell viability and an increase in the fatality rate of cancer cells in the future, royal jelly can be used as a potential candidate for treatment of leukemia. Keywords: Royal jelly, K562, peripheral blood mononuclear cell

  11. In Silico Design of Robust Bolalipid Membranes

    NARCIS (Netherlands)

    Bulacu, Monica; Periole, Xavier; Marrink, Siewert J.; Périole, Xavier

    The robustness of microorganisms used in industrial fermentations is essential for the efficiency and yield of the production process. A viable tool to increase the robustness is through engineering of the cell membrane and especially by incorporating lipids from species that survive under harsh

  12. Langerhans cells are required for UVR-induced immunosuppression

    NARCIS (Netherlands)

    Schwarz, Agatha; Noordegraaf, Madelon; Maeda, Akira; Torii, Kan; Clausen, Björn E.; Schwarz, Thomas

    2010-01-01

    Painting of haptens onto UVR-exposed skin does not result in sensitization but induces regulatory T cells (Treg). This was explained by UVR-mediated depletion of Langerhans cells (LCs). Furthermore, migration of UVR-damaged but still viable LCs into lymph nodes appears to be essential to induce

  13. Estimated yield of double-strand breaks from internal exposure to tritium

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Jing [Health Canada, Radiation Protection Bureau, Ottawa, ON (Canada)

    2012-08-15

    Internal exposure to tritium may result in DNA lesions. Of those, DNA double-strand breaks (DSBs) are believed to be important. However, experimental and computational data of DSBs induction by tritium are very limited. In this study, microdosimetric characteristics of uniformly distributed tritium were determined in dimensions of critical significance in DNA DSBs. Those characteristics were used to identify other particles comparable to tritium in terms of microscopic energy deposition. The yield of DSBs could be strongly dependent on biological systems and cellular environments. After reviewing theoretically predicted and experimentally determined DSB yields available in the literature for low-energy electrons and high-energy protons of comparable microdosimetric characteristics to tritium in the dimensions relevant to DSBs, it is estimated that the average DSB yields of 2.7 x 10{sup -11}, 0.93 x 10{sup -11}, 2.4 x 10{sup -11} and 1.6 x 10{sup -11} DSBs Gy{sup -1} Da{sup -1} could be reasonable estimates for tritium in plasmid DNAs, yeast cells, Chinese hamster V79 cells and human fibroblasts, respectively. If a biological system is not specified, the DSB yield from tritium exposure can be estimated as (2.3 ± 0.7) x 10{sup -11} DSBs Gy{sup -1} Da{sup -1}, which is a simple average over experimentally determined yields of DSBs for low-energy electrons in various biological systems without considerations of variations caused by different techniques used and obvious differences among different biological systems where the DSB yield was measured. (orig.)

  14. X-radiation-induced transformation in a C3H mouse embryo-derived cell line

    International Nuclear Information System (INIS)

    Terzaghi, M.; Little, J.B.

    1976-01-01

    Reproducible x-ray-induced oncogenic transformation has been demonstrated in an established cell line of mouse embryo fibroblasts. Cells derived from transformed foci formed malignant tumors when injected into syngeneic hosts. An exponential increase in the number of transformants per viable cell occurred with doses of up to 400 rads of x-radiation. The transformation frequency in exponentially growing cultures remained constant at 2.3 x 10 -3 following doses of 400 to 1500 rads. There was little change in survival following x-ray doses up to 300 rads. Doses greater than 300 rads were associated with an exponential decline in survival; the D 0 for the survival curve was 175 rads. Transformation frequency varied with changes in the number of viable cells seeded per dish. There was about a 10-fold decline in the transformation frequency when the number of cells was increased from 400 to 1000 viable cells/100-mm Petri dish. Below this density range there was little change in transformation frequency. The presence of lethally preirradiated cells was not associated with an enhancement of transformation in irradiated cells or with the induction of transformation in unirradiated cell cultures. Amphotericin B (Fungizone) inhibited the appearance of transformants when added to the culture medium within 2 to 3 weeks after initiation of the experiment

  15. Lactobacillus crispatus L1: high cell density cultivation and exopolysaccharide structure characterization to highlight potentially beneficial effects against vaginal pathogens.

    Science.gov (United States)

    Donnarumma, Giovanna; Molinaro, Antonio; Cimini, Donatella; De Castro, Cristina; Valli, Vivien; De Gregorio, Vincenza; De Rosa, Mario; Schiraldi, Chiara

    2014-05-30

    Vaginal lactic acid bacteria defend the host against pathogens through a combination of competitive exclusion, competition for nutrients, production of antimicrobial substances and through the activation of the immune system. A new human isolate named Lactobacillus crispatus L1 was characterized in this work, and a preliminary evaluation of its probiotic potential is described together with a process to obtain a high productivity of viable biomass. In a simulated digestion process 1.8⋅10(10) cells∙ml(-1) survived the gastric environment with 80% viability, without being affected by small intestine juices. Experiments on six different C sources were performed to analyze growth and organic acids production and, glucose, provided the best performances. A microfiltration strategy was exploited to improve the cellular yield in 2 L-fermentation processes, reaching 27 g · l(-1) of dry biomass. Moreover, L. crispatus L1 demonstrated a greater stability to high concentrations of lactic acid, compared to other lactobacilli. The specific L. crispatus L1 exopolysaccharide was purified from the fermentation broth and characterized by NMR showing structural features and similarity to exopolysaccharides produced by pathogenic strains. Live L. crispatus L1 cells strongly reduced adhesion of a yeast pathogenic strain, Candida albicans in particular, in adherence assays. Interestingly a higher expression of the human defensin HBD-2 was also observed in vaginal cells treated with the purified exopolysaccharide, indicating a possible correlation with C. albicans growth inhibition. The paper describes the evaluation of L. crispatus L1 as potential vaginal probiotic and the fermentation processes to obtain high concentrations of viable cells.

  16. A Method for Microencapsulation of Cells and a Device for Its Realization.

    Science.gov (United States)

    Lepekhova, S A; Goldberg, O A; Kravchenko, A A; Batraks, A E; Koval, E V; Kurgansky, I S; Apartsin, K A

    2017-04-01

    The device for cell encapsulation makes it possible to fabricate microcapsules of a preset size with even smooth surface, without defects or adhesion to each other, with viable cells inside the capsule. The cells were derived from newborn piglet pancreases.

  17. A hybrid bio-jetting approach for directly engineering living cells

    International Nuclear Information System (INIS)

    Kwok, Albert; Irvine, Scott; Arumuganathar, Sumathy; Jayasinghe, Suwan N; McEwan, Jean R

    2008-01-01

    This paper reports developments on a hybrid cell-engineering protocol coupling both bio-electrosprays and aerodynamically assisted bio-jets for process-handling living cells. The current work demonstrates the ability to couple these two cell-jetting protocols for handling a wide range of cells for deposition. The post-treated cells are assessed for their viability by way of flow cytometry, which illustrates a significant population of viable cells post-treatment in comparison to those controls. This work is the first example of coupling these two protocols for the process handling of living cells. The hybrid protocol demonstrates the achievement of stable cone jetting of a cellular suspension in the single-needle configuration which was previously unachieved with single-needle bio-electrosprays. Furthermore the living cells explored in these investigations expressed GFP, thus demonstrating the ability to couple gene therapy with this hybrid protocol. Hence, this approach could one day be explored for building biologically viable tissues incorporating a therapeutic payload for combating a range of cellular/tissue-based pathologies

  18. Establishment of a Viable Population of Red-Cockaded Woodpeckers at the Savannah River Site

    International Nuclear Information System (INIS)

    Johnston, P.A.

    2002-01-01

    Report on program's objective to restore viable population of Red-cockaded woodpecker at SRS. Several management strategies were used to promote population expansion of Red-cockaded woodpecker and reduction of interspecific competition with Red-Cockaded woodpecker

  19. Comparison of viable plate count, turbidity measurement and real-time PCR for quantification of Porphyromonas gingivalis.

    Science.gov (United States)

    Clais, S; Boulet, G; Van Kerckhoven, M; Lanckacker, E; Delputte, P; Maes, L; Cos, P

    2015-01-01

    The viable plate count (VPC) is considered as the reference method for bacterial enumeration in periodontal microbiology but shows some important limitations for anaerobic bacteria. As anaerobes such as Porphyromonas gingivalis are difficult to culture, VPC becomes time-consuming and less sensitive. Hence, efficient normalization of experimental data to bacterial cell count requires alternative rapid and reliable quantification methods. This study compared the performance of VPC with that of turbidity measurement and real-time PCR (qPCR) in an experimental context using highly concentrated bacterial suspensions. Our TaqMan-based qPCR assay for P. gingivalis 16S rRNA proved to be sensitive and specific. Turbidity measurements offer a fast method to assess P. gingivalis growth, but suffer from high variability and a limited dynamic range. VPC was very time-consuming and less repeatable than qPCR. Our study concludes that qPCR provides the most rapid and precise approach for P. gingivalis quantification. Although our data were gathered in a specific research context, we believe that our conclusions on the inferior performance of VPC and turbidity measurements in comparison to qPCR can be extended to other research and clinical settings and even to other difficult-to-culture micro-organisms. Various clinical and research settings require fast and reliable quantification of bacterial suspensions. The viable plate count method (VPC) is generally seen as 'the gold standard' for bacterial enumeration. However, VPC-based quantification of anaerobes such as Porphyromonas gingivalis is time-consuming due to their stringent growth requirements and shows poor repeatability. Comparison of VPC, turbidity measurement and TaqMan-based qPCR demonstrated that qPCR possesses important advantages regarding speed, accuracy and repeatability. © 2014 The Society for Applied Microbiology.

  20. On the trace-free Einstein equations as a viable alternative to general relativity

    International Nuclear Information System (INIS)

    Ellis, George F R; Van Elst, Henk; Murugan, Jeff; Uzan, Jean-Philippe

    2011-01-01

    The quantum field theoretical prediction for the vacuum energy density leads to a value for the effective cosmological constant that is incorrect by between 60 and 120 orders of magnitude. We review an old proposal of replacing Einstein's field equations by their trace-free part (the trace-free Einstein equations), together with an independent assumption of energy-momentum conservation by matter fields. While this does not solve the fundamental issue of why the cosmological constant has the value that is observed cosmologically, it is indeed a viable theory that resolves the problem of the discrepancy between the vacuum energy density and the observed value of the cosmological constant. However, one has to check that, as well as preserving the standard cosmological equations, this does not destroy other predictions, such as the junction conditions that underlie the use of standard stellar models. We confirm that no problems arise here: hence, the trace-free Einstein equations are indeed viable for cosmological and astrophysical applications. (papers)

  1. SV40-transformed human fibroblasts: evidence for cellular aging in pre-crisis cells.

    Science.gov (United States)

    Stein, G H

    1985-10-01

    Pre-crisis SV40-transformed human diploid fibroblast (HDF) cultures have a finite proliferative lifespan, but they do not enter a viable senescent state at end of lifespan. Little is known about either the mechanism for this finite lifespan in SV40-transformed HDF or its relationship to finite lifespan in normal HDF. Recently we proposed that in normal HDF the phenomena of finite lifespan and arrest in a viable senescent state depend on two separate processes: 1) an age-related decrease in the ability of the cells to recognize or respond to serum and/or other mitogens such that the cells become functionally mitogen-deprived at the end of lifespan; and 2) the ability of the cells to enter a viable, G1-arrested state whenever they experience mitogen deprivation. In this paper, data are presented that suggest that pre-crisis SV40-transformed HDF retain the first process described above, but lack the second process. It is shown that SV40-transformed HDF have a progressively decreasing ability to respond to serum as they age, but they continue to traverse the cell cycle at the end of lifespan. Concomitantly, the rate of cell death increases steadily toward the end of lifespan, thereby causing the total population to cease growing and ultimately to decline. Previous studies have shown that when SV40-transformed HDF are environmentally serum deprived, they likewise exhibit continued cell cycle traverse coupled with increased cell death. Thus, these results support the hypothesis that pre-crisis SV40-transformed HDF still undergo the same aging process as do normal HDF, but they end their lifespan in crisis rather than in the normal G1-arrested senescent state because they have lost their ability to enter a viable, G1-arrested state in response to mitogen deprivation.

  2. YIELD AND QUALITY OF SORGHUM IN IRRIGATED AGRO LANDSCAPES OF REPUBLIC OF DAGESTAN

    Directory of Open Access Journals (Sweden)

    M G. Muslimov

    2016-01-01

    Full Text Available Aim. One of drought-resistant crops that can provide stable high yields is sorghum, which is salt-tolerant, heat-resistant and a flexible crop of versatile use (green forage, silage, hay, grass meal, grain forage. The research conducted in 2010-2013 included studies on the effectiveness of the methods and norms of sowing the sorghum, required quantities of mineral fertilizers to increase the crop yields and nutritional value of sorghum sown in the irrigated lowland areas of Dagestan. Methods. We conducted three field researches. In experiments with grain sorghum (the middle ripening group Zernogradskiy 88 we studied drill and broad-cast methods of sowing, seeding rate, the calculated doses of mineral fertilizers on programmable levels of crop yields: 6 t/ha (N160P112K70, 7 t/ha (N190P128K80 and 8 t/ha (N220P144K90. Seeding rate was 300, 350 and 400 thousand viable seeds per 1 ha; broadcast was chosen as a sowing method.A field experiment with sweet sorghum included promising hybrid crop Debut, fertilizers N140P80K70, N190P110K95 and N240P140K120 to obtain 60, 70 and 80 t/ha of green mass for two mowings, respectively. Results. The use of fertilizers based on a given level of productivity at optimum plant population can significantly improve the nutritional regime of the soil during the growing season of the sweet sorghum and create optimal conditions for nitrogen, phosphorus and potassium security for the crops and thus obtain the planned crop yield. Conclusion. The fodder quality of sweet sorghum varies depending on the nutrient status of the soil and mowing time.

  3. Cannabidiol Reduces Leukemic Cell Size – But Is It Important?

    Science.gov (United States)

    Kalenderoglou, Nikoletta; Macpherson, Tara; Wright, Karen L.

    2017-01-01

    The anti-cancer effect of the plant-derived cannabinoid, cannabidiol, has been widely demonstrated both in vivo and in vitro. However, this body of preclinical work has not been translated into clinical use. Key issues around this failure can be related to narrow dose effects, the cell model used and incomplete efficacy. A model of acute lymphoblastic disease, the Jurkat T cell line, has been used extensively to study the cannabinoid system in the immune system and cannabinoid-induced apoptosis. Using these cells, this study sought to investigate the outcome of those remaining viable cells post-treatment with cannabidiol, both in terms of cell size and tracking any subsequent recovery. The phosphorylation status of the mammalian Target of Rapamycin (mTOR) signaling pathway and the downstream target ribosomal protein S6, were measured. The ability of cannabidiol to exert its effect on cell viability was also evaluated in physiological oxygen conditions. Cannabidiol reduced cell viability incompletely, and slowed the cell cycle with fewer cells in the G2/M phase of the cell cycle. Cannabidiol reduced phosphorylation of mTOR, PKB and S6 pathways related to survival and cell size. The remaining population of viable cells that were cultured in nutrient rich conditions post-treatment were able to proliferate, but did not recover to control cell numbers. However, the proportion of viable cells that were gated as small, increased in response to cannabidiol and normally sized cells decreased. This proportion of small cells persisted in the recovery period and did not return to basal levels. Finally, cells grown in 12% oxygen (physiological normoxia) were more resistant to cannabidiol. In conclusion, these results indicate that cannabidiol causes a reduction in cell size, which persists post-treatment. However, resistance to cannabidiol under physiological normoxia for these cells would imply that cannabidiol may not be useful in the clinic as an anti-leukemic agent. PMID

  4. Simulations of DSB Yields and Radiation-induced Chromosomal Aberrations in Human Cells Based on the Stochastic Track Structure Induced by HZE Particles

    Science.gov (United States)

    Ponomarev, Artem; Plante, Ianik; George, Kerry; Wu, Honglu

    2014-01-01

    The formation of double-strand breaks (DSBs) and chromosomal aberrations (CAs) is of great importance in radiation research and, specifically, in space applications. We are presenting a new particle track and DNA damage model, in which the particle stochastic track structure is combined with the random walk (RW) structure of chromosomes in a cell nucleus. The motivation for this effort stems from the fact that the model with the RW chromosomes, NASARTI (NASA radiation track image) previously relied on amorphous track structure, while the stochastic track structure model RITRACKS (Relativistic Ion Tracks) was focused on more microscopic targets than the entire genome. We have combined chromosomes simulated by RWs with stochastic track structure, which uses nanoscopic dose calculations performed with the Monte-Carlo simulation by RITRACKS in a voxelized space. The new simulations produce the number of DSBs as function of dose and particle fluence for high-energy particles, including iron, carbon and protons, using voxels of 20 nm dimension. The combined model also calculates yields of radiation-induced CAs and unrejoined chromosome breaks in normal and repair deficient cells. The joined computational model is calibrated using the relative frequencies and distributions of chromosomal aberrations reported in the literature. The model considers fractionated deposition of energy to approximate dose rates of the space flight environment. The joined model also predicts of the yields and sizes of translocations, dicentrics, rings, and more complex-type aberrations formed in the G0/G1 cell cycle phase during the first cell division after irradiation. We found that the main advantage of the joined model is our ability to simulate small doses: 0.05-0.5 Gy. At such low doses, the stochastic track structure proved to be indispensable, as the action of individual delta-rays becomes more important.

  5. Increasing production yield of tyrosine and mevalonate through inhibition of biomass formation

    DEFF Research Database (Denmark)

    Li, Songyuan; Jendresen, Christian Bille; Nielsen, Alex Toftgaard

    2016-01-01

    , in particular, resulted in an increase in mass yield of mevalonate and tyrosine by 80% and 50%, respectively. By tracking production and biomass concentrations, it was observed that the production was maintained for more than 10 h after inhibition of cell growth, despite cell maintenance requirements...

  6. Current Perspectives on Viable but Non-culturable State in Foodborne Pathogens

    OpenAIRE

    Zhao, Xihong; Zhong, Junliang; Wei, Caijiao; Lin, Chii-Wann; Ding, Tian

    2017-01-01

    The viable but non-culturable (VBNC) state, a unique state in which a number of bacteria respond to adverse circumstances, was first discovered in 1982. Unfortunately, it has been reported that many foodborne pathogens can be induced to enter the VBNC state by the limiting environmental conditions during food processing and preservation, such as extreme temperatures, drying, irradiation, pulsed electric field, and high pressure stress, as well as the addition of preservatives and disinfectant...

  7. Improvement in Saccharification Yield of Mixed Rumen Enzymes by Identification of Recalcitrant Cell Wall Constituents Using Enzyme Fingerprinting.

    Science.gov (United States)

    Badhan, Ajay; Wang, Yu-Xi; Gruninger, Robert; Patton, Donald; Powlowski, Justin; Tsang, Adrian; McAllister, Tim A

    2015-01-01

    Identification of recalcitrant factors that limit digestion of forages and the development of enzymatic approaches that improve hydrolysis could play a key role in improving the efficiency of meat and milk production in ruminants. Enzyme fingerprinting of barley silage fed to heifers and total tract indigestible fibre residue (TIFR) collected from feces was used to identify cell wall components resistant to total tract digestion. Enzyme fingerprinting results identified acetyl xylan esterases as key to the enhanced ruminal digestion. FTIR analysis also suggested cross-link cell wall polymers as principal components of indigested fiber residues in feces. Based on structural information from enzymatic fingerprinting and FTIR, enzyme pretreatment to enhance glucose yield from barley straw and alfalfa hay upon exposure to mixed rumen-enzymes was developed. Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design. Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes. The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

  8. Corrigendum to "Sinusoidal potential cycling operation of a direct ethanol fuel cell to improving carbon dioxide yields" [J. Power Sources 268 (5 December 2014) 439-442

    Science.gov (United States)

    Majidi, Pasha; Pickup, Peter G.

    2016-09-01

    The authors regret that Equation (5) is incorrect and has resulted in errors in Fig. 4 and the efficiencies stated on p. 442. The corrected equation, figure and text are presented below. In addition, the title should be 'Sinusoidal potential cycling operation of a direct ethanol fuel cell to improve carbon dioxide yields', and the reversible cell potential quoted on p. 441 should be 1.14 V. The authors would like to apologise for any inconvenience caused.

  9. Absolute quantum yield measurements for the formation of oxygen ...

    Indian Academy of Sciences (India)

    Unknown

    Abstract. The dynamics of formation of oxygen atoms after UV photoexcitation of .... The SO2 pressure in the cell was typically 30–55 mTorr (monitored by an MKS .... With this value the quantum yield for O(3P) formation could be calculated to.

  10. Immediate natural tooth pontic: A viable yet temporary prosthetic solution: A patient reported outcome

    Directory of Open Access Journals (Sweden)

    Sudhir Bhandari

    2012-01-01

    Conclusion: The concept of immediate pontic placement is surely a viable treatment option and promises an excellent transient esthetic solution for a lost tooth as well as enables good preparation of the extraction site for future prosthetic replacement.

  11. Three-dimensional printing of stem cell-laden hydrogels submerged in a hydrophobic high-density fluid

    International Nuclear Information System (INIS)

    Duarte Campos, Daniela F; Blaeser, Andreas; Weber, Michael; Fischer, Horst; Jäkel, Jörg; Neuss, Sabine; Jahnen-Dechent, Wilhelm

    2013-01-01

    Over the last decade, bioprinting technologies have begun providing important tissue engineering strategies for regenerative medicine and organ transplantation. The major drawback of past approaches has been poor or inadequate material-printing device and substrate combinations, as well as the relatively small size of the printed construct. Here, we hypothesise that cell-laden hydrogels can be printed when submerged in perfluorotributylamine (C 12 F 27 N), a hydrophobic high-density fluid, and that these cells placed within three-dimensional constructs remain viable allowing for cell proliferation and production of extracellular matrix. Human mesenchymal stem cells and MG-63 cells were encapsulated into agarose hydrogels, and subsequently printed in high aspect ratio in three dimensional structures that were supported in high density fluorocarbon. Three-dimensional structures with various shapes and sizes were manufactured and remained stable for more than six months. Live/dead and DAPI stainings showed viable cells 24 h after the printing process, as well as after 21 days in culture. Histological and immunohistochemical analyses after 14 and 21 days revealed viable cells with marked matrix production and signs of proliferation. The compressive strength values of the printed gels consequently increased during the two weeks in culture, revealing encouraging results for future applications in regenerative medicine. (paper)

  12. Neutrophil-mediated protection of cultured human vascular endothelial cells from damage by growing Candida albicans hyphae

    International Nuclear Information System (INIS)

    Edwards, J.E. Jr.; Rotrosen, D.; Fontaine, J.W.; Haudenschild, C.C.; Diamond, R.D.

    1987-01-01

    Interactions were studied between human neutrophils and cultured human umbilical vein endothelial cells invaded by Candida albicans. In the absence of neutrophils, progressive Candida germination and hyphal growth extensively damaged endothelial cell monolayers over a period of 4 to 6 hours, as determined both by morphological changes and release of 51 Cr from radiolabeled endothelial cells. Monolayers were completely destroyed and replaced by hyphae after 18 hours of incubation. In contrast, when added 2 hours after the monolayers had been infected with Candida, neutrophils selectively migrated toward and attached to hyphae at points of hyphal penetration into individual endothelial cells (observed by time-lapse video-microscopy). Attached neutrophils spread over hyphal surfaces both within and beneath the endothelial cells; neutrophil recruitment to initial sites of leukocyte-Candida-endothelial cell interactions continued throughout the first 60 minutes of observation. Neutrophil spreading and stasis were observed only along Candida hyphae and at sites of Candida-endothelial cell interactions. These events resulted in 58.0% killing of Candida at 2 hours and subsequent clearance of Candida from endothelial cell monolayers, as determined by microcolony counts and morphological observation. On introduction of additional neutrophils to yield higher ratios of neutrophils to endothelial cells (10 neutrophils:1 endothelial cell), neutrophil migration toward hyphal elements continued. Despite retraction or displacement of occasional endothelial cells by invading Candida and neutrophils, most endothelial cells remained intact, viable, and motile as verified both by morphological observations and measurement of 51 Cr release from radiolabeled monolayers

  13. Gamma-irradiated scrub typhus immunogens: development of cell-mediated immunity after vaccination of inbred mice

    International Nuclear Information System (INIS)

    Jerrells, T.R.; Palmer, B.A.; Osterman, J.V.

    1983-01-01

    Mice immunized with three injections of gamma-irradiated Karp strain of Rickettsia tsutsugamushi were evaluated for the presence of cell-mediated immunity by using delayed-type hypersensitivity, antigen-induced lymphocyte proliferation, and antigen-induced lymphokine production. These animals also were evaluated for levels of circulating antibody after immunization as well as for the presence of rickettsemia after intraperitoneal challenge with viable Karp rickettsiae. After immunization with irradiated Karp rickettsiae, a demonstrable cell-mediated immunity was present as evidenced by delayed-type hypersensitivity responsiveness, lymphocyte proliferation, and production of migration inhibition factor and interferon by immune spleen lymphocytes. Also, a reduction in circulating rickettsiae was seen in mice immunized with irradiated rickettsiae after challenge with 1,000 50% mouse lethal doses of viable, homologous rickettsiae. All responses except antibody titer and reduction of rickettsemia were similar to the responses noted in mice immunized with viable organisms. Antibody levels were lower in mice immunized with irradiated rickettsiae than in mice immunized with viable rickettsiae. Furthermore, mice that were immunized with viable rickettsiae demonstrated markedly lower levels of rickettsemia after intraperitoneal challenge compared with either mice immunized with irradiated rickettsiae or nonimmunized mice

  14. Short-term effects of milking frequency on milk yield, milk composition, somatic cell count and milk protein profile in dairy goats

    DEFF Research Database (Denmark)

    Torres, Alexandr; Hernandez Castellano, Lorenzo E; Morales-delaNuez, Antonio

    2014-01-01

    Goats in Canary Islands are milked once a day by tradition, but in most countries with high technology on farms, goats are milked twice a day, which is known to improve milk yield. Therefore it is important to know whether the increase of milking frequency can improve the production without impai...... was returned to X2 and X1. Finally, quantitative analysis showed an increase in intensities of milk protein bands from X1 to X2, but the intensities of casein bands (αS1-CN, αS2-CN, β-CN, κ-CN) and major whey proteins (α-La, β-Lg) decreased from X2 to X3.......Goats in Canary Islands are milked once a day by tradition, but in most countries with high technology on farms, goats are milked twice a day, which is known to improve milk yield. Therefore it is important to know whether the increase of milking frequency can improve the production without...... impairing milk quality. The objective of this study was to investigate the short term effects of three milking frequencies on milk yield, milk composition, somatic cell count (SCC) and milk protein profile in dairy goats traditionally milked once a day. Twelve Majorera goats in early lactation (48±4 d...

  15. Term amniotic fluid: an unexploited reserve of mesenchymal stromal cells for reprogramming and potential cell therapy applications.

    Science.gov (United States)

    Moraghebi, Roksana; Kirkeby, Agnete; Chaves, Patricia; Rönn, Roger E; Sitnicka, Ewa; Parmar, Malin; Larsson, Marcus; Herbst, Andreas; Woods, Niels-Bjarne

    2017-08-25

    Mesenchymal stromal cells (MSCs) are currently being evaluated in numerous pre-clinical and clinical cell-based therapy studies. Furthermore, there is an increasing interest in exploring alternative uses of these cells in disease modelling, pharmaceutical screening, and regenerative medicine by applying reprogramming technologies. However, the limited availability of MSCs from various sources restricts their use. Term amniotic fluid has been proposed as an alternative source of MSCs. Previously, only low volumes of term fluid and its cellular constituents have been collected, and current knowledge of the MSCs derived from this fluid is limited. In this study, we collected amniotic fluid at term using a novel collection system and evaluated amniotic fluid MSC content and their characteristics, including their feasibility to undergo cellular reprogramming. Amniotic fluid was collected at term caesarean section deliveries using a closed catheter-based system. Following fluid processing, amniotic fluid was assessed for cellularity, MSC frequency, in-vitro proliferation, surface phenotype, differentiation, and gene expression characteristics. Cells were also reprogrammed to the pluripotent stem cell state and differentiated towards neural and haematopoietic lineages. The average volume of term amniotic fluid collected was approximately 0.4 litres per donor, containing an average of 7 million viable mononuclear cells per litre, and a CFU-F content of 15 per 100,000 MNCs. Expanded CFU-F cultures showed similar surface phenotype, differentiation potential, and gene expression characteristics to MSCs isolated from traditional sources, and showed extensive expansion potential and rapid doubling times. Given the high proliferation rates of these neonatal source cells, we assessed them in a reprogramming application, where the derived induced pluripotent stem cells showed multigerm layer lineage differentiation potential. The potentially large donor base from caesarean section

  16. CRISPR-Mediated Genomic Deletion of Sox2 in the Axolotl Shows a Requirement in Spinal Cord Neural Stem Cell Amplification during Tail Regeneration

    Directory of Open Access Journals (Sweden)

    Ji-Feng Fei

    2014-09-01

    Full Text Available The salamander is the only tetrapod that functionally regenerates all cell types of the limb and spinal cord (SC and thus represents an important regeneration model, but the lack of gene-knockout technology has limited molecular analysis. We compared transcriptional activator-like effector nucleases (TALENs and clustered regularly interspaced short palindromic repeats (CRISPRs in the knockout of three loci in the axolotl and find that CRISPRs show highly penetrant knockout with less toxic effects compared to TALENs. Deletion of Sox2 in up to 100% of cells yielded viable F0 larvae with normal SC organization and ependymoglial cell marker expression such as GFAP and ZO-1. However, upon tail amputation, neural stem cell proliferation was inhibited, resulting in spinal-cord-specific regeneration failure. In contrast, the mesodermal blastema formed normally. Sox3 expression during development, but not regeneration, most likely allowed embryonic survival and the regeneration-specific phenotype. This analysis represents the first tissue-specific regeneration phenotype from the genomic deletion of a gene in the axolotl.

  17. Yield, yield components and dry matter digestibility of alfalfa experimental populations

    Directory of Open Access Journals (Sweden)

    Katić Slobodan

    2010-01-01

    Full Text Available Alfalfa is the most important forage crop grown in the temperate regions. It is cultivated for production of vegetative aerial mass used fresh or as hay, and recently as haylage and silage. In many centres worldwide, efforts are made to breed and create new alfalfa cultivars with both higher yields and of higher nutritional value. The aim of this paper was to determine yield and digestibility of 12 experimental populations of alfalfa, and to compare their results to the yields of well-known domestic alfalfa commercial cultivars. The results show significant differences in yield of green forage and dry matter among alfalfa populations, as well as in yield components, height, proportion of leaves in yield and growth rate (tab. 1, 2 and 3. Differences between in vitro digestible dry matter (% and yields of in vitro digestible dry matter (t ha-1 were also significant (tab. 5 and 6. Yield and quality of experimental populations were at the same level or higher than of control cultivars. Synthetic SINUSA exceeded the control cutivars (NS Mediana ZMS V and Banat VS in yield and quality of dry matter. .

  18. Slope Controls Grain Yield and Climatic Yield in Mountainous Yunnan province, China

    Science.gov (United States)

    Duan, X.; Rong, L.; Gu, Z.; Feng, D.

    2017-12-01

    Mountainous regions are increasingly vulnerable to food insecurity because of limited arable land, growing population pressure, and climate change. Development of sustainable mountain agriculture will require an increased understanding of the effects of environmental factors on grain and climatic yields. The objective of this study was to explore the relationships between actual grain yield, climatic yield, and environmental factors in a mountainous region in China. We collected data on the average grain yield per unit area in 119 counties in Yunnan province from 1985 to 2012, and chose 17 environmental factors for the same period. Our results showed that actual grain yield ranged from 1.43 to 6.92 t·ha-1, and the climatic yield ranged from -0.15 to -0.01 t·ha-1. Lower climatic yield but higher grain yield was generally found in central areas and at lower slopes and elevations in the western and southwestern counties of Yunnan province. Higher climatic yield but lower grain yield were found in northwestern parts of Yunnan province on steep slopes. Annual precipation and temperature had a weak influence on the climatic yield. Slope explained 44.62 and 26.29% of the variation in grain yield and climatic yield. The effects of topography on grain and climatic yields were greater than climatic factors. Slope was the most important environmental variable for the variability in climatic and grain yields in the mountainous Yunnan province due to the highly heterogeneous topographic conditions. Conversion of slopes to terraces in areas with higher climatic yields is an effective way to maintain grain production in response to climate variability. Additionally, soil amendments and soil and water conservation measures should be considered to maintain soil fertility and aid in sustainable development in central areas, and in counties at lower slopes and elevations in western and southwestern Yunnan province.

  19. A Transient Cell-shielding Method for Viable MSC Delivery Within Hydrophobic Scaffolds Polymerized in situ

    Science.gov (United States)

    2015-03-27

    Transport phenomena. New York: Wiley; 1960. [52] Caplan AI, Dennis JE. Mesenchymal stem cells as trophic mediators. J Cell Biochem 2006;98:1076e84. [53... Caplan AI. Why are MSCs therapeutic? New data: new insight. J Pathol 2009;217:318e24. [54] Dennis JE, Cohen N, Goldberg VM, Caplan AI. Targeted delivery

  20. Massachusetts Fuel Cell Bus Project: Demonstrating a Total Transit Solution for Fuel Cell Electric Buses in Boston

    Energy Technology Data Exchange (ETDEWEB)

    2017-05-22

    The Federal Transit Administration's National Fuel Cell Bus Program focuses on developing commercially viable fuel cell bus technologies. Nuvera is leading the Massachusetts Fuel Cell Bus project to demonstrate a complete transit solution for fuel cell electric buses that includes one bus and an on-site hydrogen generation station for the Massachusetts Bay Transportation Authority (MBTA). A team consisting of ElDorado National, BAE Systems, and Ballard Power Systems built the fuel cell electric bus, and Nuvera is providing its PowerTap on-site hydrogen generator to provide fuel for the bus.

  1. Yield trends and yield gap analysis of major crops in the world

    NARCIS (Netherlands)

    Hengsdijk, H.; Langeveld, J.W.A.

    2009-01-01

    This study aims to quantify the gap between current and potential yields of major crops in the world, and the production constraints that contribute to this yield gap. Using an expert-based evaluation of yield gaps and the literature, global and regional yields and yield trends of major crops are

  2. Adipose-Derived Stem Cell Delivery into Collagen Gels Using Chitosan Microspheres

    Science.gov (United States)

    2010-02-17

    Porous CSM of uniform size and composition were prepared and used as a stem cell carrier. ASC were allowed to attach to the microspheres and infiltrate...and viable, could be retrieved from the spheres, and maintained expression of stem - cell -specific markers. Electron microscopic evaluation of the cell

  3. The influence of day/night cycles on biomass yield and composition of Neochloris oleoabundans.

    Science.gov (United States)

    de Winter, Lenneke; Cabanelas, Iago Teles Dominguez; Martens, Dirk E; Wijffels, René H; Barbosa, Maria J

    2017-01-01

    Day/night cycles regulate the circadian clock of organisms to program daily activities. Many species of microalgae have a synchronized cell division when grown under a day/night cycle, and synchronization might influence biomass yield and composition. Therefore, the aim of this study was to study the influence of day/night cycle on biomass yield and composition of the green microalgae Neochloris oleoabundans . Hence, we compared continuous turbidostat cultures grown under continuous light with cultures grown under simulated day/night cycles. Under day/night cycles, cultures were synchronized as cell division was scheduled in the night, whereas under continuous light cell division occurred randomly synchronized cultures were able to use the light 10-15% more efficiently than non-synchronized cultures. Our results indicate that the efficiency of light use varies over the cell cycle and that synchronized cell division provides a fitness benefit to microalgae. Biomass composition under day/night cycles was similar to continuous light, with the exception of starch content. The starch content was higher in cultures under continuous light, most likely because the cells never had to respire starch to cover for maintenance during dark periods. Day/night cycles were provided in a 'block' (continuous light intensity during the light period) and in a 'sine' (using a sine function to simulate light intensities from sunrise to sunset). There were no differences in biomass yield or composition between these two ways of providing light (in a 'block' or in a 'sine'). The biomass yield and composition of N. oleoabundans were influenced by day/night cycles. These results are important to better understand the relations between research done under continuous light conditions and with day/night cycle conditions. Our findings also imply that more research should be done under day/night cycles.

  4. Fluorescence Quantum Yield Measurements of Fluorescent Proteins: A Laboratory Experiment for a Biochemistry or Molecular Biophysics Laboratory Course

    Science.gov (United States)

    Wall, Kathryn P.; Dillon, Rebecca; Knowles, Michelle K.

    2015-01-01

    Fluorescent proteins are commonly used in cell biology to assess where proteins are within a cell as a function of time and provide insight into intracellular protein function. However, the usefulness of a fluorescent protein depends directly on the quantum yield. The quantum yield relates the efficiency at which a fluorescent molecule converts…

  5. Successful transportation of human corneal endothelial tissues without cool preservation in varying Indian tropical climatic conditions and in vitro cell expansion using a novel polymer.

    Science.gov (United States)

    Rao, Srinivas K; Sudhakar, John; Parikumar, Periyasamy; Natarajan, Sundaram; Insaan, Aditya; Yoshioka, Hiroshi; Mori, Yuichi; Tsukahara, Shigeo; Baskar, Subramani; Manjunath, Sadananda Rao; Senthilkumar, Rajappa; Thamaraikannan, Paramasivam; Srinivasan, Thangavelu; Preethy, Senthilkumar; Abraham, Samuel J K

    2014-02-01

    Though the transplantation of human corneal endothelial tissue (CET) separated from cadaver cornea is in practice, its transportation has not been reported. We report the successful transportation of CET in varying Indian climatic conditions without cool preservation and the in vitro expansion of Human Corneal Endothelial Precursor Cells (HCEPCs) using a novel Thermo-reversible gelation polymer (TGP). CET from cadaver corneas (n = 67), unsuitable for transplantation, were used. In phase I, CET was transported in Basal Culture Medium (Group I) and TGP (Group II) and in Phase II, in TGP cocktail alone, from three hospitals 250-2500 km away, to a central laboratory. The transportation time ranged from 6 h to 72 h and the outdoor temperature between 20°C and 41°C. On arrival, CET were processed, cells were expanded upto 30 days in basal culture medium (Group A) and TGP scaffold (Group B). Cell viability and morphology were documented and Reverse transcription polymerase chain reaction (RT-PCR) characterization undertaken. In Phase I, TGP yielded more viable cells (0.11 × 10(6) cells) than Group I (0.04 × 10(6) cells). In Phase II, the average cell count was 5.44 × 10(4) cells. During expansion, viability of HCEPCs spheres in TGP was maintained for a longer duration. The cells from both the groups tested positive for B-3 tubulin and negative for cytokeratins K3 and K12, thereby proving them to be HCEPCs. TGP preserves the CET during transportation without cool preservation and supports in vitro expansion, with a higher yield of HCEPCs, similar to that reported in clinical studies.

  6. Successful transportation of human corneal endothelial tissues without cool preservation in varying Indian tropical climatic conditions and in vitro cell expansion using a novel polymer

    Directory of Open Access Journals (Sweden)

    Srinivas K Rao

    2014-01-01

    Full Text Available Background: Though the transplantation of human corneal endothelial tissue (CET separated from cadaver cornea is in practice, its transportation has not been reported. We report the successful transportation of CET in varying Indian climatic conditions without cool preservation and the in vitro expansion of Human Corneal Endothelial Precursor Cells (HCEPCs using a novel Thermo-reversible gelation polymer (TGP. Materials and Methods: CET from cadaver corneas (n = 67, unsuitable for transplantation, were used. In phase I, CET was transported in Basal Culture Medium (Group I and TGP (Group II and in Phase II, in TGP cocktail alone, from three hospitals 250-2500 km away, to a central laboratory. The transportation time ranged from 6 h to 72 h and the outdoor temperature between 20°C and 41°C. On arrival, CET were processed, cells were expanded upto 30 days in basal culture medium (Group A and TGP scaffold (Group B. Cell viability and morphology were documented and Reverse transcription polymerase chain reaction (RT-PCR characterization undertaken. Results: In Phase I, TGP yielded more viable cells (0.11 × 10 6 cells than Group I (0.04 × 10 6 cells. In Phase II, the average cell count was 5.44 × 10 4 cells. During expansion, viability of HCEPCs spheres in TGP was maintained for a longer duration. The cells from both the groups tested positive for B-3 tubulin and negative for cytokeratins K3 and K12, thereby proving them to be HCEPCs. Conclusion: TGP preserves the CET during transportation without cool preservation and supports in vitro expansion, with a higher yield of HCEPCs, similar to that reported in clinical studies.

  7. Cell recycle batch fermentation of high-solid lignocellulose using a recombinant cellulase-displaying yeast strain for high yield ethanol production in consolidated bioprocessing.

    Science.gov (United States)

    Matano, Yuki; Hasunuma, Tomohisa; Kondo, Akihiko

    2013-05-01

    The aim of this study is to develop a scheme of cell recycle batch fermentation (CRBF) of high-solid lignocellulosic materials. Two-phase separation consisting of rough removal of lignocellulosic residues by low-speed centrifugation and solid-liquid separation enabled effective collection of Saccharomyces cerevisiae cells with decreased lignin and ash. Five consecutive batch fermentation of 200 g/L rice straw hydrothermally pretreated led to an average ethanol titer of 34.5 g/L. Moreover, the display of cellulases on the recombinant yeast cell surface increased ethanol titer to 42.2 g/L. After, five-cycle fermentation, only 3.3 g/L sugar was retained in the fermentation medium, because cellulase displayed on the cell surface hydrolyzed cellulose that was not hydrolyzed by commercial cellulases or free secreted cellulases. Fermentation ability of the recombinant strain was successfully kept during a five-cycle repeated batch fermentation with 86.3% of theoretical yield based on starting biomass. Copyright © 2012 Elsevier Ltd. All rights reserved.

  8. Viable Intrauterine Pregnancy and Coexisting Molar Pregnancy in a Bicornuate Uterus: A Rare Presentation

    Directory of Open Access Journals (Sweden)

    Kavitha Krishnamoorthy

    2016-01-01

    Full Text Available A complete hydatidiform mole with a viable coexisting fetus (CMCF is a rare occurrence. Similarly, Mullerian anomalies such as a bicornuate uterus are uncommon variants of normal anatomy. We report a case of a 40-year-old female with a known bicornuate uterus presenting at 13 weeks gestation with vaginal bleeding. Ultrasound findings showed a healthy viable pregnancy in the right horn with complete molar pregnancy in the left horn. After extensive counseling, the patient desired conservative management, however, was unable to continue due to profuse vaginal bleeding. The patient underwent suction dilation and curettage under general anesthesia and evacuation of the uterine horns. Postoperatively, the patient was followed until serum beta-human chorionic gonadotropin (β-hCG level dropped to <5 mU. This is the first case of a CMCF reported in a bicornuate uterus, diagnosed with the use of ultrasound imaging.

  9. On yield gaps and yield gains in intercropping

    NARCIS (Netherlands)

    Gou, Fang; Yin, Wen; Hong, Yu; Werf, van der Wopke; Chai, Qiang; Heerink, Nico; Ittersum, van Martin K.

    2017-01-01

    Wheat-maize relay intercropping has been widely used by farmers in northwest China, and based on field experiments agronomists report it has a higher productivity than sole crops. However, the yields from farmers’ fields have not been investigated yet. Yield gap analysis provides a framework to

  10. Gastrulation and the establishment of the three germ layers in the early horse conceptus.

    NARCIS (Netherlands)

    Gaivão, M.M.F.; Rambags, B.P.B.; Stout, Tom

    2014-01-01

    Experimental studies and field surveys suggest that embryonic loss during the first 6 weeks of gestation is a common occurrence in the mare. During the first 2 weeks of development, a number of important cell differentiation events must occur to yield a viable embryo proper containing all three

  11. A viable logarithmic f(R) model for inflation

    Energy Technology Data Exchange (ETDEWEB)

    Amin, M.; Khalil, S. [Center for Fundamental Physics, Zewail City of Science and Technology,6 October City, Giza (Egypt); Salah, M. [Center for Fundamental Physics, Zewail City of Science and Technology,6 October City, Giza (Egypt); Department of Mathematics, Faculty of Science, Cairo University,Giza (Egypt)

    2016-08-18

    Inflation in the framework of f(R) modified gravity is revisited. We study the conditions that f(R) should satisfy in order to lead to a viable inflationary model in the original form and in the Einstein frame. Based on these criteria we propose a new logarithmic model as a potential candidate for f(R) theories aiming to describe inflation consistent with observations from Planck satellite (2015). The model predicts scalar spectral index 0.9615

  12. Zn(II)-cyclam based chromogenic sensors for recognition of ATP in aqueous solution under physiological conditions and their application as viable staining agents for microorganism.

    Science.gov (United States)

    Mahato, Prasenjit; Ghosh, Amrita; Mishra, Sanjiv K; Shrivastav, Anupama; Mishra, Sandhya; Das, Amitava

    2011-05-02

    Two chromogenic complexes, L.Zn (where L is (E)-4-((4-(1,4,8,11-tetraazacyclotetradecan-1-ylsulfonyl)phenyl)diazenyl)-N,N-dimethylaniline) and its [2]pseudorotaxane form (α-CD.L.Zn), were found to bind preferentially to adenosine triphosphate (ATP), among all other common anions and biologically important phosphate (AMP, ADP, pyrophosphate, and phosphate) ions in aqueous HEPES buffer medium of pH 7.2. Studies with live cell cultures of prokaryotic microbes revealed that binding of these two reagents to intercellular ATP, produced in situ, could be used in delineating the gram-positive and the gram-negative bacteria. More importantly, these dyes were found to be nontoxic to living microbes (eukaryotes and prokaryotes) and could be used for studying the cell growth dynamics. Binding to these two viable staining agents to intercellular ATP was also confirmed by spectroscopic studies on cell growth in the presence of different respiratory inhibitors that influence the intercellular ATP generation. © 2011 American Chemical Society

  13. High-yield production of herbicidal thaxtomins and analogs in a nonpathogenic Streptomyces strain.

    Science.gov (United States)

    Jiang, Guangde; Zhang, Yucheng; Powell, Magan M; Zhang, Peilan; Zuo, Ran; Zhang, Yi; Kallifidas, Dimitrios; Tieu, Albert M; Luesch, Hendrik; Loria, Rosemary; Ding, Yousong

    2018-03-30

    forward in its wide agricultural use. Furthermore, current synthetic routes to thaxtomins and analogs are lengthy, and two thaxtomin biosynthetic intermediates produced at high yields in this work can provide precursors and building blocks to advanced synthetic routes. Importantly, the production of 5-F-thaxtomin A in engineered S. albus J1074 demonstrated a viable alternative to chemical methods in the synthesis of new thaxtomin analogs. Moreover, our work presents an attractive synthetic biology strategy to improve the supply of herbicidal thaxtomins, likely finding general applications in the discovery and production of many other bioactive natural products. Copyright © 2018 American Society for Microbiology.

  14. Virulence of thermolable haemolysi tlh, gastroenteritis related pathogenicity tdh and trh of the pathogens Vibrio Parahemolyticus in Viable but Non-Culturable (VBNC) state.

    Science.gov (United States)

    Zhong, Huamin; Zhong, Yukui; Deng, Qiulian; Zhou, Zhenwen; Guan, Xiaoshan; Yan, Muxia; Hu, Tingting; Luo, Mingyong

    2017-10-01

    In the Viable but Non-Culturable (VBNC) state, microorganisms may survive under severe external environment. In this study, the specificity and sensitivity of PMA-LAMP assay on the detection of Vibrio Parahemolyticus (V. parahemolyticus) has been developed and evaluated, with further application on a number of food-borne V. parahemolyticus strains. Six primers were designed for recognizing 8 distinct targeting on tlh, tdh and trh gene. Through specific penetration through the damaged cell membrane of dead cells and intercalating into DNA, PMA could prevent DNA amplification of dead bacteria from LAMP, which enabled the differentiation of bacteria between VBNC state and dead state. The established PMA-LAMP showed significant advantage in rapidity, sensitivity and specificity, compared with regular PCR assay. The applicability had also been verified, demonstrating the PMA-LAMP was capable of detection on V. parahaemolyticus. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Optimization of Culture Medium Enhances Viable Biomass Production and Biocontrol Efficacy of the Antagonistic Yeast, Candida diversa

    Directory of Open Access Journals (Sweden)

    Jia Liu

    2017-10-01

    Full Text Available Viable biomass production is a key determinant of suitability of antagonistic yeasts as potential biocontrol agents. This study investigated the effects of three metal ions (magnesium, ferrous, and zinc on biomass production and viability of the antagonistic yeast, Candida diversa. Using response surface methodology to optimize medium components, a maximum biomass was obtained, when the collective Mg2+, Fe2+, and Zn2+ concentrations were adjusted in a minimal mineral (MM medium. Compared with the unmodified MM, and three ion-deficient MM media, yeast cells cultured in the three ion-modified MM medium exhibited a lower level of cellular oxidative damage, and a higher level of antioxidant enzyme activity. A biocontrol assay indicated that C. diversa grown in the ion-modified MM exhibited the greatest level of control of gray mold on apple fruit. These results provide new information on culture medium optimization to grow yeast antagonists in order to improve biomass production and biocontrol efficacy.

  16. Soft sensor for monitoring biomass subpopulations in mammalian cell culture processes.

    Science.gov (United States)

    Kroll, Paul; Stelzer, Ines V; Herwig, Christoph

    2017-11-01

    Biomass subpopulations in mammalian cell culture processes cause impurities and influence productivity, which requires this critical process parameter to be monitored in real-time. For this reason, a novel soft sensor concept for estimating viable, dead and lysed cell concentration was developed, based on the robust and cheap in situ measurements of permittivity and turbidity in combination with a simple model. It could be shown that the turbidity measurements contain information about all investigated biomass subpopulations. The novelty of the developed soft sensor is the real-time estimation of lysed cell concentration, which is directly correlated to process-related impurities such as DNA and host cell protein in the supernatant. Based on data generated by two fed-batch processes the developed soft sensor is described and discussed. The presented soft sensor concept provides a tool for viable, dead and lysed cell concentration estimation in real-time with adequate accuracy and enables further applications with respect to process optimization and control.

  17. Simulations of DSB Yields and Radiation-induced Chromosomal Aberrations in Human Cells Based on the Stochastic Track Structure iIduced by HZE Particles

    Science.gov (United States)

    Ponomarev, Artem; Plante, Ianik; George, Kerry; Wu, Honglu

    2014-01-01

    The formation of double-strand breaks (DSBs) and chromosomal aberrations (CAs) is of great importance in radiation research and, specifically, in space applications. We are presenting a new particle track and DNA damage model, in which the particle stochastic track structure is combined with the random walk (RW) structure of chromosomes in a cell nucleus. The motivation for this effort stems from the fact that the model with the RW chromosomes, NASARTI (NASA radiation track image) previously relied on amorphous track structure, while the stochastic track structure model RITRACKS (Relativistic Ion Tracks) was focused on more microscopic targets than the entire genome. We have combined chromosomes simulated by RWs with stochastic track structure, which uses nanoscopic dose calculations performed with the Monte-Carlo simulation by RITRACKS in a voxelized space. The new simulations produce the number of DSBs as function of dose and particle fluence for high-energy particles, including iron, carbon and protons, using voxels of 20 nm dimension. The combined model also calculates yields of radiation-induced CAs and unrejoined chromosome breaks in normal and repair deficient cells. The joined computational model is calibrated using the relative frequencies and distributions of chromosomal aberrations reported in the literature. The model considers fractionated deposition of energy to approximate dose rates of the space flight environment. The joined model also predicts of the yields and sizes of translocations, dicentrics, rings, and more complex-type aberrations formed in the G0/G1 cell cycle phase during the first cell division after irradiation. We found that the main advantage of the joined model is our ability to simulate small doses: 0.05-0.5 Gy. At such low doses, the stochastic track structure proved to be indispensable, as the action of individual delta-rays becomes more important.

  18. Radio-sensitivity of callus and cell cultures, and RAPD characterization of variants in banana [Musa spp.

    International Nuclear Information System (INIS)

    Kulkarni, V.M.; Karmarkar, V.M.; Ganapathi, T.R.; Bapat, V.A.

    2000-01-01

    Although bananas and plantains are one of the most important fruit crops, gearing up the breeding programmes for these has always remained the most difficult task due to several inherent problems such as parthenocarpy, barriers in obtaining viable seeds and long life cycle etc. In this regard, incorporation of in vitro techniques such as shoot-tip / cell cultures along with conventional as well as non-conventional methods of genetic improvement is of utmost importance, especially in those vegetatively propagated species with long crop cycle and low in vivo proliferation rate. In order to understand the radio-sensitivity, the callus and cell cultures of banana were exposed to differential doses of gamma-rays. Growth of the callus cultures reduced with increasing dose of gamma-rays. Similar trend was noticed in irradiation of cell suspensions also where a dose of 40 Gy and more was completely lethal. The experience gained from previous and present experiments has yielded optimization of the procedures for gamma-irradiation and subsequent handling of banana in vitro cultures. The RAPD analysis of the selected variants was unable to detect adequate polymorphism, and further experimentation in these regards is being done. (author)

  19. Preliminary stochastic model for managing Vibrio parahaemolyticus and total viable bacterial counts in a Pacific oyster (Crassostrea gigas) supply chain.

    Science.gov (United States)

    Fernandez-Piquer, Judith; Bowman, John P; Ross, Tom; Estrada-Flores, Silvia; Tamplin, Mark L

    2013-07-01

    Vibrio parahaemolyticus can accumulate and grow in oysters stored without refrigeration, representing a potential food safety risk. High temperatures during oyster storage can lead to an increase in total viable bacteria counts, decreasing product shelf life. Therefore, a predictive tool that allows the estimation of both V. parahaemolyticus populations and total viable bacteria counts in parallel is needed. A stochastic model was developed to quantitatively assess the populations of V. parahaemolyticus and total viable bacteria in Pacific oysters for six different supply chain scenarios. The stochastic model encompassed operations from oyster farms through consumers and was built using risk analysis software. Probabilistic distributions and predictions for the percentage of Pacific oysters containing V. parahaemolyticus and high levels of viable bacteria at the point of consumption were generated for each simulated scenario. This tool can provide valuable information about V. parahaemolyticus exposure and potential control measures and can help oyster companies and regulatory agencies evaluate the impact of product quality and safety during cold chain management. If coupled with suitable monitoring systems, such models could enable preemptive action to be taken to counteract unfavorable supply chain conditions.

  20. Antiproliferative activity of pristimerin isolated from Maytenus ilicifolia (Celastraceae) in human HL-60 cells.

    Science.gov (United States)

    Costa, Patricia Marçal da; Ferreira, Paulo Michel Pinheiro; Bolzani, Vanderlan da Silva; Furlan, Maysa; de Freitas Formenton Macedo Dos Santos, Vânia Aparecida; Corsino, Joaquim; de Moraes, Manoel Odorico; Costa-Lotufo, Letícia Veras; Montenegro, Raquel Carvalho; Pessoa, Cláudia

    2008-06-01

    Pristimerin has been shown to be cytotoxic to several cancer cell lines. In the present work, the cytotoxicity of pristimerin was evaluated in human tumor cell lines and in human peripheral blood mononuclear cells (PBMC). This work also examined the effects of pristimerin (0.4; 0.8 and 1.7 microM) in HL-60 cells, after 6, 12 and 24h of exposure. Pristimerin reduced the number of viable cells and increased number of non-viable cells in a concentration-dependent manner by tripan blue test showing morphological changes consistent with apoptosis. Nevertheless, pristimerin was not selective to cancer cells, since it inhibited PBMC proliferation with an IC50 of 0.88 microM. DNA synthesis inhibition assessed by 5-bromo-2'-deoxyuridine (BrdU) incorporation in HL-60 cells was 70% and 83% for the concentrations of 0.4 and 0.8 microM, respectively. Pristimerin (10 and 20 microM) was not able to inhibit topoisomerase I. In AO/EB (acridine orange/ethidium bromide) staining, all tested concentrations reduced the number of HL-60 viable cells, with the occurrence of necrosis and apoptosis in a concentration-dependent manner, results in agreement with trypan blue exclusion findings. The analysis of membrane integrity and internucleosomal DNA fragmentation by flow cytometry in the presence of pristimerin indicated that treated cells underwent apoptosis. The present data point to the importance of pristimerin as representative of an emerging class of potential anticancer chemicals, exhibiting an antiproliferative effect by inhibiting DNA synthesis and triggering apoptosis.

  1. The Impact of Location and Growing Medium on Harvest Yield and Flavor of Basil and Squash Microgreens

    Science.gov (United States)

    Leal, J.; Ventura, C. J.

    2016-12-01

    The purpose of this experiment is to discover the most efficient, feasible way to produce a high yield of flavorful microgreens within the best growing medium and building location at the Stanford Educational Farm. In recent years, microgreens, young, small edible greens utilized as flavor garnishes for fine dining, have evolved into an immensely profitable market that will continue to expand and prosper. To capitalize on such opportunities, the primary metrics focused upon are harvest yield and flavor of basil and squash microgreens, as they develop in different concentrations of quick root and compost while located in either a greenhouse or lath house. It was hypothesized that if basil and squash microgreens were grown in a mixture of 50% compost and 50% quick root in the greenhouse, then they would produce the greatest harvest yield and maximum amount of flavor. The general experimental protocol includes monitoring the growth of the microgreens, then harvesting directly after the first true leaves begin to emerge. Upon harvest, each set of microgreens are separated into different clear glass jars to place on a small scale for weighing to determine the yield and tasting the leaves to analyze the flavor content. The highest yield of basil (63 grams) developed in a tray of 100% quick root in the lath house, while the greatest yield of squash (51 grams) succeeded in 100% quick root in the greenhouse. Overall however, the basil grew fastest in the greenhouse but tasted stronger in the lath house. Additionally, because the harvest yield and flavor results were so poor in all other locations and growing mediums for squash during the first trial, it cannot be considered a viable microgreen. In the future, more trials should be conducted for greater numbers of trays of microgreens to collect more data. The nutritional value of microgreens should also be explored further to obtain a holistic approach to the value of these plants from seed to harvest to consumption.

  2. Building Viable and Sustainable Regional Netchains: Case Studies of Regional Pork Netchains in Spain, Germany, and The Netherlands

    Directory of Open Access Journals (Sweden)

    Rannia Nijhoff‐Savvaki

    2012-06-01

    Full Text Available The present paper aims to give insight into critical issues contributing to the successful building of viable and sustainable regional netchains, focussing on chain governance mechanisms (coordination mechanisms, quality management systems, information systems and social embedding. It uses three representative case studies from the regional pork sector in Spain, Germany, and The Netherlands, illustrating different trajectories to commercially viable and sustainable regional netchains. By analysing the cases on each critical aspect, the present paper describes important issues and proposes specific elements of further research, including on the roles for(nongovernmental organisations.

  3. A Viable Microbial Community in a Subglacial Volcanic Crater Lake, Iceland

    Science.gov (United States)

    Gaidos, Eric; Lanoil, Brian; Thorsteinsson, Thorsteinn; Graham, Andrew; Skidmore, Mark; Han, Suk-Kyun; Rust, Terri; Popp, Brian

    2004-09-01

    We describe a viable microbial community in a subglacial lake within the Grímsvötn volcanic caldera, Iceland. We used a hot water drill to penetrate the 300-m ice shelf and retrieved lake water and volcanic tephra sediments. We also acquired samples of borehole water before and after penetration to the lake, overlying glacial ice and snow, and water from a nearby subaerial geothermal lake for comparative analyses. Lake water is at the freezing point and fresh (total dissolved solids = 260 mg L-1). Detectable numbers of cells were found in samples of the lake water column and tephra sediments: 2 × 104 ml-1 and 4 × 107 g-1, respectively. Plate counts document abundant cold-adapted cultivable organisms in the lake water, but not in the borehole (before penetration) or glacial ice. Denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments amplified from genomic DNA extracted from Gr??msv??tn samples indicates that the lake community is distinct from the assemblages of organisms in borehole water (before penetration) and the overlying ice and snow. Sequencing of selected DGGE bands revealed that many sequences are highly similar to known psychrophilic organisms or cloned DNA from other cold environments. Significant uptake of 14C-labeled bicarbonate occurred in dark, low-temperature incubations of lake water samples, indicating the presence of autotrophs. Acetylene reduction assays under similar incubation conditions showed no significant nitrogen fixation potential by lake water samples. This may be a consequence of the inhibition of diazotrophy by nitrogen in the lake.

  4. MORPHOLOGICAL CHARACTERISTIC OF SPERMATOGONIA AND TESTES DISSOCIATION : A Preliminary Study for the Germ Cell Transplantation in Giant Gouramy (Osphronemus gouramy

    Directory of Open Access Journals (Sweden)

    Irma Andriani

    2010-12-01

    Full Text Available The recent study were attempting to develop spermatogonial germ cell transplantation as a tool to preserve and propagate male germ-plasm from endangered fish species, as well as to produce surrogate broodstock of commercially valuable fish. Spermatogonia identification and testes dissociation were the first necessary steps to obtain highly amount and viable population of spermatogonia as donor cells for transplantation. Using giant gouramy testes as a model, spermatogonia was histological characterized and two methods of testes dissociations were compared (i.e. medium A contained 0.5% trypsin in PBS and medium B contained 0.5% trypsin and DNase 10 IU/μL in PBS complemented with CaCl2, Hepes and FCS. Optimal incubation times (1, 2, 3, 4 and 5 hours in dissociation medium were also determined. Freshly isolated testes of immature giant gouramy were minced in dissociation medium and then incubated to get monodisperce cell suspension. Parameters observed were number and viability of spermatogonia (ø > 10 μm. The viability was analyzed using trypan blue exclusion dye. The results showed that the average number of spermatogonia observed in medium B was higher than in medium A (P0.05. The viability of spermatogonia decreased by the increasing duration time of dissociation. The viability of spermatogonia started to decrease significantly in 2 hours incubation time in medium A and 4 hours incubation time in medium B (P<0.05. In conclusion, application of dissociation medium B yielded higher number of viable spermatogonia than dissociation medium A.

  5. Cummins L10G in Kenworth truck 'viable today'

    International Nuclear Information System (INIS)

    Anon.

    1996-01-01

    As the culmination of five years of developmental work by Cummins Engines, BC Research Inc., and BC Gas Utility Ltd., a T800 Kenworth truck was outfitted with a new Cummins L10G natural gas engine, and two lightweight fully -wrapped Dynetek cylinders; it was pronounced to be 'a viable clean truck today'. The L10G spark-ignited engine operates at a relatively high peak efficiency of 37 per cent and is commercially available to meet the current California Air Resources Board heavy duty vehicle emission standards without the use of a catalytic converter. The L10G engine produces no particulate emissions, a very significant advantage, in view of the fact that particulate emissions have been identified as major contributors to respiratory ailments

  6. The role of donor characteristics and post-granulocyte colony-stimulating factor white blood cell counts in predicting the adverse events and yields of stem cell mobilization.

    Science.gov (United States)

    Chen, Shu-Huey; Yang, Shang-Hsien; Chu, Sung-Chao; Su, Yu-Chieh; Chang, Chu-Yu; Chiu, Ya-Wen; Kao, Ruey-Ho; Li, Dian-Kun; Yang, Kuo-Liang; Wang, Tso-Fu

    2011-05-01

    Granulocyte colony-stimulating factor (G-CSF) is now widely used for stem cell mobilization. We evaluated the role of post-G-CSF white blood cell (WBC) counts and donor factors in predicting adverse events and yields associated with mobilization. WBC counts were determined at baseline, after the third and the fifth dose of G-CSF in 476 healthy donors. Donors with WBC ≥ 50 × 10(3)/μL post the third dose of G-CSF experienced more fatigue, myalgia/arthralgia, and chills, but final post-G-CSF CD34(+) cell counts were similar. Although the final CD34(+) cell count was higher in donors with WBC ≥ 50 × 10(3)/μL post the fifth G-CSF, the incidence of side effects was similar. Females more frequently experienced headache, nausea/anorexia, vomiting, fever, and lower final CD34(+) cell count than did males. Donors with body mass index (BMI) ≥ 25 showed higher incidences of sweat and insomnia as well as higher final CD34(+) cell counts. Donor receiving G-CSF ≥ 10 μg/kg tended to experience bone pain, headache and chills more frequently. Multivariate analysis indicated that female gender is an independent factor predictive of the occurrence of most side effects, except for ECOG > 1 and chills. Higher BMI was also an independent predictor for fatigue, myalgia/arthralgia, and sweat. Higher G-CSF dose was associated with bone pain, while the WBC count post the third G-CSF was associated with fatigue only. In addition, one donor in the study period did not complete the mobilization due to suspected anaphylactoid reaction. Observation for 1 h after the first injection of G-CSF is required to prevent complications from unpredictable side effects.

  7. Improving the Yield of Histological Sampling in Patients With Suspected Colorectal Cancer During Colonoscopy by Introducing a Colonoscopy Quality Assurance Program.

    Science.gov (United States)

    Gado, Ahmed; Ebeid, Basel; Abdelmohsen, Aida; Axon, Anthony

    2011-08-01

    Masses discovered by clinical examination, imaging or endoscopic studies that are suspicious for malignancy typically require biopsy confirmation before treatment is initiated. Biopsy specimens may fail to yield a definitive diagnosis if the lesion is extensively ulcerated or otherwise necrotic and viable tumor tissue is not obtained on sampling. The diagnostic yield is improved when multiple biopsy samples (BSs) are taken. A colonoscopy quality-assurance program (CQAP) was instituted in 2003 in our institution. The aim of this study was to determine the effect of instituting a CQAP on the yield of histological sampling in patients with suspected colorectal cancer (CRC) during colonoscopy. Initial assessment of colonoscopy practice was performed in 2003. A total of five patients with suspected CRC during colonoscopy were documented in 2003. BSs confirmed CRC in three (60%) patients and were nondiagnostic in two (40%). A quality-improvement process was instituted which required a minimum six BSs with adequate size of the samples from any suspected CRC during colonoscopy. A total of 37 patients for the period 2004-2010 were prospectively assessed. The diagnosis of CRC was confirmed with histological examination of BSs obtained during colonoscopy in 63% of patients in 2004, 60% in 2005, 50% in 2006, 67% in 2007, 100% in 2008, 67% in 2009 and 100% in 2010. The yield of histological sampling increased significantly ( p quality assurance and improvement program increased the yield of histological sampling in patients with suspected CRC during colonoscopy.

  8. Increased saccharification yields from aspen biomass upon treatment with enzymatically generated peracetic acid.

    Science.gov (United States)

    Duncan, Shona; Jing, Qing; Katona, Adrian; Kazlauskas, Romas J; Schilling, Jonathan; Tschirner, Ulrike; Aldajani, Waleed Wafa

    2010-03-01

    The recalcitrance of lignocellulosic biomass to enzymatic release of sugars (saccharification) currently limits its use as feedstock for biofuels. Enzymatic hydrolysis of untreated aspen wood releases only 21.8% of the available sugars due primarily to the lignin barrier. Nature uses oxidative enzymes to selectively degrade lignin in lignocellulosic biomass, but thus far, natural enzymes have been too slow for industrial use. In this study, oxidative pretreatment with commercial peracetic acid (470 mM) removed 40% of the lignin (from 19.9 to 12.0 wt.% lignin) from aspen and enhanced the sugar yields in subsequent enzymatic hydrolysis to about 90%. Increasing the amount of lignin removed correlated with increasing yields of sugar release. Unfortunately, peracetic acid is expensive, and concentrated forms can be hazardous. To reduce costs and hazards associated with using commercial peracetic acid, we used a hydrolase to catalyze the perhydrolysis of ethyl acetate generating 60-70 mM peracetic acid in situ as a pretreatment to remove lignin from aspen wood. A single pretreatment was insufficient, but multiple cycles (up to eight) removed up to 61.7% of the lignin enabling release of >90% of the sugars during saccharification. This value corresponds to a predicted 581 g of fermentable sugars from 1 kg of aspen wood. Improvements in the enzyme stability are needed before the enzymatically generated peracetic acid is a commercially viable alternative.

  9. Identification of cancer stem cell markers in human malignant mesothelioma cells

    Energy Technology Data Exchange (ETDEWEB)

    Ghani, Farhana Ishrat; Yamazaki, Hiroto; Iwata, Satoshi; Okamoto, Toshihiro [Division of Clinical Immunology, Institute of Medical Science, University of Tokyo, Tokyo (Japan); Aoe, Keisuke; Okabe, Kazunori; Mimura, Yusuke [Departments of Medical Oncology, Yamaguchi-Ube Medical Center, Yamaguchi (Japan); Fujimoto, Nobukazu; Kishimoto, Takumi [Department of Respiratory Medicine, Okayama Rosai Hospital, Okayama (Japan); Yamada, Taketo [Department of Pathology, Keio University School of Medicine, Tokyo (Japan); Xu, C. Wilson [Drug Development Program, Nevada Cancer Institute, Las Vegas, NV (United States); Morimoto, Chikao, E-mail: morimoto@ims.u-tokyo.ac.jp [Division of Clinical Immunology, Institute of Medical Science, University of Tokyo, Tokyo (Japan); Drug Development Program, Nevada Cancer Institute, Las Vegas, NV (United States)

    2011-01-14

    Research highlights: {yields} We performed serial transplantation of surgical samples and established new cell lines of malignant mesothelioma. {yields} SP cell and expressions of CD9/CD24/CD26 were often observed in mesothelioma cell lines. {yields} SP and CD24{sup +} cells proliferated by asymmetric cell division-like manner. CD9{sup +} and CD24{sup +} cells have higher potential to generate spheroid colony. {yields} The marker-positive cells have clear tendency to generate larger tumors in mice. -- Abstract: Malignant mesothelioma (MM) is an aggressive and therapy-resistant neoplasm arising from the pleural mesothelial cells and usually associated with long-term asbestos exposure. Recent studies suggest that tumors contain cancer stem cells (CSCs) and their stem cell characteristics are thought to confer therapy-resistance. However, whether MM cell has any stem cell characteristics is not known. To understand the molecular basis of MM, we first performed serial transplantation of surgical samples into NOD/SCID mice and established new cell lines. Next, we performed marker analysis of the MM cell lines and found that many of them contain SP cells and expressed several putative CSC markers such as CD9, CD24, and CD26. Interestingly, expression of CD26 closely correlated with that of CD24 in some cases. Sorting and culture assay revealed that SP and CD24{sup +} cells proliferated by asymmetric cell division-like manner. In addition, CD9{sup +} and CD24{sup +} cells have higher potential to generate spheroid colony than negative cells in the stem cell medium. Moreover, these marker-positive cells have clear tendency to generate larger tumors in mouse transplantation assay. Taken together, our data suggest that SP, CD9, CD24, and CD26 are CSC markers of MM and could be used as novel therapeutic targets.

  10. VARIABILITY OF YIELD AND YIELD COMPONENTS IN “EGUSI ...

    African Journals Online (AJOL)

    journal

    Estimate of expected genetic advance in seed yield plant-1 ranged between. 25.90-48.40%. ..... values in fruit and seed yield characters have been reported in culinary melon, ... and Khund, A. 2004. Extent of heterosis and heritability in some.

  11. Genotype x environment interactions in milk yield and quality in Angus, Brahman, and reciprocal-cross cows on different forage systems.

    Science.gov (United States)

    Brown, M A; Brown, A H; Jackson, W G; Miesner, J R

    2001-07-01

    Milk yield and quality were observed on 93 Angus, Brahman, and reciprocal-cross cows over 3 yr to evaluate the interactions of direct and maternal breed effects and heterosis with forage environment. Forage environments were common bermudagrass (BG), endophyte-infected tall fescue (E+), and a rotational system (ROT) of both forages, in which each forage (BG or E+) was grazed during its appropriate season, usually June through October for BG and November through May for E+. Milk yield was estimated each of 6 mo (April through September) via milking machine and converted to a 24-h basis. Milk fat, milk protein, and somatic cell count were analyzed by a commercial laboratory. Heterosis for milk yield was similar among forages, averaging 2.4 kg (P 0.30). Heterosis for somatic cell counts as percentages of purebred means was similar for BG (-68.3%) and E+ (-68.9%) and less favorable for ROT (-31.6%). Maternal breed effects for somatic cell count favored Angus on ROT (P < 0.10) with a similar nonsignificant trend on BG and E+. Direct breed effects for somatic cell count favored Brahman on ROT (P < 0.10) with similar nonsignificant trends on BG and E+. These results suggested that a rotation of cows from E+ to BG in the summer can partially alleviate negative effects of E+ on milk yield. Conclusions also indicated an advantage to crossbred cows in somatic cell count and provided evidence of both direct and maternal breed effects for this trait. The results also suggested that direct breed effects for milk yield, milk fat, and somatic cell count and heterosis for milk yield and somatic cell count (as percentages of purebred means) tended to vary with forage environment, indicating a potential for genotype x environment interaction for these traits.

  12. Characterization of HIT type solar cells

    NARCIS (Netherlands)

    Rath, J.K.

    2011-01-01

    The silicon heterojunction solar cell (SHJ) has made rapid progress in reaching high efficiency and it is already developed as an industrially viable product. However, much of its progress has come through process development while there is scarce knowledge on the microscopic nature of the

  13. Formulation and evaluation of Bacillus coagulans-loaded hypromellose mucoadhesive microspheres.

    Science.gov (United States)

    Alli, Sk Md Athar

    2011-01-01

    Development of a novel delivery system has been attempted to deliver viable probiotic cells into the gut for a prolonged period of time while maintaining high numbers of viable cells within the formulation throughout the shelf-life of the product and during the gastrointestinal transit. Core mucoadhesive microspheres of Bacillus coagulans were developed employing several grades of hypromellose, a mucoadhesive polymer, following coacervation and phase separation technique and were subsequently enteric-coated with hypromellose phthalate. Microspheres were evaluated for percent yield; entrapment efficiency; in vitro swelling; surface morphology; particle size, size distribution, and zeta potential; flow property, mucoadhesion property by the ex vivo mucoadhesive strength test and the in vitro wash off test; in vitro release profile and release kinetic; in vivo probiotic activity; and stability. The values for the kinetic constant and regression coefficient of model-dependent approaches and the difference factor (f(1)), the similarity factor (f(2)), and the Rescigno index (ξ(1) and ξ(2)) of model independent approaches were determined for comparing in vitro dissolution profiles. Freeze dried B. coagulans cells were successfully formulated as enteric-coated mucoadhesive microspheres with satisfactory physical structure and yield. The viability of B. coagulans was maintained in the simulated gastric conditions and during processing; in simulated intestinal conditions exhibiting mucoadhesion, and controlling and extending the viable cell release following zero-order; and was satisfactorily stable at room temperature. Test results depict statistically significant effects of the hypromellose grade and their concentration on the performance and release profile of formulations.

  14. Fission product yields

    International Nuclear Information System (INIS)

    Valenta, V.; Hep, J.

    1978-01-01

    Data are summed up necessary for determining the yields of individual fission products from different fissionable nuclides. Fractional independent yields, cumulative and isobaric yields are presented here for the thermal fission of 235 U, 239 Pu, 241 Pu and for fast fission (approximately 1 MeV) of 235 U, 238 U, 239 Pu, 241 Pu; these values are included into the 5th version of the YIELDS library, supplementing the BIBFP library. A comparison is made of experimental data and possible improvements of calculational methods are suggested. (author)

  15. Larger yield of cyclobutane dimers than 8-oxo-7,8-dihydroguanine in the DNA of UVA-irradiated human skin cells

    International Nuclear Information System (INIS)

    Courdavault, Sophie; Baudouin, Caroline; Charveron, Marie; Favier, Alain; Cadet, Jean; Douki, Thierry

    2004-01-01

    Exposure to solar ultraviolet light is the major cause of most skin cancers. While the genotoxic properties of UVB radiation are now well understood, the DNA damaging processes triggered by less energetic but more abundant UVA photons remain to be elucidated. Evidence has been provided for the induction of oxidative lesions to cellular DNA including strand breaks and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo). Formation of cyclobutane pyrimidine dimers (CPDs) has also been reported, mostly in rodent cells. In order to gain insights into the relevance of the latter photoproducts in UVA-mutagenesis of human skin, we quantified the level of 8-oxodGuo and CPDs within primary cultures of normal fibroblasts and keratinocytes using specific chromatographic assays. The yield of formation of CPDs was found to be higher than that of 8-oxodGuo in both cell types. In addition, CPDs were mostly TT derivatives, and neither (6-4) photoproducts nor Dewar valence isomers were detected. These observations are reminiscent of results obtained in rodent cells and suggest that a photosensitized triplet energy transfer occurs and that this reaction is more efficient than photooxidation of DNA components. The predominant formation of CPDs with respect to oxidative damage within normal human skin cells exposed to UVA radiation should be taken into account in photoprotection strategies

  16. Larger yield of cyclobutane dimers than 8-oxo-7,8-dihydroguanine in the DNA of UVA-irradiated human skin cells

    Energy Technology Data Exchange (ETDEWEB)

    Courdavault, Sophie [Laboratoire Lesions des Acides Nucleiques, Service de Chimie Inorganique et Biologique, CEA/DSM/Departement de Recherche Fondamentale sur la Matiere Condensee, CEA-Grenoble, 17, avenue des Martyrs, 38054 Grenoble Cedex 9 (France); Baudouin, Caroline [Institut de Recherche Pierre Fabre, Laboratoire de Biologie Cellulaire, Toulouse (France); Charveron, Marie [Institut de Recherche Pierre Fabre, Laboratoire de Biologie Cellulaire, Toulouse (France); Favier, Alain [Laboratoire Lesions des Acides Nucleiques, Service de Chimie Inorganique et Biologique, CEA/DSM/Departement de Recherche Fondamentale sur la Matiere Condensee, CEA-Grenoble, 17, avenue des Martyrs, 38054 Grenoble Cedex 9 (France); Cadet, Jean [Laboratoire Lesions des Acides Nucleiques, Service de Chimie Inorganique et Biologique, CEA/DSM/Departement de Recherche Fondamentale sur la Matiere Condensee, CEA-Grenoble, 17, avenue des Martyrs, 38054 Grenoble Cedex 9 (France); Douki, Thierry [Laboratoire Lesions des Acides Nucleiques, Service de Chimie Inorganique et Biologique, CEA/DSM/Departement de Recherche Fondamentale sur la Matiere Condensee, CEA-Grenoble, 17, avenue des Martyrs, 38054 Grenoble Cedex 9 (France)]. E-mail: tdouki@cea.fr

    2004-11-22

    Exposure to solar ultraviolet light is the major cause of most skin cancers. While the genotoxic properties of UVB radiation are now well understood, the DNA damaging processes triggered by less energetic but more abundant UVA photons remain to be elucidated. Evidence has been provided for the induction of oxidative lesions to cellular DNA including strand breaks and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo). Formation of cyclobutane pyrimidine dimers (CPDs) has also been reported, mostly in rodent cells. In order to gain insights into the relevance of the latter photoproducts in UVA-mutagenesis of human skin, we quantified the level of 8-oxodGuo and CPDs within primary cultures of normal fibroblasts and keratinocytes using specific chromatographic assays. The yield of formation of CPDs was found to be higher than that of 8-oxodGuo in both cell types. In addition, CPDs were mostly TT derivatives, and neither (6-4) photoproducts nor Dewar valence isomers were detected. These observations are reminiscent of results obtained in rodent cells and suggest that a photosensitized triplet energy transfer occurs and that this reaction is more efficient than photooxidation of DNA components. The predominant formation of CPDs with respect to oxidative damage within normal human skin cells exposed to UVA radiation should be taken into account in photoprotection strategies.0.

  17. 9 CFR 113.27 - Detection of extraneous viable bacteria and fungi in live vaccines.

    Science.gov (United States)

    2010-01-01

    ... bacteria and fungi in live vaccines. 113.27 Section 113.27 Animals and Animal Products ANIMAL AND PLANT... bacteria and fungi in live vaccines. Unless otherwise specified by the Administrator or elsewhere exempted... Seed Bacteria shall be tested for extraneous viable bacteria and fungi as prescribed in this section. A...

  18. Estimation of Viable Biomass In Wastewater And Activated Sludge By Determination of ATP, Oxygen Utilization Rate And FDA Hydrolysis

    DEFF Research Database (Denmark)

    Jørgensen, Poul-Erik; Eriksen, T.; Jensen, B.K.

    1992-01-01

    ATP content, oxygen utilization rate (OUR) and fluorescein diacetate (FDA) hydrolysis were tested for the ability to express the amount of viable biomass in wastewater and activated sludge. The relationship between biomass and these activity parameters was established in growth cultures made...... with biomass, while FDA hydrolysis in the sludge failed to show any such correlation. Conversion factors of 3 mg ATP/g dw, 300 mg O2/h g dw and 0.4 A/h (mg dw/ml) for ATP, OUR and FDA methods, respectively, were calculated. When the methods were applied for in situ determinations in four different wastewater...... plants, it was found that ATP content and respiration rate estimated viable biomass to range from 81 to 293 mg dw/g SS for raw wastewater and from 67 to 187 mg dw/g SS for activated sludge with a rather weak correlation between ATP and respiration measurements. The FDA hydrolysis estimated viable biomass...

  19. yield and yield componemts of extra early maize (zea mays l.)

    African Journals Online (AJOL)

    SHARIFAI

    maize crop and improve the soil structures and chemical nutrients of the soil. The significant interaction between intra-row spacing and poultry manure on cob diameter, 100 grain weight and grain yield showed the importance of poultry manure on yield and yield components of maize crop. Poultry manure increases both ...

  20. An adapted yield criterion for the evolution of subsequent yield surfaces

    Science.gov (United States)

    Küsters, N.; Brosius, A.

    2017-09-01

    In numerical analysis of sheet metal forming processes, the anisotropic material behaviour is often modelled with isotropic work hardening and an average Lankford coefficient. In contrast, experimental observations show an evolution of the Lankford coefficients, which can be associated with a yield surface change due to kinematic and distortional hardening. Commonly, extensive efforts are carried out to describe these phenomena. In this paper an isotropic material model based on the Yld2000-2d criterion is adapted with an evolving yield exponent in order to change the yield surface shape. The yield exponent is linked to the accumulative plastic strain. This change has the effect of a rotating yield surface normal. As the normal is directly related to the Lankford coefficient, the change can be used to model the evolution of the Lankford coefficient during yielding. The paper will focus on the numerical implementation of the adapted material model for the FE-code LS-Dyna, mpi-version R7.1.2-d. A recently introduced identification scheme [1] is used to obtain the parameters for the evolving yield surface and will be briefly described for the proposed model. The suitability for numerical analysis will be discussed for deep drawing processes in general. Efforts for material characterization and modelling will be compared to other common yield surface descriptions. Besides experimental efforts and achieved accuracy, the potential of flexibility in material models and the risk of ambiguity during identification are of major interest in this paper.

  1. Dentin and dental pulp regeneration by the patient’s endogenous cells

    OpenAIRE

    KIM, SAHNG G.; ZHENG, YING; ZHOU, JIAN; CHEN, MO; EMBREE, MILDRED C.; SONG, KAREN; JIANG, NAN; MAO, JEREMY J.

    2013-01-01

    The goal of regenerative endodontics is to restore the functions of the dental pulp–dentin complex. Two approaches are being applied toward dental pulp–dentin regeneration: cell transplantation and cell homing. The majority of previous approaches are based on cell transplantation by delivering ex vivo cultivated cells toward dental pulp or dentin regeneration. Many hurdles limit the clinical translation of cell transplantation such as the difficulty of acquiring and isolating viable cells, un...

  2. Cardiac-Derived Extracellular Matrix Enhances Cardiogenic Properties of Human Cardiac Progenitor Cells

    NARCIS (Netherlands)

    Gaetani, Roberto; Yin, Christopher; Srikumar, Neha; Braden, Rebecca; Doevendans, Pieter A; Sluijter, Joost P G; Christman, Karen L

    2016-01-01

    The use of biomaterials has been demonstrated as a viable strategy to promote cell survival and cardiac repair. However, limitations on combinational cell-biomaterial therapies exist, as cellular behavior is influenced by the microenvironment and physical characteristics of the material. Among the

  3. Inactivation of viable Ascaris eggs by reagents during enumeration.

    Science.gov (United States)

    Nelson, K L; Darby, J L

    2001-12-01

    Various reagents commonly used to enumerate viable helminth eggs from wastewater and sludge were evaluated for their potential to inactivate Ascaris eggs under typical laboratory conditions. Two methods were used to enumerate indigenous Ascaris eggs from sludge samples. All steps in the methods were the same except that in method I a phase extraction step with acid-alcohol (35% ethanol in 0.1 N H(2)SO(4)) and diethyl ether was used whereas in method II the extraction step was avoided by pouring the sample through a 38-microm-mesh stainless steel sieve that retained the eggs. The concentration of eggs and their viability were lower in the samples processed by method I than in the samples processed by method II by an average of 48 and 70%, respectively. A second set of experiments was performed using pure solutions of Ascaris suum eggs to elucidate the effect of the individual reagents and relevant combination of reagents on the eggs. The percentages of viable eggs in samples treated with acid-alcohol alone and in combination with diethyl ether or ethyl acetate were 52, 27, and 4%, respectively, whereas in the rest of the samples the viability was about 80%. Neither the acid nor the diethyl ether alone caused any decrease in egg viability. Thus, the observed inactivation was attributed primarily to the 35% ethanol content of the acid-alcohol solution. Inactivation of the eggs was prevented by limiting the direct exposure to the extraction reagents to 30 min and diluting the residual concentration of acid-alcohol in the sample by a factor of 100 before incubation. Also, the viability of the eggs was maintained if the acid-alcohol solution was replaced with an acetoacetic buffer. None of the reagents used for the flotation step of the sample cleaning procedure (ZnSO(4), MgSO(4), and NaCl) or during incubation (0.1 N H(2)SO(4) and 0.5% formalin) inactivated the Ascaris eggs under the conditions studied.

  4. Cover crop rotations in no-till system: short-term CO2 emissions and soybean yield

    Directory of Open Access Journals (Sweden)

    João Paulo Gonsiorkiewicz Rigon

    Full Text Available ABSTRACT: In addition to improving sustainability in cropping systems, the use of a spring and winter crop rotation system may be a viable option for mitigating soil CO2 emissions (ECO2. This study aimed to determine short-term ECO2 as affected by crop rotations and soil management over one soybean cycle in two no-till experiments, and to assess the soybean yields with the lowest ECO2. Two experiments were carried out in fall-winter as follows: i triticale and sunflower were grown in Typic Rhodudalf (TR, and ii ruzigrass, grain sorghum, and ruzigrass + grain sorghum were grown in Rhodic Hapludox (RH. In the spring, pearl millet, sunn hemp, and forage sorghum were grown in both experiments. In addition, in TR a fallow treatment was also applied in the spring. Soybean was grown every year in the summer, and ECO2 were recorded during the growing period. The average ECO2 was 0.58 and 0.84 g m2 h–1 with accumulated ECO2 of 5,268 and 7,813 kg ha–1 C-CO2 in TR and RH, respectively. Sunn hemp, when compared to pearl millet, resulted in lower ECO2 by up to 12 % and an increase in soybean yield of 9% in TR. In RH, under the winter crop Ruzigrazz+Sorghum, ECO2 were lower by 17%, although with the same soybean yield. Soil moisture and N content of crop residues are the main drivers of ECO2 and soil clay content seems to play an important role in ECO2 that is worthy of further studies. In conclusion, sunn hemp in crop rotation may be utilized to mitigate ECO2 and improve soybean yield.

  5. Design for manufacturability and yield for nano-scale CMOS

    CERN Document Server

    Chiang, Charles C

    2007-01-01

    Talks about the various aspects of manufacturability and yield in a nano-CMOS process and how to address each aspect at the proper design step starting with the design and layout of standard cells. This book is suitable for practicing IC designer and for graduate students intent on having a career in IC design or in EDA tool development.

  6. Inheritance of grain yield and its correlation with yield components in ...

    African Journals Online (AJOL)

    SAM

    2014-03-19

    Mar 19, 2014 ... 7 × 7 incomplete diallel cross of seven wheat parents during the crop season of 2009 to 2010. Mean square of general ... Genetic background and yield traits of the seven parents. Parent. Pedigree. Released year ..... Correlation and path analysis for yield and yield contributing characters in wheat (Triticum ...

  7. Growth and viability of Aedes albopictus cell line in vitro after cesium-137 gamma irradiation

    International Nuclear Information System (INIS)

    Blakely, E.A.

    1975-01-01

    The radiosensitivity of the cultured mosquito cell line Aedes albopictus (Skuse) was investigated. Population growth was followed by total cell counts and by viable cell counts on aliquots of cultures exposed to various doses of gamma radiation during exponential growth. Viable cell determinations were based on the cellular exclusion of the dye, alcian blue, in a procedure adapted to the insect cells in culture. Viability determinations in the irradiated exponential cultures indicated that initially there was some increase in the gestation, suggesting that gonadal steroids may have unusual effects on uterine physiology and biochemistry in this species. Consequently, studies were undertaken to elucidate some of the basic responses of hamster uteri to estradiol benzoate and progesterone under conditions of protein malnutrition, actinomycin D administration and corticosterone injection. Furthermore, the effects of gonadal steroids on uteri of pregnant ovariectomized hamsters were studied

  8. Yield and economic performance of organic and conventional cotton-based farming systems--results from a field trial in India.

    Science.gov (United States)

    Forster, Dionys; Andres, Christian; Verma, Rajeev; Zundel, Christine; Messmer, Monika M; Mäder, Paul

    2013-01-01

    The debate on the relative benefits of conventional and organic farming systems has in recent time gained significant interest. So far, global agricultural development has focused on increased productivity rather than on a holistic natural resource management for food security. Thus, developing more sustainable farming practices on a large scale is of utmost importance. However, information concerning the performance of farming systems under organic and conventional management in tropical and subtropical regions is scarce. This study presents agronomic and economic data from the conversion phase (2007-2010) of a farming systems comparison trial on a Vertisol soil in Madhya Pradesh, central India. A cotton-soybean-wheat crop rotation under biodynamic, organic and conventional (with and without Bt cotton) management was investigated. We observed a significant yield gap between organic and conventional farming systems in the 1(st) crop cycle (cycle 1: 2007-2008) for cotton (-29%) and wheat (-27%), whereas in the 2(nd) crop cycle (cycle 2: 2009-2010) cotton and wheat yields were similar in all farming systems due to lower yields in the conventional systems. In contrast, organic soybean (a nitrogen fixing leguminous plant) yields were marginally lower than conventional yields (-1% in cycle 1, -11% in cycle 2). Averaged across all crops, conventional farming systems achieved significantly higher gross margins in cycle 1 (+29%), whereas in cycle 2 gross margins in organic farming systems were significantly higher (+25%) due to lower variable production costs but similar yields. Soybean gross margin was significantly higher in the organic system (+11%) across the four harvest years compared to the conventional systems. Our results suggest that organic soybean production is a viable option for smallholder farmers under the prevailing semi-arid conditions in India. Future research needs to elucidate the long-term productivity and profitability, particularly of cotton and

  9. Existence of both culturable and viable but non culturable (VNC) E. coli populations with distinct settling velocities in karst aquifer

    Science.gov (United States)

    Petit, F.; Ratajczak, M.; Massei, N.; Lafite, R.; Clermont, O.; Denamur, E.; Berthe, T.

    2012-12-01

    associated to "non settleable" particles, decreased from the sinkhole to the spring. In contrast in dry period while the turbidity and the contamination by culturable E. coli is low, the three populations of viable, culturable and VNC E. coli are introduced in the karst. Within the population of culturable E. coli, persistent strains mainly belonging to B1 phylogroup and growing at 7°C were introduced in karst aquifer. At the spring, whatever the class of particles up to 96% of viable cells were in VNC state suggesting that a physiological change occurred within aquifer, independently of the association of E. coli with particles. At the well, the pumping of the water induced an input of resident VNC E. coli consecutively to a resuspension of particles previously settled within the karstic network, during a past turbid event.

  10. Higgs production in vector boson fusion in the H{yields} {tau}{tau} {yields} ll + 4{nu} final state with ATLAS. A sensitivity study

    Energy Technology Data Exchange (ETDEWEB)

    Schmitz, Martin

    2011-05-15

    A study of the expected sensitivity of the ATLAS experiment to discover the Standard Model Higgs boson produced via vector boson fusion (VBF) and its decay to H{yields} {tau}{tau} {yields} ll + 4{nu} is presented. The study is based on simulated proton-proton collisions at a centre-of-mass energy of 14 TeV. For the rst time the discovery potential is evaluated in the presence of additional proton-proton interactions (pile-up) to the process of interest in a complete and consistent way. Special emphasis is placed on the development of background estimation techniques to extract the main background processes Z{yields} {tau}{tau} and t anti t production using data. The t anti t background is estimated using a control sample selected with the VBF analysis cuts and the inverted b-jet veto. The dominant background process Z {yields} {tau}{tau} is estimated using Z{yields} {mu}{mu} events. Replacing the muons of the Z{yields} {mu}{mu} event with simulated {tau}-leptons, Z {yields} {tau}{tau} events are modelled to high precision. For the replacement of the Z boson decay products a dedicated method based on tracks and calorimeter cells is developed. Without pile-up a discovery potential of 3{sigma} to 3.4{sigma} in the mass range 115 GeV

  11. Corporate strategy and viable future land use: Planning for closure from the outset of mining

    International Nuclear Information System (INIS)

    Warhurst, A.

    2000-01-01

    This paper provides an overview of the environmental impact of mining on viable future land use and underlines the imperative of improved environmental management and closure planning. It argues that pollution prevention, through planning for closure, can lead to cost-effective strategies for sustainable minerals development and viable future land use. This seems to be most true for greenfield sites since, generally, the earlier closure planning and pollution prevention is built into a project, the more cost-effective and environmentally benign closure will be. Further, for greenfield sites, pollution prevention techniques can be employed from the outset, at the stages of exploration and mine development, and then monitored and improved through the operation stage to closure, and can be kept in place to manage future land use. The paper discusses how global changes in the industry, following the liberalisation of investment regimes, and mergers and strategic alliances between key firms, has, by virtue of the diffusion of new technology, led to further opportunities to prevent pollution and optimise future land use through planning for closure from the outset. The objectives and components of closure plans are also reviewed as the paper draws on case studies to highlight some of the possible constraints and challenges to pollution prevention that may be faced at the level of both public policy and corporate strategy. The article concludes by suggesting a forward-looking approach to integrated environmental management and viable future land-use planning based on a dynamic model for environmental management. (author)

  12. Effects of cutting frequency on alfalfa yield and yield components in ...

    African Journals Online (AJOL)

    Effects of cutting frequency on alfalfa yield and yield components in Songnen Plain, Northeast China. J Chen, F Tang, R Zhu, C Gao, G Di, Y Zhang. Abstract. The productivity and quality of alfalfa (Medicago sativa L.) is strongly influenced by cutting frequency (F). To clarify that the yield and quality of alfalfa if affected by F, ...

  13. Resuscitation of the viable but non-culturable state of Salmonella enterica serovar Oranienburg by recombinant resuscitation-promoting factor derived from Salmonella Typhimurium strain LT2.

    Science.gov (United States)

    Panutdaporn, N; Kawamoto, K; Asakura, H; Makino, S-I

    2006-02-15

    A gene encoding the resuscitation-promoting factor (Rpf) from Salmonella Typhimurium LT2 was cloned and characterized. The amino acid sequence encoded by S. Typhimurium LT2 rpf gene shares 24.2% homology with Micrococcus luteus Rpf, which is secreted by growing cells, and required to resuscitate from viable but non-culturable (VNC) state. The S. Typhimurium LT2 rpf gene is 696 bp long, and shared a conserved segment with Salmonella enterica serovar Oranienburg (99.4%). Recombinant Rpf (rRpf) proteins of S. Typhimurium LT2 after expression in E. coli BL21 harboring the pET15-b plasmid was approximately 25 kDa. Since S. Oranienburg cells are relatively quick to enter the VNC state just after incubating in the presence of 7% NaCl at 37 degrees C for 3 days, we evaluated the biological effect of rRpf by using S. Oranienburg VNC cells. The rRpf not only promoted proliferation but also induced resuscitation of VNC cells to the culturable state in a dose-dependent manner. Therefore, rRpf may be useful for detection of bacterial contaminants present in the VNC form in food samples and the environment.

  14. Cell death induced by hydroxyapatite on L929 fibroblast cells.

    Science.gov (United States)

    Inayat-Hussain, S H; Rajab, N F; Roslie, H; Hussin, A A; Ali, A M; Annuar, B O

    2004-05-01

    Biomaterials intended for end-use application as bone-graft substitutes have to undergo safety evaluation. In this study, we investigated the in vitro cytotoxic effects especially to determine the mode of death of two hydroxyapatite compounds (HA2, HA3) which were synthesized locally. The methods used for cytotoxicity was the standard MTT assay whereas AO/PI staining was performed to determine the mode of cell death in HA treated L929 fibroblasts. Our results demonstrated that both HA2 and HA3 were not significantly cytotoxic as more than 75% cells after 72 hours treatment were viable. Furthermore, we found that the major mode of cell death in HA treated cells was apoptosis. In conclusion, our results demonstrated that these hydroxyapatite compounds are not cytotoxic where the mode of death was primarily via apoptosis.

  15. Performance of Pd on activated carbon as hydrogen electrode with respect to hydrogen yield in a single cell proton exchange membrane (PEM) water electrolyser

    Energy Technology Data Exchange (ETDEWEB)

    Naga Mahesh, K.; Sarada Prasad, J.; Venkateswer Rao, M.; Himabindu, V. [Centre for Environment, Institute of Science and Technology, Jawaharlal Nehru Technological University Hyderabad, Kukatpally, Hyderabad 500085 (A.P.) (India); Yerramilli, Anjaneyulu [TLGVRC, JSU Box 18739, Jackson State University, Jackson, MS 32917 - 0939 (United States); Raghunathan Rao, P. [Fuel cell section, Heavy Water Division, Bhabha Atomic Research Centre, Trombay, Mumbai - 400 085 (India)

    2009-08-15

    Palladium (Pd) on activated carbon is used as electrocatalyst coated on Nafion 115 membrane as Hydrogen electrode and RuO{sub 2} is coated on other side of membrane used as oxygen electrode. 5 wt% and 10 wt% Pd on activated carbon is prepared as membrane electrode assembly (MEA) and investigated the performance of the same using inhouse prepared 10 cm{sup 2} single cell. The performance of the single cell assembly and the hydrogen yield are reported during electrolysis operation at temperatures 27 C, 45 C and 65 C at 0.1, 0.2, 0.3, 0.4, 0.5 A/cm{sup 2} current densities with respect to voltages. (author)

  16. Kinetics of the cells of the hematopoietic system in the course of postradiation regeneration. [Mice

    Energy Technology Data Exchange (ETDEWEB)

    Kondratenko, N F

    1975-01-01

    Twenty-four hours after sublethal whole-body irradiation (200 rad) of mice, only 4 percent of the stem cells were viable and continued to differentiate. Exponential growth of these cells started a day later and by week 2 the number was completely restored and stabilized. The kinetics of committed precursors of granulocytes and macrophages was different. Some 20 percent of these cells were viable 24 hours after irradiation and the granulocytes in particular continued to differentiate into mature cellular elements. The total number remained at the minimal level for a week, after which the logarithmic phase began, with complete recovery achieved toward the end of week 2 after irradiation. Thus, the restoration of hematopoiesis in sublethally irradiated mice is due primarily to the exponential growth of surviving stem cells in bone marrow.

  17. High-Yield Purification, Preservation, and Serial Transplantation of Human Satellite Cells

    Directory of Open Access Journals (Sweden)

    Steven M. Garcia

    2018-03-01

    Full Text Available Summary: Investigation of human muscle regeneration requires robust methods to purify and transplant muscle stem and progenitor cells that collectively constitute the human satellite cell (HuSC pool. Existing approaches have yet to make HuSCs widely accessible for researchers, and as a result human muscle stem cell research has advanced slowly. Here, we describe a robust and predictable HuSC purification process that is effective for each human skeletal muscle tested and the development of storage protocols and transplantation models in dystrophin-deficient and wild-type recipients. Enzymatic digestion, magnetic column depletion, and 6-marker flow-cytometric purification enable separation of 104 highly enriched HuSCs per gram of muscle. Cryostorage of HuSCs preserves viability, phenotype, and transplantation potential. Development of enhanced and species-specific transplantation protocols enabled serial HuSC xenotransplantation and recovery. These protocols and models provide an accessible system for basic and translational investigation and clinical development of HuSCs. : Garcia and colleagues report methods for efficient purification of satellite cells from human skeletal muscle. They use their approaches to demonstrate stem cell functions of endogenous satellite cells and to make human satellite cells accessible for sharing among researchers. Keywords: human satellite cell purification, serial transplantation, satellite cell cryopreservation

  18. Organized Hole Conducting Polymer/Nanoparticle Composite Solar Cell Fiber, Phase I

    Data.gov (United States)

    National Aeronautics and Space Administration — Recent innovations in noncrystalline solar cells are beginning to emerge as viable replacements to traditional CVD grown single crystal counterparts in applications...

  19. Cell-type-specific predictive network yields novel insights into mouse embryonic stem cell self-renewal and cell fate.

    Directory of Open Access Journals (Sweden)

    Karen G Dowell

    Full Text Available Self-renewal, the ability of a stem cell to divide repeatedly while maintaining an undifferentiated state, is a defining characteristic of all stem cells. Here, we clarify the molecular foundations of mouse embryonic stem cell (mESC self-renewal by applying a proven Bayesian network machine learning approach to integrate high-throughput data for protein function discovery. By focusing on a single stem-cell system, at a specific developmental stage, within the context of well-defined biological processes known to be active in that cell type, we produce a consensus predictive network that reflects biological reality more closely than those made by prior efforts using more generalized, context-independent methods. In addition, we show how machine learning efforts may be misled if the tissue specific role of mammalian proteins is not defined in the training set and circumscribed in the evidential data. For this study, we assembled an extensive compendium of mESC data: ∼2.2 million data points, collected from 60 different studies, under 992 conditions. We then integrated these data into a consensus mESC functional relationship network focused on biological processes associated with embryonic stem cell self-renewal and cell fate determination. Computational evaluations, literature validation, and analyses of predicted functional linkages show that our results are highly accurate and biologically relevant. Our mESC network predicts many novel players involved in self-renewal and serves as the foundation for future pluripotent stem cell studies. This network can be used by stem cell researchers (at http://StemSight.org to explore hypotheses about gene function in the context of self-renewal and to prioritize genes of interest for experimental validation.

  20. Identification of p63+ keratinocyte progenitor cells in circulation and their matrix-directed differentiation to epithelial cells

    OpenAIRE

    Nair, Renjith P; Krishnan, Lissy K

    2013-01-01

    Introduction In the event of chronic diabetes or burn wounds, accomplishing skin regeneration is a major concern. Autologous skin grafting is the most effective remedy, but the tissue harvest may create more nonhealing wounds. Currently available skin substitutes have a limited clinical outcome because of immune reactions arising from the xenobiotic scaffold or allogenous cells. Autologous stem cells that can be collected without an additional injury may be a viable option for skin-tissue eng...

  1. Analysis of cell flow and cell loss following X-irradiation using sequential investigation of the total number of cells in the various parts of the cell cycle

    International Nuclear Information System (INIS)

    Skog, S.; Tribukait, B.

    1985-01-01

    The cell flow and cell loss of an in vivo growing Ehrlich ascites tumour were calculated by sequential estimation of changes in total number of cells in the cell cycle compartments. Normal growth was compared with the grossly disturbed cell flow evident after a 5 Gy X-irradiation. The doubling time of normal, exponentially growing cells was 24 hr. The generation time was 21 hr and the potential doubling time was 21 hr. Thus, the growth fraction was 1.0 and the cell loss rate about 0.5%/hr. Following irradiation, a transiently increased relative outflow rate from all cell cycle compartments was found at about 3 and 40 hr, and from S phase at 24 hr after irradiation. Increase in cell loss as well as non-viable cells was observed at 24 hr after irradiation at the time of release of the irradiation-induced G 2 blockage. The experiments show the applicability and limitations of cell flow and cell loss calculations by sequential analysis of the total number of cells in the various parts of the cell cycle. (author)

  2. Isolation, Characterization, and Differentiation of Stem Cells for Cartilage Regeneration

    OpenAIRE

    Beane, Olivia S.; Darling, Eric M.

    2012-01-01

    The goal of tissue engineering is to create a functional replacement for tissues damaged by injury or disease. In many cases, impaired tissues cannot provide viable cells, leading to the investigation of stem cells as a possible alternative. Cartilage, in particular, may benefit from the use of stem cells since the tissue has low cellularity and cannot effectively repair itself. To address this need, researchers are investigating the chondrogenic capabilities of several multipotent stem cell ...

  3. Genetic options for improving fodder yield and quality in forage sorghum

    Directory of Open Access Journals (Sweden)

    C. Aruna

    2015-01-01

    Full Text Available Improving yield and quality of fodder from forage sorghum is important, especially in the semi-arid tropics, where sorghum is a major source of fodder. The aim of this work was to understand the genetic basis of fodder yield and quality traits, and character associations, and to estimate combining ability of the parents. The experiment was carried out during 2 successive rainy seasons using 10 parents crossed in a half-diallel design. Significant differences among the genotypes for fodder yield, quality and cell wall constituents were observed. Important quality traits, crude protein and digestibility (IVOMD, were not correlated with fodder yield, indicating the potential to improve yield and quality simultaneously in forage sorghum. General combining ability and specific combining ability variances showed that, for almost all characters, both additive and non-additive gene effects were important, with a predominance of non-additive effects. Parental lines SEVS4, HC308 and UPMC503 were good general combiners for yield and quality. The brown midrib lines, EC582508 and EC582510, were good general combiners for low lignin and high IVOMD. Strategies for improving forage sorghum to suit animal and biofuel industries are discussed.Keywords: Digestibility, crude protein, ADL, diallel analysis, gene effects.DOI: 10.17138/TGFT(349-58

  4. Live cell imaging reveals different modes of cytotoxic action of extracts derived from commonly used luting cements.

    Science.gov (United States)

    Trumpaitė-Vanagienė, Rita; Čebatariūnienė, Alina; Tunaitis, Virginijus; Pūrienė, Alina; Pivoriūnas, Augustas

    2018-02-01

    To compare cytotoxicity of extracts derived from commonly used luting cements: Hoffmann's Zinc Phosphate (ZPC), GC Fuji Plus Resin Modified Glass Ionomer (RMGIC) and 3M ESPE RelyX Unicem Resin Cement (RC) on primary human gingival fibroblasts (HGFs). HGFs were exposed to different concentrations of the ZPC, RMGIC and RC extracts. The cytotoxicity was assessed with the PrestoBlue Cell Viability Reagent and viable cells were counted by a haemocytometer using the trypan blue exclusion test. In order to determine the primary mechanism of the cell death induced by extracts from different luting cements, the real-time monitoring of caspase-3/-7 activity and membrane integrity of cells was employed. The extracts from the RMGIC and ZPC decreased the metabolic activity and numbers of viable cells. Unexpectedly, the extracts from the RC evoked only small effects on the metabolic activity of HGFs with a decreasing number of viable cells in a dose-and time-dependent manner. The live cell imaging revealed that the apoptosis was the primary mechanism of a cell death induced by the extracts derived from the RMGIC, whereas the extracts from the RC and ZPC induced a cell death through a necrotic and caspase-independent pathway. The apoptosis was the primary mechanism of the cell death induced by the extracts derived from the RMGIC, whereas the extracts from the RC and ZPC induced a cell death via a necrotic pathway. We suggest that metabolic assays commonly used to assess the cytotoxicity of luting cements should be validated by alternative methods. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Electrical Characterization of HIT type solar cells

    NARCIS (Netherlands)

    Rath, J.K.

    2012-01-01

    The silicon heterojunction solar cell (SHJ) has made rapid progress in reaching high efficiency and it is already developed as an industrially viable product. However, much of its progress has come through process development while there is scarce knowledge on the microscopic nature of the

  6. cAMP-CRP acts as a key regulator for the viable but non-culturable state in Escherichia coli.

    Science.gov (United States)

    Nosho, Kazuki; Fukushima, Hiroko; Asai, Takehiro; Nishio, Masahiro; Takamaru, Reiko; Kobayashi-Kirschvink, Koseki Joseph; Ogawa, Tetsuhiro; Hidaka, Makoto; Masaki, Haruhiko

    2018-03-01

    A variety of bacteria, including Escherichia coli, are known to enter the viable but non-culturable (VBNC) state under various stress conditions. During this state, cells lose colony-forming activities on conventional agar plates while retaining signs of viability. Diverse environmental stresses including starvation induce the VBNC state. However, little is known about the genetic mechanism inducing this state. Here, we aimed to reveal the genetic determinants of the VBNC state of E. coli. We hypothesized that the VBNC state is a process wherein specific gene products important for colony formation are depleted during the extended period of stress conditions. If so, higher expression of these genes would maintain colony-forming activities, thereby restraining cells from entering the VBNC state. From an E. coli plasmid-encoded ORF library, we identified genes that were responsible for maintaining high colony-forming activities after exposure to starvation condition. Among these, cpdA encoding cAMP phosphodiesterase exhibited higher performance in the maintenance of colony-forming activities. As cpdA overexpression decreases intracellular cAMP, cAMP or its complex with cAMP-receptor protein (CRP) may negatively regulate colony-forming activities under stress conditions. We confirmed this using deletion mutants lacking adenylate cyclase or CRP. These mutants fully maintained colony-forming activities even after a long period of starvation, while wild-type cells lost most of this activity. Thus, we concluded that the lack of cAMP-CRP effectively retains high colony-forming activities, indicating that cAMP-CRP acts as a positive regulator necessary for the induction of the VBNC state in E. coli.

  7. Yield and oil composition of 38 basil (Ocimum basilicum L.) accessions grown in Mississippi.

    Science.gov (United States)

    Zheljazkov, Valtcho D; Callahan, Amber; Cantrell, Charles L

    2008-01-09

    A field experiment was conducted to assess yield, oil content, and composition of 38 genotypes of sweet basil ( Ocimum basilicum L.). Overall, biomass yields were high and comparable to those reported in the literature. However, basil genotypes differed significantly with respect to oil content and composition. Oil content of the tested accessions varied from 0.07% to 1.92% in dry herbage. On the basis of the oil composition, basil accessions were divided into seven groups: (1) high-linalool chemotype [19-73% (-)-linalool], (2) linalool-eugenol chemotype [six chemotypes with 28-66% (-)-linalool and 5-29% eugenol], (3) methyl chavicol chemotype [six accessions with 20-72% methyl chavicol and no (-)-linalool], (4) methyl chavicol-linalool chemotype [six accessions with 8-29% methyl chavicol and 8-53% (-)-linalool], (5) methyl eugenol-linalool chemotype [two accessions with 37% and 91% methyl eugenol and 60% and 15% (-)-linalool], (6) methyl cinnamate-linalool chemotype [one accession with 9.7% methyl cinnamate and 31% (-)-linalool], and (7) bergamotene chemotype [one accession with bergamotene as major constituent, 5% eucalyptol, and basil could be a viable essential oil crop in Mississippi. The availability of various chemotypes offers the opportunity for production of basil to meet the market requirements of specific basil oils or individual compounds such as (-)-linalool, eugenol, methyl chavicol, methyl cinnamate, or methyl eugenol.

  8. Fake Journals: Their Features and Some Viable Ways to Distinguishing Them

    DEFF Research Database (Denmark)

    Hemmat Esfe, Mohammad; Wongwises, Somchai; Asadi, Amin

    2015-01-01

    In this paper, we aim to discuss the fake journals and their advertisement and publication techniques. These types of journals mostly start and continue their activities by using the name of some indexed journals and establishing fake websites. The fake journals and publishers, while asking...... the authors for a significant amount of money for publishing their papers, have no peer-review process, publish the papers without any revision on the fake sites, and put the scientific reputation and prestige of the researchers in jeopardy. In the rest of the paper, we present some viable techniques in order...

  9. Effects of Land-Applied Ammonia Scrubber Solutions on Yield, Nitrogen Uptake, Soil Test Phosphorus, and Phosphorus Runoff.

    Science.gov (United States)

    Martin, Jerry W; Moore, Philip A; Li, Hong; Ashworth, Amanda J; Miles, Dana M

    2018-03-01

    Ammonia (NH) scrubbers reduce amounts of NH and dust released from animal rearing facilities while generating nitrogen (N)-rich solutions, which may be used as fertilizers. The objective of this study was to determine the effects of various NH scrubber solutions on forage yields, N uptake, soil-test phosphorus (P), and P runoff. A small plot study was conducted using six treatments: (i) an unfertilized control, (ii) potassium bisulfate (KHSO) scrubber solution, (iii) aluminum sulfate [Al(SO) ⋅14HO, alum] scrubber solution, (iv) sodium bisulfate (NaHSO) scrubber solution, (v) sulfuric acid (HSO) scrubber solution, and (vi) ammonium nitrate (NHNO) fertilizer. The scrubber solutions were obtained from ARS Air Scrubbers attached to commercial broiler houses. All N sources were applied at a rate of 112 kg N ha. Plots were harvested approximately every 4 wk and soil-test P measurements were made, then a rainfall simulation study was conducted. Cumulative forage yields were greater ( scrubber solutions than for alum (6.7 Mg ha) or HSO (6.5 Mg ha) scrubber solutions or for NHNO (6.9 Mg ha). All N sources resulted in higher yields than the control (5.1 Mg ha). The additional potassium in the KHSO treatment likely resulted in higher yields. Although Mehlich-III-extractable P was not affected, water-extractable P in soil was lowered by the alum-based scrubber solution, which also resulted in lower P runoff. This study demonstrates that N captured using NH scrubbers is a viable N fertilizer. Copyright © by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America, Inc.

  10. Bond yield curve construction

    Directory of Open Access Journals (Sweden)

    Kožul Nataša

    2014-01-01

    Full Text Available In the broadest sense, yield curve indicates the market's view of the evolution of interest rates over time. However, given that cost of borrowing it closely linked to creditworthiness (ability to repay, different yield curves will apply to different currencies, market sectors, or even individual issuers. As government borrowing is indicative of interest rate levels available to other market players in a particular country, and considering that bond issuance still remains the dominant form of sovereign debt, this paper describes yield curve construction using bonds. The relationship between zero-coupon yield, par yield and yield to maturity is given and their usage in determining curve discount factors is described. Their usage in deriving forward rates and pricing related derivative instruments is also discussed.

  11. Evidence that cell surface charge reduction modifes capillary red cell velocity-flux relationships in hamster cremaster muscle

    NARCIS (Netherlands)

    Vink, H.; Wieringa, P. A.; Spaan, J. A.

    1995-01-01

    1. From capillary red cell velocity (V)-flux (F) relationships of hamster cremaster muscle a yield velocity (VF = 0) can be derived at which red cell flux is zero. Red cell velocity becomes intermittent and/or red blood cells come to a complete standstill for velocities close to this yield velocity,

  12. Measuring the external quantum efficiency of two-terminal polymer tandem solar cells

    NARCIS (Netherlands)

    Gilot, J.; Wienk, M.M.; Janssen, R.A.J.

    2010-01-01

    Tandem configurations, in which two cells are stacked and connected in series, offer a viable approach to further increase the power conversion efficiency (PCE) of organic solar cells. To enable the future rational design of new materials it is important to accurately assess the contributions of

  13. Cosmic constraint on massive neutrinos in viable f(R) gravity with producing ΛCDM background expansion

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Jianbo; Wu, Yabo; Wang, Yan; Yang, Weiqiang [Liaoning Normal University, Department of Physics, Dalian (China); Liu, Molin [Xinyang Normal University, Department of Physics, Xinyang (China)

    2016-12-15

    Tensions between several cosmic observations were found recently, such as the inconsistent values of H{sub 0} (or σ{sub 8}) were indicated by the different cosmic observations. Introducing the massive neutrinos in ΛCDM could potentially solve the tensions. Viable f(R) gravity producing ΛCDM background expansion with massive neutrinos is investigated in this paper. We fit the current observational data: Planck-2015 CMB, RSD, BAO, and SNIa to constrain the mass of neutrinos in viable f(R) theory. The constraint results at 95% confidence level are: Σm{sub ν} < 0.202 eV for the active-neutrino case, m{sub ν,sterile}{sup eff} < 0.757 eV with N{sub eff} < 3.22 for the sterile neutrino case. For the effects due to the mass of the neutrinos, the constraint results on model parameter at 95% confidence level become f{sub R0} x 10{sup -6} > -1.89 and f{sub R0} x 10{sup -6} > -2.02 for two cases, respectively. It is also shown that the fitting values of several parameters much depend on the neutrino properties, such as the cold dark matter density, the cosmological quantities at matter-radiation equality, the neutrino density and the fraction of baryonic mass in helium. Finally, the constraint result shows that the tension between direct and CMB measurements of H{sub 0} gets slightly weaker in the viable f(R) model than that in the base ΛCDM model. (orig.)

  14. Variations in Volatile Oil Yield and Composition of "Xin-yi" (Magnolia biondii Pamp. Flower Buds) at Different Growth Stages.

    Science.gov (United States)

    Hu, Mingli; Bai, Mei; Ye, Wei; Wang, Yaling; Wu, Hong

    2018-06-01

    Dried flower buds of Magnolia biondii Pamp. are the main ingredient in "Xin-yi" in China, and the volatile oils of M. biondii flower buds are the principal medicinal component. Gas chromatographymass spectrometry (GC-MS) and microscopic techniques were employed to detect the volatile yields of M. biondii flowers at various growth stages. The volatile oil yields of M. biondii flowers differed significantly at different growth stages and were closely related to flower dry weight, oil cell density and degree of oil accumulation. In February 2016, flower buds had the highest dry weight, the maximum percentage of oil cells at the oil saturation stage and the highest density of oil cells, which coincided with the highest oil yield. In March 2016, flower buds had a lower dry weight, a higher percentage of oil cells at the oil-degrading stage and the lowest oil cell density, resulting in decreased oil yields. The total amounts of the major medicinal components in the M. biondii flower also showed regular changes at different growth stages. In January and February of 2016, M. biondii flowers had a higher dry weight, volatile oil yield and total content of medicinal ingredients, which was the best time for harvesting high-quality medicinal components. Our study reveals that volatile oil content and chemical composition are closely related to the growth stage of M. biondii flower buds. The results provide a scientific morphology and composition index for evaluating the medicinal value and harvesting of high-quality M. biondii medicinal herbs.

  15. 7755 EFFECT OF NPK FERTILIZER ON FRUIT YIELD AND YIELD ...

    African Journals Online (AJOL)

    Win7Ent

    2013-06-03

    Jun 3, 2013 ... peasant farmers in Nigeria. With the increased ... did not significantly (p=0.05) increase the fruit yield nor the seed yield. Key words: NPK fertilizer, Fruit ..... SAS (Statistical Analysis System) Version 9.1. SAS Institute Inc., Cary, ...

  16. Time-course correlation of biofilm properties and electrochemical performance in single-chamber microbial fuel cells

    KAUST Repository

    Ren, Zhiyong; Ramasamy, Ramaraja P.; Cloud-Owen, Susan Red; Yan, Hengjing; Mench, Matthew M.; Regan, John M.

    2011-01-01

    The relationship between anode microbial characteristics and electrochemical parameters in microbial fuel cells (MFCs) was analyzed by time-course sampling of parallel single-bottle MFCs operated under identical conditions. While voltage stabilized within 4. days, anode biofilms continued growing during the six-week operation. Viable cell density increased asymptotically, but membrane-compromised cells accumulated steadily from only 9% of total cells on day 3 to 52% at 6. weeks. Electrochemical performance followed the viable cell trend, with a positive correlation for power density and an inverse correlation for anode charge transfer resistance. The biofilm architecture shifted from rod-shaped, dispersed cells to more filamentous structures, with the continuous detection of Geobacter sulfurreducens-like 16S rRNA fragments throughout operation and the emergence of a community member related to a known phenazine-producing Pseudomonas species. A drop in cathode open circuit potential between weeks two and three suggested that uncontrolled biofilm growth on the cathode deleteriously affects system performance. © 2010 Elsevier Ltd.

  17. Social Networking and Smart Technology: Viable Environmental Communication Tools…?

    Science.gov (United States)

    Montain, J.; Byrne, J. M.

    2010-12-01

    To what extent do popular social networking channels represent a viable means for disseminating information regarding environmental change to the general public? Are new forms of communication such as YouTube™, Facebook™, MySpace™ and Twitter™ and smart devices such as iPhone™ and BlackBerry™ useful and effective in terms motivating people into social action and behavioural modification; or do they simply pay ‘lip service’ to these pressing environmental issues? This project will explore the background connections between social networking and environmental communication and education; and outline why such tools might be an appropriate way to connect to a broad audience in an efficient and unconventional manner. Further, research will survey the current prevalence of reliable environmental change information on social networking Internet-based media; and finally, suggestions for improved strategies and new directions will be provided.

  18. Stem-Cell Inactivation on Transplantation of Haemopoietic Cell Suspensions from Genetically Different Donors

    Energy Technology Data Exchange (ETDEWEB)

    Petrov, R. V. [Institute of Biophysics, Ministry of Public Health of the USSR, Moscow, USSR (Russian Federation)

    1969-07-15

    The transplantation of a mixture of haemopoietic or lymphoid cells from two genetically different mice into lethally irradiated F{sub 1} recipients results in marked or total inactivation of the colony-forming units of the graft. This phenomenon is observed following transplantation of mixtures of spleen cells or bone-marrow cells from animals of different genotypes: CBA + C57BL, A + CBA, A + C57BL, C3H + C57BL, CBA + (CBA x C57BL) F{sub 1}. Maximum inactivation is observed when lymph-node cells of one genotype are transplanted with spleen or bone-marrow cells of another genotype. Use of non-syngenic kidney cells or lymphoid cells inactivated by irradiation as one component of the mixture shows that inactivation of genetically heterogeneous stem cells requires the participation of viable lymphoid cells. The inactivation phenomenon is also observed with Jerne's method. This shows that inactivation affects not only colony-forming cells but also the immunologically competent precursors of antibody-producing cells. (author)

  19. Development of a high yielding E. coli periplasmic expression system for the production of humanized Fab' fragments.

    Science.gov (United States)

    Ellis, Mark; Patel, Pareshkumar; Edon, Marjory; Ramage, Walter; Dickinson, Robert; Humphreys, David P

    2017-01-01

    Humanized Fab' fragments may be produced in the periplasm of Escherichia coli but can be subject to degradation by host cell proteases. In order to increase Fab' yield and reduce proteolysis we developed periplasmic protease deficient strains of E. coli. These strains lacked the protease activity of Tsp, protease III and DegP. High cell density fermentations indicated Tsp deficient strains increased productivity two fold but this increase was accompanied by premature cell lysis soon after the induction of Fab' expression. To overcome the reduction in cell viability we introduced suppressor mutations into the spr gene. The mutations partially restored the wild type phenotype of the cells. Furthermore, we coexpressed a range of periplasmic chaperone proteins with the Fab', DsbC had the most significant impact, increasing humanized Fab' production during high cell density fermentation. When DsbC coexpression was combined with a Tsp deficient spr strain we observed an increase in yield and essentially restored "wild type" cell viability. We achieved a final periplasmic yield of over 2.4g/L (final cell density OD 600 105), 40 h post Fab' induction with minimal cell lysis.The data suggests that proteolysis, periplasm integrity, protein folding and disulphide bond formation are all potential limiting steps in the production of Fab' fragments in the periplasm of E. coli. In this body of work, we have addressed these limiting steps by utilizing stabilized protease deficient strains and chaperone coexpression. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:212-220, 2017. © 2016 American Institute of Chemical Engineers.

  20. 40 CFR 180.1011 - Viable spores of the microorganism Bacillus thuringiensis Berliner; exemption from the...

    Science.gov (United States)

    2010-07-01

    ... EXEMPTIONS FOR PESTICIDE CHEMICAL RESIDUES IN FOOD Exemptions From Tolerances § 180.1011 Viable spores of the... characteristics of the parent strain or contamination by other microorganisms. (3) Each lot of spore preparation... production is a Bacillus thuringiensis strain which does not produce β-exotoxin under standard manufacturing...

  1. Response of Yield and Yield Components of Tef [Eragrostis Tef ...

    African Journals Online (AJOL)

    The partial budget analysis also indicates that applications of 46 kg. N ha-1 and 10 kg P ha-1 are ..... (1994) indicated that where the grain yield response is negative, yield reduction is primarily caused by a .... An Economic Training. Manual.

  2. Cell biology, MRI and geometry: insight into a microscopic/macroscopic marriage.

    Science.gov (United States)

    de Oliveira, Sérgio Almeida; Gowdak, Luís Henrique W; Buckberg, Gerald; Krieger, José Eduardo

    2006-04-01

    The concept of cell therapy as an adjunctive therapy to myocardial surgical revascularization for patients with severe coronary artery disease is illustrated by two case reports of ischemic cardiac disease that were unsuitable for revascularization by coronary grafting. The potential interaction of cell therapy, magnetic resonance imaging (MRI) of viability, and left ventricle (LV) restoration is described. Each patient had an ejection fraction below 30%, a relatively conical heart, and MRI gadolinium scan showing predominantly viable muscle. Intramyocardial injections of autologous bone marrow-derived cells (BMC) were performed along with either incomplete coronary artery bypass grafting (CABG) (to mother regions) or with transmyocardial laser revascularization (TMLR). An improvement in contractile function was seen at 6-12-month intervals after the procedure. The implications of possible underlying mechanisms of improvement in both myocardial perfusion and contractility suggest the striking importance of both micro- and macroenvironment for any cell-based therapeutic strategy. These observations imply that the interaction of cell biology, viability by MRI and geometry may be important in the future, as geometry can be restored surgically, and the new architectural form may develop enhanced function if it contains viable tissue and cell-based treatment can be delivered.

  3. Higher antibiotic yielding mutants of bacillus subtilis by gamma radiation

    International Nuclear Information System (INIS)

    Ahmad, M.S.; Shaukat, G.A.; Malik, M.A.

    1987-01-01

    When Bacillus Subtilis AECL69 was grown in malt extract-pepetone-molasses-sugar (MPMS) medium, it could produce antibiotic substance(s) with antibacterial and antifungal properties in the culture fluid. The bacterial cells grown in MPMS medium were washed and suspended into distilled water and irradiated with gamma rays in Gammacell 220 at different doses. Higher antibiotic yielding isolates (plus mutants) were obtained from cell pollutions irradiated at 15 Kr. These gamma rays-induced plus mutants showed simultaneous higher production of antibacterial as well as antifungal activity. (author)

  4. Natural recharge to sustainable yield from the barind aquifer: a tool in preparing effective management plan of groundwater resources.

    Science.gov (United States)

    Monirul Islam, Md; Kanungoe, P

    2005-01-01

    This paper presents the results of water balance study and aquifer simulation modeling for preliminary estimation of the recharge rate and sustainable yield for the semi arid Barind Tract region of Bangladesh. The outcomes of the study are likely to be useful for planning purposes. It is found from detailed water balance study for the area that natural recharge rates in the Barind Tract vary widely year to year. It may have resulted from the method used for the calculation. If the considered time interval had been smaller than the monthly rainfall, the results could have been different. Aquifer Simulation Modeling (ASM) for the Barind aquifer is used to estimate long-term sustainable yield of the groundwater considering limiting drawdown from the standpoint of economic pumping cost. In managing a groundwater basin efficiently and effectively, evaluation of the maximum annual groundwater yield of the basin that can be withdrawn and used without producing any undesirable effect is one of the most important issues. In investigating such recharge rate, introduction of certain terms such as sustainable yield and safe yield has been accompanied. Development of this area involves proper utilization of this vast land, which is possible only through ensured irrigation for agriculture. The Government of Bangladesh has a plan to develop irrigation facilities by optimum utilization of available ground and surface water. It is believed that the groundwater table is lowering rapidly and the whole region is in an acute state of deforestation. Indiscriminate groundwater development may accelerate deforestation trend. In this context estimation of actual natural recharge rate to the aquifer and determination of sustainable yield will assist in proper management and planning of environmentally viable abstraction schemes. It is revealed from the study that the sustainable yield of ground water (204 mm/y) is somewhat higher than the long-term annual average recharge (152.7 mm) to the

  5. Economically Viable Components from Jerusalem Artichoke (Helianthus tuberosus L.) in a Biorefinery Concept

    Science.gov (United States)

    Johansson, Eva; Prade, Thomas; Angelidaki, Irini; Svensson, Sven-Erik; Newson, William R.; Gunnarsson, Ingólfur Bragi; Persson Hovmalm, Helena

    2015-01-01

    Biorefinery applications are receiving growing interest due to climatic and waste disposal issues and lack of petroleum resources. Jerusalem artichoke (Helianthus tuberosus L.) is suitable for biorefinery applications due to high biomass production and limited cultivation requirements. This paper focuses on the potential of Jerusalem artichoke as a biorefinery crop and the most viable products in such a case. The carbohydrates in the tubers were found to have potential for production of platform chemicals, e.g., succinic acid. However, economic analysis showed that production of platform chemicals as a single product was too expensive to be competitive with petrochemically produced sugars. Therefore, production of several products from the same crop is a must. Additional products are protein based ones from tubers and leaves and biogas from residues, although both are of low value and amount. High bioactive activity was found in the young leaves of the crop, and the sesquiterpene lactones are of specific interest, as other compounds from this group have shown inhibitory effects on several human diseases. Thus, future focus should be on understanding the usefulness of small molecules, to develop methods for their extraction and purification and to further develop sustainable and viable methods for the production of platform chemicals. PMID:25913379

  6. Automated Manufacturing of Potent CD20-Directed Chimeric Antigen Receptor T Cells for Clinical Use.

    Science.gov (United States)

    Lock, Dominik; Mockel-Tenbrinck, Nadine; Drechsel, Katharina; Barth, Carola; Mauer, Daniela; Schaser, Thomas; Kolbe, Carolin; Al Rawashdeh, Wael; Brauner, Janina; Hardt, Olaf; Pflug, Natali; Holtick, Udo; Borchmann, Peter; Assenmacher, Mario; Kaiser, Andrew

    2017-10-01

    The clinical success of gene-engineered T cells expressing a chimeric antigen receptor (CAR), as manifested in several clinical trials for the treatment of B cell malignancies, warrants the development of a simple and robust manufacturing procedure capable of reducing to a minimum the challenges associated with its complexity. Conventional protocols comprise many open handling steps, are labor intensive, and are difficult to upscale for large numbers of patients. Furthermore, extensive training of personnel is required to avoid operator variations. An automated current Good Manufacturing Practice-compliant process has therefore been developed for the generation of gene-engineered T cells. Upon installation of the closed, single-use tubing set on the CliniMACS Prodigy™, sterile welding of the starting cell product, and sterile connection of the required reagents, T cells are magnetically enriched, stimulated, transduced using lentiviral vectors, expanded, and formulated. Starting from healthy donor (HD) or lymphoma or melanoma patient material (PM), the robustness and reproducibility of the manufacturing of anti-CD20 specific CAR T cells were verified. Independent of the starting material, operator, or device, the process consistently yielded a therapeutic dose of highly viable CAR T cells. Interestingly, the formulated product obtained with PM was comparable to that of HD with respect to cell composition, phenotype, and function, even though the starting material differed significantly. Potent antitumor reactivity of the produced anti-CD20 CAR T cells was shown in vitro as well as in vivo. In summary, the automated T cell transduction process meets the requirements for clinical manufacturing that the authors intend to use in two separate clinical trials for the treatment of melanoma and B cell lymphoma.

  7. Hybrid cell adhesive material for instant dielectrophoretic cell trapping and long-term cell function assessment.

    Science.gov (United States)

    Reyes, Darwin R; Hong, Jennifer S; Elliott, John T; Gaitan, Michael

    2011-08-16

    Dielectrophoresis (DEP) for cell manipulation has focused, for the most part, on approaches for separation/enrichment of cells of interest. Advancements in cell positioning and immobilization onto substrates for cell culture, either as single cells or as cell aggregates, has benefited from the intensified research efforts in DEP (electrokinetic) manipulation. However, there has yet to be a DEP approach that provides the conditions for cell manipulation while promoting cell function processes such as cell differentiation. Here we present the first demonstration of a system that combines DEP with a hybrid cell adhesive material (hCAM) to allow for cell entrapment and cell function, as demonstrated by cell differentiation into neuronlike cells (NLCs). The hCAM, comprised of polyelectrolytes and fibronectin, was engineered to function as an instantaneous cell adhesive surface after DEP manipulation and to support long-term cell function (cell proliferation, induction, and differentiation). Pluripotent P19 mouse embryonal carcinoma cells flowing within a microchannel were attracted to the DEP electrode surface and remained adhered onto the hCAM coating under a fluid flow field after the DEP forces were removed. Cells remained viable after DEP manipulation for up to 8 d, during which time the P19 cells were induced to differentiate into NLCs. This approach could have further applications in areas such as cell-cell communication, three-dimensional cell aggregates to create cell microenvironments, and cell cocultures.

  8. Synthesis and biologic studies of iodinated (125I/127I) ethidium

    International Nuclear Information System (INIS)

    Ho Nanhui; Tumeh, Paul C.; Kassis, Amin I.

    2001-01-01

    An iodinated ( 125 I/ 127 I) ethidium derivative (3,8-diamino-5-[6'-(p-iodobenzoylamino)-4'-azahexyl]-6 -phenylphenanthridinium chloride hydrochloride) was synthesized and characterized. The labeling yield of the 125 I-labeled derivative was 75% for carrier-free 125 I, with a radiochemical purity of 95%. The incubation of iodoethidium with calf thymus DNA resulted in a substantial enhancement of fluorescence yield, indicating the intercalation of this compound into DNA. In the presence of iodoethidium, the nuclei of methanol-treated mammalian cells fluoresced, while those of viable cells did not (since the plasma membrane is impermeable to iodoethidium). When viable cells were incubated with the reduced form of the derivative, 125 I/ 127 I-dihydroethidium traversed the plasma membrane, was oxidized in the cytoplasm, and intercalated into nuclear DNA. Finally, we tested the hypothesis that larger malignant solid tumors, containing a relatively greater percentage of degenerating permeable cells, can be targeted with 125 I-ethidium. In-vivo studies demonstrated a small but positive correlation (R = 0.72) between tumor volume and the uptake of the derivative. Because of the ubiquitous presence of abnormal permeable cells and necrosis in tumors, our results support the belief that radiolabeled DNA-intercalating or DNA-binding molecules may be of diagnostic and therapeutic value for a variety of solid tumors in humans

  9. Synthesis and biologic studies of iodinated ({sup 125}I/{sup 127}I) ethidium

    Energy Technology Data Exchange (ETDEWEB)

    Ho Nanhui; Tumeh, Paul C.; Kassis, Amin I. E-mail: amin_kassis@hms.harvard.edu

    2001-11-01

    An iodinated ({sup 125}I/{sup 127}I) ethidium derivative (3,8-diamino-5-[6'-(p-iodobenzoylamino)-4'-azahexyl]-6 -phenylphenanthridinium chloride hydrochloride) was synthesized and characterized. The labeling yield of the {sup 125}I-labeled derivative was 75% for carrier-free {sup 125}I, with a radiochemical purity of 95%. The incubation of iodoethidium with calf thymus DNA resulted in a substantial enhancement of fluorescence yield, indicating the intercalation of this compound into DNA. In the presence of iodoethidium, the nuclei of methanol-treated mammalian cells fluoresced, while those of viable cells did not (since the plasma membrane is impermeable to iodoethidium). When viable cells were incubated with the reduced form of the derivative, {sup 125}I/{sup 127}I-dihydroethidium traversed the plasma membrane, was oxidized in the cytoplasm, and intercalated into nuclear DNA. Finally, we tested the hypothesis that larger malignant solid tumors, containing a relatively greater percentage of degenerating permeable cells, can be targeted with {sup 125}I-ethidium. In-vivo studies demonstrated a small but positive correlation (R = 0.72) between tumor volume and the uptake of the derivative. Because of the ubiquitous presence of abnormal permeable cells and necrosis in tumors, our results support the belief that radiolabeled DNA-intercalating or DNA-binding molecules may be of diagnostic and therapeutic value for a variety of solid tumors in humans.

  10. Correlation and path-cofficient analysis of seed yield and yield ...

    African Journals Online (AJOL)

    This study was undertaken in order to determine the association among yield components and their direct and indirect effects on the seed yield of confectionery sunflower. 36 confectionery sunflower populations originated from different regions of Northwest Iran were characterized using 11 agromorphological traits ...

  11. Cell-to-cell communication and cellular environment alter the somatostatin status of delta cells

    Energy Technology Data Exchange (ETDEWEB)

    Kelly, Catriona, E-mail: catriona.kelly@qub.ac.uk [SAAD Centre for Pharmacy and Diabetes, School of Biomedical Sciences, University of Ulster, Coleraine (United Kingdom); Flatt, Peter R.; McClenaghan, Neville H. [SAAD Centre for Pharmacy and Diabetes, School of Biomedical Sciences, University of Ulster, Coleraine (United Kingdom)

    2010-08-20

    Research highlights: {yields} TGP52 cells display enhanced functionality in pseudoislet form. {yields} Somatostatin content was reduced, but secretion increased in high glucose conditions. {yields} Cellular interactions and environment alter the somatostatin status of TGP52 cells. -- Abstract: Introduction: Somatostatin, released from pancreatic delta cells, is a potent paracrine inhibitor of insulin and glucagon secretion. Islet cellular interactions and glucose homeostasis are essential to maintain normal patterns of insulin secretion. However, the importance of cell-to-cell communication and cellular environment in the regulation of somatostatin release remains unclear. Methods: This study employed the somatostatin-secreting TGP52 cell line maintained in DMEM:F12 (17.5 mM glucose) or DMEM (25 mM glucose) culture media. The effect of pseudoislet formation and culture medium on somatostatin content and release in response to a variety of stimuli was measured by somatostatin EIA. In addition, the effect of pseudoislet formation on cellular viability (MTT and LDH assays) and proliferation (BrdU ELISA) was determined. Results: TGP52 cells readily formed pseudoislets and showed enhanced functionality in three-dimensional form with increased E-cadherin expression irrespective of the culture environment used. However, culture in DMEM decreased cellular somatostatin content (P < 0.01) and increased somatostatin secretion in response to a variety of stimuli including arginine, calcium and PMA (P < 0.001) when compared with cells grown in DMEM:F12. Configuration of TGP52 cells as pseudoislets reduced the proliferative rate and increased cellular cytotoxicity irrespective of culture medium used. Conclusions: Somatostatin secretion is greatly facilitated by cell-to-cell interactions and E-cadherin expression. Cellular environment and extracellular glucose also significantly influence the function of delta cells.

  12. Recellularization via the bile duct supports functional allogenic and xenogenic cell growth on a decellularized rat liver scaffold.

    Science.gov (United States)

    Hassanein, Wessam; Uluer, Mehmet C; Langford, John; Woodall, Jhade D; Cimeno, Arielle; Dhru, Urmil; Werdesheim, Avraham; Harrison, Joshua; Rivera-Pratt, Carlos; Klepfer, Stephen; Khalifeh, Ali; Buckingham, Bryan; Brazio, Philip S; Parsell, Dawn; Klassen, Charlie; Drachenberg, Cinthia; Barth, Rolf N; LaMattina, John C

    2017-01-02

    Recent years have seen a proliferation of methods leading to successful organ decellularization. In this experiment we examine the feasibility of a decellularized liver construct to support growth of functional multilineage cells. Bio-chamber systems were used to perfuse adult rat livers with 0.1% SDS for 24 hours yielding decellularized liver scaffolds. Initially, we recellularized liver scaffolds using a human tumor cell line (HepG2, introduced via the bile duct). Subsequent studies were performed using either human tumor cells co-cultured with human umbilical vein endothelial cells (HUVECs, introduced via the portal vein) or rat neonatal cell slurry (introduced via the bile duct). Bio-chambers were used to circulate oxygenated growth medium via the portal vein at 37C for 5-7 days. Human HepG2 cells grew readily on the scaffold (n = 20). HepG2 cells co-cultured with HUVECs demonstrated viable human endothelial lining with concurrent hepatocyte growth (n = 10). In the series of neonatal cell slurry infusion (n = 10), distinct foci of neonatal hepatocytes were observed to repopulate the parenchyma of the scaffold. The presence of cholangiocytes was verified by CK-7 positivity. Quantitative albumin measurement from the grafts showed increasing albumin levels after seven days of perfusion. Graft albumin production was higher than that observed in traditional cell culture. This data shows that rat liver scaffolds support human cell ingrowth. The scaffold likewise supported the engraftment and survival of neonatal rat liver cell slurry. Recellularization of liver scaffolds thus presents a promising model for functional liver engineering.

  13. Advantageous direct quantification of viable closely related probiotics in petit-suisse cheeses under in vitro gastrointestinal conditions by Propidium Monoazide--qPCR.

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    Martha Lissete Morales Villarreal

    Full Text Available Species-specific Quantitative Real Time PCR (qPCR alone and combined with the use of propidium monoazide (PMA were used along with the plate count method to evaluate the survival of the probiotic strains Lactobacillus acidophilus La-5 and Bifidobacterium animalis subsp. lactis Bb-12, and the bacteriocinogenic and potentially probiotic strain Lactobacillus sakei subsp. sakei 2a in synbiotic (F1 and probiotic (F2 petit-suisse cheeses exposed throughout shelf-life to in vitro simulated gastrointestinal tract conditions. The three strains studied showed a reduction in their viability after the 6 h assay. Bb-12 displayed the highest survival capacity, above 72.6 and 74.6% of the initial populations, respectively, by plate count and PMA-qPCR, maintaining population levels in the range or above 6 log CFU/g. The prebiotic mix of inulin and FOS did not offer any additional protection for the strains against the simulated gastrointestinal environment. The microorganisms' populations were comparable among the three methods at the initial time of the assay, confirming the presence of mainly viable and culturable cells. However, with the intensification of the stress induced throughout the various stages of the in vitro test, the differences among the methods increased. The qPCR was not a reliable enumeration method for the quantification of intact bacterial populations, mixed with large numbers of injured and dead bacteria, as confirmed by the scanning electron microscopy results. Furthermore, bacteria plate counts were much lower (P<0.05 than with the PMA-qPCR method, suggesting the accumulation of stressed or dead microorganisms unable to form colonies. The use of PMA overcame the qPCR inability to differentiate between dead and alive cells. The combination of PMA and species-specific qPCR in this study allowed a quick and unequivocal way of enumeration of viable closely related species incorporated into probiotic and synbiotic petit-suisse cheeses and

  14. Isolation of murine postnatal brain microglia for phenotypic characterization using magnetic cell separation technology.

    Science.gov (United States)

    Harms, Ashley S; Tansey, Malú G

    2013-01-01

    To shorten the time between brain harvesting and microglia isolation, and characterization, we utilized the MACS(®) neural dissociation kit followed by OctoMACS(®) CD11b magnetic bead isolation technique to positively select for brain microglia expressing the pan-microglial marker CD11b, a key subunit of the membrane attack complex (MAC). This protocol yields a viable and highly pure (>95%) microglial population of approximately 500,000 cells per pup that is amenable for in vitro characterization within hours or days after being harvested from brain tissue. Primary microglia from C57Bl/6 mice were plated for next-day analyses of morphology and cellular markers by immunocytochemistry or for analysis of gene expression under resting or LPS-stimulated conditions. The ease of isolation enables investigators to perform molecular and cellular analyses without having to wait 1-2 weeks to isolate microglia by conventional methods involving mechanical agitation to dislodge these from astrocyte beds.

  15. Soviet test yields

    Science.gov (United States)

    Vergino, Eileen S.

    Soviet seismologists have published descriptions of 96 nuclear explosions conducted from 1961 through 1972 at the Semipalatinsk test site, in Kazakhstan, central Asia [Bocharov et al., 1989]. With the exception of releasing news about some of their peaceful nuclear explosions (PNEs) the Soviets have never before published such a body of information.To estimate the seismic yield of a nuclear explosion it is necessary to obtain a calibrated magnitude-yield relationship based on events with known yields and with a consistent set of seismic magnitudes. U.S. estimation of Soviet test yields has been done through application of relationships to the Soviet sites based on the U.S. experience at the Nevada Test Site (NTS), making some correction for differences due to attenuation and near-source coupling of seismic waves.

  16. Cell proliferation, viability, and in vitro differentiation of equine mesenchymal stem cells seeded on bacterial cellulose hydrogel scaffolds

    International Nuclear Information System (INIS)

    Favi, Pelagie M.; Benson, Roberto S.; Neilsen, Nancy R.; Hammonds, Ryan L.; Bates, Cassandra C.; Stephens, Christopher P.; Dhar, Madhu S.

    2013-01-01

    The culture of multipotent mesenchymal stem cells on natural biopolymers holds great promise for treatments of connective tissue disorders such as osteoarthritis. The safety and performance of such therapies relies on the systematic in vitro evaluation of the developed stem cell-biomaterial constructs prior to in vivo implantation. This study evaluates bacterial cellulose (BC), a biocompatible natural polymer, as a scaffold for equine-derived bone marrow mesenchymal stem cells (EqMSCs) for application in bone and cartilage tissue engineering. An equine model was chosen due to similarities in size, load and types of joint injuries suffered by horses and humans. Lyophilized and critical point dried BC hydrogel scaffolds were characterized using scanning electron microscopy (SEM) to confirm nanostructure morphology which demonstrated that critical point drying induces fibre bundling unlike lyophilisation. EqMSCs positively expressed the undifferentiated pluripotent mesenchymal stem cell surface markers CD44 and CD90. The BC scaffolds were shown to be cytocompatible, supporting cellular adhesion and proliferation, and allowed for osteogenic and chondrogenic differentiation of EqMSCs. The cells seeded on the BC hydrogel were shown to be viable and metabolically active. These findings demonstrate that the combination of a BC hydrogel and EqMSCs are promising constructs for musculoskeletal tissue engineering applications. - Highlights: ► Critical point drying induces fibre bundling unlike lyophilisation. ► Cells positively expressed undifferentiated pluripotent stem cell markers. ► BCs were cytocompatible, supported cell adhesion, proliferation and differentiation ► Cells seeded on BC scaffolds were viable and metabolically active. ► Findings demonstrate that BC and EqMSCs are promising tissue engineered constructs

  17. Cell proliferation, viability, and in vitro differentiation of equine mesenchymal stem cells seeded on bacterial cellulose hydrogel scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Favi, Pelagie M.; Benson, Roberto S. [Department of Materials Science and Engineering, College of Engineering, University of Tennessee, Knoxville, TN 37996 (United States); Neilsen, Nancy R. [Department of Biomedical and Diagnostic Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996 (United States); Hammonds, Ryan L. [Department of Materials Science and Engineering, College of Engineering, University of Tennessee, Knoxville, TN 37996 (United States); Bates, Cassandra C. [Department of Large Animal Clinical Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996 (United States); Stephens, Christopher P. [Department of Surgery, Graduate School of Medicine, University of Tennessee, Knoxville, TN 37996 (United States); Center for Materials Processing, University of Tennessee, Knoxville, TN 37996 (United States); Dhar, Madhu S., E-mail: mdhar@utk.edu [Department of Large Animal Clinical Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996 (United States)

    2013-05-01

    The culture of multipotent mesenchymal stem cells on natural biopolymers holds great promise for treatments of connective tissue disorders such as osteoarthritis. The safety and performance of such therapies relies on the systematic in vitro evaluation of the developed stem cell-biomaterial constructs prior to in vivo implantation. This study evaluates bacterial cellulose (BC), a biocompatible natural polymer, as a scaffold for equine-derived bone marrow mesenchymal stem cells (EqMSCs) for application in bone and cartilage tissue engineering. An equine model was chosen due to similarities in size, load and types of joint injuries suffered by horses and humans. Lyophilized and critical point dried BC hydrogel scaffolds were characterized using scanning electron microscopy (SEM) to confirm nanostructure morphology which demonstrated that critical point drying induces fibre bundling unlike lyophilisation. EqMSCs positively expressed the undifferentiated pluripotent mesenchymal stem cell surface markers CD44 and CD90. The BC scaffolds were shown to be cytocompatible, supporting cellular adhesion and proliferation, and allowed for osteogenic and chondrogenic differentiation of EqMSCs. The cells seeded on the BC hydrogel were shown to be viable and metabolically active. These findings demonstrate that the combination of a BC hydrogel and EqMSCs are promising constructs for musculoskeletal tissue engineering applications. - Highlights: ► Critical point drying induces fibre bundling unlike lyophilisation. ► Cells positively expressed undifferentiated pluripotent stem cell markers. ► BCs were cytocompatible, supported cell adhesion, proliferation and differentiation ► Cells seeded on BC scaffolds were viable and metabolically active. ► Findings demonstrate that BC and EqMSCs are promising tissue engineered constructs.

  18. Dissolvable tattoo sensors: from science fiction to a viable technology

    Science.gov (United States)

    Cheng, Huanyu; Yi, Ning

    2017-01-01

    Early surrealistic painting and science fiction movies have envisioned dissolvable tattoo electronic devices. In this paper, we will review the recent advances that transform that vision into a viable technology, with extended capabilities even beyond the early vision. Specifically, we focus on the discussion of a stretchable design for tattoo sensors and degradable materials for dissolvable sensors, in the form of inorganic devices with a performance comparable to modern electronics. Integration of these two technologies as well as the future developments of bio-integrated devices is also discussed. Many of the appealing ideas behind developments of these devices are drawn from nature and especially biological systems. Thus, bio-inspiration is believed to continue playing a key role in future devices for bio-integration and beyond.

  19. Dissolvable tattoo sensors: from science fiction to a viable technology

    International Nuclear Information System (INIS)

    Cheng, Huanyu; Yi, Ning

    2017-01-01

    Early surrealistic painting and science fiction movies have envisioned dissolvable tattoo electronic devices. In this paper, we will review the recent advances that transform that vision into a viable technology, with extended capabilities even beyond the early vision. Specifically, we focus on the discussion of a stretchable design for tattoo sensors and degradable materials for dissolvable sensors, in the form of inorganic devices with a performance comparable to modern electronics. Integration of these two technologies as well as the future developments of bio-integrated devices is also discussed. Many of the appealing ideas behind developments of these devices are drawn from nature and especially biological systems. Thus, bio-inspiration is believed to continue playing a key role in future devices for bio-integration and beyond. (invited comment)

  20. An injectable calcium phosphate-alginate hydrogel-umbilical cord mesenchymal stem cell paste for bone tissue engineering

    Science.gov (United States)

    Zhao, Liang; Weir, Michael D.; Xu, Hockin H. K.

    2010-01-01

    The need for bone repair has increased as the population ages. Stem cell-scaffold approaches hold immense promise for bone tissue engineering. However, currently, preformed scaffolds for cell delivery have drawbacks including the difficulty to seed cells deep into the scaffold, and inability for injection in minimally invasive surgeries. Current injectable polymeric carriers and hydrogels are too weak for load-bearing orthopedic application. The objective of this study was to develop an injectable and mechanically-strong stem cell construct for bone tissue engineering. Calcium phosphate cement (CPC) paste was combined with hydrogel microbeads encapsulating human umbilical cord mesenchymal stem cells (hUCMSCs). The hUCMSC-encapsulating composite paste was fully injectable under small injection forces. Cell viability after injection matched that in hydrogel without CPC and without injection. Mechanical properties of the construct matched the reported values of cancellous bone, and were much higher than previous injectable polymeric and hydrogel carriers. hUCMSCs in the injectable constructs osteodifferentiated, yielding high alkaline phosphatase, osteocalcin, collagen type I, and osterix gene expressions at 7 d, which were 50–70 fold higher than those at 1 d. Mineralization by the hUCMSCs at 14 d was 100-fold that at 1 d. In conclusion, a fully-injectable, mechanically-strong, stem cell-CPC scaffold construct was developed. The encapsulated hUCMSCs remained viable, osteodifferentiated, and synthesized bone minerals. The new injectable stem cell construct with load-bearing capability may enhance bone regeneration in minimally-invasive and other orthopedic surgeries. PMID:20570346