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Sample records for hybridoma culture supernatants

  1. On-line characterization of a hybridoma cell culture process.

    Science.gov (United States)

    Zhou, W; Hu, W S

    1994-06-20

    The on-line determination of the physiological state of a cell culture process requires reliable on-line measurements of various parameters and calculations of specific rates from these measurements. The cell concentration of a hybridoma culture was estimated on-line by measuring optical density (OD) with a laser turbidity probe. The oxygen uptake rate (OUR) was determined by monitoring dynamically dissolved oxygen concentration profiles and closing oxygen balances in the culture. The base addition for neutralizing lactate produced by cells was also monitored on-line via a balance. Using OD and OUR measurements, the specific growth and specific oxygen consumption rates were determined on-line. By combining predetermined stoichiometric relationships among oxygen and glucose consumption and lactate production, the specific glucose consumption and lactate production rates were also calculated on-line. Using these on-line measurements and calculations, the hybridoma culture process was characterized on-line by identifying the physiological states. They will also facilitate the implementation of nutrient feeding strategies for fed-batch and perfusion cultures. (c) 1994 John Wiley & Sons, Inc.

  2. Label-free hybridoma cell culture quality control by a chip-based impedance flow cytometer.

    Science.gov (United States)

    Pierzchalski, Arkadiusz; Hebeisen, Monika; Mittag, Anja; Bocsi, Jozsef; Di Berardino, Marco; Tarnok, Attila

    2012-11-07

    Impedance flow cytometry (IFC) was evaluated as a possible alternative to fluorescence-based methods for on-line quality monitoring of hybridoma cells. Hybridoma cells were cultured at different cell densities and viability was estimated by means of IFC and fluorescence-based flow cytometry (FCM). Cell death was determined by measuring the impedance phase value at high frequency in low conductivity buffer. IFC data correlate well with reference FCM measurements using AnnexinV and 7-AAD staining. Hybridoma cells growing at different densities in cell culture revealed a density-dependent subpopulation pattern. Living cells of high density cultures show reduced impedance amplitudes, indicating particular cellular changes. Dead cell subpopulations become evident in cultures with increasing cell densities. In addition, a novel intermediate subpopulation, which most probably represents apoptotic cells, was identified. These results emphasize the extraordinary sensitivity of high frequency impedance measurements and their suitability for hybridoma cell culture quality control.

  3. A Proteomic Study of Clavibacter Michiganensis Subsp. Michiganensis Culture Supernatants

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    Eva Hiery

    2015-11-01

    Full Text Available Clavibacter michiganensis, subsp. michiganensis is a Gram-positive plant pathogen infecting tomato (Solanum lycopersicum. Despite a considerable economic importance due to significant losses of infected plants and fruits, knowledge about virulence factors of C. michiganensis subsp. michiganensis and host-pathogen interactions on a molecular level are rather limited. In the study presented here, the proteome of culture supernatants from C. michiganensis subsp. michiganensis NCPPB382 was analyzed. In total, 1872 proteins were identified in M9 and 1766 proteins in xylem mimicking medium. Filtration of supernatants before protein precipitation reduced these to 1276 proteins in M9 and 976 proteins in the xylem mimicking medium culture filtrate. The results obtained indicate that C. michiganensis subsp. michiganensis reacts to a sucrose- and glucose-depleted medium similar to the xylem sap by utilizing amino acids and host cell polymers as well as their degradation products, mainly peptides, amino acids and various C5 and C6 sugars. Interestingly, the bacterium expresses the previously described virulence factors Pat-1 and CelA not exclusively after host cell contact in planta but already in M9 minimal and xylem mimicking medium.

  4. A Proteomic Study of Clavibacter Michiganensis Subsp. Michiganensis Culture Supernatants.

    Science.gov (United States)

    Hiery, Eva; Poetsch, Ansgar; Moosbauer, Tanja; Amin, Bushra; Hofmann, Jörg; Burkovski, Andreas

    2015-11-12

    Clavibacter michiganensis, subsp. michiganensis is a Gram-positive plant pathogen infecting tomato (Solanum lycopersicum). Despite a considerable economic importance due to significant losses of infected plants and fruits, knowledge about virulence factors of C. michiganensis subsp. michiganensis and host-pathogen interactions on a molecular level are rather limited. In the study presented here, the proteome of culture supernatants from C. michiganensis subsp. michiganensis NCPPB382 was analyzed. In total, 1872 proteins were identified in M9 and 1766 proteins in xylem mimicking medium. Filtration of supernatants before protein precipitation reduced these to 1276 proteins in M9 and 976 proteins in the xylem mimicking medium culture filtrate. The results obtained indicate that C. michiganensis subsp. michiganensis reacts to a sucrose- and glucose-depleted medium similar to the xylem sap by utilizing amino acids and host cell polymers as well as their degradation products, mainly peptides, amino acids and various C5 and C6 sugars. Interestingly, the bacterium expresses the previously described virulence factors Pat-1 and CelA not exclusively after host cell contact in planta but already in M9 minimal and xylem mimicking medium.

  5. Methods for reducing the ammonia in hybridoma cell cultures.

    Science.gov (United States)

    Capiaumont, J; Legrand, C; Carbonell, D; Dousset, B; Belleville, F; Nabet, P

    1995-02-21

    The factors which limit the proliferation of eukaryotic cells in vitro are still not well known. Ammonia is believed to be toxic for mammalian cell proliferation and secretion. We have tried two approaches to reducing the ammonia in the medium. We first limited the ammonia produced by the cells by replacing glutamine by glutamate. Then, we used two chemical engineering methods to eliminate accumulated ammonia. In one the used medium was passed through a natural cation exchanger: the clinoptilolite. In the other, the culture medium was passed through a hydrophobic microporous hollow fiber module. Replacing the glutamine by glutamate reduced the medium ammonia concentration. The physicochemical removal of ammonia induced a better cell growth, but not a better specific antibody secretion.

  6. GADD153 expression does not necessarily correlate with changes in culture behavior of hybridoma cells

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    Chartrand Kevin

    2007-12-01

    Full Text Available Abstract Background The acute sensitivity of some hybridoma cell lines to culture-related stresses severely limits their productivity. Recent developments in the characterization of the stress signals modulating the cellular phenotype revealed that the pro-apoptotic transcription factor Gadd153 could be used as a marker to facilitate the optimization of mammalian cell cultures. In this report, we analyzed the expression of Gadd153 in Sp2/0-Ag14 murine hybridoma cells grown in stationary batch culture and subjected to two different culture optimization paradigms: L-glutamine supplementation and ectopic expression of Bcl-xL, an anti-apoptotic gene. Results The expression of Gadd153 was found to increase in Sp2/0-Ag14 cells in a manner which coincided with the decline in cell viability. L-glutamine supplementation prolonged Sp2/0-Ag14 cell survival and greatly suppressed Gadd153 expression both at the mRNA and protein level. However, Gadd153 levels remained low after L-glutamine supplementation even as cell viability declined. Bcl-xL overexpression also extended Sp2/0-Ag14 cell viability, initially delayed the induction of Gadd153, but did not prevent the increase in Gadd153 protein levels during the later phase of the culture, when cell viability was declining. Interestingly, L-glutamine supplementation prevented Gadd153 up-regulation in cells ectopically expressing Bcl-xL, but had no effect on cell viability. Conclusion This study highlights important limitations to the use of Gadd153 as an indicator of cell stress in hybridoma cells.

  7. In vitro culture of hybridoma cells in agarose beads producing antibody secretion for two weeks.

    Science.gov (United States)

    Cadic, C; Dupuy, B; Pianet, I; Merle, M; Margerin, C; Bezian, J H

    1992-01-05

    A new process for embedding cells in agarose is described. Beads were obtained by extruding an ultralow gelling temperature agarose solution in a capillary containing a hydrophobic medium flowthrough. The toxicity of the procedure has been evaluated by monitoring the energy status of agarose-embedded C(6) glioma cells with (31)P nuclear magnetic resonance (NMR). Suspension and microbead cultures of hybridoma cell line were compared. In suspension culture the number of cells and the antibody concentrations increased for 5 days before the stationary phase began, when the cultures were stopped. In agarose bead cultures, the gel provided an enormous support surface area (50 m(2)/ mL of gel). It was possible to seed 20-fold more cells. The gel pressure modified the proliferative process and antibody pattern secretion. In particular, the antibodies could be harvested for two weeks.

  8. Optimization and robustness analysis of hybridoma cell fed-batch cultures using the overflow metabolism model.

    Science.gov (United States)

    Amribt, Z; Dewasme, L; Vande Wouwer, A; Bogaerts, Ph

    2014-08-01

    The maximization of biomass productivity in fed-batch cultures of hybridoma cells is analyzed based on the overflow metabolism model. Due to overflow metabolism, often attributed to limited oxygen capacity, lactate and ammonia are formed when the substrate concentrations (glucose and glutamine) are above a critical value, which results in a decrease in biomass productivity. Optimal feeding rate, on the one hand, for a single feed stream containing both glucose and glutamine and, on the other hand, for two separate feed streams of glucose and glutamine are determined using a Nelder-Mead simplex optimization algorithm. The optimal multi exponential feed rate trajectory improves the biomass productivity by 10 % as compared to the optimal single exponential feed rate. Moreover, this result is validated by the one obtained with the analytical approach in which glucose and glutamine are fed to the culture so as to control the hybridoma cells at the critical metabolic state, which allows maximizing the biomass productivity. The robustness analysis of optimal feeding profiles obtained with different optimization strategies is considered, first, with respect to parameter uncertainties and, finally, to model structure errors.

  9. Actinobacillus pleuropneumoniae culture supernatants interfere with killing of Pasteurella multocida by swine pulmonary alveolar macrophages.

    OpenAIRE

    Chung, W. B.; Bäckström, L; McDonald, J.; Collins, M T

    1993-01-01

    The effect of Actinobacillus pleuropneumoniae culture supernatant on swine pulmonary alveolar macrophage (PAM) functions was studied. The A. pleuropneumoniae culture supernatant was toxic to PAMs when tested by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and lactate dehydrogenase (LDH) release assays. Biological activity of the supernatant was ascribed to cytotoxins. Both the LDH and MTT assays were used for measurement of crude A. pleuropneumoniae cytotoxin concentrati...

  10. Early stationary phase culture supernatant accelerates growth of sputum cultures collected after initiation of anti-tuberculosis treatment.

    Science.gov (United States)

    Kolwijck, E; Friedrich, S O; Karinja, M N; van Ingen, J; Warren, R M; Diacon, A H

    2014-07-01

    We investigated the effect of Mycobacterium tuberculosis culture supernatant added to sputum cultures collected during the first 8 weeks of anti-tuberculosis treatment. With ongoing treatment duration, time to culture positivity decreased significantly in supernatant-enriched cultures, possibly due to stimulation of dormant or slowly metabolizing M. tuberculosis cells.

  11. [On-line monitoring of oxygen uptake rate and its application in hybridoma culture].

    Science.gov (United States)

    Feng, Qiang; Mi, Li; Li, Ling; Wang, Xian-Hui; Chen, Zhi-Nan

    2003-09-01

    On-line analysis and control are critical for the optimization of product yields in animal cell culture. The close monitor of viable cell number helps to gain a better insight into the metabolism and to refine culture strategy. In this study, we use the oxygen uptake rate (OUR) to estimate the number of viable cell and the OUR-based feed-back control strategy for nutrients feeding to improve the efficiency of cell culture. A hybridoma cell line (HAb18) was cultured in fed-batch and perfusion model using serum free medium in 5L CelliGen Plus bioreactor (NBS Co., American) and 5L Biostat B bioreactor (Braun Co., Germany). The system and the method for online monitoring OUR in bioreactors, based on the dynamic measurement of dissolved oxygen (DO), were developed. The method of on-line cell concentration estimation was established based on the relationship between the growth of the hybridoma and the uptake rate of oxygen. This method was then used to determine OUR and the concentrations of cell, antibody, glucose, lactate, glutamine and ammonia in the bioreactors at given times. The relationship between OUR and nutrients metabolism was studied and OUR-based feed-back control strategy, which used the state deltaOUR = 0 as the regulation point, was established and used to control the rates of nutrients or medium feeding rate in the perfusion culture. The results showed that there was close relationship between OUR, concentration of live cells, productivity of antibody and consumption of glutamine. The sudden decrease in OUR may be caused by glutamine depletion, and with different delay times, the viable cell concentration and antibody productivity also decreased. The further analysis revealed the linear relationship between OUR and the density of live cells in the exponential growth phase as qOUR = (0.103 +/- 0.028) x 10(-12) mol/cell/h. These findings can be applied to the on-line detection of live cell density. Our study also indicated that by adjusting the perfusion

  12. Dialysis cultures with immobilized hybridoma cells for effective production of monoclonal antibodies.

    Science.gov (United States)

    Pörtner, R; Lüdemann, I; Märkl, H

    1997-01-01

    An industrial scale reactor concept for continuous cultivation of immobilized animal cells (e.g. hybridoma cells) in a radial-flow fixed bed is presented, where low molecular weight metabolites are removed via dialysis membrane and high molecular products (e.g. monoclonal antibodies) are enriched. In a new "nutrient-split" feeding strategy concentrated medium is fed directly to the fixed bed unit, whereas a buffer solution is used as dialysis fluid. This feeding strategy was investigated in a laboratory scale reactor with hybridoma cells for production of monoclonal antibodies. A steady state monoclonal antibody concentration of 478 mg l(-1) was reached, appr. 15 times more compared to the concentration reached in chemostat cultures with suspended cells. Glucose and glutamine were used up to 98%. The experiments were described successfully with a kinetic model for immobilized growing cells. Conclusions were drawn for scale-up and design of the large scale system. c(Glc) - glucose concentration, mmol l(-1); c(Gln) - glutamine concentration, mmol l(-1); c(Amm) - ammonia concentration, mmol l(-1); c(Lac) - lactate concentration, mmol l(-1); c(MAb) - MAb concentration, mg l(-1); D - dilution rate, d(-1); D(i) - dilution rate in the inner chamber of the membrane dialysis reactor, d(-1); D(0) - dilution rate in the outer chamber of the membrane dialysis reactor, d(-1); q*(FB,Glc) - volume specific glucose uptake rate related to the fixed bed volume, mmol l(FB) (-1) h(-1); q*(FB,Gln) - volume specific glutamine uptake rate related to the fixed bed volume, mmol l(FB) (-1) h(-1).

  13. Dynamic metabolic flux analysis using a convex analysis approach: Application to hybridoma cell cultures in perfusion.

    Science.gov (United States)

    Fernandes de Sousa, Sofia; Bastin, Georges; Jolicoeur, Mario; Vande Wouwer, Alain

    2016-05-01

    In recent years, dynamic metabolic flux analysis (DMFA) has been developed in order to evaluate the dynamic evolution of the metabolic fluxes. Most of the proposed approaches are dedicated to exactly determined or overdetermined systems. When an underdetermined system is considered, the literature suggests the use of dynamic flux balance analysis (DFBA). However the main challenge of this approach is to determine an appropriate objective function, which remains valid over the whole culture. In this work, we propose an alternative dynamic metabolic flux analysis based on convex analysis, DMFCA, which allows the determination of bounded intervals for the fluxes using the available knowledge of the metabolic network and information provided by the time evolution of extracellular component concentrations. Smoothing splines and mass balance differential equations are used to estimate the time evolution of the uptake and excretion rates from this experimental data. The main advantage of the proposed procedure is that it does not require additional constraints or objective functions, and provides relatively narrow intervals for the intracellular metabolic fluxes. DMFCA is applied to experimental data from hybridoma HB58 cell perfusion cultures, in order to investigate the influence of the operating mode (batch and perfusion) on the metabolic flux distribution.

  14. The culture of Chlorella vulgaris in a recycled supernatant: Effects on biomass production and medium quality

    KAUST Repository

    Hadj-Romdhane, F.

    2013-03-01

    Reusing supernatant of microalgae culture medium can have inhibitory or toxic effects on the biomass production because of the release of organic metabolites by cells in the culture medium during their growth. This work investigated the impact of Chlorella vulgaris medium recycling on culture productivity, cells quality and accumulation of excreted metabolites in the culture medium. No significant impact on the C. vulgaris growth was observed after 63days of recycling, the productivity remained stable at around 0.55kgm-3day-1. Organic matters accumulated in supernatant were identified as biopolymers (BP) poor in nitrogen and with a size above 40kDa (probably polysaccharides), and small organic molecules (SOM) richer in nitrogen with a molecular size ranging from 1 to 3kDa. The concentration of biopolymers in the supernatant increased till to a maximum and then decreased, possibly consumed by bacteria, whereas small organic compounds accumulated in the medium. © 2013 Elsevier Ltd.

  15. Bioactivities of Culture Supernatants from Retroviral Packaging Cells Carrying the Mouse Fas Ligand Gene

    Institute of Scientific and Technical Information of China (English)

    LIU Lingbo; ZOU Ping; GUO Rong; XIAO Juan; XU Zhiliang

    2001-01-01

    The bioactivities of culture supernatants from retroviral packaging cells carrying the mouse Fas ligand (mFasL) gene was investigated. FasLcDNA was cloned into PLXIN with an internal ribosome entry site to link two cistrons through gene recombination technology, PLXIN and the recombinant vector PLFIN were separately transfected into PA317 retrovirus packing cell line by lipofectamine 2000, and the resistant clones were selected with G418 selective medium. The integration of genome DNA was assayed by genomic DNA PCR. NIH3T3 cells were transduced by the culture supernatants from PA317 carrying the mFasLcDNA gene, and were selected with G418 selective medium, so as to select the PLFIN-PA317 clone capable of producing higher titer of supernatants. The levels of mFasL protein on NIH3T3 cells membrane were assayed by flow cytometry (FCM). The biological activity of mFasL on NIH3T3 cells membrane was investigated by the inducing apoptosis of Fas+ Yac-1 cells co-cultured with NIH3T3 cells expressing Fas ligand. To explore the direct mFasL cytotoxicity of culture supernatants from retroviral packaging cells carrying the mFasL gene, the culture supernatants from PLFIN-PA317 and PLXIN-PA317 were separately co-cultured with Yac-1cells in parallel. The recombinant PLFIN was successfully constructed. The highest titer of supernatants from twelve resistant clones was 8. 5 × 105 colony-forming-unit (CFU)/ml. The NIH3T3cells transfected by above supernatants had a higher level of mFasL (53.81±6.9 %), and significantly induced the apoptosis of Fas+ Yac-1 cells (56. 78±4.5 %), as both were cocultured for 5 h at1 : 1 ratio, whereas it is 7. 08±3.4 % in control group (P<0. 01). Supernatant from PLFINPA317 could also directly induce the apoptosis of Yac-1 within 5 h of incubation. Thus, the culture supernatants from PLFIN-PA317 possessed both infectivity and cytotoxicity of mFasL.

  16. Treatment of serum with supernatants from cultures of Candida albicans reduces its serum-dependent phagocytosis

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    Santos Aderbal Antonio dos

    2002-01-01

    Full Text Available Candida albicans is a potent activator of the complement system, and heat labile opsonins produced by activation of C3 (C3b and iC3b enhance phagocytosis of C. albicans mediated by complement receptors. In this study we treated mouse serum with supernatants from cultures of a protease producer strain of C. albicans and evaluated the ability of this serum to enhance phagocytosis of C. albicans. Cell-free supernatants from cultures of C. albicans were concentrated 5 fold and added to mouse serum for 30 min at 37ºC, before using this serum for opsonization of glutaraldehyde-fixed yeast cells. We observed that normal mouse serum increased about 3 fold the phagocytosis of C. albicans by mice peritoneal macrophages, whereas supernatant-treated serum did not increase phagocytosis. This effect of supernatants on serum was prevented by addition of pepstatin (5 µg/ ml; an inhibitor of C. albicans acid proteases to the medium. Serum treated with supernatants from cultures of a protease-deficient mutant of C. albicans also increased about 3 fold phagocytosis of the yeast. These results suggest that a protease produced by C. albicans causes proteolysis of serum opsonins, thereby reducing the phagocytosis of the yeast.

  17. Dialysis cultures with immobilized hybridoma cells for effective production of monoclonal antibodies

    OpenAIRE

    Pörtner, Ralf; Lüdemann, Ines; Märkl, Herbert

    1997-01-01

    An industrial scale reactor concept for continuous cultivation of immobilized animal cells (e.g. hybridoma cells) in a radial-flow fixed bed is presented, where low molecular weight metabolites are removed via dialysis membrane and high molecular products (e.g. monoclonal antibodies) are enriched. In a new “nutrient-split” feeding strategy concentrated medium is fed directly to the fixed bed unit, whereas a buffer solution is used as dialysis fluid. This feeding strategy was investigated in a...

  18. EFFECT OF GLYCOSYLATION AT ASN302 OF PRO-UROKINASE ON ITS STABILITY IN CULTURE SUPERNATANT

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Objective To investigate the effect of glycosylation at Asn302 of pro-urokinase (pro-UK) on the stability in culture supernatant.Methods Nonglycosylated pro-UK was constructed by site-directed mutagenesis of Asn302 to Ala302. The proUK mutant and native pro-UK were transfected into dhfr-CHO cells, and serum-free culture supernatant was harvested and incubated at 4℃ and 37℃, respectively. The pro-UK activity in culture supernatant was measured by the optical density (OD) increase with time ( 12 hours) at 405 nm. Without thermolysin activation, the percentage of single chain pro-UK was measured.Results After 48 hours of incubation at 4℃, the activities of pro-UK mutant and native pro-UK decreased 3.7%and 2. 9% respectively, and at 37℃ decreased 37.9 % and 23.5 %, respectively. The total activity of native pro-UK was significantly higher than that of nonglycosylated mutant at 37℃. The single-chain percentage of native pro-UK was higher than that of nonglycosylated mutant at both 4℃ and 37℃.Conclusion Higher temperature increases the proteolysis of pro-UK. The glycosylation site on Asn302 is beneficial to pro-UK stability in culture supernatant.

  19. Dormant cells of Staphylococcus aureus are resuscitated by spent culture supernatant.

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    Ben Pascoe

    Full Text Available We describe the first in vitro model of dormancy in Staphylococcus aureus, showing that cells are generated which can be resuscitated by addition of spent medium supernatant taken from cultures of the same organism. Over 30 days, culturable counts in dormant cultures of S. aureus SH1000 fell from 10(6-10(7 cfu/ml to 600-fold increase in bacterial growth. Resuscitation was a specific effect, greatly reduced by boiling or addition of trypsin to the spent supernatant. Supernatant also effected a reduction in lag phase of dormant cultures. SEM demonstrated the presence of small coccoid cells in dormant cultures. The results are similar to those seen with resuscitation promoting factors (Rpfs in actinobacteria. This is the first time resuscitation has been demonstrated in Staphylococcus aureus, which is an important human pathogen. A better understanding of control and reactivation of dormant cells could lead to major improvements in managing staphylococcal infections; resuscitation could be an important step in restoring susceptibility to antibiotic treatment.

  20. Heterogeneous conditions in dissolved oxygen affect N-glycosylation but not productivity of a monoclonal antibody in hybridoma cultures.

    Science.gov (United States)

    Serrato, J Antonio; Palomares, Laura A; Meneses-Acosta, Angélica; Ramírez, Octavio T

    2004-10-20

    It is known that heterogeneous conditions exist in large-scale animal cell cultures. However, little is known about how heterogeneities affect cells, productivities, and product quality. To study the effect of non-constant dissolved oxygen tension (DOT), hybridomas were subjected to sinusoidal DOT oscillations in a one-compartment scale-down simulator. Oscillations were forced by manipulating the inlet oxygen partial pressure through a feedback control algorithm in a 220-mL bioreactor maintained at a constant agitation. Such temporal DOT oscillations simulate spatial DOT gradients that can occur in large scales. Different oscillation periods, in the range of 800 to 12,800 s (axis of 7% (air saturation) and amplitude of 7%), were tested and compared to constant DOT (10%) control cultures. Oscillating DOT decreased maximum cell concentrations, cell growth rates, and viability indexes. Cultures at oscillating DOT had an increased glycolytic metabolism that was evidenced by a decrease in yield of cells on glucose and an increase in lactate yield. DOT gradients, even several orders of magnitude higher than those expected under practical large-scale conditions, did not significantly affect the maximum concentration of an IgG(1) monoclonal antibody (MAb). The glycosylation profile of the MAb produced at a constant DOT of 10% was similar to that reported in the literature. However, MAb produced under oscillating culture conditions had a higher amount of triantennary and sialylated glycans, which can interfere with effector functions of the antibody. It was shown that transient excursions of hybridomas to limiting DOT, as occurs in deficiently mixed large-scale bioreactors, is important to culture performance as the oscillation period, and thus the time cells spent at low DOT, affected cell growth, metabolism, and the glycosylation pattern of MAb. Such results underline the importance of monitoring protein characteristics for the development of large-scale processes.

  1. Use of spray-dried zirconia microspheres in the separation of immunoglobulins from cell culture supernatant.

    Science.gov (United States)

    Subramanian, A; Carr, P W; McNeff, C V

    2000-08-18

    A method suitable for the isolation of monoclonal antibodies (MAbs) on novel zirconia microspheres (20-30 microm) is described. Zirconia microspheres were generated by spray drying colloidal zirconia. Spray-dried zirconia microspheres were further classified and characterized by X-ray diffraction, BET porosimetry and scanning electron microscopy. Spray-dried zirconia microspheres were modified with ethylenediamine-N,N'-tetra(methylenephosphonic) acid (EDTPA) to create a cation-exchange chromatographic support. The chromatographic behavior of a semi-preparative column packed with EDTPA-modified zirconia microspheres was evaluated and implications for scale-up are provided. EDTPA-modified zirconia microspheres were further used to purify MAbs from cell culture supernatant. Analysis by enzyme linked immunosorbent assay and gel electrophoresis demonstrate that MAbs can be recovered from a cell culture supernatant at high yield (92-98%) and high purity (>95%) in a single chromatographic step.

  2. Rheological properties of mammalian cell culture suspensions: Hybridoma and HeLa cell lines.

    Science.gov (United States)

    Shi, Y; Ryu, D D; Ballica, R

    1993-03-25

    Data on viscous (eta') and elastic (eta'') components of the complex viscosity versus oscillatory angular frequency (0.01 to 4.0 rad/s) with increasing strains were obtained for hybridoma cell (62'D3) and HeLa cell (S3) suspensions in PBS at 0.9 (mL/mL) cell volume fraction using a Weissenberg rheogoniometer equipped with two parallel plate geometry at ambient temperature. Both cell suspensions exhibited shear thinning behavior. From the measured viscoelastic properties, the yield stress was calculated. Hybridoma cell suspension (15 microm as the mean diameter of cells) showed the yield stress at 550 dyne/cm(2) that was 1.8 times higher than the value of HeLa cell suspension (22 microm mean diameter) as measured at the oscillatory angular frequency, 4.0 rad/s. The apparent viscosities of HeLa cell suspension at four concentrations and varying steady shear rate were also determined using the Brookfield rotational viscometer. The yield stress to steady shear test was about 130 dyne/cm(2) for HeLa cell suspension at 0.9 (mL/mL) cell volume fraction. The apparent viscosity was in the range about 1 approximately 1000 Poise depending on the cell concentration and shear rate applied. A modified semiempirical Mooney equation, eta = eta(0) exp[K gamma(.)(-beta)phi(c)(1 - K'' sigmaphi(c) /D)] was derived based on the cell concentration, the cell morphology, and the steady shear rate. The beta, shear rate index, was estimated as 0.159 in the range of shear rate, 0.16 to 22.1 s(-1), for the cell volume fractions from 0.6 to 0.9 (mL/mL). In this study, the methods of determining the shear sensitivity and the viscous and the elastic components of mammalian cell suspensions are described under the steady shear field.

  3. An efficient process of generating bispecific antibodies via controlled Fab-arm exchange using culture supernatants.

    Science.gov (United States)

    Paul, Suparna; Connor, Judy; Nesspor, Tom; Haytko, Peter; Boakye, Ken; Chiu, Mark L; Jiang, Haiyan

    2016-05-01

    Bispecific antibody generation is actively pursued for therapeutic and research antibody development. Although there are multiple strategies for generating bispecific antibodies (bsAbs); the common challenge is to develop a scalable method to prepare bsAbs with high purity and yield. The controlled Fab-arm exchange (cFAE) method combines two parental monoclonal antibodies (mAbs), each with a matched point mutation, F405L and K409R in the respective CH3 domains. The conventional process employs two steps: the purification of two parental mAbs from culture supernatants followed by cFAE. Following a reduction/oxidation reaction, the bispecific mAb is formed with greater than 95% heterodimerization efficiency. In this study, cFAE was initiated in culture supernatants expressing the two parental mAbs, thereby eliminating the need to first purify the parental mAbs. The bsAbs formed in culture supernatant was then purified using a Protein A affinity chromatography. The BsAbs generated in this manner had efficiency comparable to the conventional method using purified parental mAbs. BsAbs prepared by two different routes showed indistinguishable characteristics by SDS capillary electrophoresis, analytical size exclusion, and cation exchange chromatography. This alternative method significantly shortened timelines and reduced resources required for bsAb generation, providing an improved process with potential benefits in large-scale bsAb preparation, as well as for HTP small-scale bsAb matrix selection.

  4. Flow Cytometry Sorting to Separate Viable Giant Viruses from Amoeba Co-culture Supernatants

    Science.gov (United States)

    Khalil, Jacques Y. B.; Langlois, Thierry; Andreani, Julien; Sorraing, Jean-Marc; Raoult, Didier; Camoin, Laurence; La Scola, Bernard

    2017-01-01

    Flow cytometry has contributed to virology but has faced many drawbacks concerning detection limits, due to the small size of viral particles. Nonetheless, giant viruses changed many concepts in the world of viruses, as a result of their size and hence opened up the possibility of using flow cytometry to study them. Recently, we developed a high throughput isolation of viruses using flow cytometry and protozoa co-culture. Consequently, isolating a viral mixture in the same sample became more common. Nevertheless, when one virus multiplies faster than others in the mixture, it is impossible to obtain a pure culture of the minority population. Here, we describe a robust sorting system, which can separate viable giant virus mixtures from supernatants. We tested three flow cytometry sorters by sorting artificial mixtures. Purity control was assessed by electron microscopy and molecular biology. As proof of concept, we applied the sorting system to a co-culture supernatant taken from a sample containing a viral mixture that we couldn't separate using end point dilution. In addition to isolating the quick-growing Mimivirus, we sorted and re-cultured a new, slow-growing virus, which we named “Cedratvirus.” The sorting assay presented in this paper is a powerful and versatile tool for separating viral populations from amoeba co-cultures and adding value to the new field of flow virometry. PMID:28111619

  5. Identification and control of dissolved oxygen in hybridoma cell culture in a shear sensitive environment.

    Science.gov (United States)

    Simon, L; Karim, M N

    2001-01-01

    The productivity of mammalian cells can be enhanced by facilitating adequate oxygen transfer into the cultivation medium. However, current methods of controlling dissolved oxygen (DO) fail to account for alterations in medium composition during the course of the fermentation. These changes, which directly affect gas solubility and overall mass transfer coefficient, may be significant and deteriorate controller's performance in the long run. In this paper, the applications of Generalized Predictive Controllers (GPC) to DO control were investigated in a shear sensitive environment and compared to PID and Model Predictive Controllers (MPC). Input and output data for system identification were initially generated by varying the composition of oxygen fed into the bioreactor from 0 to 0.21 mol % while keeping the total inlet gas flow rate at 8.75 vvm. The process was identified using an AutoRegressive model with eXogeneous inputs (ARX) model and tested on different data sets. The model parameters were then correlated with the overall mass transfer coefficients. In simulation tests, the output of the PID controller switched from minimum to maximum values while more continuous control signals were obtained with the MPC and GPC controllers. When tested in a cell-free medium, all three controllers were able to track setpoint changes with some chattering observed in the control signals. The GPC outperformed the MPC and PID controllers when applied to the cultivation of hybridoma cells.

  6. Viability and proliferation of L929, tumour and hybridoma cells in the culture media containing sericin protein as a supplement or serum substitute.

    Science.gov (United States)

    Cao, Ting-Ting; Zhang, Yu-Qing

    2015-09-01

    Cell cultures often require the addition of animal serum and other supplements. In this study, silk sericin, a bioactive protein, recovered from the waste of silk floss production was hydrolysed into three pepsin-degraded sericin peptides with different ranges of molecular mass. Normal animal cells, tumour cells and hybridoma cells were cultured systematically in FBS culture media containing sericin as a supplement or serum substitute. The culture test and microscopic observation of L929 cells showed that the smaller molecular weight of the degraded sericin is most suitable for cell culture. The cell culture results showed that with the degradation of sericin, for normal mouse fibroblast L929 cells, addition of 0.75 % sericin into FBS culture medium yields cell viability that is superior to FBS culture medium alone. When all serum was replaced by sericin, cell viability in the sericin medium could reach about one half of that in FBS medium. When in a medium containing a mixture of FBS: sericin (6:4, v/v), the cell culture effect is about 80 %. For the cultures of four tumour and one hybridoma cells, regardless of the molecular weight range, these degraded sericin peptides could substitute all serum in FBS media. The cell viability and proliferation of these tumour and hybridoma cells are equivalent or superior to that in FBS medium. In other words, cell viability and proliferation of these tumour and hybridoma cells in sericin media are more preferable to serum media. The mechanism of the sericin protein to promote cell growth and proliferation will be further investigated later.

  7. Purification of monoclonal antibodies, IgG1, from cell culture supernatant by use of metal chelate convective interaction media monolithic columns.

    Science.gov (United States)

    Rajak, Poonam; Vijayalakshmi, M A; Jayaprakash, N S

    2012-12-01

    Monoclonal antibodies (MAbs) have diverse applications in diagnostics and therapeutics. The recent advancement in hybridoma technology for large-scale production of MAbs in bioreactors demands rapid and efficient purification methods. Conventional affinity purification systems have drawbacks of low flow rates and denaturation of antibodies owing to harsh elution conditions. Here, we attempted purification of MAbs by use of a high-throughput metal-chelate methacrylate monolithic system. Monolithic macroporous convective interaction media-iminodiacetate (CIM-IDA) disks immobilized with four different metal ions (Cu²⁺, Ni²⁺, Zn²⁺ and Co²⁺) were used and evaluated for purification of anti-human serum albumin IgG1 mouse MAbs from cell culture supernatant after precipitation with 50% ammonium sulfate. Elution with 10 mM imidazole in the equilibration buffer (25 mM MMA = MOPS (Morpholino propane sulfonic acid) + MES (Morpholino ethane sulfonic acid) + Acetate + 0.5 M NaCl, pH 7.4) resulted in a purification of 25.7 ± 2.9-fold and 32.5 ± 2.6-fold in experiments done using Zn²⁺ and Co²⁺ metal ions, respectively. The highest recovery of 85.4 ± 1.0% was obtained with a CIM-IDA-Zn(II) column. SDS-PAGE, ELISA and immuno-blot showed that the antibodies recovered were pure, with high antigen-binding efficiency. Thus, metal chelate CIM monoliths could be a potential alternative to conventional systems for fast and efficient purification of MAbs from the complex cell culture supernatant.

  8. Bioleaching of metals from steel slag by Acidithiobacillus thiooxidans culture supernatant.

    Science.gov (United States)

    Hocheng, Hong; Su, Cheer; Jadhav, Umesh U

    2014-12-01

    The generation of 300–500 kg of slag per ton of the steel produced is a formidable amount of solid waste available for treatment. They usually contain considerable quantities of valuable metals. In this sense, they may become either important secondary resource if processed in eco-friendly manner for secured supply of contained metals or potential pollutants, if not treated properly. It is possible to recover metals from steel slag by applying bioleaching process. Electric arc furnace (EAF) slag sample was used for bioleaching of metals. In the present study, before bioleaching experiment water washing of an EAF slag was carried out. This reduced slag pH from 11.2 to 8.3. Culture supernatants of Acidithiobacillus thiooxidans (At. thiooxidans), Acidithiobacillus ferrooxidans (At. ferrooxidans), and Aspergillus niger (A. niger) were used for metal solubilization. At. thiooxidans culture supernatant containing 0.016 M sulfuric acid was found most effective for bioleaching of metals from an EAF slag. Maximum metal extraction was found for Mg (28%), while it was least for Mo (0.1%) in six days. Repeated bioleaching cycles increased metal recovery from 28% to 75%, from 14% to 60% and from 11% to 27%, for Mg, Zn and Cu respectively.

  9. Analysis of IL-2-like factor in lymphocyte culture supernatant of olive flounder, Paralichthys oliveaceus

    Institute of Scientific and Technical Information of China (English)

    WU Riqin; ZHANG Peijun; LI Jun; XU Yongli

    2005-01-01

    To study immune mechanism of fish lymphocyte we performed a proliferation assay and ELISA using monoclonal antibody against human IL-2. The result showed that an interleukin-2 (IL-2)-like factor was detected in the supernatant of plant haemoglutinin (PHA)-stimulated lymphocyte culture from peripheral blood,spleen and head kidney of olive flounder, Paralichthys olivaceus. The quantities of IL-2-1ike factor in the supematant from different lymphoid tissues were quite different. The IL-2 like factor in the supernatant from cultured head kidney lymphocytes was much higher than those of peripheral blood lymphocytes and spleen lymphocytes (P<0.01). The IL-2 activity was found in either mouse thymocyte proliferation assay or flounder head kidney lymphocyte proliferation assay and shown to have obvious enhancing effect on proliferation of the above two types of cell. The recombinant human IL-2 (rhIL-2) was able to stimulate flounder thymocyte proliferation and used to detect the IL-2 receptor (IL-2R) on the surface of flounder lymphocyte. The cross-reaction between the lymphocytes of flounder peripheral blood and CD25(IL-2R) was detected with flow cytometry and shown that the percentage of CD25-positive cell in peripheral blood was 7.74± 0.67%.

  10. Combined data preprocessing and multivariate statistical analysis characterizes fed-batch culture of mouse hybridoma cells for rational medium design.

    Science.gov (United States)

    Selvarasu, Suresh; Kim, Do Yun; Karimi, Iftekhar A; Lee, Dong-Yup

    2010-10-01

    We present an integrated framework for characterizing fed-batch cultures of mouse hybridoma cells producing monoclonal antibody (mAb). This framework systematically combines data preprocessing, elemental balancing and statistical analysis technique. Initially, specific rates of cell growth, glucose/amino acid consumptions and mAb/metabolite productions were calculated via curve fitting using logistic equations, with subsequent elemental balancing of the preprocessed data indicating the presence of experimental measurement errors. Multivariate statistical analysis was then employed to understand physiological characteristics of the cellular system. The results from principal component analysis (PCA) revealed three major clusters of amino acids with similar trends in their consumption profiles: (i) arginine, threonine and serine, (ii) glycine, tyrosine, phenylalanine, methionine, histidine and asparagine, and (iii) lysine, valine and isoleucine. Further analysis using partial least square (PLS) regression identified key amino acids which were positively or negatively correlated with the cell growth, mAb production and the generation of lactate and ammonia. Based on these results, the optimal concentrations of key amino acids in the feed medium can be inferred, potentially leading to an increase in cell viability and productivity, as well as a decrease in toxic waste production. The study demonstrated how the current methodological framework using multivariate statistical analysis techniques can serve as a potential tool for deriving rational medium design strategies. Copyright © 2010 Elsevier B.V. All rights reserved.

  11. Comparison among three anion exchange chromatographic supports to capture erythropoietin from cell culture supernatant

    Institute of Scientific and Technical Information of China (English)

    Lourdes HERNNDEZ; Diobel STEWART; Lourdes ZUMALACRREGUI; Daniel AMARO

    2015-01-01

    Affinity and ion exchange conventional chromatography have been used to capture erythropoietin ( EPO)from mammalian cell culture supernatant. Currently,chromatographic adsorbent perfusion is available, however a limited number of applications have been found in the literature. In this work,three anion exchange chromatographic supports( gel,membrane and monolithic)were evaluated in the capture step of the recombi-nant erythropoietin purification process. The influences of load and flow rate on each support performance were analyzed. Also the purity of the EPO molecules was determined. A productivity analysis,as a decision tool for larger scale implementation,was done. As a conclusion,the evaluated supports are technically suitable to cap-ture EPO with adequate recovery and good purity. However,the monolithic column admits high operating velocity,showing the highest adsorption capacity and productivity.

  12. Evaluation of selected control strategies for fed-batch cultures of a hybridoma cell line.

    Science.gov (United States)

    Pörtner, Ralf; Schwabe, Jan-Oliver; Frahm, Björn

    2004-08-01

    While fed-batch suspension culture of animal cells continues to be of industrial importance for the large-scale production of pharmaceutical products, existing control concepts are still insufficient. The present paper illustrates the advantages and disadvantages of different fed-batch strategies, including fixed-feed trajectories, control via OUR (oxygen uptake rate) (stoichiometric feeding), a priori determination of feed trajectories based on a kinetic model and the model-based adaptive OLFO (open-loop-feedback-optimal) control strategy. A recommendation as to which control strategy should be used for a specific process has to consider the respective process. For an established process with a well characterized and stable production cell line, probably the application of a fixed feed trajectory should be recommended. An adaptive, model-based control strategy could be the method of choice during cell-line development or for rapid production of small amounts of product for clinical trials, owing to its universal character and because it does not require intensive process development.

  13. Quantitative analysis of particles, genomes and infectious particles in supernatants of haemorrhagic fever virus cell cultures

    Directory of Open Access Journals (Sweden)

    Hedlund Kjell-Olof

    2011-02-01

    Full Text Available Abstract Information on the replication of viral haemorrhagic fever viruses is not readily available and has never been analysed in a comparative approach. Here, we compared the cell culture growth characteristics of haemorrhagic fever viruses (HFV, of the Arenaviridae, Filoviridae, Bunyaviridae, and Flavivridae virus families by performing quantitative analysis of cell culture supernatants by (i electron microscopy for the quantification of virus particles, (ii quantitative real time PCR for the quantification of genomes, and (iii determination of focus forming units by coating fluorescent antibodies to infected cell monolayers for the quantification of virus infectivity. The comparative analysis revealed that filovirus and RVFV replication results in a surplus of genomes but varying degrees of packaging efficiency and infectious particles. More efficient replication and packaging was observed for Lassa virus, and Dengue virus resulting in a better yield of infectious particles while, YFV turned out to be most efficient with only 4 particles inducing one FFU. For Crimean-Congo haemorrhagic fever virus (CCHFV a surplus of empty shells was observed with only one in 24 particles equipped with a genome. The complete particles turned out to be extraordinarily infectious.

  14. Purification and characterization of a novel fibrinolytic enzyme from culture supernatant of Pleurotus ostreatus.

    Science.gov (United States)

    Liu, Xiao-Lan; Zheng, Xi-Qun; Qian, Peng-Zhi; Kopparapu, Narasimha-Kumar; Deng, Yong-Ping; Nonaka, Masanori; Harada, Naoki

    2014-02-28

    A fibrinolytic enzyme was produced by an edible mushroom of Pleurotus ostreatus using submerged culture fermentation. The enzyme was purified from the culture supernatant by applying a combination of freeze-thaw treatment, ammonium sulfate precipitation, hydrophobic interaction, and gel filtration chromatographies. The enzyme was purified by a 147-fold, with a yield of 7.54%. The molecular masses of the enzyme an determined by gel filtration and SDSPAGE were 13.6 and 18.2 kDa, respectively. The isoelectric point of the enzyme was 8.52. It hydrolyzed fibrinogen by cleaving the α and β chains of fibrinogen followed by the γ chains, and also activated plasminogen into plasmin. The enzyme was optimally active at 45°C and pH 7.4. The enzyme activity was completely inhibited by EDTA, whereas protease inhibitors of TPCK, SBTI, PMSF, aprotinin and pepstatin showed no inhibition on its activity. The partial amino acid sequences of the enzyme as determined by Q-TOF2 were ATFVGCSATR, GGTLIHESSHFTR, and YTTWFGTFVTSR. These sequences showed a high degree of homology with those of metallo-endopeptidases from P. ostreatus and Armillaria mellea. The purified enzyme can also be applied as a natural agent for oral fibrinolytic therapy or prevention of thrombosis.

  15. Two-dimensional gel electrophoresis for controlling and comparing culture supernatants of mammalian cell culture productions systems.

    Science.gov (United States)

    Wimmer, K; Harant, H; Reiter, M; Blüml, G; Gaida, T; Katinger, H

    1994-01-01

    A recombinant Chinese hamster ovary cell line, producing human erythropoietin, was cultivated in a continuous mode in a stirred tank reactor applying different dilution rates. In order to monitor the stability of this expression system, product and non-product proteins of the cell culture supernatant were analyzed by two-dimensional electrophoresis. The consistency of the isoforms of the recombinant product was determined by western blot combined with specific staining. The same cell line was propagated in a high cell density cultivation system based on macro-cell-aggregates. The patterns of secreted proteins of the cell line cultivated in the different systems were compared in order to detect modifications in protein expression of the product and of non product proteins relevant for cell culture supernatant. Hardly any alterations in two-dimensional pattern were detectable. The isoforms of erythropoietin, as well as the overall pattern of secreted proteins, detectable with the two-dimensional electrophoresis method were remarkably stable under different cultivation conditions.

  16. Isolation and Purification of an Early Pregnancy Factor–Like Molecule from Culture Supernatants Obtained from Lymphocytes of Pregnant Women

    OpenAIRE

    1998-01-01

    Purpose:Our purpose was to determine whether lymphocytes synthesize proteins during pregnancy, to observe whether one of the proteins synthesized has early pregnancy factor (EPF)–like activity and to isolate and purify this molecule from culture supernatants obtained from stimulated lymphocytes of pregnant women.

  17. Overcoming bottlenecks of enzymatic biofuel cell cathodes: crude fungal culture supernatant can help to extend lifetime and reduce cost.

    Science.gov (United States)

    Sané, Sabine; Jolivalt, Claude; Mittler, Gerhard; Nielsen, Peter J; Rubenwolf, Stefanie; Zengerle, Roland; Kerzenmacher, Sven

    2013-07-01

    Enzymatic biofuel cells (BFCs) show great potential for the direct conversion of biochemically stored energy from renewable biomass resources into electricity. However, enzyme purification is time-consuming and expensive. Furthermore, the long-term use of enzymatic BFCs is hindered by enzyme degradation, which limits their lifetime to only a few weeks. We show, for the first time, that crude culture supernatant from enzyme-secreting microorganisms (Trametes versicolor) can be used without further treatment to supply the enzyme laccase to the cathode of a mediatorless BFC. Polarization curves show that there is no significant difference in the cathode performance when using crude supernatant that contains laccase compared to purified laccase in culture medium or buffer solution. Furthermore, we demonstrate that the oxygen reduction activity of this enzymatic cathode can be sustained over a period of at least 120 days by periodic resupply of crude culture supernatant. This is more than five times longer than control cathodes without the resupply of culture supernatant. During the operation period of 120 days, no progressive loss of potential is observed, which suggests that significantly longer lifetimes than shown in this work may be possible. Our results demonstrate the possibility to establish simple, cost efficient, and mediatorless enzymatic BFC cathodes that do not require expensive enzyme purification procedures. Furthermore, they show the feasibility of an enzymatic BFC with an extended lifetime, in which self-replicating microorganisms provide the electrode with catalytically active enzymes in a continuous or periodic manner.

  18. Rapid Extracellular Biosynthesis of Silver Nanoparticles by Cunninghamella phaeospora Culture Supernatant

    Science.gov (United States)

    Ghareib, Mohamed; Tahon, Medhat Abu; Saif, Mona Mostafa; El-Sayed Abdallah, Wafaa

    2016-01-01

    The development of green approaches for the biosynthesis of silver nanoparticles (AgNPs) is of prime significance in the field of nanotechnology research. A fast and eco-friendly protocol for the biosynthesis of extracellular AgNPs using culture supernatant (CS) from the fungus Cunninghamella phaeospora was studied in this work. This CS was proved as a potential new source for the extracellular biosynthesis of AgNPs. The AgNPs were formed at 100 oC and pH 9 within four min of contact between CS and 1mM silver nitrate (AgNO3) solution. Nitrate reductase (NR) was confirmed to play a pivotal role in the biosynthesis of AgNPs. The enzyme expressed its highest activity at 80 oC and pH 9. At 100 oC the enzyme retained 70% of its original activity for one hour. The half-life (T1/2) of the enzyme activity was calculated to be 1.55 h confirming its thermostability. The produced AgNPs were characterized by UV-Vis spectroscopy, high resolution-transmission electron microscope (HR-TEM) and x-ray diffraction (XRD). These NPs showed an absorption peak at 415 nm in UV-Vis spectrum corresponding to the plasmon resonance of AgNPs. Transmission electron micrographs revealed the production of monodispersed spherical NPs with average particle size 14 nm. XRD spectrum of the NPs confirmed the formation of metallic crystalline silver. It was also suggested that the aromatic amino acids play a role in the biosynthesis process. The current research provided an insight on the green biosynthesis of AgNPs including some mechanistic aspects using a new mycogenic source.

  19. [Effects of culture supernatant of human amnion mesenchymal stem cells on biological characteristics of human fibroblasts].

    Science.gov (United States)

    Wu, Qi'er; Lyu, Lu; Xin, Haiming; Luo, Liang; Tong, Yalin; Mo, Yongliang; Yue, Yigang

    2016-06-01

    To investigate the effects of culture supernatant of human amnion mesenchymal stem cells (hAMSCs-CS) on biological characteristics of human fibroblasts. (1) hAMSCs were isolated from deprecated human fresh amnion tissue of placenta and then sub-cultured. The morphology of hAMSCs on culture day 3 and hAMSCs of the third passage were observed with inverted phase contrast microscope. (2) Two batches of hAMSCs of the third passage were obtained, then the expression of vimentin of cells was observed with immunofluorescence method, and the expression of cell surface marker CD90, CD73, CD105, and CD45 was detected by flow cytometer. (3) hAMSCs-CS of the third passage at culture hour 72 were collected, and the content of insulin-like growth factor Ⅰ (IGF-Ⅰ), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), and basic fibroblast growth factor (bFGF) were detected by enzyme-linked immunosorbent assay. (4) Human fibroblasts were isolated from deprecated human fresh prepuce tissue of circumcision and then sub-cultured. Human fibroblasts of the third passage were used in the following experiments. Cells were divided into blank control group and 10%, 30%, 50%, and 70% hAMSCs-CS groups according to the random number table (the same grouping method below), with 48 wells in each group. Cells in blank control group were cultured with DMEM/F12 medium containing 2% fetal bovine serum (FBS), while cells in the latter 4 groups were cultured with DMEM/F12 medium containing corresponding volume fraction of hAMSCs-CS and 2% FBS. The proliferation activity of cells was detected by cell counting kit 8 and microplate reader at culture hour 12, 24, 48, and 72, respectively, and corresponding volume fraction of hAMSCs-CS which causing the best proliferation activity of human fibroblasts was used in the following experiments. (5) Human fibroblasts were divided into blank control group and 50% hAMSCs-CS group and treated as in (4), with 4 wells in each group, at post

  20. Comparison of EBV DNA viral load in whole blood, plasma, B-cells and B-cell culture supernatant.

    Science.gov (United States)

    Ouedraogo, David Eric; Bollore, Karine; Viljoen, Johannes; Foulongne, Vincent; Reynes, Jacques; Cartron, Guillaume; Vendrell, Jean-Pierre; Van de Perre, Philippe; Tuaillon, Edouard

    2014-05-01

    Epstein-Barr virus (EBV) genome quantitation in whole blood is used widely for therapeutic monitoring of EBV-associated disorders in immunosuppressed individuals and in patients with EBV-associated lymphoma. However, the most appropriate biological material to be used for EBV DNA quantitation remains a subject of debate. This study compare the detection rate and levels of EBV DNA from whole blood, plasma, enriched B-cells, and B-cell short-term culture supernatant using quantitative real-time PCR. Samples were collected from 33 subjects with either HIV infection or B-cell lymphoma. Overall, EBV DNA was detected in 100% of enriched B-cell samples, in 82% of B-cell culture supernatants, in 57% of plasma, and 42% of whole blood samples. A significant correlation for EBV viral load was found between enriched B-cell and B-cell culture supernatant material (ρ = 0.92; P cells (ρ = -0.02; P = 0.89), whole blood and plasma (ρ = 0.24; P = 0.24), or enriched B-cells and plasma (ρ = 0.08; P = 0.77). Testing of enriched B-cells appeared to be the most sensitive method for detection of EBV DNA as well as for exploration of the cellular reservoir. Quantitation of EBV DNA in plasma and B-cell culture supernatant may be of interest to assess EBV reactivation dynamics and response to treatment as well as to decipher EBV host-pathogen interactions in various clinical scenarios.

  1. Improving the performance of a biofuel cell cathode with laccase-containing culture supernatant from Pycnoporus sanguineus.

    Science.gov (United States)

    Fokina, Oleksandra; Eipper, Jens; Winandy, Lex; Kerzenmacher, Sven; Fischer, Reinhard

    2015-01-01

    Laccases are multicopper oxidoreductases that can be used in biofuel cells to improve cathode performance by cathodic oxygen reduction. Here we present a laccase from the ligninolytic white-rot fungus Pycnoporus sanguineus that, in contrast to the Trametes versicolor laccase, can be produced in the absence of inducers in a standard culture medium. After 7days of cultivation the activity of this laccase in culture supernatant reached 2.5U/ml, which is high enough for direct application of the supernatant in biofuel cells. The highest current density of 115.0±3.5μA/cm(2) at 400mV vs. SCE was obtained at pH 5 with a buckypaper cathode with a laccase-containing culture supernatant. The enzyme also showed electrocatalytic activity at pH 6 and 7. These results not only present a new cost-efficient laccase for improving cathode performance, but also show that new laccases with different catalytic properties can be suitable for biofuel cells.

  2. Increasing the culture efficiency of hybridoma cells by the use of integrated metabolic control of glucose and glutamine at low levels.

    Science.gov (United States)

    Li, Ling; Mi, Li; Feng, Qiang; Liu, Rong; Tang, Hao; Xie, Li; Yu, Xiaoling; Chen, Zhinan

    2005-08-01

    The metabolism of HAb18 hybridoma cells was shifted to decrease metabolite accumulation and to improve culture efficiency by integrated metabolic control of glucose and glutamine at low levels. When glucose and glutamine levels were decreased to 0.5 and 0.3 mM respectively, lactate and ammonia production were reduced by 62.6 and 74% respectively, glucose-to-cell yield was increased from 0.23x10(9) to 0.66x10(9) cells.mmol-1, and glutamine-to-cell yield from 0.18x10(9) to 1.95x10(9) cells.mmol-1. Compared with high-level glucose and glutamine fed-batch cultures, low-level glucose and glutamine led to higher cell density (1.0x10(7) versus 0.3x10(7) cells.ml-1), longer culture span (14 as opposed to 8 days) and higher antibody yield (250 as against 150 mg.l-1). These results indicate that hybridoma culture efficiency would be increased by the integrated control of glucose and glutamine at 0.5 and 0.3 mM respectively. In contrast with previously reported glucose-and/or-glutamine-level-controlled fed-batch cultures, we demonstrated an efficient strategy of nutrient level selection and amino acid feeding. More importantly, our accurately and well-distributed Equable Feeding Control System opens a new avenue for reducing metabolites to low levels by controlling nutrients at low levels.

  3. Improved fusion technique. I. Human umbilical cord serum, a new and potent growth promoter, compared with other B cell and hybridoma activators.

    Science.gov (United States)

    Westerwoudt, R J; Blom, J; Naipal, A M; Van Rood, J J

    1983-08-12

    Accelerated proliferation of hybridoma cells was observed in the presence of human umbilical cord serum (HUCS). This had very strong growth-promoting activity, even at a concentration of 2%. A comparison was made between HUCS and other B cell growth promoters, such as lipopolysaccharide (LPS) and dextran sulfate (DxS), macrophage supernatant, and human endothelial culture supernatant (HECS). The growth-promoting effect of HUCS was superior. Using a microcytotoxicity assay, we found no significant differences in the number of antibody producing clones with the various culture media, except for fetal calf serum.

  4. THE EFFECT OF ANTISENSE OLIGONUCLEOTIDE ON THE INTERLEUKIN-5 IN THE SUPERNATANTS OF SPLEEN CELL CULTURES OF ASTHMATIC MICE

    Institute of Scientific and Technical Information of China (English)

    王美琴; 白春学; 钮善福; 方晓惠; 陈常庆; 陈波

    2001-01-01

    To explore the effect of antisense oligonucleotide on the production of IL-5 by mouse spleen T lymphocytes.Methods Based on the IL-5 cDNA sequence of mouse, a segment of antisense oligonucleotide was designed and synthesized. 5’-labeling of antisense oligonucleotide was signed by T4 PNK in order that the efficiency of stearylamine liposome in transfecting antisense oligonucleotide can be evaluated. Asthma model was duplicated with ovalbumin(OVA) absorbed to aluminum hydroxide. T lymphocytes of mice were separated by nylon fiber method, then T lymphocytes transfected with different concentration of antisense oligonucleotide with cation stearylamine liposme were incubated respectively in order to observe the effect of antisense oligonucleotide on Il-5 production by T lymphocytes. IL-5 levels in the supernatants of T lymphocyte cultures were determined by ELISA.Results Stearylamine liposome could markedly increase the efficiency of antisense oligonucleotide transfection. The transfection efficiency of antisense oligouncleotide increased approximately 12 times at a ratio of 1: 15m/m (antisense oligonucleotide to SA liposome). In healthy and asthma Balb/c mice, IL-5 was not detectable in the supernatants of T lymphocyte cultures without stimulated with OVA; however, IL-5 was increased markedly in the supernatants of T lymphocyte cultures stimulated with OVA. After transfection with different concentrations of antisense oligonucleotide, IL-5 levels in the supernatants of T lymphocyte cultures were significantly lower than those in control cultured without antisense oligonucleotide transfection. IL-5 levels decreased from 44.60±6.23 pg/ml to 30.70±7.362 pg/ml, 17.20±6.181 pg/ml and 8.16±2.34 pg/ml respectively. And IL-5 synthesis was inhibited by 31.17%, 61.43% and 81.7% respectively.Conclusion IL-5 synthesis could be obviously inhibited by antisense oligonucleotide and showed a markedly correlation between dose and effectiveness. It suggests the production

  5. Anion-exchange purification of recombinant factor IX from cell culture supernatant using different chromatography supports.

    Science.gov (United States)

    Ribeiro, Daniel A; Passos, Douglas F; Ferraz, Helen C; Castilho, Leda R

    2013-11-01

    Both recombinant and plasma-derived factor IX concentrates are used in replacement therapies for the treatment of haemophilia B. In the present work, the capture step for a recombinant FIX (rFIX) purification process was investigated. Different strong anion-exchange chromatography media (the resins Q Sepharose(®) FF and Fractogel(®) TMAE, the monolith CIM(®) QA and the membrane adsorber Sartobind(®) Q) were tested for their rFIX binding capacity under dynamic conditions. In these experiments, crude supernatant from CHO cells was used, thus in the presence of supernatant contaminants and mimicking process conditions. The highest dynamic binding capacity was obtained for the monolith, which was then further investigated. To study pseudoaffinity elution of functional rFIX with Ca(2+) ions, a design of experiments to evaluate the effects of pH, NaCl and CaCl2 on yield and purification factor was carried out. The effect of pH was not statistically significant, and a combination of no NaCl and 45mM CaCl2 yielded a good purification factor combined with a high yield of active rFIX. Under these conditions, activity yield of rFIX was higher than the mass yield, confirming selective elution of functional, γ-carboxylated rFIX. Scaling-up of this process 8 fold resulted in very similar process performance. Monitoring of the undesired activated FIX (FIXa) revealed that the FIXa/FIX ratio (1.94%) was higher in the eluate than in the loaded sample, but was still within an acceptable range. HCP and DNA clearances were high (1256 and 7182 fold, respectively), indicating that the proposed process is adequate for the intended rFIX capture step. Copyright © 2013 Elsevier B.V. All rights reserved.

  6. Effect of Physical and Chemical Changes on the Antimicrobial Activity of Culture Supernatant Fluid of Lactic Acid Bacteria

    Directory of Open Access Journals (Sweden)

    Leila Goudarzi (MSc

    2016-04-01

    Full Text Available Background and Objective: Lactic acid bacteria are Gram-positive, catalase-negative, nonsporulating, either rod- or coccus-shaped bacteria that have beneficial effects on their hosts by producing antimicrobial substances such as lactic acid, hydrogen peroxide, bacteriocins and biosurfactants. Bacteriocins are antimicrobial peptides that are produced by bacteria and can inhibit the growth of other bacteria. Methods: In this experimental study, bacteriocin production by Lactobacilli as known probiotic strains was evaluated in different physicochemical conditions. Antagonistic activity was evaluated using quantitative method of Microscale Optical Density Assay (MODA. After neutralization of acid and treatment with various enzymes, temperature, pH and NaCl conditions, the antimicrobial activity of culture supernatant fluid of Lactobacillus acidophilus and L. plantarum was investigated against pathogenic Proteus. Results: The culture supernatant fluid of Lactobacilli was sensitive to proteolytic enzymes with relatively good stability to temperature. The antimicrobial activity was also present due to production of bacteriocin under different NaCl conditions (1 to 4% NaCl and pH range of 5 to 8. Conclusion: It seems that the antimicrobial liquid of Lactobacillus strains contains bacteriocin, which shows antimicrobial effects against pathogenic strains of Proteus. To investigate further this effect, some complementary studies should be performed.

  7. Antifungal performance of extracellular chitinases and culture supernatants of Streptomyces galilaeus CFFSUR-B12 against Mycosphaerella fijiensis Morelet.

    Science.gov (United States)

    Castillo, Benjamín Moreno; Dunn, Michael F; Navarro, Karina Guillén; Meléndez, Francisco Holguín; Ortiz, Magdalena Hernández; Guevara, Sergio Encarnación; Palacios, Graciela Huerta

    2016-03-01

    The tropical and mycoparasite strain Streptomyces galilaeus CFFSUR-B12 was evaluated as an antagonist of Mycosphaerella fijiensis Morelet, causal agent of the Black Sigatoka Disease (BSD) of banana. On zymograms of CFFSUR-B12 culture supernatants, we detected four chitinases of approximately 32 kDa (Chi32), 20 kDa (Chi20), and two with masses well over 170 kDa (ChiU) that showed little migration during denaturing electrophoresis at different concentrations of polyacrylamide. The thymol-sulphuric acid assay showed that the ChiU were glycosylated chitinases. Moreover, matrix assisted laser desorption ionization time-of-flight MS analysis revealed that the ChiU are the same protein and identical to a family 18 chitinase from Streptomyces sp. S4 (gi|498328075). Chi32 was similar to an extracellular protein from Streptomyces albus J1074 (gi|478687481) and Chi20 was non-significantly similar to chitinases from five different strains of Streptomyces (P > 0.05). Subsequently, Chi32 and Chi20 were partially purified by anion exchange and hydrophobic interaction chromatography and tested against M. fijiensis. Chitinases failed to inhibit ascospore germination, but inhibited up to 35 and 62% of germ tube elongation and mycelial growth, respectively. We found that crude culture supernatant and living cells of S. galilaeus CFFSUR-B12 were the most effective in inhibiting M. fijiensis and are potential biocontrol agents of BSD.

  8. Continuous precipitation of process related impurities from clarified cell culture supernatant using a novel coiled flow inversion reactor (CFIR).

    Science.gov (United States)

    Kateja, Nikhil; Agarwal, Harshit; Saraswat, Aditya; Bhat, Manish; Rathore, Anurag S

    2016-10-01

    Coiled Flow Inverter Reactor (CFIR) has recently been explored for facilitating continuous operation of several unit operations involved in downstream processing of biopharmaceuticals such as viral inactivation and protein refolding. The application of CFIR for continuous precipitation of clarified cell culture supernatant has been explored. The pH based precipitation is optimized in the batch mode and then in the continuous mode in CFIR using a design of experiments (DOE) study. Improved clearance of host cell DNA (52× vs. 39× in batch), improved clearance of host cell proteins (HCP) (7× vs. 6× in batch) and comparable recovery (90 vs. 91.5 % in batch) are observed along with six times higher productivity. To further demonstrate wider applicability of CFIR in performing continuous precipitation, two more case studies involving use of two different precipitation protocols (CaCl2 based and caprylic acid based) are also performed. In both cases, clearance of host cell DNA, HCP, and product recovery are found to be comparable or better in CFIR than in batch operations. Moreover, increase in productivity of 16 times (CaCl2 based) and eight times (caprylic acid based) is obtained for the two precipitation protocols, respectively. The data clearly demonstrate that CFIR can be seamlessly integrated into a continuous bioprocess train for performing continuous precipitation of clarified cell culture supernatant. To our knowledge this is the first report of such use. Copyright © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Protein A is released into the Staphylococcus aureus culture supernatant with an unprocessed sorting signal.

    Science.gov (United States)

    O'Halloran, Dara P; Wynne, Kieran; Geoghegan, Joan A

    2015-04-01

    The immunoglobulin binding protein A (SpA) of Staphylococcus aureus is synthesized as a precursor with a C-terminal sorting signal. The sortase A enzyme mediates covalent attachment to peptidoglycan so that SpA is displayed on the surface of the bacterium. Protein A is also found in the extracellular medium, but the processes involved in its release are not fully understood. Here, we show that a portion of SpA is released into the supernatant with an intact sorting signal, indicating that it has not been processed by sortase A. Release of SpA was reduced when the native sorting signal of SpA was replaced with the corresponding region of another sortase-anchored protein (SdrE). Similarly, a reporter protein fused to the sorting signal of SpA was released to a greater extent than the same polypeptide fused to the SdrE sorting signal. Released SpA protected bacteria from killing in human blood, indicating that it contributes to immune evasion.

  10. Vip3A is responsible for the potency of Bacillus thuringiensis 9816C culture supernatant against Helicoverpa armigera and Spodoptera exigua.

    Science.gov (United States)

    Cai, Jun; Xiao, Liang; Yan, Bing; Bin, Guan; Chen, Yuehua; Ren, Gaixin

    2006-04-01

    Culture supernatant of Bacillus thuringiensis 9816C had high toxicity against Helicoverpa armigera and Spodoptera exigua. However, it lost insecticidal activities after being bathed in boiling water for 5 min. Acrystalliferous mutants of Bt9816C (Bt9816C-NP1 and Bt9816C-NP2) cured of its endogenous plasmids no longer possessed vip3A gene and toxicity. The 89 kD protein which existed in Bt9816C supernatant disappeared in the two mutants' supernatant; nevertheless, the two mutants still exhibited hemolytic and phospholipase C activity as Bt9816C did. The vip3A gene of Bt9816C, vip3Aa18, was cloned and expressed in Escherichia coli BL21. Bioassay demonstrated that the recombinant E. coli had high toxicity against S. exigua. Taken together, it suggested that Vip3A protein was responsible for the toxicity of Bt9816C culture supernatants.

  11. Differences in the glycosylation profile of a monoclonal antibody produced by hybridomas cultured in serum-supplemented, serum-free or chemically defined media.

    Science.gov (United States)

    Serrato, J Antonio; Hernández, Vanessa; Estrada-Mondaca, Sandino; Palomares, Laura A; Ramírez, Octavio T

    2007-06-01

    SFM (serum-free medium) is preferred to media containing animal-derived components when culturing mammalian cells for the production of therapeutic recombinant proteins and mAbs (monoclonal antibodies). Nonetheless, eliminating animal-derived components from media can strongly modify culture performance and alter protein glycosylation. In the present study, mAb glycosylation profiles, extracellular exoglycosidase activities, hybridoma growth and mAb production in traditional medium containing 10% (v/v) FBS (fetal bovine serum) [DMEM (Dulbecco's modified Eagle's medium)/FBS] were compared with those obtained in either SFM or CDM (chemically defined medium). SFM and CDM supported higher cell and mAb concentrations than did DMEM/FBS; however, CE (capillary electrophoresis) analyses revealed important changes in mAb glycosylation patterns. Glycosylation patterns showed a broad microheterogeneity in all the media, ranging from complex to high-mannose and paucimannosidic glycans. mAb produced in DMEM/FBS presented 26 glycan structures, whereas a lower glycan microheterogeneity was found for cultures in CDM or SFM, which presented 24 and 22 structures respectively. In DMEM/FBS and CDM, complex glycans without terminal galactose (G0) represented 28 and 32% of the total glycans respectively and 42 and 46% corresponded to galactosylated structures (G1 plus G2) respectively. In contrast, G0 glycans in SFM accounted for 58%, whereas only 28% corresponded to G1 and G2 structures. Extracellular beta-galactosidase activity increased approx. 3-fold in SFM, which can explain the higher G0 content compared with cultures in the other two media. A desirable decrease in sialylated structures, but an undesirable increase in fucosylated forms, was observed in mAb produced in SFM and CDM media. Approxi. 80% of potential mAb glycosylation sites were occupied, regardless of the culture medium used.

  12. Platelet-Rich Gel Supernatants Stimulate the Release of Anti-Inflammatory Proteins on Culture Media of Normal Equine Synovial Membrane Explants.

    Science.gov (United States)

    Ríos, Diana L; López, Catalina; Carmona, Jorge U

    2015-01-01

    The aims were as follows: (1) to evaluate the effects at 48 and 96 h of two concentrations (25 and 50%) of leukocyte and platelet-rich gel (L-PRG) and pure PRG (P-PRG) supernatants on the production/degradation in normal equine synovial membrane explants (SME) of platelet derived growth factor isoform BB, transforming growth factor beta-1, tumor necrosis factor alpha, interleukin (IL-) 4 (IL-4), IL-1 receptor antagonist (IL-1ra), and hyaluronan (HA) synthesis and (2) to correlate these molecules with their respective PRG supernatant treatments. SME from 6 horses were cultured for 96 h with L-PRG and P-PRG supernatants at 25 and 50% concentrations, respectively. SME culture media were changed each 48 h and used for determination by ELISA of the molecules, which were also determined in synovial fluid. 25% L-PRG supernatant produced a sustained release over time of IL-1ra and a gradual release of HA, whereas 50% L-PRG supernatant produced a sustained increase over time of IL-4 and HA. 50% P-PRG supernatant produced an increased and sustained production of IL-1ra and IL-4. The cellular composition and the articular concentration (volume) of a platelet-rich plasma preparation could affect the anti-inflammatory and anabolic joint responses in horses with osteoarthritis.

  13. Biological synthesis of very small silver nanoparticles by culture supernatant of Klebsiella pneumonia: The effects of visible-light irradiation and the liquid mixing process

    Energy Technology Data Exchange (ETDEWEB)

    Mokhtari, Narges [Department of Pharmaceutical Biotechnology and Biotechnology Research Center, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Daneshpajouh, Shahram; Seyedbagheri, Seyedali; Atashdehghan, Reza [Hydrometallurgy Research Unit, Research and Development Center, National Iranian Copper Industries Company, Sarcheshmeh, Rafsanjan (Iran, Islamic Republic of); Abdi, Khosro [Department of Pharmaceutical Biotechnology and Biotechnology Research Center, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Sarkar, Saeed [Research Center for Science and Technology in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Minaian, Sara [Department of Pharmaceutical Biotechnology and Biotechnology Research Center, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Shahverdi, Hamid Reza [Department of Material Science, Faculty of Engineering, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Shahverdi, Ahmad Reza, E-mail: shahverd@sina.tums.ac.ir [Department of Pharmaceutical Biotechnology and Biotechnology Research Center, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of)

    2009-06-03

    This study has investigated different visible-light irradiation's effect on the formation of silver nanoparticles from silver nitrate using the culture supernatant of Klebsiella pneumonia. Our study shows that visible-light emission can significantly prompt the synthesis of silver nanoparticles. Also, the study experimentally investigated the liquid mixing process effect on silver nanoparticle synthesis by visible-light irradiation. This study successfully synthesized uniformly dispersed silver nanoparticles with a uniform size and shape in the range of 1-6 nm with an average size of 3 nm. Furthermore, the study investigated the mechanism of the reduction of silver ions by culture supernatant of K. pneumonia, and used X-ray diffraction to characterize silver chloride as an intermediate compound. Silver chloride was prepared synthetically and used as a substrate for the synthesis of silver nanoparticles by culture supernatant of K. pneumonia. The silver nanoparticles have been prepared from silver chloride during this investigation for the first time.

  14. Modulation of the Culture Supernatant of Decidual Cells with Exogenous Cytokines on Killing Activity of Natural Killer Cells in Early Pregnancy

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To investigate the important function of cytokines in early pregnancy and to provide basic and experimental evidence for understanding the mechanism of their action. Methods Add interferon-γ (IFN-γ) , interleukin- 2(IL- 2) , interleukin- 6(IL-6) and epidermal growth factor (EGF) to the confluent culturing decidual cells with three different concentrations and harvest the culture supernatant after 12, 24 and 48 h separately. Observe the effect of the supernatant on killing activity of NK cells with radioimmunological assay of 51Cr immersion. Results The culture supernatant of decidual cells can promote the killing activity of NK cells in various degrees, and the effect is independent of the type, concentration and acting time of cytokines. Conclusion In normal pregnancy, decidual cytokine network is in a dynamic equilibri um. Exogenous cytokines would be harm to normal pregnancy by interfering the equi librium state, but the exact mechanism needs further study.

  15. Modulation of the Culture Supernatant of Decidual Cells with Exogenous Cytokines on Killing Activity of Natural Killer Cells in Early Pregnancy

    Institute of Scientific and Technical Information of China (English)

    胡冬梅; 王丽莉; 何援利

    2000-01-01

    Objective To investigate the important function of cytohines in early pregnancy and to provide basic and experimental evidence for understanding the mechanism of their action.Methods Add interferon-γ (IFN-γ) ,interleuhin-2(IL-2) , interleuhin-6(IL-6) andepidermal growth factor(EGF) to the confluent culturing decidual cells with three different concentrations and harvest the culture supernatant after 12, 24 and 48 h separately. Observe the effect of the supernatant on killing activity of NK cells with radioimmunological assay of 51Cr immersion.Results The culture supernatant of decidual cells can promote the killing activity of NK cells in various degrees, and the effect is independent of the type, concentration and acting time of cytokines.Conclusion In normal pregnancy, decidual cytokine network is in a dynamic equilibri-um. Exogenous cytokines would be harm to normal pregnancy by interfering the equi-librium state, but the exact mechanism needs further study.

  16. Quantitative Determination of Lethal Toxin Proteins in Culture Supernatant of Human Live Anthrax Vaccine Bacillus anthracis A16R.

    Science.gov (United States)

    Zai, Xiaodong; Zhang, Jun; Liu, Ju; Liu, Jie; Li, Liangliang; Yin, Ying; Fu, Ling; Xu, Junjie; Chen, Wei

    2016-02-25

    Bacillus anthracis (B. anthracis) is the etiological agent of anthrax affecting both humans and animals. Anthrax toxin (AT) plays a major role in pathogenesis. It includes lethal toxin (LT) and edema toxin (ET), which are formed by the combination of protective antigen (PA) and lethal factor (LF) or edema factor (EF), respectively. The currently used human anthrax vaccine in China utilizes live-attenuated B. anthracis spores (A16R; pXO1+, pXO2-) that produce anthrax toxin but cannot produce the capsule. Anthrax toxins, especially LT, have key effects on both the immunogenicity and toxicity of human anthrax vaccines. Thus, determining quantities and biological activities of LT proteins expressed by the A16R strain is meaningful. Here, we explored LT expression patterns of the A16R strain in culture conditions using another vaccine strain Sterne as a control. We developed a sandwich ELISA and cytotoxicity-based method for quantitative detection of PA and LF. Expression and degradation of LT proteins were observed in culture supernatants over time. Additionally, LT proteins expressed by the A16R and Sterne strains were found to be monomeric and showed cytotoxic activity, which may be the main reason for side effects of live anthrax vaccines. Our work facilitates the characterization of anthrax vaccines components and establishment of a quality control standard for vaccine production which may ultimately help to ensure the efficacy and safety of the human anthrax vaccine A16R.

  17. Putative new heat-stable cytotoxic and enterotoxic factors in culture supernatant of Escherichia coli isolated from drinking water

    Directory of Open Access Journals (Sweden)

    DA Ribeiro

    2011-01-01

    Full Text Available Enteric infections caused by the ingestion of contaminated water, especially by Escherichia coli, are important to define the virulence properties of these bacteria. Due to frequent infantile diarrhea in the city of Ouro Preto, Minas Gerais state, Brazil, the phenotypic and genotypic diarrheagenic properties of E. coli isolated from drinking water were studied. The culture supernatants of 39 (40% among a total of 97 E. coli isolates from drinking water were positive by suckling mouse assay and induced cytotoxic effects on Vero cells. The enterotoxic and cytotoxic activities were present in the fraction with less than 10 kDa and were not lost when heated up to 60°C and 100°C for 30 minutes. PCR assays showed that among these 39 Vero cytotoxigenic E. coli, four (10.2% were positive for ST II (estB and two (5% positive for αHly (hlyA. Gene amplification of SLT (stx 1, stx 2, ST I (estA, LT (eltI, eltII, EAST1 (astA, EHly (enhly and plasmid-encoded enterotoxin (pet were not observed. This heat-stable cytotoxic enterotoxin of E. coli is probably a new putative diarrheagenic virulence factor, as a toxin presenting these characteristics has not yet been described.

  18. A novel toxin homologous to large clostridial cytotoxins found in culture supernatant of Clostridium perfringens type C.

    Science.gov (United States)

    Amimoto, Katsuhiko; Noro, Taichi; Oishi, Eiji; Shimizu, Mitsugu

    2007-04-01

    An unknown cytotoxin was identified in the culture supernatant of Clostridium perfringens type C. The cytotoxin, named TpeL, which was purified using mAb-based affinity chromatography, had a lethal activity of 62 minimum lethal dose (MLD) mg(-1) in mice and a cytotoxic activity of 6.2x10(5) cytotoxic units (CU) mg(-1) in Vero cells. The nucleotide sequence of TpeL was determined. The entire ORF had a length of 4953 bases, and the same nucleotide sequence was not recorded in the GenBank/EMBL/DDBJ databases. The molecular mass calculated from the deduced amino acid sequence was 191 kDa, and a signal peptide region was not found within the ORF. The deduced amino acid sequence exhibited 30-39 % homology to Clostridium difficile toxins A (TcdA) and B (TcdB), Clostridium sordellii lethal toxin (TcsL) and Clostridium novyi alpha-toxin (TcnA). The amino acid sequence of TpeL is shorter than these toxins, and the homologous region was located at the N-terminal site. Eighteen strains of C. perfringens types A, B and C were surveyed for the presence of the tpeL gene by PCR. The tpeL gene was detected in all type B (one strain) and C strains (five strains), but not in any type A strains (12 strains). TpeL was detected in culture filtrates of the five type C strains by dot-blot analysis, but not in the type B strain. It was concluded that TpeL is a novel toxin similar to the known large clostridial cytotoxins. Furthermore, the data indicated that TpeL is produced by many C. perfringens type C strains.

  19. [Effects of combined application of culture supernatant of human umbilical cord mesenchymal stem cells and ciprofloxacin on Staphylococcus aureus in vitro].

    Science.gov (United States)

    Zhou, B; Tu, H L; Ba, T; Wang, L F; Wang, S J; Nie, S Y

    2017-06-20

    Objective: To explore the effects of combined application of culture supernatant of human umbilical cord mesenchymal stem cells (hUCMSCs) and ciprofloxacin on Staphylococcus aureus (SA) in vitro. Methods: hUCMSCs were isolated from umbilical cord tissue of full-term healthy fetus after cesarean section and cultured. Cells in the third passage were used in the experiments after identification. SA strains isolated from wounds of burn patients in our burn wards were used in the experiments. Cells were divided into 0, 10, 100, and 1 000 ng/mL lipopolysaccharide (LPS) groups according to the random number table (the same dividing method below). Cells were cultured with culture medium of mesenchymal stem cells (MSCs) after being treated with medium containing the corresponding mass concentrations of LPS for 12 h. At post culture hour (PCH) 6, 12, and 24, 6 wells of culture supernatant of cells in each group were obtained to measure the content of LL-37 with enzyme-linked immunosorbent assay. Ninety blood agar plates were divided into ciprofloxacin control group (CC), ciprofloxacin+ supernatant group (CS), and ciprofloxacin+ supernatant+ LL-37 antibody group (CSL), with 30 blood agar plates in each group. Blood agar plates in group CC were coated with 1.5×10(8) colony forming unit (CFU)/mL bacteria solution prepared with normal saline. Blood agar plates in group CS were coated with 1.5×10(8) CFU/mL bacteria solution prepared with normal saline and culture supernatant of hUCMSCs (cultured by culture medium of MSCs, the same below) in double volume of normal saline. Blood agar plates in group CSL were coated with 1.5×10(8) CFU/mL bacteria solution prepared with normal saline, culture supernatant of hUCMSCs in double volume of normal saline, and 2.6 μL LL-37 antibody in the concentration of 2 μg/mL. At PCH 12, 24, and 48, 10 blood agar plates of each group were harvested to observe the distribution of SA colony on blood agar plate and to measure the diameter of

  20. Understanding ForteBio's Sensors for High-Throughput Kinetic and Epitope Screening for Purified Antibodies and Yeast Culture Supernatant.

    Science.gov (United States)

    Yu, Yao; Mitchell, Scott; Lynaugh, Heather; Brown, Michael; Nobrega, R Paul; Zhi, Xiaoyong; Sun, Tingwan; Caffry, Isabelle; Cao, Yuan; Yang, Rong; Burnina, Irina; Xu, Yingda; Estep, Patricia

    2016-01-01

    Real-time and label-free antibody screening systems are becoming more popular because of the increasing output of purified antibodies and antibody supernatant from many antibody discovery platforms. However, the properties of the biosensor can greatly affect the kinetic and epitope binning results generated by these label-free screening systems. ForteBio human-specific ProA, anti-human IgG quantitation (AHQ), anti-human Fc capture (AHC) sensors, and custom biotinylated-anti-human Fc capture (b-AHFc) sensors were evaluated in terms of loading ability, regeneration, kinetic characterization, and epitope binning with both purified IgG and IgG supernatant. AHC sensors proved unreliable for kinetic or binning assays at times, whereas AHQ sensors showed poor loading and regeneration abilities. ProA sensors worked well with both purified IgG and IgG supernatant. However, the interaction between ProA sensors and the Fab region of the IgG with VH3 germline limited the application of ProA sensors, especially in the epitope binning experiment. In an attempt to generate a biosensor type that would be compatible with a variety of germlines and sample types, we found that the custom b-AHFc sensors appeared to be robust working with both purified IgG and IgG supernatant, with little evidence of sensor-related artifacts.

  1. Non-covalent pomegranate (Punica granatum) hydrolyzable tannin-protein complexes modulate antigen uptake, processing and presentation by a T-cell hybridoma line co-cultured with murine peritoneal macrophages.

    Science.gov (United States)

    Madrigal-Carballo, Sergio; Haas, Linda; Vestling, Martha; Krueger, Christian G; Reed, Jess D

    2016-12-01

    In this work we characterize the interaction of pomegranate hydrolyzable tannins (HT) with hen egg-white lysozyme (HEL) and determine the effects of non-covalent tannin-protein complexes on macrophage endocytosis, processing and presentation of antigen. We isolated HT from pomegranate and complex to HEL, the resulting non-covalent tannin-protein complex was characterized by gel electrophoresis and MALDI-TOF MS. Finally, cell culture studies and confocal microscopy imaging were conducted on the non-covalent pomegranate HT-HEL protein complexes to evaluate its effect on macrophage antigen uptake, processing and presentation to T-cell hybridomas. Our results indicate that non-covalent pomegranate HT-HEL protein complexes modulate uptake, processing and antigen presentation by mouse peritoneal macrophages. After 4 h of pre-incubation, only trace amounts of IL-2 were detected in the co-cultures treated with HEL alone, whereas a non-covalent pomegranate HT-HEL complex had already reached maximum IL-2 expression. Pomegranate HT may increase rate of endocytose of HEL and subsequent expression of IL-2 by the T-cell hybridomas.

  2. Characterization of monoclonal antibodies by a fast and easy liquid chromatography-mass spectrometry time-of-flight analysis on culture supernatant.

    Science.gov (United States)

    Henninot, Antoine; Terrier, Aurelie; Charton, Julie; Urbain, Rémi; Fontayne, Alexandre; Deprez, Benoit; Beghyn, Terence

    2015-12-15

    Rapid and efficient structural analysis is key to the development of new monoclonal antibodies. We have developed a fast and easy process to obtain mass spectrometry profiles of antibodies from culture supernatant. Treatment of the supernatant with IdeS generates three fragments of 25 kDa that can be analyzed by liquid chromatography-mass spectrometry time-of-flight (LC-MS TOF) in one run: LC, Fd, and Fc/2. This process gives rapid access to isoform and glycoform profiles. To specifically measure the fucosylation yield, we included a one-pot treatment with EndoS that removes the distal glycan heterogeneity. Our process was successfully compared with high-performance capillary electrophoresis with laser-induced fluorescence detection (HPCE-LIF), currently considered as the "gold standard" method. Copyright © 2015 Elsevier Inc. All rights reserved.

  3. Effect of amino acid supplementation on titer and glycosylation distribution in hybridoma cell cultures-Systems biology-based interpretation using genome-scale metabolic flux balance model and multivariate data analysis.

    Science.gov (United States)

    Reimonn, Thomas M; Park, Seo-Young; Agarabi, Cyrus D; Brorson, Kurt A; Yoon, Seongkyu

    2016-09-01

    Genome-scale flux balance analysis (FBA) is a powerful systems biology tool to characterize intracellular reaction fluxes during cell cultures. FBA estimates intracellular reaction rates by optimizing an objective function, subject to the constraints of a metabolic model and media uptake/excretion rates. A dynamic extension to FBA, dynamic flux balance analysis (DFBA), can calculate intracellular reaction fluxes as they change during cell cultures. In a previous study by Read et al. (2013), a series of informed amino acid supplementation experiments were performed on twelve parallel murine hybridoma cell cultures, and this data was leveraged for further analysis (Read et al., Biotechnol Prog. 2013;29:745-753). In order to understand the effects of media changes on the model murine hybridoma cell line, a systems biology approach is applied in the current study. Dynamic flux balance analysis was performed using a genome-scale mouse metabolic model, and multivariate data analysis was used for interpretation. The calculated reaction fluxes were examined using partial least squares and partial least squares discriminant analysis. The results indicate media supplementation increases product yield because it raises nutrient levels extending the growth phase, and the increased cell density allows for greater culture performance. At the same time, the directed supplementation does not change the overall metabolism of the cells. This supports the conclusion that product quality, as measured by glycoform assays, remains unchanged because the metabolism remains in a similar state. Additionally, the DFBA shows that metabolic state varies more at the beginning of the culture but less by the middle of the growth phase, possibly due to stress on the cells during inoculation. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1163-1173, 2016.

  4. Cell-free culture supernatant of Bifidobacterium breve CNCM I-4035 decreases pro-inflammatory cytokines in human dendritic cells challenged with Salmonella typhi through TLR activation.

    Science.gov (United States)

    Bermudez-Brito, Miriam; Muñoz-Quezada, Sergio; Gomez-Llorente, Carolina; Matencio, Esther; Bernal, Maria J; Romero, Fernando; Gil, Angel

    2013-01-01

    Dendritic cells (DCs) constitute the first point of contact between gut commensals and our immune system. Despite growing evidence of the immunomodulatory effects of probiotics, the interactions between the cells of the intestinal immune system and bacteria remain largely unknown. Indeed,, the aim of this work was to determine whether the probiotic Bifidobacterium breve CNCM I-4035 and its cell-free culture supernatant (CFS) have immunomodulatory effects in human intestinal-like dendritic cells (DCs) and how they respond to the pathogenic bacterium Salmonella enterica serovar Typhi, and also to elucidate the molecular mechanisms involved in these interactions. Human DCs were directly challenged with B. breve/CFS, S. typhi or a combination of these stimuli for 4 h. The expression pattern of genes involved in Toll-like receptor (TLR) signaling pathway and cytokine secretion was analyzed. CFS decreased pro-inflammatory cytokines and chemokines in human intestinal DCs challenged with S. typhi. In contrast, the B. breve CNCM I-4035 probiotic strain was a potent inducer of the pro-inflammatory cytokines and chemokines tested, i.e., TNF-α, IL-8 and RANTES, as well as anti-inflammatory cytokines including IL-10. CFS restored TGF-β levels in the presence of Salmonella. Live B.breve and its supernatant enhanced innate immune responses by the activation of TLR signaling pathway. These treatments upregulated TLR9 gene transcription. In addition, CFS was a more potent inducer of TLR9 expression than the probiotic bacteria in the presence of S. typhi. Expression levels of CASP8 and IRAK4 were also increased by CFS, and both treatments induced TOLLIP gene expression. Our results indicate that the probiotic strain B. breve CNCM I-4035 affects the intestinal immune response, whereas its supernatant exerts anti-inflammatory effects mediated by DCs. This supernatant may protect immune system from highly infectious agents such as Salmonella typhi and can down-regulate pro

  5. Food preservative potential of gassericin A-containing concentrate prepared from a cheese whey culture supernatant from Lactobacillus gasseri LA39.

    Science.gov (United States)

    Nakamura, Kiyoshi; Arakawa, Kensuke; Kawai, Yasushi; Yasuta, Narimi; Chujo, Takahiro; Watanabe, Masamichi; Iioka, Hiroyuki; Tanioka, Masashi; Nishimura, Junko; Kitazawa, Haruki; Tsurumi, Koichi; Saito, Tadao

    2013-02-01

    Gassericin A (GA) is a circular bacteriocin produced by Lactobacillus gasseri LA39. In this study, GA-containing concentrate was prepared using a cross-flow membrane filtration device (30 kDa cut-off) from the culture supernatant of Lb. gasseri LA39 cultivated in a cheese whey-based food-grade medium. The bacteriocin activity titer in the concentrate was 16 times as high as that of the culture supernatant and was completely maintained through each incubation at 4°C for 3 months, 37°C for 2 months, 60°C for 5 h, and 100°C for 30 min. The GA-containing concentrate was used with glycine powder to make custard creams, and then four representative strains of custard cream spoilage bacteria (Bacillus cereus, Lactococcus lactis subsp. lactis, Achromobacter denitrificans and Pseudomonas fluorescens) were individually inoculated at c. 10(3) colony forming units/g in the custard creams. Throughout 30 days of incubation at 30°C, all of the inoculated bacteria were completely inhibited by the combination of 5% (w/w) of the GA-containing concentrate and 0.5% (w/w) glycine. This is the first highly practical application of GA to foods as a biopreservative, and the concentration method and the bacteriocin concentrate would contribute to biopreservation of several foods.

  6. High-efficient n-butanol production by co-culturing Clostridium acetobutylicum and Saccharomyces cerevisiae integrated with butyrate fermentative supernatant addition.

    Science.gov (United States)

    Luo, Hongzhen; Zeng, Qingwei; Han, Shuo; Wang, Zhaoyu; Dong, Qing; Bi, Yanhong; Zhao, Yuping

    2017-04-01

    Butanol is not only an important chemical intermediate and solvent in pharmaceutical and cosmetics industries, but also considered as an advanced biofuel. Although species of the natural host Clostridium have been engineered, butanol titers in the anaerobe seem to be limited by its intolerance to butanol less than 13 g/L. Here we aimed to develop a technology for enhancing butanol production by a co-culture system with butyrate fermentative supernatant addition. First, when adding 4.0 g/L butyrate into the acetone-butanol-ethanol (ABE) fermentation broth with single-shot at 24 h, the "acid crash" phenomenon occurred and the ABE fermentation performance deteriorated. Subsequently, we found that adding certain amino acids could effectively enhance butyrate re-assimilation, butanol tolerance and titer (from 11.1 to 14.8 g/L). Additionally, in order to decrease the raw material cost, butyrate fermentative supernatant produced by Clostridium tyrobutyricum was applied to butanol production in the Clostridium acetobutylicum/Saccharomyces cerevisiae co-culture system, instead of adding synthetic butyrate. Final butanol and total ABE concentrations reached higher levels of 16.3 and 24.8 g/L with increments of 46.8 and 37.8%, respectively. These results show that the proposed fermentation strategy has great potential for efficiently butanol production with an economic approach.

  7. Distinct galactofuranose antigens in the cell wall and culture supernatants as a means to differentiate Fusarium from Aspergillus species.

    Science.gov (United States)

    Wiedemann, Annegret; Kakoschke, Tamara Katharina; Speth, Cornelia; Rambach, Günter; Ensinger, Christian; Jensen, Henrik Elvang; Ebel, Frank

    2016-09-01

    Detection of carbohydrate antigens is an important means for diagnosis of invasive fungal infections. For diagnosis of systemic Aspergillus infections, galactomannan is commonly used, the core antigenic structure of which consists of chains of several galactofuranose moieties. In this study, we provide evidence that Fusarium produces at least two distinct galactofuranose antigens: Smaller amounts of galactomannan and larger quantities of a novel antigen recognized by the monoclonal antibody AB135-8. In A. fumigatus, only minor amounts of the AB135-8 antigen are found in supernatants and in the apical regions of hyphae. A galactofuranose-deficient A. fumigatus mutant lacks the AB135-8 antigen, which strongly suggests that galactofuranose is an essential constituent of this antigen. Using a combination of AB135-8 and a galactomannan-specific antibody, we were able to unambiguously differentiate A. fumigatus and Fusarium hyphae in immunohistology. Moreover, since Fusarium releases the AB135-8 antigen, it appears to be a promising target antigen for a serological detection of Fusarium infections. Copyright © 2016 Elsevier GmbH. All rights reserved.

  8. Culture supernatants of breast cancer cell line MDA-MB-231 treated with parthenolide inhibit the proliferation, migration, and lumen formation capacity of human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    LI Cai-juan; GUO Su-fen; SHI Tie-mei

    2012-01-01

    Background Parthenolide has been tested for anti-tumor activities,such as anti-proliferation and pro-apoptosis in recent studies.However,little is known about its role in the process of tumor angiogenesis.This study aims to investigate the effects and potential mechanisms of parthenolide on the proliferation,migration and lumen formation capacity of human umbilical vein endothelial cells.Methods Different concentrations of parthenolide were applied to the human breast cancer cell line MDA-MB-231 cells.After 24-hour incubation,the culture supematants were harvested and used to treat human umbilical vein endothelial cells for 24 hours.Then an inverted fluorescence phase contrast microscope was used to evaluate the human umbilical vein endothelial cells.The secretion of vascular endothelial growth factor (VEGF),interleukin (IL)-8 and matrix metalloproteinases (MMP)-9 in the culture supernatant of the MDA-MB-231 cells was then measured with enzyme-linked immunosorbent assay (ELISA) assays.Results Suppression of proliferation,migration,and the lumen formation capacity of human umbilical vein endothelial cells was observed in the presence of the culture supernatants from the breast cancer cell line treated with different concentrations of parthenolide.Parthenolide decreased the levels of the angiogenic factors MMP-9,VEGF,and IL-8secreted by the MDA-MB-231 cells.Conclusions Parthenolide may suppress angiogenesis through decreasing angiogenic factors secreted by breast cancer cells to interfere with the proliferation,migration and lumen-like structure formation of endothelial cells,thereby inhibiting tumor growth.It is a promising potential anti-angiogenic drug.

  9. Performance of multiplex commercial kits to quantify cytokine and chemokine responses in culture supernatants from Plasmodium falciparum stimulations.

    Directory of Open Access Journals (Sweden)

    Gemma Moncunill

    Full Text Available BACKGROUND: Cytokines and chemokines are relevant biomarkers of pathology and immunity to infectious diseases such as malaria. Several commercially available kits based on quantitative suspension array technologies allow the profiling of multiple cytokines and chemokines in small volumes of sample. However, kits are being continuously improved and information on their performance is lacking. METHODOLOGY/PRINCIPAL FINDINGS: Different cytokine/chemokine kits, two flow cytometry-based (eBioscience® FlowCytomix™ and BD™ Cytometric Bead Array Human Enhanced Sensitivity and four Luminex®-based (Invitrogen™ Human Cytokine 25-Plex Panel, Invitrogen™ Human Cytokine Magnetic 30-Plex Panel, Bio-Rad® Bio-Plex Pro™ Human Cytokine Plex Assay and Millipore™ MILLIPLEX® MAP Plex Kit were compared. Samples tested were supernatants of peripheral blood mononuclear cells of malaria-exposed children stimulated with Plasmodium falciparum parasite lysates. Number of responses in range that could be detected was determined and reproducibility of duplicates was evaluated by the Bland-Altman test. Luminex® kits performed better than flow cytometry kits in number of responses in range and reproducibility. Luminex® kits were more reproducible when magnetic beads were used. However, within each methodology overall performance depended on the analyte tested in each kit. Within the Luminex® kits, the Invitrogen™ with polystyrene beads had the poorer performance, whereas Invitrogen™ with magnetic beads had the higher percentage of cytokines/chemokines with both readings in range (40%, followed by Bio-Rad® with magnetic beads (35%. Regarding reproducibility, the Millipore™ kit had the highest percentage (60% of cytokines/chemokines with acceptable limits of agreement (<30%, followed by the Invitrogen™ with magnetic beads (40% that had tighter limits of agreement. CONCLUSIONS/SIGNIFICANCE: Currently available kits for cytokine and chemokine

  10. Screening for epitope specificity directly on culture supernatants in the early phase of monoclonal antibody production by an ELISA with biotin-labeled antigen

    DEFF Research Database (Denmark)

    Andersen, Ditte C; Jensen, Charlotte H; Gregersen, Annemette

    2004-01-01

    This report describes an assay for comparison of epitope specificity in groups of monoclonal antibodies against a given antigen. The only prerequisite is the biotin-labeled antigen. One of the monoclonal antibodies is captured onto a plastic surface via a rabbit anti-mouse Ig, and the other...... preincubated with biotinylated antigen. When the two antibodies react with the same epitope subsequent binding of the biotin-labeled antigen is abolished (inhibition). In the cases where no inhibition was observed, the two antibodies were considered to react with distinct, independent epitopes. The obvious...... advantages using this assay, are that it can be performed directly on culture supernatants in the early phase of monoclonal antibody production, and also works for antigens with repetitive epitopes. Moreover, the bonus effect, i.e., a signal in excess of the reference signal when sets of monoclonal...

  11. Comprehensive supernatant treatment

    Energy Technology Data Exchange (ETDEWEB)

    Egan, Z. [Oak Ridge National Lab., TN (United States)

    1996-10-01

    This task involves testing of sorbent materials for removing cesium, strontium, and technetium from the saline solutions in DOE storage tank supernatant at Oak Ridge and other sites. Staff at Oak Ridge National Laboratory (ORNL) are recovering and treating the liquid (supernatant) portions of Melton Valley Storage Tank (MVST) waste in a hot cell to separate and remove the radionuclides. Batch tests will be used to evaluate and select the most promising materials for supernatant treatment to reduce the amount of waste for final disposal. Small column tests will be made on selected sorbents to verify the batch data and to obtain additional data for process design. Efforts will be made to obtain samples of tank supernatant from Hanford for comparison.

  12. Lactobacillus rhamnosus and its cell-free culture supernatant differentially modulate inflammatory biomarkers in Escherichia coli-challenged human dendritic cells.

    Science.gov (United States)

    Bermudez-Brito, Miriam; Muñoz-Quezada, Sergio; Gomez-Llorente, Carolina; Romero, Fernando; Gil, Angel

    2014-05-28

    The intestinal immune system maintains a delicate balance between immunogenicity against invading pathogens and tolerance to the commensal microbiota and food antigens. Different strains of probiotics possess the ability to finely regulate the activation of dendritic cells (DC), polarising the subsequent activity of T-cells. Nevertheless, information about their underlying mechanisms of action is scarce. In the present study, we investigated the immunomodulatory effects of a potentially probiotic strain, Lactobacillus rhamnosus CNCM I-4036, and its cell-free culture supernatant (CFS) on human DC challenged with Escherichia coli. The results showed that the levels of pro-inflammatory cytokines such as IL-1β, IL-6, IL-8 and IL-12p70 were higher in the cells treated with live L. rhamnosus than in the cells treated with the CFS. In the presence of E. coli, the supernatant was more effective than the probiotic bacteria in reducing the secretion of pro-inflammatory cytokines. In addition, live L. rhamnosus potently induced the production of transforming growth factor (TGF)-β1 and TGF-β2, whereas the CFS increased the secretion of TGF-β1. However, in the presence of E. coli, both treatments restored the levels of TGF-β. The probiotic strain L. rhamnosus CNCM I-4036 and its CFS were able to activate the Toll-like receptor signalling pathway, enhancing innate immunity. The two treatments induced gene transcription of TLR-9. Live L. rhamnosus activated the expression of TLR-2 and TLR-4 genes, whereas the CFS increased the expression of TLR-1 and TLR-5 genes. In response to the stimulation with probiotic/CFS and E. coli, the expression of each gene tested was notably increased, with the exception of TNF-α and NFKBIA. In conclusion, the CFS exhibited an extraordinary ability to suppress the production of pro-inflammatory cytokines by DC, and may be used as an effective and safer alternative to live bacteria.

  13. Calcium carbonate mediates higher lignin peroxidase activity in the culture supernatant of Streptomyces Viridosporus T7A

    Directory of Open Access Journals (Sweden)

    J. M. B. MACEDO

    1999-06-01

    Full Text Available Lignin peroxidase (LiP production has been extensively studied due to the potential use of this enzyme in environmental pollution control. Important aspects of the production of the enzyme by S. viridosporus T7A which have been studied include the improvement of yield and enzyme stabilization. In experiments performed in agitated flasks containing culture media composed of yeast extract as the source of nitrogen, mineral salts and different carbon sources, the use of glucose resulted in the highest values for LiP activity (350 U/L, specific LiP activity (450 U/g and productivity (7 U/L/h. As the profile obtained with glucose-containing medium suggested enzyme instability, the effect of calcium carbonate was evaluated. The addition of CaCO3 in two different concentrations, 0.5% and 5.0%, resulted in higher values of maximum LiP activity, 600 and 900 U/L, respectively. The presence of this salt also anticipated enzyme activity peaks and allowed the detection of higher enzyme activities in the extracellular medium for longer periods of time. These results indicate a positive effect of calcium carbonate on LiP production, which is extremely relevant for industrial processes.

  14. Generation of human hybridomas producing migration inhibitory factor (MIF) and of murine hybridomas secreting monoclonal antibodies to human MIF.

    Science.gov (United States)

    Weiser, W Y; Remold, H G; David, J R

    1985-01-01

    Human T-cell hybridomas were established by hybridization of concanavalin A (Con A)-stimulated human peripheral blood T lymphocytes with cells from a 6-thioguanine-resistant, aminopterin-sensitive mutant line designated CEM-WH4, derived from the continuously growing human T cell line, CEM. High levels of MIF activity were demonstrated in the supernatants of two hybridoma lines, T-CEMA and T-CEMB but not of CEM-WH4 when stimulated with phorbol myristate acetate and phytohemagglutinin. In comparison, MIF derived from Con A-stimulated peripheral blood mononuclear cells showed 100 times less activity. Upon isoelectrofocusing, MIF activity of T-CEMB was found exclusively between pH 4.6 and 5.3 whereas MIF derived from T-CEMA showed heterogeneity with a major peak of MIF recovered at pH 4.6-5.3 and a minor peak at pH 2.4-3.3. These molecules, however, were all found to have an apparent MW of 68,000 and were resistant to trypsin. Most of these characteristics are in accordance with second day pH 3- and pH 5-MIF derived from peripheral blood mononuclear cells. When spleen cells from BALB/c mice immunized with T-CEMB-MIF were used to fuse with NS-1 mouse myeloma cells, nine hybridomas secreting antibodies to human MIF were obtained. Clone D112 which demonstrated the highest MIF-neutralizing activity was found to neutralize MIF derived from T-CEMA, peripheral blood mononuclear cells, and a T cell line, Mo.

  15. Analysis of metal Bioleaching from thermal power plant fly ash by Aspergillus niger 34770 culture supernatant and reduction of phytotoxicity during the process.

    Science.gov (United States)

    Jadhav, Umesh U; Hocheng, Hong

    2015-01-01

    Aspergillus niger culture supernatant is used for bioleaching process. Before starting bioleaching process, fly ash was washed with distilled water. This removed 100 % sodium, 47 % (±0.45) boron, 38.07 % (±0.12) calcium, 29.89 % (±0.78) magnesium, and 11.8 % (±0.05) potassium. The pH was reduced from 10.5 to 8.5 after water washing. During bioleaching process, around 100 % metal removal was achieved in 4 h for all metals except chromium 93 % (±1.18), nickel 83 % (±0.32), arsenic 78 % (±0.52), and lead 70 % (±0.20). The process parameters including temperature, shaking speed, and solid/liquid ratio were optimized for bioleaching process. Experiments were conducted to evaluate effect of fly ash on growth of mung bean (Vigna radiata). At 20 g/100 ml fly ash concentration no germination of V. radiata seeds was observed. With an increasing concentration of untreated fly ash, a gradual decrease in root/shoot length was observed. After bioleaching process 78 % (±0.19) germination of V. radiata was observed with 20 g/100 ml fly ash. This study will help to develop an efficient process to remove the toxic metals from fly ash.

  16. Inhibition of miR122a by Lactobacillus rhamnosus GG culture supernatant increases intestinal occludin expression and protects mice from alcoholic liver disease.

    Science.gov (United States)

    Zhao, Haiyang; Zhao, Cuiqing; Dong, Yuanyuan; Zhang, Min; Wang, Yuhua; Li, Fengyuan; Li, Xiaokun; McClain, Craig; Yang, Shulin; Feng, Wenke

    2015-05-05

    Alcoholic liver disease (ALD) has a high morbidity and mortality. Chronic alcohol consumption causes disruption of intestinal microflora homeostasis, intestinal tight junction barrier dysfunction, increased endotoxemia, and eventually liver steatosis/steatohepatitis. Probiotic Lactobacillus rhamnosus GG (LGG) and the bacteria-free LGG culture supernatant (LGGs) have been shown to promote intestinal epithelial integrity and protect intestinal barrier function in ALD. However, little is known about how LGGs mechanistically works to increase intestinal tight junction proteins. Here we show that chronic ethanol exposure increased intestinal miR122a expression, which decreased occludin expression leading to increased intestinal permeability. Moreover, LGGs supplementation decreased ethanol-elevated miR122a level and attenuated ethanol-induced liver injury in mice. Similar to the effect of ethanol exposure, overexpression of miR122a in Caco-2 monolayers markedly decreased occludin protein levels. In contrast, inhibition of miR122a increased occludin expression. We conclude that LGGs supplementation functions in intestinal integrity by inhibition of miR122a, leading to occludin restoration in mice exposed to chronic ethanol.

  17. The Effect of Lactobacillus crispatus and Lactobacillus rhamnosusCulture Supernatants on Expression of Autophagy Genes and HPV E6 and E7 Oncogenes in The HeLa Cell Line.

    Science.gov (United States)

    Motevaseli, Elahe; Azam, Rosa; Akrami, Seyed Mohammad; Mazlomy, Mohammadali; Saffari, Mojtaba; Modarressi, Mohammad Hossein; Daneshvar, Maryam; Ghafouri-Fard, Soudeh

    2016-01-01

    The aim of this study was to clarify the mechanism by which lactobacilli exert their cytotoxic effects on cervical cancer cells. In addition, we aimed to evalu- ate the effect of lactobacilli on the expression of human papilloma virus (HPV) onco- genes. In this experimental study, using quantitative real-time polymer- ase chain reaction (PCR), we analyzed the expression of CASP3 and three autophagy genes [ATG14, BECN1 and alpha 2 catalytic subunit of AMPK (PRKAA2)] along with HPV18 E6 and E7 genes in HeLa cells before and after treatment with Lactobacillus crispatus and Lactobacillus rhamnosus culture supernatants. The expression of CASP3 and autophagy genes in HeLa cells was de- creased after treatment with lactobacilli culture supernatants. However, this de- crease was not significant for PRKAA2 when compared with controls. In addition, expression of HPV E6 was significantly decreased after treatment with lactobacilli culture supernatants. Lactobacilli culture supernatants can decrease expression of ATG14 and BECN1 as well as the HPV E6 oncogene. It has been demonstrated that the main changes occurring during cervical carcinogenesis in cell machinery can be reversed by suppression of HPV oncogenes. Therefore, downregulation of HPV E6 by lacto- bacilli may have therapeutic potential for cervical cancer. As the role of autophagy in cancer is complicated, further work is required to clarify the link between downregula- tion of autophagy genes and antiproliferative effects exerted by lactobacilli.

  18. Detoxification of azo dyes mediated by cell-free supernatant culture with manganese-dependent peroxidase activity: effect of Mn2+ concentration and H2O2 dose.

    Science.gov (United States)

    Contreras, Elsa; Urra, Johana; Vásquez, Carlos; Palma, Carolyn

    2012-01-01

    White-rot fungi (WRF) are capable of degrading complex organic compounds such as lignin, and the enzymes that enable these processes can be used for the detoxification of recalcitrant organopollutants. The aim of this study is to evaluate a system based on the use of an in vitro ligninolytic enzyme for the detoxification of recalcitrant dye pollutants. The dyes selected for investigation were the anionic and cationic commercial azo dyes, basic blue 41 (BB41), acid black 1 (AB1), and reactive black 5 (RB5). A supernatant, cell-free culture of WRF with manganese peroxidase activity was used to investigate its degradative capacity under various conditions, and concentrations of cofactors, H(2)O(2) and Mn(2+). The assays were carried out using a 2(2) experimental designs whose variables were concentration of Mn(2+) (33 and 1,000 μM) and semicontinuous dosage of the H(2)O(2) (0.02 and 0.10 μmol) added at a frequency of 0.2 min(-1). The response variables analyzed were the efficiency and the initial rate of the decolorization process. The dye concentrations considered ranged from 10 to 200 mg L(-1). AB1 and RB5 were decolorized over the entire interval of concentrations studied; reaching efficiencies between 15 and 95%. Decolorization of up to 100 mg L(-1), BB41 had less than 30% efficiency. The decay of the concentration of AB1 was interpreted by two-stage kinetics model, with the exception of the condition of 33 μM Mn(2+)-0.02 μmol of H(2)O(2) in which only one stage was observed. For all assays performed with 33 μM Mn(2+), the initial rate of the decolorization process was found to be dependent on the dosage of H(2)O(2). The results of this study can be applied to the development bioreactors for the degradation of recalcitrant pollutants from the textile industry and may be used as a model for expanding the use of extracellular enzyme supernatants in bioremediation. Copyright © 2011 American Institute of Chemical Engineers (AIChE).

  19. A human-human hybridoma producing cytotoxic antibody to HLA-B15, cross-reacting with B17, B5, B35 and B18.

    Science.gov (United States)

    Hansen, T; Kolstad, A; Thorsby, E; Hannestad, K

    1987-05-01

    Mononuclear blood cells from a multiparous woman were transformed with Epstein Barr virus, and a cell line (Tr2D8) producing anti-HLA antibody was obtained. This cell line was immortalized by hybridization to the human fusion partners KR4 and KR12. While the EBV line died after 7 months, the hybridomas have remained stable for 13 months. The EBV line supernatant (40 micrograms IgM/ml) lysed peripheral blood mononuclear cells (PBMC) bearing B15, B17, B5 and B35. Consistent lysis of B18 bearing cells was only observed with lymphoblastoid cell lines. The supernatant from the Tr2D8 (EBV line X KR4) hybridoma (2.7 micrograms IgM/ml) only lysed B15 bearing PBMC. At a concentration of 13.5 micrograms IgM/ml, the hybridoma antibody lysed lymphoblastoid cell lines bearing B15, B17, B5, B35 and B18.

  20. Growth kinetics of hybridoma cells: (2) The effects of varying energy source concentrations.

    Science.gov (United States)

    Low, K; Harbour, C

    1985-01-01

    Commercial media used for the growth of hybridoma cells contain different glucose concentrations. We have attempted to establish the optimal levels of glucose required for maximum hybridoma cell yields by studying the energy source metabolism of two murine hybridoma cell lines in several commercial media in static 25 cm2 flask cultures. The glucose and lactate quotients and growth yields from glucose of the two cell lines were similar in all media. Glucose limited growth in DME containing lg/L. Glucose supplementation to 2g/L in DME significantly increased cell and antibody yields. When fructose was used as a substitute for glucose the fructose quotient of one cell line was found to be similar to its glucose quotient whereas that of the other cell line was significantly reduced compared to its glucose quotient.

  1. Monoclonal antibodies based on hybridoma technology.

    Science.gov (United States)

    Yagami, Hisanori; Kato, Hiroshi; Tsumoto, Kanta; Tomita, Masahiro

    2013-03-01

    Based on the size and scope of the present global market for medicine, monoclonal antibodies (mAbs) have a very promising future, with applications for cancers through autoimmune ailments to infectious disease. Since mAbs recognize only their target antigens and not other unrelated proteins, pinpoint medical treatment is possible. Global demand is dramatically expanding. Hybridoma technology, which allows production of mAbs directed against antigens of interest is therefore privileged. However, there are some pivotal points for further development to generate therapeutic antibodies. One is selective generation of human mAbs. Employment of transgenic mice producing human antibodies would overcome this problem. Another focus is recognition sites and conformational epitopes in antigens may be just as important as linear epitopes, especially when membrane proteins such as receptors are targeted. Recognition of intact structures is of critical importance for medical purposes. In this review, we describe patent related information for therapeutic mAbs based on hybridoma technology and also discuss new advances in hybridoma technology that facilitate selective production of stereospecific mAbs.

  2. Enhancement of hybridoma formation, clonability and cell proliferation in a nanoparticle-doped aqueous environment

    Directory of Open Access Journals (Sweden)

    Karnieli Ohad

    2008-01-01

    Full Text Available Abstract Background The isolation and production of human monoclonal antibodies is becoming an increasingly important pursuit as biopharmaceutical companies migrate their drug pipelines away from small organic molecules. As such, optimization of monoclonal antibody technologies is important, as this is becoming the new rate-limiting step for discovery and development of new pharmaceuticals. The major limitations of this system are the efficiency of isolating hybridoma clones, the process of stabilizing these clones and optimization of hybridoma cell secretion, especially for large-scale production. Many previous studies have demonstrated how perturbations in the aqueous environment can impact upon cell biology. In particular, radio frequency (RF irradiation of solutions can have dramatic effects on behavior of solutions, cells and in particular membrane proteins, although this effect decays following removal of the RF. Recently, it was shown that nanoparticle doping of RF irradiated water (NPD water produced a stabilized aqueous medium that maintained the characteristic properties of RF irradiated water for extended periods of time. Therefore, the ordering effect in water of the RF irradiation can now be studied in systems that required prolonged periods for analysis, such as eukaryotic cell culture. Since the formation of hybridoma cells involves the formation of a new membrane, a process that is affected by the surrounding aqueous environment, we tested these nanoparticle doped aqueous media formulations on hybridoma cell production. Results In this study, we tested the entire process of isolation and production of human monoclonal antibodies in NPD water as a means for further enhancing human monoclonal antibody isolation and production. Our results indicate an overall enhancement of hybridoma yield, viability, clonability and secretion. Furthermore, we have demonstrated that immortal cells proliferate faster whereas primary human fibroblasts

  3. Human lymphocyte markers defined by antibodies derived from somatic cell hybrids. II. A hybridoma secreting antibody against an antigen expressed by human B and null lymphocytes.

    Science.gov (United States)

    Beckman, I G; Bradley, J; Brooks, D A; Kupa, A; McNamara, P J; Thomas, M E; Zola, H

    1980-06-01

    A hybridoma (FMC4) has been derived which secretes antibody showing selective reaction with human B lymphocytes, monocytes and some null lymphocytes. Few, if any, T lymphocytes in normal blood are stained, although stimulation of lymphocytes with PHA leads to an increase in the proportion of cells reacting with the hybridoma antibody. The antibody reacts with B and null lymphoblastoid cell lines but not with T cell lines. B chronic lymphocytic leukaemia (CLL) cells but not T-CLLs are stained and null-type acute lymphoblastic leukaemia (ALL) cells but not T-type ALL also react. Normal blood myeloid cells do not react with FMC4 supernatant whilst some myeloid leukaemias do. The expression of the antigen reacting with FMC4 supernatant suggests that FMC4 may secrete an antibody against the human equivalent of the Ia antigen.

  4. Development of an indirect ELISA to detect Corynebacterium pseudotuberculosis specific antibodies in sheep employing T1 strain culture supernatant as antigen

    Directory of Open Access Journals (Sweden)

    Miriam F. Rebouças

    2013-11-01

    Full Text Available Corynebacterium pseudotuberculosis is the etiologic agent of caseous lymphadenitis (CLA, a chronic disease that affects goats and sheep, characterized by granuloma formation in subcutaneous and internal lymph nodes. CLA causes significant economic losses to commercial goat herds. In this study, we aimed to test secreted antigens secreted from T1 strain bacteria grown in brain heart infusion (BHI broth in an indirect ELISA system to determine the presence of specific immunoglobulins against C. pseudotuberculosis. We analyzed the BHI antigen electrophoretic profile and the recognition pattern by infected sheep sera samples. The ELISA results were compared with multiplex PCR assay and IFN-gamma production. The ELISA was able to discriminate between negative and positive animals, with a sensitivity of 89% and a specificity of 99%, using microbiological isolation as gold standard. When this assay was compared with multiplex PCR and specific IFN-gamma quantification, six discrepant results were found among thirty-two samples. We concluded that the ELISA using antigens secreted from C. pseudotuberculosis T1 strain growth in BHI broth culture can be used for the serodiagnosis of CLA in sheep.

  5. Regulatory T-Cell (Treg) Hybridoma as a Novel Tool to Study Foxp3 Regulation and Treg Fate1

    Science.gov (United States)

    Sharma, Rahul; Sung, Sun-Sang J.; Ju, Chiao-Ying A.; Umesh, S. Deshmukh; Fu, Shu Man; Ju, Shyr-Te

    2011-01-01

    The CD25+Foxp3+ regulatory T-cells (Treg) that had lost CD25 and Foxp3 in vivo (ex-Treg) exist but are difficult to study. We generated antigen (Ag)-specific Treg hybridomas from iTreg clones (iTreg-hyb) using iTreg of DO11.10.Foxp3-GFP mice and presented evidence that they behave like ex-Treg. The iTreg-hyb displayed little CD25 and Foxp3-GFP but strong expression could be induced with OVA323–339 in the presence of Ag-presenting cells, rIL-2 and rTGF-β1. They displayed all of the iTreg-associated markers examined except CTLA-4, the latter was also absent in the ex-Treg. They lacked the Helios transcription factor, suggesting they were derived from iTreg. Similar to ex-Treg, the iTreg-hyb produced high level of IL-2 and Foxp3 under specific activation conditions. Two unusual properties were observed. First, the ability to induce Foxp3-GFP upon activation is progressively lost in culture over a period of 2 to 4 weeks. Second, Rag2−/− spleen cells alone selectively induced Foxp3-GFP expression albeit 30 times less efficient than Ag-specific activation. We identified cell-free supernatant, IL-6, IL-9, and IL-27 as Foxp3-inducing factors. Our study has significant implications to the stability, plasticity and fate of Treg. The usefulness and limitation of iTreg-hyb as a novel tool to study Foxp3 regulation and the fate of specific Treg subsets are discussed. PMID:21621978

  6. Cybernetic modeling and regulation of metabolic pathways in multiple steady states of hybridoma cells.

    Science.gov (United States)

    Guardia, M J; Gambhir, A; Europa, A F; Ramkrishna, D; Hu, W S

    2000-01-01

    Hybridoma cells utilize a pair of complementary and partially substitutable substrates, glucose and glutamine, for growth. It has been shown that cellular metabolism shifts under different culture conditions. When those cultures at different metabolic states are switched to a continuous mode, they reach different steady states under the same operating conditions. A cybernetic model was constructed to describe the complementary and partial substitutable nature of substrate utilization. The model successfully predicted the metabolic shift and multiple steady-state behavior. The results are consistent with the experimental observation that the history of the culture affects the resulting steady state.

  7. Organization and expression of immunoglobulin genes in fetal liver hybridomas.

    Science.gov (United States)

    Perry, R P; Kelley, D E; Coleclough, C; Kearney, J F

    1981-01-01

    The organization and expression of immunoglobulin genes were studied in a series of six hybridomas derived from the fusion of a nonproducing myeloma cell with cells from mouse fetal liver. These hybridomas, which exhibit several phenotypic characteristics of immature B lymphocytes, all have productively rearranged mu heavy chain genes and produce both the membrane and secreted forms of mu mRNA in a ratio of about 1:10. Significantly, none of the hybridomas has an unrearranged (germ line) allelic mu gene. Examination of the kappa light chain genes revealed that all six of the hybridomas contain unrearranged kappa loci and produce 8.4-kilobase transcripts containing kappa constant region sequences. None of the five hybridomas that exhibit a mu-only phenotype contains a rearranged kappa gene other than that derived from the myeloma parent. One hybridoma, which actively secretes kappa immunoglobulin, contains a rearranged kappa gene of fetal liver origin and synthesizes a distinctive kappa mRNA precursor in addition to the 8.4-kilobase transcript. These results demonstrate that rearrangement of heavy chain immunoglobulin genes normally occurs prior to that of light chain genes and further indicate that the transcriptional competence of the kappa constant region locus is established prior to the time of its rearrangement.

  8. Cell culture supernatants for detection perforin ELISA

    African Journals Online (AJOL)

    Najwa

    2014-02-19

    Feb 19, 2014 ... suggested by Whitaker (1972), the resin was packaged gently in glass column, the .... crystals and incubated in shaker at room temperature for 30 min. ... a well used test for the medical statistics; P value <0.01 is considered.

  9. Metabolic Flux Analysis Model of Hybridoma Cell and Its Application to Batch Culture%杂交瘤细胞代谢流分析模型及其在批式培养中的应用

    Institute of Scientific and Technical Information of China (English)

    仝志明; 曹竹安

    2001-01-01

    An improved mathematical model of metabolic flux analysis bymaterial balances and linear programming method was established, which was first applied to batch culture process in this paper. The analysis results indicate that the metabolic efficiency of substrates such as glucose is very low. It showed that the response of cell metabolism to the step change of glutamine and other amino acid concentration was time-delayed. Metabolic efficiency was higher both in lag phase with highest substrate concentrations and glutamine-limited phase than that in exponential phase. It was also concluded that restraining glutamine from entering energy metabolism could reduce the accumulation of both ammonia and lactate.%通过物料衡算和线性规划方法建立了进行代谢流分析的数学模型,并首次应用于批式培养过程.分析结果表明,细胞对葡萄糖等底物的代谢效率很低,细胞对谷氨酰胺等氨基酸浓度的变化响应迟缓,在底物充裕的迟滞期及谷氨酰胺限制条件下的代谢效率高于对数生长期.另外限制谷氨酰胺等氨基酸进入能量代谢,既可减少氨的积累,也会减少乳酸的生成.

  10. Growth Inhibition of Cronobacter sakazakii in Experimentally Contaminated Powdered Infant Formula by Kefir Supernatant.

    Science.gov (United States)

    Kim, Dong-Hyeon; Chon, Jung-Whan; Kang, Il-Byeong; Kim, Hyunsook; Kim, Hong-Seok; Song, Kwang-Young; Seo, Kun-Ho

    2015-09-01

    Kefir is a type of fermented milk containing lactic and acetic acid bacteria and yeast. In this study, we evaluated the antimicrobial activity of kefir supernatant against Cronobacter sakazakii in powdered infant formula (PIF). In a spot-on-lawn test, the growth of 20 C. sakazakii strains, including 10 clinical and 10 food isolates, was completely inhibited in the presence of kefir supernatant. Significant differences in the diameters of inhibition zones were observed upon treatment with kefir compared with the results for Lactobacillus kefiri and Candida kefyr culture supernatants or solutions of lactic and acetic acid and ethyl alcohol in the agar well diffusion test (P < 0.05). The addition of 100 μl of kefir supernatant to 1 ml of nutrient broth completely inhibited the growth of C. sakazakii, as evaluated by spectrophotometry. The antimicrobial activity of kefir supernatant in experimentally contaminated PIF was also tested; we found no viable C. sakazakii cells remaining in PIF rehydrated with 30% kefir supernatant solution for 1 h, demonstrating that the antimicrobial activity of kefir supernatant against C. sakazakii could be applied in real food samples.

  11. Use of Taguchi's methods as a basis to optimize hybridoma cell line growth and antibody production in a spinner flask

    OpenAIRE

    Kallel, Héla; Zaïri, Hind; Rourou, Samia; Essafi, Makram; Barbouche, Ridha; Dellagi, Koussay; Fathallah, Dahmani M.

    2002-01-01

    Taguchi’s methods were used for the design of an experimental strategy aimed at optimizing cell density and monoclonal antibody (mAb) production from a spinner flask hybridoma culture. 23G11 is an antibody to the human leukocyte adhesion molecule, CR3 or β 2 integrin (CD11b/CD18). It recognizes specifically the A-domain of the α subunit CD11b. Anti β 2 integrin monoclonal antibodies hold a great potential for preventing inflammation mediated tissue injuries. An L8 orthogonal experimental desi...

  12. Use of Taguchi's methods as a basis to optimize hybridoma cell line growth and antibody production in a spinner flask.

    Science.gov (United States)

    Kallel, Héla; Zaïri, Hind; Rourou, Samia; Essafi, Makram; Barbouche, Ridha; Dellagi, Koussay; Fathallah, Dahmani M

    2002-05-01

    Taguchi's methods were used for the design of an experimental strategy aimed at optimizing cell density and monoclonal antibody (mAb) production from a spinner flask hybridoma culture. 23G11 is an antibody to the human leukocyte adhesion molecule, CR3 or beta 2 integrin (CD11b/CD18). It recognizes specifically the A-domain of the alpha subunit CD11b. Anti beta 2 integrin monoclonal antibodies hold a great potential for preventing inflammation mediated tissue injuries. An L8 orthogonal experimental design was used to investigate four different culture components: stirring speed, nature of serum, concentration of serum and nature of media (RPMI 1640 or RPMI 1640 supplemented with glucose and glutamine). The experiments were conducted using two levels for each factor studied and a direct ELISA test was used to estimate the level of antibody production. Statistical analysis of the collected data pointed to the stirring speed and serum concentration, and the interaction between these parameters, as the components that affected cell growth. Antibody production was affected by these factors and by the nature of medium but also by the following two interactions: stirring speed/nature of serum and stirring speed/concentration of serum. This study emphasizes the value of using Taguchi's methods as a basis for optimization of mAb production from a hybridoma culture, in cost effective and significantly less labor intensive ways.

  13. Effects of LPS-stimulated monocyte culture supernatant on the osteogenic function of osteoblastic cells in vitro%脂多糖刺激单核巨噬细胞培养上清液对成骨细胞成骨特性的影响

    Institute of Scientific and Technical Information of China (English)

    王海燕; 邓辉; 徐春燕; 叶洁; 胡荣党

    2011-01-01

    目的 探讨建立牙周炎症的体外实验模型的可行性,为后续进行牙周炎症状态下应力对牙槽骨改建的研究奠定基础.方法 以牙龈卟啉单胞菌(Porphyromonas gingivalis,P.g)脂多糖(lipopolysaccharide,LPS)Pg-LPS刺激小鼠单核巨噬细胞(RAW264.7)的培养上清液作用于体外培养的小鼠成骨细胞(MC3T3-E1)24 h和48 h后,分别通过噻唑蓝(MTT)法和碱性磷酸酶(ALP)试剂盒检测MC3T3-E1的增殖活性和碱性磷酸酶活性,观察脂多糖刺激的小鼠单核巨噬细胞培养上清液对小鼠成骨细胞成骨功能的影响.结果 Pg-LPS刺激RAW264.7后其上清液含L-1β、IL-6 和TNF-α等炎症因子,且与LPS的浓度和刺激时间呈依赖性.此上清液对MC3T3-E1的增殖和ALP活性均有抑制作用,亦与上清液中以上炎症因子的浓度呈依赖性,与体内牙周炎症状态相似.结论 Pg-LPS体外刺激RAW264.7的培养上清液可以应用于建立体外牙周炎症模型,这将为后续研究牙周炎症状态下应力对牙槽骨改建的影响奠定初步的基础.%Objective To investigate the influence of Pg-LPS stimulated monocyte (RAW264.7) culture supernatant on the proliferation and alkaline phosphatase (ALP) activity of osteoblastic cells (MC3T3-E1) in vitro. Methods The culture supernatant of monocytes stimulated with Pg-LPS was applied to osteoblasts MC3T3-E1 with different diluted concentrations(10%、20%、30%、40% and 50%) simultaneously for 24 h and 48 h in vitro.The cellular proliferation was assessed by MTT assay, Cellular ALP activity was examined using ALP measurement kit, and the results were statistically analyzed using SPSS 12.0 software package. Results LPS stimulated RAW264.7 to release inflammatory cytokines including IL-1β, IL-6 and TNF-α, whose secretion was in a time and dose dependent manner with the concentration of LPS. After the stimulation of different concentrations of inflammatory supernatant, the proliferation and ALP activity of MC3T3-El

  14. The Immunostimulation of F4ac ETEC-Culture Supernatant to Porcine Small Intestinal Epithelial Cells%F4ac型产肠毒素大肠杆菌菌液上清对仔猪小肠上皮细胞的免疫刺激作用

    Institute of Scientific and Technical Information of China (English)

    周传丽; 刘铮铸; 俞英; 张勤

    2014-01-01

    [Objective]Porcine enterotoxigenic Escherichia coli (ETEC) is a worldwide cause of bacteria induced diarrhoea in piglets. In the veterinary practices of pig production, serological identification of ETEC shows that F4ac is the most common serological type expressed in ETEC strains isolated from diarrheic piglets. So far, the mechanism by which ETEC produces diarrhoea in piglets has been clearly elucidated. However, the immunostimulation of ETEC-culture to host target cells has not been studied or described. In the present study, the immunostimulation of the supernatant of F4ac ETEC-culture to IPEC-J2 cells was studied.[Method]The culture of F4ac ETEC strain 200 was collected and centrifuged (4℃, 4 000 r/min for 15 min) after 12 hours in culture, and the supernatant was sterilized by passing it through a 0.22μm filter. The solution combined the sterilized supernatant with equivalent DMEM/F12 medium was used to challenge IPEC-J2 cells for 3 hours. The IPEC-J2 cells co-cultured with fresh LB medium and equivalent DMEM/F12 medium were as the control group. For both treatments, each experiment was repeated three times. Total RNAs of the stimulated and control IPEC-J2 cells were extracted using TRIZOL Reagent following the manufacturer’s instructions. According to the manufacturer’s instructions, complementary DNA (cDNA) was synthesized from RNA using the Prime Script® RT reagent Kit with gDNA Eraser (Perfect Real Time). The cDNA samples were then analyzed with real time RT-PCR using a LightCycler® 480 Real-Time PCR System. The real time RT-PCR reactions were performed in a final volume of 20μL with the Roche SYBR Green PCR Kit according to the manufacturer’s instructions. The pig house-keeping gene β-actin was used as the internal standards to correct the input of cDNA. Triplicate qRT-PCRs were performed on each cDNA and the average Ct was used for further analysis. The relative quantification values were calculated using the 2-ΔΔCt. Differential m

  15. Growth kinetics of hybridoma cells: (1) The effects of varying foetal calf serum levels.

    Science.gov (United States)

    Low, K; Harbour, C

    1985-01-01

    The growth kinetics, i.e. growth rate, cell yield and antibody production, of two murine hybridoma cell lines have been studied in several commercial media at different FCS levels in static 25 cm2 flask cultures. Reduction of FCS levels from 10% to 5% did not affect significantly the antibody yield whereas at 2% and 1% FCS levels growth rate, cell and antibody yield were reduced significantly in all media. Considerable differences were noted in the maximum cell populations obtained in the different media, with IMDM producing the highest cell and antibody yields; IMDM greater than HiGem greater than DME greater than RPMI. The cell lines did not grow in the absence of FCS but did grow in the presence of basal medium supplemented with insulin and transferrin at 10 mg per L. Both cell lines were stable during several months' passage in this medium. A supplement containing human albumin or BSA at 1 g per L combined with the insulin and transferrin (10 mg per L) could replace 1% FCS in DME without significantly affecting the cell yields of B6.

  16. Effects of the supernatant from ankylosing spondylitis synovial cells culture on the osteogenic differentiation of ligament fibroblasts%强直性脊柱炎滑膜细胞培养上清液对韧带成纤维细胞成骨分化的影响

    Institute of Scientific and Technical Information of China (English)

    徐卫东; 梁志民; 袁恒锋

    2012-01-01

    Objective To investigate the effects of synovial cells culture fluid on the differentiation of ligament fibroblasts into osteoblasts. Methods Specimens of ligament and synovium were obtained from the hip joints with ankylosing spondylitis respectively. The primary cell culture of ligament fibroblasts and synovial cells were processed and then the supernatant of synovial cells in the first passage was collected and used to induce ligament cells. Osteogenic phenotype and alkaline phosphatase (ALP) activity were determined at the days of 0, 5,10,15 and 20 respectively. Calcium deposit was detected by Von Kossa staining at the 25th day. Results With time extending, the expression of ALP and osteocalcin increased significantly at day 10 and reached a peak at day 15. Calcium deposit could be detected at day 25. Conclusions The supernatant of synovial cells cuture plays a certain role on the differentiation of ligament fibroblasts into osteoblasts.%目的 探讨强直性脊柱炎(AS)滑膜细胞培养上清液对韧带成纤维细胞成骨分化的影响.方法 分别进行AS患者滑膜和韧带成纤维细胞原代培养,收集第1代的滑膜细胞培养液上清液,应用此上清液对韧带成纤维细胞进行诱导,分别于0、5、10、15、20 d检测碱性磷酸酶和骨钙素表达水平,并于25 d时进行钙结节染色.结果 在应用AS滑膜细胞培养上清液对韧带成纤维细胞进行诱导的第10天,碱性磷酸酶(ALP)和骨钙素(OC)表达水平明显升高,并在第15天达到高峰.第25天时进行钙结节Von Kossa染色,发现有钙结节形成.结论 AS滑膜细胞培养上清液对韧带成纤维细胞具有成骨诱导分化的能力.

  17. Data of rational process optimization for the production of a full IgG and its Fab fragment from hybridoma cells

    OpenAIRE

    Martina Röhm; Alina Handl; Maria König; Chrystelle Mavoungou; René Handrick; Katharina Schindowski

    2016-01-01

    This data article focuses on the production of monoclonal antibodies (mAb) and their fragments Fab and F(ab′)2. Here, we present the data of an optimization protocol to improve the product yield of a hybridoma cell process using a Design of Experiment (DoE) strategy. Furthermore, the data of the evaluated conditions were used to test feeding strategies in shake flasks. They were verified in controlled 2 L fed-batch bioreactor processes. Supplementing the culture medium with human insulin-like...

  18. MHC CLASS-II-RESTRICTED T-CELL HYBRIDOMAS RECOGNIZING THE NUCLEOCAPSID PROTEIN OF AVIAN CORONAVIUS IBV

    NARCIS (Netherlands)

    BOOTS, AMH; VANLIEROP, MJ; KUSTERS, JG; VANKOOTEN, PJS; VANDERZELIST, BAM; HENSEN, EJ; Boots, Annemieke

    1991-01-01

    Mice were immunized with purified infectious bronchitis virus (IBV), strain M41. Spleen cells, expanded in vitro by stimulation with M41, were immortalized by fusion to obtain T-cell hybridomas, and two major histocompatability complex (MHC) class II (I-E)-restricted T-cell hybridomas were selected

  19. Detection of interleukin-10 and transforming growth factor-β1 in the culture supernatant of CD4+CD25+ T cells from patients with alopecia areata%斑秃患者外周血CD4+CD25+T细胞培养上清液白介素10和转化生长因子β1检测

    Institute of Scientific and Technical Information of China (English)

    马新华; 邵文俊; 金宛宛; 高宇

    2014-01-01

    Objective To evaluate the potential association of CD4+CD25+ T cells with alopecia areata.Methods Totally,this study enrolled 23 patients with progressive alopecia areata,25 patients with stable alopecia areata,and 25 healthy controls.Peripheral blood was isolated from these subjects followed by isolation of CD4+ CD25+ regulatory T cells,which were then cuhured with the presence of anti-CD3 and-CD28 monoclonal antibodies for four days.Subsequently,enzyme-linked immunosorbent assay was performed to measure the levels of interleukin (IL)-10 and transforming growth factor (TGF)-β1 in the culture supematant of these T cells.Results The levels of IL-10 and TGF-β1 were (31.68 ± 6.78) pg/ml and (32.29 ± 6.8) pg/ml respectively in the culture supernatant of CD4+CD25+ regulatory T cells from patients with progressive alopecia areata,significantly lower than those from the healthy controls ((57.34 ± 14.15) pg/ml and (57.43 ± 15.16) pg/ml,both P < 0.05) and patients with stable alopecia areata ((52.56 ± 13.02) pg/ml and (61.75 ± 14.10) pg/ml,both P < 0.05).However,no significant difference was observed in the supernatant levels of IL-10 or TGF-β1 between the healthy controls and patients with stable alopecia areata.Conclusions The secretion of IL-10 and TGF-β1 by CD4+CD25+ T cells is decreased in patients with progressive alopecia areata,which may contribute to the pathogenesis of alopecia areata.%目的 探讨CD4+CD25+T细胞与斑秃发病之间的关系.方法 收集了3组研究对象,其中健康对照组25例、稳定期斑秃患者25例、进展期斑秃患者23例.抽取所有对象外周血,提取CD4+CD25+T细胞,培养4d,收集培养上清液,ELISA法检测上清液IL-10和TGF-β1水平.结果 进展期斑秃患者外周血CD4+CD25+T细胞培养的IL-10和TGF-β1分别为(31.68±6.78) pg/ml和(32.29±6.80) pg/ml,明显低于健康对照组(57.34±14.15) pg/ml、(57.43±15.16) pg/ml和稳定期斑秃患者(52.56±13.02) pg/ml和(61.75±14.10) pg

  20. Modelling antagonic effect of lactic acid eacteria supernatants on some pathogenic bacteria

    OpenAIRE

    2009-01-01

    This work presents a statistical model of survival analysis for three pathogenic bacterial strains (Escherichia coli, Listeria monocytogenes and Staphylococcus aureus), when treated with neutralized and non-neutralized filtered supernatants broth from cultures of Lactobacillus acidhophilus, Lactobacillus rhamnosus and Lactobacillus sake. Survival analysis is a method employed to determine the period of time from an initial stage up to the occurrence of a particular event of interest, as death...

  1. On chip electrofusion of single human B cells and mouse myeloma cells for efficient hybridoma generation

    NARCIS (Netherlands)

    Kemna, Evelien; Wolbers, F.; van den Berg, Albert; Vermes, I.

    2011-01-01

    This article describes the development and full characterization of a microfluidic chip for electrofusion of human peripheral blood B-cells and mouse myeloma (NS-1) cells to generate hybridomas. The chip consists of an array of 783 traps, with dimensions that were optimized to obtain a final cell

  2. 乳酸菌发酵液中DL-3-苯乳酸高效液相色谱快速检测方法的建立%Development of HPLC Method to Assay DL-3-Phenyllactic Acid in Culture Supernatant from Lactic Acid Bacteria

    Institute of Scientific and Technical Information of China (English)

    张忠华; 李晓然; 李洪梅; 李海舟; 龚福明; 柳陈坚

    2013-01-01

    DL-3-Phenyllactic acid (PLA) is a novel organic acid produced by some lactic acid bacteria,which can inhibit the growth of a wide variety of food-borne pathogens.On the other hand,PLA is safe and does not affect the odor of food at low concentrations.However,its further application was limited by its high equipment cost and complicated process so that the screening and identifying the high-PLA-producing LAB strains is restricted.A rapid method is developed in this paper for the determination of DL-3-Phenyllactic acid in MRS culture supernatant from lactic acid bacteria by high performance liquid chromatography.The determination is carried out on Agilent Zorbax SB-C18 (4.6 mm × 150 mm,5 μm).0.5% H3PO4 (V/V) and 0.5%H3PO4-CH3CN solution (V/V) is used as mobile phase at a flow rate of 1mL/min.Detection wavelength is set at 210 nm.The calibration curve is linear when DL-3-phenyllactic acid concentration is in the range of 60 ~800 mg/L.The corresponding regression equation is y =14.723x + 0.132 5,r =0.999 9.The method is then used to recover MRS fermented supernatant at 50 ~ 150 mg/L.The effectiveness is proven by the results as the recoveries are all above 94.0% and the relative standard deviation of samples is 0.58%.%DL-3苯乳酸是近年来发现可以由部分乳酸菌分泌产生的一种新型有机酸,它能够有效抑制多种食源性致病菌,并且具有安全、无毒副作用、低浓度添加时不影响食品风味等优点.然而现有苯乳酸检测方法存在所需检测设备投入成本高、操作步骤繁琐等缺陷,从而严重限制高产苯乳酸菌株的筛选和利用.本研究建立乳酸菌发酵液中苯乳酸高效液相色谱法快速检测方法,采用Agilent Zorbax SB-C18(4.6 mm×150 mm,5μm)色谱柱;流动相A:0.5%磷酸水溶液(V/V),B:0.5%磷酸乙腈溶液(V/V).流速1 mL/min,检测波长210 nm.DL-3-苯乳酸浓度在60~800 mg/L之间线性回归良好(其方程为y=14.723x +0.132 5,r=0.9999).应用本

  3. Characterization of hybridoma cell responses to elevated pCO(2) and osmolality: intracellular pH, cell size, apoptosis, and metabolism.

    Science.gov (United States)

    deZengotita, Vivian M; Schmelzer, Albert E; Miller, William M

    2002-02-15

    CO(2) partial pressure (pCO(2)) in industrial cell culture reactors may reach 150-200 mm Hg, which can significantly inhibit cell growth and recombinant protein production. The inhibitory effects of elevated pCO(2) at constant pH are due to a combination of the increases in pCO(2) and [HCO(-) (3)], per se, and the associated increase in osmolality. To decouple the effects of pCO(2) and osmolality, low-salt basal media have been used to compensate for this associated increase in osmolality. Under control conditions (40 mm Hg-320 mOsm/kg), hybridoma cell growth and metabolism was similar in DMEM:F12 with 2% fetal bovine serum and serum-free HB GRO. In both media, pCO(2) and osmolality made dose-dependent contributions to the inhibition of hybridoma cell growth and synergized to more extensively inhibit growth when combined. Elevated osmolality was associated with increased apoptosis. In contrast, elevated pCO(2) did not increase apoptotic cell death. Specific antibody production also increased with osmolality although not with pCO(2). In an effort to understand the mechanisms through which elevated pCO(2) and osmolality affect hybridoma cells, glucose metabolism, glutamine metabolism, intracellular pH (pHi), and cell size were monitored in batch cultures. Elevated pCO(2) (with or without osmolality compensation) inhibited glycolysis in a dose-dependent fashion in both media. Osmolality had little effect on glycolysis. On the other hand, elevated pCO(2) alone had no effect on glutamine metabolism, whereas elevated osmolality increased glutamine uptake. Hybridoma mean pHi was approximately 0.2 pH units lower than control at 140 mm Hg pCO(2) (with or without osmolality compensation) but further increases in pCO(2) did not further decrease pHi. Osmolality had little effect on pHi. Cell size was smaller than control at elevated pCO(2) at 320 mOsm/kg, and greater than control in hyperosmotic conditions at 40 mm Hg.

  4. Cell vacuolation induced by Haemophilus influenzae supernatants in HEp-2 cells

    Directory of Open Access Journals (Sweden)

    Maria del Rosario Espinoza-Mellado

    2013-12-01

    Full Text Available Haemophilus influenzae belongs to respiratory tract microbiota. We observed vacuoles formation in previous studies with H. influenzae culture supernatants, so in this work we characterised that cytotoxic effect. We observed an abundant production of acidic cytoplasmic vacuoles due to the presence of a “vacuolating factor” in H. influenzae supernatants which was characterised as thermolabile. Greatest vacuolating activity was observed when utilizing the fraction > 50 kDa. The presence of a large number of vacuoles in HEp-2 cells was verified by transmission electron microscopy and some vacuoles were identified with a double membrane and/or being surrounded by ribosomes. These results suggest similar behaviour to that of vacuolating effects described by autotransporter proteins an undescribed cytotoxic effect induced by H. influenzae .

  5. Screening a hybridoma producing a specific monoclonal antibody to HLA-A24+Bw4 antigen by cytotoxicity inhibition assay.

    Science.gov (United States)

    Hiroishi, S; Kaneko, T; Arita, J

    1987-02-01

    A hybridoma secreting a monoclonal antibody (Tsa-1, IgG3) reacting specifically to HLA-A24+Bw4 was screened by cytotoxicity inhibition assay and micrototoxicity test. The R value of the antibody was 0.843.

  6. Development of a bispecific monoclonal antibody to pesticide carbofuran and triazophos using hybrid hybridomas.

    Science.gov (United States)

    Jin, R Y; Guo, Y R; Wang, C M; Wu, J X; Zhu, G N

    2009-01-01

    A mouse hybrid hybridoma (tetradoma) was derived from fusing hybridomas producing monoclonal antibody to N-methylcarbamate pesticide carbofuran with hybridomas producing monoclonal antibody to organophosphorus pesticide Triazophos. The prepared tetradoma line (12C1 to 2H12) secreted hybrid immunoglobulin exhibiting parental and bispecific binding characteristics. The effect of relevant physicochemical factors on the immunoassay based on the 12C1 to 2H12 bispecific monoclonal antibody had been studied to optimize the ELISA performance. The developed immunoassay showed that the detection limit (I(20)) were 0.36 and 1.89 ng/mL for triazophos and carbofuran, respectively, without obvious cross-reactivity to other related compounds. Water samples spiked with triazophos at 0.5, 1, and 5 ng/mL or carbofuran at 5, 10, and 20 ng/mL were directly analyzed by the developed ELISA format. The mean recovery of triazophos and carbofuran were 108.1% and 107.5%, with variation coefficient of 15.9% and 17.7%, respectively.

  7. Surface Enhanced Raman Spectroscopy detection of p-coumaric acid from cell supernatant using gold-capped silicon nanopillar substrates

    DEFF Research Database (Denmark)

    Morelli, Lidia; Jendresen, Christian Bille; Burger, Robert;

    HCA spiked in culture medium, in the same concentration range (10-4 – 10-5 M) commonly found in cell supernatant. For supernatant analysis, triplicate cultures of FjTAL modified (P strains) and control (C strains) E.coli strains were carried out according to the methods described by[5] and shown in Fig.1....... Samples of cell supernatant were extracted from each culture at 0, 3, 24 and 48 h post seeding and their pHCA content was measured with HPLC[5]. For SERS analysis, aliquots of supernatant were diluted 10-fold with MilliQ water, and 1 μL droplets were dried on the SERS substrates. A MatLab analysis......A standard protocol for analysis of microbial factories requires the screening of several populations in order to find the best performing ones. This is done with standard analytical methods (e.g. HPLC) with an expensive and time-consuming process. Surface Enhanced Raman Spectroscopy (SERS...

  8. Detection of p-coumaric acid from cell supernatant using surfaceenhanced Raman spectroscopy

    DEFF Research Database (Denmark)

    Morelli, Lidia; Jendresen, Christian Bille; Zor, Kinga

    2016-01-01

    A standard protocol for analysis of microbial factories requires the screening of several populations in order to find the bestperforming ones. Standard analytical methods usually include high performance liquid chromatography (HPLC), thin layerchromatography (TLC) or spectrophotometry, which...... on detection ofp-coumaric acid (pHCA) in cell supernatant.SERS active substrates, based on leaning gold-capped silicon nanopillars, were used for detection. They were successfullyused to detect culture medium spiked with pHCA, and the effect of medium dilution was studied. For analysis of biologicalproduction...

  9. 人羊膜间充质干细胞培养上清液对人成纤维细胞生物学功能的影响%Effects of culture supernatant of human amnion mesenchymal stem cells on biological characteristics of human fibroblasts

    Institute of Scientific and Technical Information of China (English)

    吴骐而; 吕璐; 辛海明; 罗亮; 童亚林; 莫永亮; 岳毅刚

    2016-01-01

    Objective To investigate the effects of culture supernatant of human amnion mesenchymal stem cells (hAMSCs-CS) on biological characteristics of human fibroblasts.Methods (1) hAMSCs were isolated from deprecated human fresh amnion tissue of placenta and then sub-cultured.The morphology of hAMSCs on culture day 3 and hAMSCs of the third passage were observed with inverted phase contrast microscope.(2) Two batches of hAMSCs of the third passage were obtained,then the expression of vimentin of cells was observed with immunofluorescence method,and the expression of cell surface marker CD90,CD73,CD105,and CD45 was detected by flow cytometer.(3) hAMSCs-CS of the third passage at culture hour 72 were collected,and the content of insulin-like growth factor Ⅰ (IGF-Ⅰ),vascular endothelial growth factor (VEGF),epidermal growth factor (EGF),and basic fibroblast growth factor (bFGF) were detected by enzyme-linked immunosorbent assay.(4) Hunan fibroblasts were isolated from deprecated human fresh prepuce tissue of circumcision and then sub-cultured.Human fibroblasts of the third passage were used in the following experiments.Cells were divided into blank control group and 10%,30%,50%,and 70% hAMSCs-CS groups according to the random number table (the same grouping method below),with 48 wells in each group.Cells in blank control group were cultured with DMEM/F12 medium containing 2% fetal bovine serum (FBS),while cells in the latter 4 groups were cultured with DMEM/F12 medium containing corresponding volume fraction of hAMSCs-CS and 2% FBS.The proliferation activity of cells was detected by cell counting kit 8 and microplate reader at culture hour 12,24,48,and 72,respectively,and corresponding volume fraction of hAMSCs-CS which causing the best proliferation activity of human fibroblasts was used in the following experiments.(5) Human fibroblasts were divided into blank control group and 50% hAMSCs-CS group and treated as in (4),with 4 wells in each group,at post

  10. Data of rational process optimization for the production of a full IgG and its Fab fragment from hybridoma cells

    Directory of Open Access Journals (Sweden)

    Martina Röhm

    2016-09-01

    Full Text Available This data article focuses on the production of monoclonal antibodies (mAb and their fragments Fab and F(ab′2. Here, we present the data of an optimization protocol to improve the product yield of a hybridoma cell process using a Design of Experiment (DoE strategy. Furthermore, the data of the evaluated conditions were used to test feeding strategies in shake flasks. They were verified in controlled 2 L fed-batch bioreactor processes. Supplementing the culture medium with human insulin-like growth factor-I (IGF-I and Pluronic F-68, as well as a nutrient rich additive for fed-batch, resulted in improved cell growth correlating with a 7 day elongated process time and a 4.5 fold higher product titer. Finally, a rapid Fab generation protocol and the respective data are presented using different papain digestion and a camelid anti-kappa light chain VHH affinity ligand.

  11. Data of rational process optimization for the production of a full IgG and its Fab fragment from hybridoma cells.

    Science.gov (United States)

    Röhm, Martina; Handl, Alina; König, Maria; Mavoungou, Chrystelle; Handrick, René; Schindowski, Katharina

    2016-09-01

    This data article focuses on the production of monoclonal antibodies (mAb) and their fragments Fab and F(ab')2. Here, we present the data of an optimization protocol to improve the product yield of a hybridoma cell process using a Design of Experiment (DoE) strategy. Furthermore, the data of the evaluated conditions were used to test feeding strategies in shake flasks. They were verified in controlled 2 L fed-batch bioreactor processes. Supplementing the culture medium with human insulin-like growth factor-I (IGF-I) and Pluronic F-68, as well as a nutrient rich additive for fed-batch, resulted in improved cell growth correlating with a 7 day elongated process time and a 4.5 fold higher product titer. Finally, a rapid Fab generation protocol and the respective data are presented using different papain digestion and a camelid anti-kappa light chain VHH affinity ligand.

  12. Photodynamic response of an endothelial hybridoma cell line using zinc(II) tetrasubstituted phthalocyanines

    Science.gov (United States)

    Cruse-Sawyer, Janet E.; Dixon, B.; Roberts, David J.; Griffiths, John; Brown, Stanley B.

    1995-03-01

    The EAhy 926 cell is a hybridoma line derived from human endothelium and A549/8 cells. They display stable endothelial characteristics and may provide an indication of how endothelial cells respond to photodynamic therapy. Cells were grown as monolayers, seeded at a density of 104 cells/35 mm dish, and then incubated with zinc (II) tetrasubstituted phthalocyanines (carboxylated, sulphonated, pyridinium or diethanolamine sulphonamide). After 24 hours, the cells were illuminated with laser light at 680 nm or 692 nm as appropriate. The response to each photosensitizer was evaluated using cell proliferation, clonogenicity, and release of tissue factor.

  13. A luminescent hybridoma-based biosensor for rapid detection of V. cholerae upon induction of calcium signaling pathway.

    Science.gov (United States)

    Zamani, Parichehr; Sajedi, Reza H; Hosseinkhani, Saman; Zeinoddini, Mehdi; Bakhshi, Bita

    2016-05-15

    In this study, a hybridoma based biosensor was developed for rapid, sensitive and selective detection of Vibrio cholerae O1 which converts the antibody-antigen binding to bioluminescence light. After investigation on hybridoma performance, the biosensor was constructed by transfecting specific hybridoma cells with aequorin reporter gene and the bioluminescence activities of stable biosensor were measured. The sensitivity of biosensor was as few as 50 CFU/ml and it showed no responses to other entric bacteria. Moreover, the response time of biosensor was estimated in 7th second which means this method is considerably faster than many available detection assays. In addition, this biosensor was successfully applied to V. cholerae detection in environmental samples with no significant loss in sensitivity, demonstrating our proposed biosensor provides a sensitive and reliable method for detection of V. cholerae in natural samples. The application of whole hybridoma cell directly as a sensing element in biosensor construction which mentioned for the first time in present study suggests that hybridoma cells could provide a valuable tool for future studies in both basic and diagnostic sciences and could be considered as a fast and specific sensing element for detection of other pathogens in different applications. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Mimicking the germinal center reaction in hybridoma cells to isolate temperature-selective anti-PEG antibodies.

    Science.gov (United States)

    Su, Yu-Cheng; Al-Qaisi, Talal S; Tung, Hsin-Yi; Cheng, Tian-Lu; Chuang, Kuo-Hsiang; Chen, Bing-Mae; Roffler, Steve R

    2014-01-01

    Modification of antibody class and binding properties typically requires cloning of antibody genes, antibody library construction, phage or yeast display and recombinant antibody expression. Here, we describe an alternative "cloning-free" approach to generate antibodies with altered antigen-binding and heavy chain isotype by mimicking the germinal center reaction in antibody-secreting hybridoma cells. This was accomplished by lentiviral transduction and controllable expression of activation-induced cytidine deaminase (AID) to generate somatic hypermutation and class switch recombination in antibody genes coupled with high-throughput fluorescence-activated cell sorting (FACS) of hybridoma cells to detect altered antibody binding properties. Starting from a single established hybridoma clone, we isolated mutated antibodies that bind to a low-temperature structure of polyethylene glycol (PEG), a polymer widely used in nanotechnology, biotechnology and pharmaceuticals. FACS of AID-infected hybridoma cells also facilitated rapid identification of class switched variants of monoclonal IgM to monoclonal IgG. Mimicking the germinal center reaction in hybridoma cells may offer a general method to identify and isolate antibodies with altered binding properties and class-switched heavy chains without the need to carry out DNA library construction, antibody engineering and recombinant protein expression.

  15. The use of syngeneic cells producing human somatotropic hormone (hSTH) for immunization of mice and development of hybridomas.

    Science.gov (United States)

    Panyutich, A V; Prassolov, V S; Shydlovskaya, E A; Reznikov, M V; Chumakov, P M; Voitenok, N N

    1990-08-01

    We studied the possibility of syngeneic cells expressing heterologous protein being used for sensitization of mice and production of hybridomas. Recombinant retroviral vector containing cloned human somatotropic hormone (hSTH) gene was used to express hSTH in BALB/3T3 cells. BALB/c mice were injected intrasplenically (i/s) or combination of intraperitoneally (i/p) and intrasplenically with hSTH-producing cells. Sensitized splenocytes were fused with myeloma cell P3X63-AgB.653. Screening for anti-hSTH hybridomas was performed by enzyme-linked immunoassay. Both single i/s injection of producer cells as well as combined i/s and i/p injections were effective for sensitization of splenocytes. Combined injection was effective for production of IgG and IgM secreting hybridomas. Single i/s injection led to generation of only IgM producing hybridomas. The results proved that syngeneic cells expressing genes of heterologous proteins can be used for splenocyte sensitization and hybridoma preparation.

  16. A human-mouse hybridoma producing monoclonal antibody against human sperm coating antigen.

    Science.gov (United States)

    Kyurkchiev, S D; Shigeta, M; Koyama, K; Isojima, S

    1986-01-01

    Since anti-sperm antibodies were first discovered in the sera of women, the relationship of these antibodies to sterility has been studied by many investigators. In order to determine the antigens of spermatozoa responsible for raising antibodies to spermatozoa in humans, many studies have been carried out by purifying human spermatozoa cell membrane and seminal plasma components. Since it was found that the purification was difficult by physiochemical procedures, the immunoaffinity chromatography bound monoclonal antibody (Mab) to spermatozoa antigens was attempted for this purpose. The establishment of hybridomas producing Mabs to human seminal plasma and human spermatozoa was reported by Shigeta et al. (1980), Isojima, Koyoma & Fujiwara (1982), Lee et al. (1982) and Isahakia & Alexander (1984). The ordinary approaches to obtain the Mabs consisted of xenogenic immunization with human semen and cell fusion of immunized spleen cells with mouse myeloma cells. However, the antigenic epitopes of human spermatozoa, which induced antibody production, are xenogenic for the mouse, and therefore there is a possibility that there is a difference in recognized antigenic epitopes in humans as isotypic and in mice as xenogenic. In order to study these antigenic epitopes, which correspond to antibodies against spermatozoa in women, the establishment of human-mouse hybridomas, which produced anti-semen antibodies as produced in sterile women, became essential. In these studies, we used recently developed cell fusion techniques to fuse immunized human peripheral lymphocytes with mouse myeloma cells. PMID:3456978

  17. Acoustic perfusion processes for hybridoma cultures: viability, cell cycle and metabolic analysis

    NARCIS (Netherlands)

    Dalm, M.C.F.

    2007-01-01

    For the production of glycosylated proteins, such as monoclonal antibodies, hormones, and blood clothing factors, generally mammalian cells are used. Mammalian cells are preferred over other expression systems, such as bacteria or yeast, because they are able to glycosylate proteins in a human-like

  18. Acoustic perfusion processes for hybridoma cultures: viability, cell cycle and metabolic analysis

    NARCIS (Netherlands)

    Dalm, M.C.F.

    2007-01-01

    For the production of glycosylated proteins, such as monoclonal antibodies, hormones, and blood clothing factors, generally mammalian cells are used. Mammalian cells are preferred over other expression systems, such as bacteria or yeast, because they are able to glycosylate proteins in a human-like

  19. Hybridoma cell-culture and glycan profile dataset at various bioreactor conditions.

    Science.gov (United States)

    Bhatia, Hemlata; Read, Erik; Agarabi, Cyrus; Brorson, Kurt; Lute, Scott; Yoon, Seongkyu

    2016-12-01

    This is an "11 factor-2 level-12 run" Plackett-Burman experimental design dataset. The dataset includes 11 engineering bioreactor parameters as input variables. These 11 factors were varied at 2 levels and 23 response variables that are glycan profile attributes, were measured "A Design Space Exploration for Control of Critical Quality Attributes of mAb" (H. Bhatia, E.K. Read, C.D. Agarabi, K.A. Brorson, S.C. Lute, S. Yoon S, 2016) [2].

  20. Hybridoma cell-culture and glycan profile dataset at various bioreactor conditions

    Directory of Open Access Journals (Sweden)

    Hemlata Bhatia

    2016-12-01

    Full Text Available This is an “11 factor-2 level-12 run” Plackett-Burman experimental design dataset. The dataset includes 11 engineering bioreactor parameters as input variables. These 11 factors were varied at 2 levels and 23 response variables that are glycan profile attributes, were measured “A Design Space Exploration for Control of Critical Quality Attributes of mAb” (H. Bhatia, E.K. Read, C.D. Agarabi, K.A. Brorson, S.C. Lute, S. Yoon S, 2016 [2].

  1. Radically altered T cell receptor signaling in glycopeptide-specific T cell hybridoma induced by antigen with minimal differences in the glycan group

    DEFF Research Database (Denmark)

    Jensen, T; Nielsen, M; Gad, Monika;

    2001-01-01

    A T cell hybridoma raised against the synthetic glycopeptide T(72)(Tn) was used to study whether the initial TCR signaling events are markedly different when the hybridoma is stimulated with glycopeptides closely related to the cognate glycopeptide antigen. T(72)(Tn) has an alpha-D-GalNAc group O...

  2. Immunoproteomic analysis of antibody in lymphocyte supernatant in patients with typhoid fever in Bangladesh.

    Science.gov (United States)

    Charles, Richelle C; Liang, Li; Khanam, Farhana; Sayeed, M Abu; Hung, Chris; Leung, Daniel T; Baker, Stephen; Ludwig, Albrecht; Harris, Jason B; Larocque, Regina C; Calderwood, Stephen B; Qadri, Firdausi; Felgner, Philip L; Ryan, Edward T

    2014-03-01

    We have previously shown that an assay based on detection of anti-Salmonella enterica serotype Typhi antibodies in supernatant of lymphocytes harvested from patients presenting with typhoid fever (antibody in lymphocyte supernatant [ALS] assay) can identify 100% of patients with blood culture-confirmed typhoid fever in Bangladesh. In order to define immunodominant proteins within the S. Typhi membrane preparation used as antigen in these prior studies and to identify potential biomarkers unique to S. Typhi bacteremic patients, we probed microarrays containing 2,724 S. Typhi proteins with ALS collected at the time of clinical presentation from 10 Bangladeshis with acute typhoid fever. We identified 62 immunoreactive antigens when evaluating both the IgG and IgA responses. Immune responses to 10 of these antigens discriminated between individuals with acute typhoid infection and healthy control individuals from areas where typhoid infection is endemic, as well as Bangladeshi patients presenting with fever who were subsequently confirmed to have a nontyphoid illness. Using an ALS enzyme-linked immunosorbent assay (ELISA) format and purified antigen, we then confirmed that immune responses against the antigen with the highest immunoreactivity (hemolysin E [HlyE]) correctly identified individuals with acute typhoid or paratyphoid fever in Dhaka, Bangladesh. These observations suggest that purified antigens could be used with ALS and corresponding acute-phase activated B lymphocytes in diagnostic platforms to identify acutely infected patients, even in areas where enteric fever is endemic.

  3. Acaricidal activities of whole cell suspension, cell-free supernatant,and crude cell extract of Xenorhabdus stokiae against mushroom mite (Luciaphorus sp.)

    Institute of Scientific and Technical Information of China (English)

    Prapassom BUSSAMAN; Chirayu SA-UTH; Paweena RATTANAS ENA; Angsumarn CHANDRAPATYA

    2012-01-01

    Xenorhabdus bacterium has been used as a biological control agent against Luciaphorus sp.,a mushroom mite endemic in Thailand.To develop an effective formulation of Xenorhabdus stokiae,treatments using different parts of X.stokiae isolate PB09 culture,including whole cell suspension,cell-free supernatant,and crude cell extract,were performed.The results show that different parts ofX.stokiae isolate PB09 culture could induce variable effects on mite mortality and fecundity.Application with cell-free supernatant of X.stokiae culture resulted in both the highest mite mortality rate [(89.00+3.60)%] and the lowest mite fecundity [(41.33+23.69) eggs/gravid female].Whole cell suspension of X.stokiae isolate PB09 culture was found to be slightly less effective than its cell-free supernatant,suggesting that X.stokiae was more likely to release its metabolites with acaricidal activities to the surrounding culture media.Crude cell extract of X.stokiae was not effective against mites.Cell-free supernatant of X.stokiae isolate PB09 was the most effective biological control agent and it could be conveniently used in future formulations instead of live bacteria.

  4. Culture supernatants from V. cholerae O1 ElTor strains isolated from different geographic areas induce cell vacuolation and cytotoxicity Cepas de V. cholerae O1 biotipo ElTor aisladas de diferente origen geográfico inducen vacuolización celular y citotoxicidad

    Directory of Open Access Journals (Sweden)

    Jorge E Vidal

    2009-02-01

    Full Text Available OBJECTIVE: To investigate whether the HlyA-induced vacuolating effect is produced by V. cholerae O1 ElTor strains isolated from different geographic origins, including Mexico. MATERIAL AND METHODS: Supernatant-induced haemolysis, vacuolating activity and cytotoxicity in Vero cells were recorded. PCR, RFLP analysis and molecular cloning were performed. RESULTS: All ElTor strains analyzed induced cellular vacuolation. Ribotype 2 strains isolates from the U.S. gulf coast yielded the highest titer of vacuolating activity. Eight of nine strains were haemolytic, while all strains were PCR positive for the hlyA gene. We cloned the hlyA gene from two ElTor strains, a toxigenic (2514-88, ctxAB+ and a non-toxigenic Mexican strain (CM 91-3, ctxAB-. Supernatant from those recombinant E. coli strains induced haemolysis, cell vacuolation and cytotoxicity. RFLP-PCR analysis revealed similarities in the hlyA gene from all strains tested. CONCLUSION: The HlyA-induced vacuolating effect is a widespread phenotype of epidemic V. cholerae O1 ElTor strains.OBJETIVO: Analizar el efecto vacuolizante de cepas de V. cholerae O1 ElTor aisladas de diferente origen geográfico, incluyendo México. MATERIAL Y MÉTODOS: Se realizaron pruebas de hemolisis, vacuolización y citotoxicidad en células Vero, así como PCR, análisis por RFLP y clonación molecular. RESULTADOS: Todas las cepas indujeron el efecto vacuolizante. Las cepas del ribotipo 2, aisladas de las costas del Golfo en Estados Unidos, presentaron títulos altos de vacuolización. El gen hlyA fue amplificado en las nueve cepas mediante PCR, aunque sólo ocho fueron hemolíticas. Se clonó el gen hlyA de una cepa toxigénica (2514-88, ctxAB+ y de una cepa no toxigénica aislada en México (CM 91-3, ctxAB-. El sobrenadante de las clonas recombinantes indujo hemólisis, efecto vacuolizante y citotoxicidad. El RFLP mostró alta similitud del gen hlyA de las cepas estudiadas. CONCLUSIÓN: El efecto vacuolizante es un

  5. Preformed purified peptide/major histocompatibility class I complexes are potent stimulators of class I-restricted T cell hybridomas

    DEFF Research Database (Denmark)

    Stryhn, A; Pedersen, L O; Ortiz-Navarrete, V;

    1994-01-01

    and quantitated. Latex particles were subsequently coated with known amounts of preformed complexes and used to stimulate the T cell hybridomas. Stimulation was specific, i.e. only the appropriate peptide/class I combination were stimulatory, and quite sensitive, i.e. as little as 300 complexes per bead could...

  6. Specific amplification by PCR of rearranged genomic variable regions of immunoglobulin genes from mouse hybridoma cells.

    Science.gov (United States)

    Berdoz, J; Monath, T P; Kraehenbuhl, J P

    1995-04-01

    We have designed a novel strategy for the isolation of the rearranged genomic fragments encoding the L-VH-D-JH and L-V kappa/lambda-J kappa/lambda regions of mouse immunoglobulin genes. This strategy is based on the PCR amplification of genomic DNA from mouse hybridomas using multiple specific primers chosen in the 5'-untranslated region and in the intron downstream of the rearranged JH/J kappa/lambda sequences. Variable regions with intact coding sequences, including full-length leader peptides (L) can be obtained without previous DNA sequencing. Our strategy is based on a genomic template that produces fragments that do not need to be adapted for recombinant antibody expression, thus facilitating the generation of chimeric and isotype-switched immunoglobulins.

  7. Monoclonal antibodies directed against protoplasts of soybean cells : Generation of hybridomas and characterization of a monoclonal antibody reactive with the cell surface.

    Science.gov (United States)

    Villanueva, M A; Metcalf, T N; Wang, J L

    1986-09-01

    Splenocytes, derived from mice that had been immunized with protoplasts prepared from suspension cultures of root cells of Glycine max (L.) Merr. (SB-1 cell line), were fused with a murine myeloma cell line. The resulting hybridoma cultures were screened for the production of antibodies directed against the soybean protoplasts and were then cloned. One monoclonal antibody, designated MVS-1, was found to bind to the outer surface of the plasma membrane on the basis of several criteria: (a) agglutination of the protoplasts; (b) binding of fluorescence-labeled immunoglobulin on protoplasts yielding a ring staining pattern with prominent intensity at the edges; and (c) saturable binding by protoplasts of (125)I-labeled Antibody MVS-1. The antigenic target of Antibody MVS-1, identified by immunoblotting techniques, contained a polypeptide of relative molecular mass (Mr) approx. 400000 under both reducing and non-reducing conditions. When the antigenic target of Antibody MVS-1 was chromatographed in potassium phosphate buffer, the position of elution corresponded to that of a high-molecular-weight species (Mr 400000). These results provide the protein characterization required for the analysis of the mobility of Antibody MVS-1 bound to the plasma membrane of SB-1 cells.

  8. Purification of monoclonal antibodies from whole hybridoma fermentation broth by fluidized bed adsorption.

    Science.gov (United States)

    Thömmes, J; Halfar, M; Lenz, S; Kula, M R

    1995-02-01

    To achive the coarse purification of a monoclonal antibody from whole hybridoma fermentation broth a fluidized bed cation exchange process was used. The procedure consisted of application of the crude sample and washing of the bed in a fluidized mode and elution in a fixed bed mode. A completely clarified eluate was obtained with purification factors between 4 and 8 and a concentration of the desired product (monoclonal antibody) by a factor of more than 3 was achived. Thus, a combination of the three early steps of the downstream process clarification, concentration and coarse purification was possible. Two different materials were tested: a commercially available agarose-based matrix (Stream-line-SP), and a self-derivatized material based on controlled-pore glass (Bioran). Initial experiments were performed to describe the fluidization of the glass material. Comparison with the agarose material showed several differences, the agarose matrix allowing liquid flow closer to plug flow than the glass material. Increased backmixing in the liquid phase was detected when fluidizing the glass adsorbent compared with the agarose-based matrix. Despite this fact, comparison of the two materials with respect to antibody binding and elution demonstrated a similar performance. (c) 1995 John Wiley & Sons, Inc.

  9. A cell sorter with modified bamboo charcoal for the efficient selection of specific antibody-producing hybridomas.

    Science.gov (United States)

    Lin, Chien-Chen; Ni, Mei-Hui; Chang, Yu-Chung; Yeh, Hsiu-Lun; Lin, Feng-Huei

    2010-11-01

    Monoclonal antibodies (mAbs) have been proven useful in research and clinical applications. However, the generation of mAbs by conventional hybridoma technology is time-, cost- and labor-consuming. Here we developed a simplified procedure for efficient generation and selection of antibody-producing hybridomas within 1 h, using a particular cell sorter design, a cytoflow reactor-based cell sorter (CBCS) which consists mainly of the "cytoflow reactor" that comprises two components, a reaction chamber and a glass tubing for air and medium exchange by gravity, and the "sorting material", human EGFR-conjugated bamboo charcoal, for specific B-cell enrichment. The high surface area and porous structure of bamboo charcoal greatly increased cell density and protein production. Moreover, from Raman, FT-IR spectroscopy and IFA analysis, the carboxylation and immobilization of bamboo charcoal can be introduced easily by nitric acid treatment and conjugated handily with human EGFR using EDC/NHS. Other evidences, such as IFA, showed that the specific hybridomas generated in this study could secrete specific anti-human EGFR antibodies. Our design allows the production of mAbs while avoiding time-consuming steps, such as large numbers of limiting dilutions and screening assays, and demonstrates that the CBCS could be a powerful tool for monoclonal antibody production.

  10. Fluorescence Spectrum and Decay Measurement for Hsil VS Normal Cytology Differentiation in Liquid Pap Smear Supernatant

    Science.gov (United States)

    Vaitkuviene, A.; Gegzna, V.; Juodkazis, S.; Jursenas, S.; Miasojedovas, S.; Kurtinaitiene, R.; Rimiene, J.; Vaitkus, J.

    2009-06-01

    Cervical smear material contains endo and exocervical cells, mucus and inflammative, immune cells in cases of pathology. Just not destroyed keratinocytes lay on the glass for microscopy. Liquid cytology supernatant apart other diagnostics could be used for photodiagnostic. The spectroscopic parameters suitable for Normal and HSIL cytology groups supernatant differentiation are demonstrated. The dried liquid PAP supernatant fractions—sediment and liquid were investigated. Excitation and emission matrices (EEM), supernatant fluorescence decay measured under 280 nm diode short pulse excitation and fluorescence spectroscopy by excitation with 355 nm laser light were analyzed. The differences between Normal and HSIL groups were statistically proven in the certain spectral regions. Fluorescence decay peculiarities show spectral regions consisting of few fluorophores. Obtained results on fluorescence differences in Normal and HSIL groups' supernatant shows the potency of photodiagnosis application in cervical screening.

  11. Performance of a membrane-dialysis bioreactor with a radial-flow fixed bed for the cultivation of a hybridoma cell line.

    Science.gov (United States)

    Bohmann, A; Pörtner, R; Märkl, H

    1995-10-01

    A bioreactor system for the continuous cultivation of animal cells with a high potential for scale-up is presented. This reactor system consists of radial-flow fixed-bed units coupled with a dialysis module The dialysis membrane enables the supply of low-molecular-weight nutrients and removal of toxic metabolites, while high-molecular-weight nutrients and products (e.g., monoclonal antibodies) are retained and accumulated. This concept was investigated on the laboratory scale in a bioreactor with an integrated dialysis membrane. The efficiency of the reactor system and the reproducibility of the cell activity (hybridoma cells) under certain process conditions could be demonstrated in fermentations up to 77 days. Based on model calculations, an optimized fermentation strategy was formulated and experimentally confirmed. Compared to chemostat cultures with suspended cells, a ten-times higher mAb concentration (383 mg1(-1)) could be obtained. The highest volumetric specific mAb production rate determined was 6.1 mg mAb (1 fixed bed)-1h-1.

  12. Establishment and characterization of mouse bone marrow-derived mast cell hybridomas

    Energy Technology Data Exchange (ETDEWEB)

    Kawahara, Takeshi, E-mail: tkawafb@shinshu-u.ac.jp [Integrated Department of Sciences of Functional Foods, Graduate School of Agriculture, Shinshu University, Nagano (Japan)

    2012-11-01

    Interleukin (IL)-3-dependent mouse bone marrow-derived mast cells (BMMCs) are an important model for studying the function of mucosal-type mast cells. In the present study, BMMCs were successfully immortalized by cell fusion using a hypoxanthine-aminopterin-thymidine medium-sensitive variant of P815 mouse mastocytoma (P815-6TgR) as a partner cell line. The established mouse mast cell hybridomas (MMCHs) expressed {alpha}, {beta}, and {gamma} subunits of high-affinity immunoglobulin E (IgE) receptor (Fc{epsilon}RI) and possessed cytoplasmic granules devoid of or partially filled with electron-dense material. Four independent MMCH clones continuously proliferated without supplemental exogenous IL-3 and showed a degranulation response on stimulation with IgE+antigen. Furthermore, histamine synthesis and release by degranulation were confirmed in MMCH-D5, a MMCH clone that showed the strongest degranulation response. MMCH-D5 exhibited elevated levels of IL-3, IL-4, IL-13, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor (TNF)-{alpha}, and cyclooxygenase 2, and production of prostaglandin D{sub 2} and leukotriene C{sub 4} in response to IgE-induced stimulation. MMCH clones also expressed Toll-like receptors (TLRs) 1, 2, 4, and 6 and showed elevated levels of TNF-{alpha} expression in response to stimulation with TLR2 and TLR4 ligands. The MMCHs established using this method should be suitable for studies on Fc{epsilon}RI- and TLR-mediated effector functions of mast cells.

  13. HYBRIDOMA PRODUCTION USING IMMUNE LYMPHOCYTES AGAINST BRUGIA MALAYI ANTIGEN WITH MYELOMA CELLS

    Directory of Open Access Journals (Sweden)

    Soeyoko Soeyoko

    2012-09-01

    Full Text Available Wuchereria bancrofti, Brugia malayi, and Brugia timori are the causative agents of lymphatic hiariasis in Indonesia, but in some endemic areas, Brugia malayi is the most commonly found,Diagnosis of hiariasis is normally based on clinical and parasitological examinations, but both have limitations. Therefore now the immunological examination plays an important role in the diagnosis of filariasis. The discovery of monoclonal antibodies recently may probably give a firm scientific basis in immunology and add a new dimension to the efforts of developing a specific and sensitive immunological test for various stages of filarial infection. In this study, the production of hybridoma cells to develop monoclonal antibodies against B. malayi integrated a number of techniques: preparations of B. malayi surface antigen, immune lymphocyte cells, NSI myeloma cells and macrophage feeder layers, and a fusion of immune lymphocytes with myeloma cells. The results of this study can be concluded in three points: Protein analysis of the surface antigen was examined by Biureet and SDS-PAGE. A total of fourteen examinations were conducted by using 4000 L3 for each experiment. Three were not detected by Biureet method, but showed five protein fractions by SDS-PAGE. The protein concentration of the surface L3 was varied from 85.0/µg/ml to 769.23/µg/ml, with an average of 297.04/µg/ml.The immunoreactivifies of Balb/c mice antibodies to B. malayi L3 surface antigen were tested by ELISA and showed a gradual increase after four times immunizations at two weeks interval. The optical density (OD after four times immunizations was varied from 0.363 to 0.878 each mouse, where as the positive control sera OD was 0.570.Hybridization using immune lymphocytes against B. malayi L3 surface antigen with myeloma cells yielded 60.41% hybrid cells and none of them produced monoclonal antibodies tested of ELISA.

  14. Beneficial effects of BV2 cell on proliferation and neuron-differentiating of mesenchymal stem cells in the circumstance of injured PC12 cell supernatant

    Institute of Scientific and Technical Information of China (English)

    Xiao-Guang LUO; Hong WANG; Jin ZHOU; Rong YAN; Zhe WU; Chao-Dong ZHANG; Qiu-Shuang WANG

    2006-01-01

    Objective The microglias is the representative of immune cells in the brain. It plays dual roles of both repairing and damaging in injured nervous system, and works as an inevitable component of the circumstance of injured neurons. This study was aiming at the effects of the microglias on the biological activities of mesenchymal stem cells (MSCs) inthe circumstance of injured neurons. Methods MSCs were obtained by primary culture. We adopted PC12 cells (PC12) and BV2 cells (BV2) to substitute for neurons and microglias, respectively. PC12 were injured by aged Aβ1-40 and the supernatant of the injured PC12 was used to set up the circumstance of injured neurons. Transwells were used for co-culture of BV2 and MSCs, which allowed the independent detection of cells after co-culture. Immunofluorescence was used to identify MSCs and neuron-differentiating cells with CD44 and neuron specific enolase (NSE) staining, respectively. MTT assay was adopted to measure the proliferation. Results In the circumstance of both BV2 presence and injured PC12 supernatant incubation, either the proliferation or the differentiation of MSCs reached the highest, which seemed to be contradictory, but we gave our explanations. With the BV2 co-culture, the proliferation of MSCs tend to be higher, but the neuron-differentiating MSCs were similar to those incubated without BV2 co-culture either in normal or injured in PC12 supernatant. With the incubation of injured PC12 supernatant, the neuron-differentiating cells were significantly higher than that of control (P < 0.05). Conclusion In the circumstance of injured neurons, microlgias tend to promote the MSCs proliferation. Although not helpful in neuron-differentiating, microglias did not exert any negative effect either.

  15. Cell-free fetal DNA in amniotic fluid supernatant for prenatal diagnosis.

    Science.gov (United States)

    Soltani, M; Nemati, M; Maralani, M; Estiar, M A; Andalib, S; Fardiazar, Z; Sakhinia, E

    2016-04-30

    In widespread conviction, amniotic fluid is utilized for prenatal diagnosis. Amniotic fluid supernatant is usually discarded, notwithstanding being a good source of fetal DNA. The aim of the present study was to assess cell-free fetal DNA extracted from amniotic fluid supernatant for application in prenatal diagnosis such as gender determination and early diagnosis of β-thalassemia. Samples of amniotic fluid of 70 pregnant women were collected and went through routine tests along with tests for cell-free fetal DNA from amniotic fluid supernatant. The DNA in the amniotic fluid supernatant was extracted and analyzed for gender determination by PCR and Real-time PCR. ARMS-PCR was applied to test early diagnosis of IVS II-I mutation (common β-thalassemia mutation) and E7V mutation for sickle cell anemia using DNA extracted from the amniotic fluid supernatant. Using the cell-free fetal DNA extracted from the amniotic fluid supernatant, the sensitivity of PCR and Real-time PCR for gender detection was compared with the routine cytogenetic method. The fetus tested for sickle cell anemia and β-thalassemia was observed to be healthy but heterozygous for IVS II-I mutation. The findings indicated that cell-free fetal DNA from amniotic fluid supernatant can be a good source of fetal DNA and be used in early prenatal diagnosis since because of its fast and accurate application. Therefore, it would be suggested that the amniotic fluid supernatant's disposal is prevented because if the tests needs to be repeated, cell-free fetal DNA extracted from the amniotic fluid supernatant can be used as an alternative source for prenatal diagnosis.

  16. Effect of Lactobacillus sp. isolates supernatant on Escherichia coli O157:H7 enhances the role of organic acids production as a factor for pathogen control

    Directory of Open Access Journals (Sweden)

    Larissa B. Poppi

    2015-04-01

    Full Text Available Many attempts have been made to establish the control of foodborne pathogens through Lactobacillus isolates and their metabolism products with success being obtained in several situations. The aim of this study was to investigate the antagonistic effect of eight Lactobacillus isolates, including L. casei subsp. pseudoplantarum, L. plantarum, L. reuteri and L. delbrueckii subsp. delbrueckii, on the pathogenic Escherichia colistrain O157:H7. The inhibitory effect of pure cultures and two pooled cultures supernatants of Lactobacillus on the growth of pathogenic bacteria was evaluated by the spot agar method and by monitoring turbidity. Antimicrobial activity was confirmed for L. reuteri and L. delbrueckii subsp. delbrueckii and for a pool of lactic acid bacteria. The neutralized supernatant of the pool exerted a higher antimicrobial activity than that of the individual strains. Furthermore, D-lactic acid and acetic acid were produced during growth of the Lactobacillus isolates studied.

  17. How does the supernatant of Lactobacillus acidophilus affect the proliferation and differentiation activities of rat bone marrow-derived stromal cells?

    Science.gov (United States)

    Samadikuchaksaraei, A; Gholipourmalekabadi, M; Saberian, M; Abdollahpour Alitappeh, M; Shahidi Delshad, E

    2016-08-31

    Low proliferation rate and unwanted differentiation of bone marrow-derived stromal cells (rBMSCs) during the frequent passages have limited the use of such cells in clinical cell therapy. Recently, the researchers have focused on the effects of the components produced by some bacteria on proliferation of the stem cells. In this study, we discussed the possible effects of the Lactobacillus acidophilus supernatant on proliferation and differentiation of the rBMSCs. For this aim, the cells were isolated from rat bone marrow, characterized by culturing on tissue specific differentiation media and stained. The cells (passage two) were treated with different concentrations of the L. acidophilus supernatant (0, 0.1, 0.3, 0.9, 3, 9 and 30 &mgr;l/ml) for 14 days. The proliferation and differentiation capacity of the cells were then determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT assay) and tissue specific staining. The results showed a positive effect of the supernatant on the cell proliferation in 3 and 9 &mgr;l/ml concentrations, while did not affect the differentiation capacity of the rBMSCs. The current study strongly suggests the L. acidophilus supernatant as an alternative material that could be added to the media with aim of improvement in the proliferation rate of the rBMSCs without affecting their differentiation capacity.

  18. Test procedures and instructions for Hanford tank waste supernatant cesium removal

    Energy Technology Data Exchange (ETDEWEB)

    Hendrickson, D.W., Westinghouse Hanford

    1996-05-31

    This document provides specific test procedures and instructions to implement the test plan for the preparation and conduct of a cesium removal test using Hanford Double-Shell Slurry Feed supernatant liquor from tank 251-AW-101 in a bench-scale column.Cesium sorbents to be tested include resorcinol-formaldehyde resin and crystalline silicotitanate. The test plan for which this provides instructions is WHC-SD-RE-TP-022, Hanford Tank Waste Supernatant Cesium Removal Test Plan.

  19. Test procedures and instructions for Hanford complexant concentrate supernatant cesium removal using CST

    Energy Technology Data Exchange (ETDEWEB)

    Hendrickson, D.W.

    1997-01-08

    This document provides specific test procedures and instructions to implement the test plan for the preparation and conduct of a cesium removal test, using Hanford Complexant Concentrate supernatant liquor from tank 241-AN-107, in a bench-scale column. The cesium sorbent to be tested is crystalline silicotitanate. The test plan for which this provides instructions is WHC-SD-RE-TP-023, Hanford Complexant Concentrate Supernatant Cesium Removal Test Plan.

  20. Global Profiling of Metabolite and Lipid Soluble Microbial Products in Anaerobic Wastewater Reactor Supernatant Using UPLC-MS(E).

    Science.gov (United States)

    Tipthara, Phornpimon; Kunacheva, Chinagarn; Soh, Yan Ni Annie; Wong, Stephen C C; Pin, Ng Sean; Stuckey, David C; Boehm, Bernhard O

    2017-02-03

    Identification of soluble microbial products (SMPs) released during bacterial metabolism in mixed cultures in bioreactors is essential to understanding fundamental mechanisms of their biological production. SMPs constitute one of the main foulants (together with colloids and bacterial flocs) in membrane bioreactors widely used to treat and ultimately recycle wastewater. More importantly, the composition and origin of potentially toxic, carcinogenic, or mutagenic SMPs in renewable/reused water supplies must be determined and controlled. Certain classes of SMPs have previously been studied by GC-MS, LC-MS, and MALDI-ToF MS; however, a more comprehensive LC-MS-based method for SMP identification is currently lacking. Here we develop a UPLC-MS approach to profile and identify metabolite SMPs in the supernatant of an anaerobic batch bioreactor. The small biomolecules were extracted into two fractions based on their polarity, and separate methods were then used for the polar and nonpolar metabolites in the aqueous and lipid fractions, respectively. SMPs that increased in the supernatant after feed addition were identified primarily as phospholipids, ceramides, with cardiolipins in the highest relative abundance, and these lipids have not been previously reported in wastewater effluent.

  1. The effect of moderately halophilic bacteria supernatant on proliferation and apoptosis of cancer cells and mesenchymal stem cells.

    Science.gov (United States)

    Sarvari, S; Seyedjafari, E; Amoozgar, M A; Bakhshandeh, B

    2015-06-10

    Many drug discoveries and developing of their applications has originated from microbial metabolites. The most efforts in development of new drugs are concerned with anti—cancer agents that cause better treatment results, less side effects, and more economical production. Several anti—tumor drugs have been recently extracted from natural microbial products. Among these various microbial diversity, Marin bacteria and Archaea have been considered as important and efficient organisms to serve as manufacturers of diverse bioactive compounds. Moderately halophilic microorganisms isolated from saline ponds and lakes of Iran show high capability for production of bioactive compounds like enzymes, dyes and anti—cancer agents. In this research, nine moderately halophilic bacteria isolates were screened to evaluate their anti—cancer agent productivity. After five days of culture on suitable mediums, supernatant samples were tested for in vitro anti—proliferative activity against Human Umbilical Vein Endothelial Cells (HUVEC) while same concentrations of supernatants were examined for evaluating of proliferative activity against Adipose—derived Mesenchymal stem cells (MSCs). Both assessments were carried out by MTT assay and double PI and DAPI staining. GASX17, GBWy6 and GBPX3 isolates just induced HUVEC cell deaths and exhibited anti—proliferative activity while R2S12 not only reduced HUVEC cell proliferation but also enhanced proliferation of MSCs. R2S12 , GASX17, GBWy6 and GBPX3 isolates were characterized biochemically and six hydrophilic components were detected. This research established new bioactive compounds that could be used as an effective treatment in chemotherapy.

  2. Complete dissection of the Hb(64-76) determinant using T helper 1, T helper 2 clones, and T cell hybridomas

    DEFF Research Database (Denmark)

    Evavold, B D; Williams, S G; Hsu, B L

    1992-01-01

    We have generated cloned Th1 cells, Th2 cells, and T cell hybridomas specific for the single immunogenic peptide from the beta-chain of murine hemoglobin (Hb(64-76)). The availability of these various types of T cells provided us an unique opportunity to examine and dissect the T cell response...... to an immunogenic peptide. A panel of altered Hb peptides was made by replacing each amino acid in the Hb peptide (positions 64-76) with a conservative amino acid substitution or an alanine. Although none of the eleven T cell clones and hybridomas tested exhibited the same pattern of reactivity to the substituted...

  3. [The Supernatant Obtained from Cultured Anip973 Cells Enhances the Biological Activities of HUVEC].

    Science.gov (United States)

    Zhang, Cuicui; Li, Yueya; Wang, Jing; Li, Kai

    2015-11-01

    背景与目的 肿瘤微血管内皮细胞与正常组织来源的微血管内皮细胞相比,具有对生长因子反应性高、粘附因子表达高等特点,造成肿瘤血管通透性高且新生旺盛,导致肿瘤的快速生长和转移。因此了解肿瘤微环境中内皮细胞发生、形态及功能上的异质性特点,对进行合理的、个性化的、以血管内皮细胞为靶点的抗血管新生治疗有一定的指导作用。本实验旨在研究高转移肺腺癌Anip973细胞培养上清液对人脐静脉内皮细胞(human umbilical vein endothelial cell, HUVEC)表面标记及其生物学行为的影响。方法 以含有不同浓度Anip973细胞上清液的培养基(RPMI-1640+10%胎牛血清)培养的HUVEC为实验组、单纯培养基培养的HUVEC为对照组,以CCK-8检测各组细胞增殖率、基质胶成管实验检测成管能力、Transwell检测迁移能力、流式细胞术检测细胞表面标记CD105、CD31以及凋亡标记Annexin V的表达,并分析细胞表面标记的表达与生物学行为之间的关系。结果 与对照组相比,经Anip973细胞上清液培养的HUVEC增殖更多、且在浓度为250 µL/mL作用24 h最明显(P=0.002);成管及迁移能力亦均较对照组增强,分别在浓度为125 µL/mL(P<0.001)、250 µL/mL(P=0.002)时达最强;细胞表面标记CD105表达阳性率升高、浓度为250 µL/mL(P=0.028)最明显;CD31表达阳性率呈浓度依赖性升高;凋亡细胞比例降低。相关性分析显示:CD105的表达阳性率与细胞增殖及迁移能力呈正相关关系、CD31的表达与生物学行为并无明显相关性。结论 Anip973细胞上清液能促进HUVEC细胞增殖、迁移、血管形成,CD105也随之变化、并可在一定程度上反应其生物学行为。.

  4. Characterization of tetanus toxin, neat and in culture supernatant, by electrospray mass spectrometry

    NARCIS (Netherlands)

    Baar, B.L.M. van; Hulst, A.G.; Roberts, B.; Wils, E.R.J.

    2002-01-01

    A method was developed for the liquid chromatographic-mass spectrometric (LC-MS) identification of extremely neurotoxic toxins. The method combines sample treatment in a safety containment and analysis of detoxified material in a common laboratory facility. The method was applied to the

  5. Characterization of tetanus toxin, neat and in culture supernatant, by electrospray mass spectrometry

    NARCIS (Netherlands)

    Baar, B.L.M. van; Hulst, A.G.; Roberts, B.; Wils, E.R.J.

    2002-01-01

    A method was developed for the liquid chromatographic-mass spectrometric (LC-MS) identification of extremely neurotoxic toxins. The method combines sample treatment in a safety containment and analysis of detoxified material in a common laboratory facility. The method was applied to the characteriza

  6. Residual urinary extracellular vesicles in ultracentrifugation supernatants after hydrostatic filtration dialysis enrichment

    Science.gov (United States)

    Musante, Luca; Tataruch-Weinert, Dorota; Kerjaschki, Dontscho; Henry, Michael; Meleady, Paula; Holthofer, Harry

    2017-01-01

    ABSTRACT Urinary extracellular vesicles (UEVs) appear an ideal source of biomarkers for kidney and urogenital diseases. The majority of protocols designed for their isolation are based on differential centrifugation steps. However, little is still known of the type and amount of vesicles left in the supernatant. Here we used an isolation protocol for UEVs which uses hydrostatic filtration dialysis as first pre-enrichment step, followed by differential centrifugation. Transmission electron microscopy (TEM), mass spectrometry (MS), western blot, ELISA assays and tuneable resistive pulse sensing (TRPS) were used to characterise and quantify UEVs in the ultracentrifugation supernatant. TEM showed the presence of a variety of small size vesicles in the supernatant while protein identification by MS matched accurately with the protein list available in Vesiclepedia. Screening and relative quantification for specific vesicle markers showed that the supernatant was preferentially positive for CD9 and TSG101. ELISA tests for quantification of exosome revealed that 14%, was left in the supernatant with a particle diameter of 110 nm and concentration of 1.54 × 1010/ml. Here we show a comprehensive characterisation of exosomes and other small size urinary vesicles which the conventional differential centrifugation protocol may lose.

  7. Evaluation of Hanford Tank Supernatant Availability for Technetium Management Project Studies in FY16

    Energy Technology Data Exchange (ETDEWEB)

    Rapko, Brian M. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2015-09-30

    This report examines the need for actual Hanford tank waste solutions to support tasks in the Technetium Management Program in fiscal year (FY) 2016. One key need is to identify both samples where a majority of the soluble technetium is present as pertechnetate and samples where it is not. The total amount of tank supernatant needed from any given tank waste supernatant was determined by polling the tasks leaders for their technology testing needs in FY16 and then arbitrarily ascribing a 10% process loss associated with consolidation and the Cs-137 removal needed to reduce the dose to a level suitable for testing in radiological fumehoods. These polling results identified a need for approximately 2.1 to 3.6 kg of any particular targeted Hanford tank waste supernatant.

  8. Modelling antagonic effect of lactic acid eacteria supernatants on some pathogenic bacteria

    Directory of Open Access Journals (Sweden)

    Augustus Caeser Franke Portella

    2009-11-01

    Full Text Available This work presents a statistical model of survival analysis for three pathogenic bacterial strains (Escherichia coli, Listeria monocytogenes and Staphylococcus aureus, when treated with neutralized and non-neutralized filtered supernatants broth from cultures of Lactobacillus acidhophilus, Lactobacillus rhamnosus and Lactobacillus sake. Survival analysis is a method employed to determine the period of time from an initial stage up to the occurrence of a particular event of interest, as death or a particular culture growth failure. In order to evaluate the potential efficacy of the ahead mentioned lactic acid bacteria when used as bioprotective starters in foods, experimental data were statistically treated and expressed by simple representative curves. Following the methodology of Cox and Kaplan-Meier, it was possible to make the selection of the best bioprotective lactic starter, as a predictive tool for evaluation of shelf life and prevention of eventual risks in fresh sausages and other similar food products.Este trabalho apresenta um modelo estatístico de análise de sobrevivência para três bactérias patogénicas (Escherichia coli, Listeria monocytogenes e Staphylococcus aureus, quando tratados com sobrenadantes filtrados neutralizado e não neutralizado provenientes de culturas de Lactobacillus acidhophilus, Lactobacillus rhamnosus e Lactobacillus sake. A Análise de sobrevivência é um método utilizado para determinar o período de tempo a partir de uma fase inicial até a ocorrência de um determinado evento de interesse, como a morte ou a inibição de uma particular cultura, a fim de avaliar a eficácia potencial das referidas bactérias lácticas quando usadas como bioproteção em alimentos. Os dados experimentais foram tratados estatisticamente, seguindo a metodologia de Cox e Kaplan-Meier e foi possível fazer a seleção dos melhores fermentos láticos bioprotectivos, como uma ferramenta para avaliação preditiva, vida de

  9. Long acting β2-agonist and corticosteroid restore airway glandular cell function altered by bacterial supernatant

    Directory of Open Access Journals (Sweden)

    Nawrocki-Raby Béatrice

    2010-01-01

    Full Text Available Abstract Background Staphylococcus aureus releases virulence factors (VF that may impair the innate protective functions of airway cells. The aim of this study was to determine whether a long-acting β2 adrenergic receptor agonist (salmeterol hydroxynaphthoate, Sal combined with a corticosteroid (fluticasone propionate, FP was able to regulate ion content and cytokine expression by airway glandular cells after exposure to S. aureus supernatant. Methods A human airway glandular cell line was incubated with S. aureus supernatant for 1 h and then treated with the combination Sal/FP for 4 h. The expression of actin and CFTR proteins was analyzed by immunofluorescence. Videomicroscopy was used to evaluate chloride secretion and X-ray microanalysis to measure the intracellular ion and water content. The pro-inflammatory cytokine expression was assessed by RT-PCR and ELISA. Results When the cells were incubated with S. aureus supernatant and then with Sal/FP, the cellular localisation of CFTR was apical compared to the cytoplasmic localisation in cells incubated with S. aureus supernatant alone. The incubation of airway epithelial cells with S. aureus supernatant reduced by 66% the chloride efflux that was fully restored by Sal/FP treatment. We also observed that Sal/FP treatment induced the restoration of ion (Cl and S and water content within the intracellular secretory granules of airway glandular cells and reduced the bacterial supernatant-dependent increase of pro-inflammatory cytokines IL8 and TNFα. Conclusions Our results demonstrate that treatment with the combination of a corticosteroid and a long-acting β2 adrenergic receptor agonist after bacterial infection restores the airway glandular cell function. Abnormal mucus induced by defective ion transport during pulmonary infection could benefit from treatment with a combination of β2 adrenergic receptor agonist and glucocorticoid.

  10. Neuronal activation by mucosal biopsy supernatants from irritable bowel syndrome patients is linked to visceral sensitivity.

    Science.gov (United States)

    Buhner, Sabine; Braak, Breg; Li, Qin; Kugler, Eva Maria; Klooker, Tamira; Wouters, Mira; Donovan, Jemma; Vignali, Sheila; Mazzuoli-Weber, Gemma; Grundy, David; Boeckxstaens, Guy; Schemann, Michael

    2014-10-01

    Based on the discomfort/pain threshold during rectal distension, irritable bowel syndrome (IBS) patients may be subtyped as normo- or hypersensitive. We previously showed that mucosal biopsy supernatants from IBS patients activated enteric and visceral afferent neurons. We tested the hypothesis that visceral sensitivity is linked to the degree of neuronal activation. Normo- and hypersensitive IBS patients were distinguished by their discomfort/pain threshold to rectal balloon distension with a barostat. Using potentiometric and Ca(2+) dye imaging, we recorded the response of guinea-pig enteric submucous and mouse dorsal root ganglion (DRG) neurons, respectively, to mucosal biopsy supernatants from normosensitive (n = 12 tested in enteric neurons, n = 9 tested in DRG) and hypersensitive IBS patients (n = 9, tested in both types of neurons). In addition, we analysed the association between neuronal activation and individual discomfort/pain pressure thresholds. The IBS supernatants evoked Ca(2+) transients in DRG neurons and spike discharge in submucous neurons. Submucous and DRG neurons showed significantly stronger responses to supernatants from hypersensitive IBS patients as reflected by higher spike frequency or stronger [Ca(2+)]i transients in a larger proportion of neurons. The neuroindex as a product of spike frequency or [Ca(2+)]i transients and proportion of responding neurons correlated significantly with the individual discomfort/pain thresholds of the IBS patients. Supernatants from hypersensitive IBS patients caused stronger activation of enteric and DRG neurons. The level of activation correlated with the individual discomfort/pain threshold pressure values. These findings support our hypothesis that visceral sensitivity is linked to activation of peripheral neurons by biopsy supernatants.

  11. Boildown Study on Supernatant Liquid Retrieved from AW-106 in December 2012

    Energy Technology Data Exchange (ETDEWEB)

    Page, Jason S.

    2013-06-04

    This document reports the results of a boildown study using a composite created from supernatant liquid grab samples retrieved from tank 241-AW-106 in December of 2012. The composite was made using predetermined volumes of the grab samples which accounted for layering of the supernatant liquid in the tank. The finished composite was a clear, yellow liquid containing no visible solids at hot cell ambient temperatures (24 - 27 °C). The density of the test composite was measured in the hot cell immediately before the boildown study and was 1.266 g/mL at 27.1 °C.

  12. 15th International Conference on Human Antibodies and Hybridomas. 14-16 April 2010, Tiara Park Atlantico Hotel, Porto, Portugal.

    Science.gov (United States)

    Kotlan, Beatrix

    2010-11-01

    Antibodies and antibody conjugates are currently one of the largest classes of new drug entities under development. These versatile molecules are being investigated for the treatment of many pathological conditions, such as cancer and infectious, inflammatory and autoimmune diseases. Antibodies can exert biological effects as naked antibodies by themselves, or can be used as delivery agents conjugated with various drugs (e.g., immunoconjugates) and as tools of multistep targeting. Site-specific delivery of therapeutic agents has been the ultimate goal of the pharmaceutical industry, as it has the potential to maximize drug efficiency while minimizing side effects. Antibodies have much potential for this objective. Thus, it is useful to summarize some of the main strategies currently being employed for the development of these diverse therapeutic molecules and to highlight the recent novelties in the field. These goals were the focus of the 15th International Conference on Human Antibodies and Hybridomas, held during 14-16 April 2010 in Porto, Portugal.

  13. The effect of NaCl substitution with KCl on proteinase activities of cell-free extract and cell-free supernatant at different pH levels and salt concentrations: Lactobacillus acidophilus and Lactobacillus casei.

    Science.gov (United States)

    Ayyash, M M; Sherkat, F; Shah, N P

    2013-01-01

    The aim of this study was to investigate the effect of substitution of NaCl with KCl at different pH levels and salt concentrations on proteinase activity of cell-free extract and cell-free supernatant of the probiotics Lactobacillus acidophilus and Lactobacillus casei. de Man, Rogosa, and Sharpe broth aliquots were mixed with 2 pure salts (NaCl and KCl) and 2 salt concentrations at 2 concentration levels (5 and 10%), inoculated with Lactobacillus acidophilus or Lactobacillus casei, and incubated aerobically at 37°C for 22 h. The cultures were then centrifuged at 4,000×g for 30 min, and the collected cell pellets were used to prepare cell-wall proteinases and the supernatants used as a source of supernatant (extracellular) proteinases. The proteolytic activity and protein content of both portions were determined. After incubation of both portions with 3 milk caseins (α-, β-, κ-casein), the supernatants were individually subjected to analysis of angiotensin-converting enzyme (ACE)-inhibitory activity and proteolytic activity using the o-phthalaldehyde method. Significant differences were observed in ACE-inhibitory and proteolytic activities between salt substitution treatments of cell-free extract and cell-free supernatant from both probiotic strains at the same salt concentration and pH level.

  14. Fate of cyanobacteria in drinking water treatment plant lagoon supernatant and sludge

    Energy Technology Data Exchange (ETDEWEB)

    Pestana, Carlos J.; Reeve, Petra J.; Sawade, Emma [Australian Water Quality Centre, South Australian Water Corporation, Adelaide, SA 5000 (Australia); Voldoire, Camille F. [Australian Water Quality Centre, South Australian Water Corporation, Adelaide, SA 5000 (Australia); École Européenne de Chimie, Polymères et Matériaux (ECPM), Strasbourg 67087 (France); Newton, Kelly; Praptiwi, Radisti [Australian Water Quality Centre, South Australian Water Corporation, Adelaide, SA 5000 (Australia); Collingnon, Lea [Australian Water Quality Centre, South Australian Water Corporation, Adelaide, SA 5000 (Australia); École Européenne de Chimie, Polymères et Matériaux (ECPM), Strasbourg 67087 (France); Dreyfus, Jennifer [Allwater, Adelaide Services Alliance, Wakefield St, Adelaide, SA 5001 (Australia); Hobson, Peter [Australian Water Quality Centre, South Australian Water Corporation, Adelaide, SA 5000 (Australia); Gaget, Virginie [University of Adelaide, Ecology and Environmental Sciences, School of Biological Sciences, Adelaide, SA 5005 (Australia); Newcombe, Gayle, E-mail: gayle.newcombe@sawater.com.au [Australian Water Quality Centre, South Australian Water Corporation, Adelaide, SA 5000 (Australia)

    2016-09-15

    In conventional water treatment processes, where the coagulation and flocculation steps are designed to remove particles from drinking water, cyanobacteria are also concentrated into the resultant sludge. As a consequence, cyanobacteria-laden sludge can act as a reservoir for metabolites such as taste and odour compounds and cyanotoxins. This can pose a significant risk to water quality where supernatant from the sludge treatment facility is returned to the inlet to the plant. In this study the complex processes that can take place in a sludge treatment lagoon were investigated. It was shown that cyanobacteria can proliferate in the conditions manifest in a sludge treatment lagoon, and that cyanobacteria can survive and produce metabolites for at least 10 days in sludge. The major processes of metabolite release and degradation are very dependent on the physical, chemical and biological environment in the sludge treatment facility and it was not possible to accurately model the net effect. For the first time evidence is provided to suggest that there is a greater risk associated with recycling sludge supernatant than can be estimated from the raw water quality, as metabolite concentrations increased by up to 500% over several days after coagulation, attributed to increased metabolite production and/or cell proliferation in the sludge. - Highlights: • Cyanobacteria in water treatment sludge significantly impact supernatant quality • Cyanobacteria can survive, and thrive, in sludge lagoon supernatant and in treatment sludge • Metabolite concentrations in cyanobacteria in sludge can increase up to 500% • The risk associated with supernatant recycling was assessed relative to available treatment barriers.

  15. Faecalibacterium prausnitzii supernatant improves intestinal barrier function in mice DSS colitis.

    Science.gov (United States)

    Carlsson, Anders H; Yakymenko, Olena; Olivier, Isabelle; Håkansson, Fathima; Postma, Emily; Keita, Asa V; Söderholm, Johan D

    2013-10-01

    OBJECTIVE. The intestinal microbiota plays a substantial role in the pathogenesis of inflammatory bowel disease (IBD). Faecalibacterium prausnitzii (FP) is underrepresented in IBD patients and have been suggested to have anti-inflammatory effects in mice. Increased intestinal permeability is common in IBD but the relationship between FP and intestinal barrier function has not been investigated. Our aim was to study treatment with FP supernatant on intestinal barrier function in a dextran sodium sulfate (DSS) colitis mice model. MATERIAL AND METHODS. C57BL/6 mice received 3% DSS in tap water ad libitum during five days to induce colitis. From day 3 the mice received a daily gavage with FP supernatant or broth during seven days. Ileum and colon were mounted in Ussing chambers for permeability studies with (51)Cr-EDTA and Escherichia coli K-12. Colon was saved for Western blot analyses of tight junction proteins. RESULTS. DSS-treated mice showed significant weight loss and colon shortening. Gavage with FP supernatant resulted in a quicker recovery after DSS treatment and less extensive colonic shortening. Ileal mucosa of DSS mice showed a significant increase in (51)Cr-EDTA-passage compared to controls. (51)Cr-EDTA passage was significantly decreased in mice receiving FP supernatant. No significant differences were observed in passage of E. coli K12. Western blots showed a trend to increased claudin-1 and claudin-2 expressions in DSS mice. CONCLUSIONS. Supernatant of FP enhances the intestinal barrier function by affecting paracellular permeability, and may thereby attenuate the severity of DSS-induced colitis in mice. These findings suggest a potential role of FP in the treatment of IBD.

  16. Comparison of excretory-secretory antigen and positive faecal supernatant antigen in the detection of Echinococcus granulosus infection in dogs by CIEP

    Directory of Open Access Journals (Sweden)

    P. R. Prathiush

    Full Text Available Coproantigen detection of Echinococcosis in dogs by counter immunoelectrophoresis was standardized. Adult Echinococcus granulosus worms were obtained from intestine of a necropsied positive dog. Excretory-secretory antigen was prepared by culturing adult worms in Medium 199 (pH 7.4. Faeces of positive dog were collected and fecal supernatant was prepared and used for coproantigen detection. CIEP was carried out using tris-borate buffer (pH 8.0 at a constant current of 8mA/slide for 60 minutes. CIEP detected infection with both the antigens. [Vet World 2009; 2(11.000: 421-422

  17. Thalidomide and pentoxifylline block the renal effects of supernatants of macrophages activated with Crotalus durissus cascavella venom

    Directory of Open Access Journals (Sweden)

    A.M.C. Martins

    2004-10-01

    Full Text Available Because thalidomide and pentoxifylline inhibit the synthesis and release of tumor necrosis factor-alpha (TNF-alpha, we determined the effect of these drugs on the renal damage induced by supernatants of macrophages activated with Crotalus durissus cascavella venom in order to identify the role of TNF-alpha in the process. Rat peritoneal macrophages were collected with RPMI medium and stimulated in vitro with C.d. cascavella venom (10 µg/ml in the absence and presence of thalidomide (15 µM or pentoxifylline (500 µM for 1 h and washed and kept in culture for 2 h. Supernatant (1 ml was tested on an isolated perfused rat kidney (N = 6 for each group. The first 30 min of each experiment were used as control. The supernatant was added to the perfusion system. All experiments lasted 120 min. The toxic effect of the preparation of venom-stimulated macrophages on renal parameters was determined. At 120 min, thalidomide (Thalid and pentoxifylline (Ptx inhibited (P < 0.05 the increase in perfusion pressure caused by the venom (control = 114.0 ± 1.3; venom = 137.1 ± 1.5; Thalid = 121.0 ± 2.5; Ptx = 121.4 ± 4.0 mmHg, renal vascular resistance (control = 4.5 ± 0.2; venom = 7.3 ± 0.6; Thalid = 4.5 ± 0.9; Ptx = 4.8 ± 0.6 mmHg/ml g-1 min-1, urinary flow (control = 0.23 ± 0.001; venom = 0.44 ± 0.01; Thalid = 0.22 ± 0.007; Ptx = 0.21 ± 0.009 ml g-1 min-1, glomerular filtration rate (control = 0.72 ± 0.06; venom = 1.91 ± 0.11; Thalid = 0.75 ± 0.04; Ptx = 0.77 ± 0.05 ml g-1 min-1 and the decrease in percent tubular sodium transport (control = 77.0 ± 0.9; venom = 73.9 ± 0.66; Thalid = 76.6 ± 1.1; Ptx = 81.8 ± 2.0%, percent tubular chloride transport (control = 77.1 ± 1.2; venom = 71.4 ± 1.1; Thalid = 77.6 ± 1.7; Ptx = 76.8 ± 1.2%, and percent tubular potassium transport (control = 72.7 ± 1.1; venom = 63.0 ± 1.1; Thalid = 72.6 ± 1.0; Ptx = 74.8 ± 1.0%, 30 min before and during the stimulation of macrophages with C.d. cascavella venom

  18. Optmization of culture media for Bifidobacterium bifidum and Bifidobacterium crudilactis and study of the antimicrobial effect of culture supernatants

    OpenAIRE

    Bondue, Pauline; Delcenserie, Véronique

    2015-01-01

    Complex oligosaccharides from human milk (HMO) contribute to infant health. Bifidobacteriumbifidum mainly found in breast-fed infant microbiota has all the enzymatic machinery for degradation of HMO. On the other hand, whey is rich in complex bovin milk oligosaccharides (BMO) very similar to HMO, including 3’-sialyllactose (3’SL). They are very likely to be metabolised by B. bifidum too, but also by B. crudilactis, a bovine origin strain. Fermentation of HMO or BMO by bifidobacteria can resul...

  19. Inhibition of Candida albicans biofilm formation and modulation of gene expression by probiotic cells and supernatant.

    Science.gov (United States)

    James, K M; MacDonald, K W; Chanyi, R M; Cadieux, P A; Burton, J P

    2016-04-01

    Oral candidiasis is a disease caused by opportunistic species of Candida that normally reside on human mucosal surfaces. The transition of Candida from budding yeast to filamentous hyphae allows for covalent attachment to oral epithelial cells, followed by biofilm formation, invasion and tissue damage. In this study, combinations of Lactobacillus plantarum SD5870, Lactobacillus helveticus CBS N116411 and Streptococcus salivarius DSM 14685 were assessed for their ability to inhibit the formation of and disrupt Candida albicans biofilms. Co-incubation with probiotic supernatants under hyphae-inducing conditions reduced C. albicans biofilm formation by >75 % in all treatment groups. Likewise, combinations of live probiotics reduced biofilm formation of C. albicans by >67 %. When live probiotics or their supernatants were overlaid on preformed C. albicans biofilms, biofilm size was reduced by >63 and >65 % respectively. Quantitative real-time PCR results indicated that the combined supernatants of SD5870 and CBS N116411 significantly reduced the expression of several C. albicans genes involved in the yeast-hyphae transition: ALS3 (adhesin/invasin) by 70 % (P biofilm formation) by >99 % (P formation of and removing preformed C. albicans biofilms. Our novel results point to the downregulation of several Candida genes critical to the yeast-hyphae transition, biofilm formation, tissue invasion and cellular damage.

  20. The Effect of Anaerobic and Aerobic Fish Sludge Supernatant on Hydroponic Lettuce

    Directory of Open Access Journals (Sweden)

    Simon Goddek

    2016-06-01

    Full Text Available The mobilization of nutrients from fish sludge (i.e., feces and uneaten feed plays a key role in optimizing the resource utilization and thus in improving the sustainability of aquaponic systems. While several studies have documented the aerobic and anaerobic digestion performance of aquaculture sludge, the impact of the digestate on plant growth has yet to be understood. The present study examines the impact of either an aerobic or an anaerobic digestion effluent on lettuce plant growth, by enriching a mixture of aquaculture and tap water with supernatants from both aerobic and anaerobic batch reactors. The lettuce plants grown in the hydroponic system supplied with supernatant from an anaerobic reactor had significantly better performance with respect to weight gain than both, those in the system where supernatant from the aerobic reactor was added, as well as the control system. It can be hypothesized that this effect was caused by the presence of NH4+ as well as dissolved organic matter, plant growth promoting rhizobacteria and fungi, and humic acid, which are predominantly present in anaerobic effluents. This study should therefore be of value to researchers and practitioners wishing to further develop sludge remineralization in aquaponic systems.

  1. Migration stimulation factor (MStF), from murine B cells, constitutively produced by a T-B hybridoma.

    Science.gov (United States)

    Gauthier-Rahman, S; el-Gharbi, N; Bouvet, J P; Goodhart, M; Decreusefond, C; Couderc, J

    1992-10-01

    Hybridomas were established between murine spleen B cells and the thymoma cell line BW5147, to purify the migration stimulation factor (MStF), a molecule likely involved in immunosuppression. The parental B cells were from Lo/PHA mice previously shown to produce high levels of MStF after immunization by appropriate (tolerogenic) doses of ovalbumin. Among the positive clones, B9 was selected, since it produced high levels of MStF constitutively, and no immunoglobulin. This clone was shown to contain the genome of the B-cell fusion partner, since one of its L chain genes had undergone a VK-JK rearrangement. Isolation of MStF by size-exclusion chromatography showed 2 major peaks of activity, one of which eluted in a 20-kDa, almost protein-free fraction. This elution is unlikely to correspond to the true molecular mass, since MStF was found not to be a protein. Indeed, MStF was TCA-soluble, thermoresistant, highly hydrophobic and protease-resistant, but activity was abolished by neuraminidase digestion. The possibility of its being a small molecule transported by a protein carrier was also ruled out. These results suggest that MStF is a complex molecule containing both sialic residues and a lipid moiety. Experiments are planned to further investigate the chemical structure of this unusual B-cell factor.

  2. A flexible state-space approach for the modeling of metabolic networks II: advanced interrogation of hybridoma metabolism.

    Science.gov (United States)

    Baughman, Adam C; Sharfstein, Susan T; Martin, Lealon L

    2011-03-01

    Having previously introduced the mathematical framework of topological metabolic analysis (TMA) - a novel optimization-based technique for modeling metabolic networks of arbitrary size and complexity - we demonstrate how TMA facilitates unique methods of metabolic interrogation. With the aid of several hybridoma metabolic investigations as case-studies (Bonarius et al., 1995, 1996, 2001), we first establish that the TMA framework identifies biologically important aspects of the metabolic network under investigation. We also show that the use of a structured weighting approach within our objective provides a substantial modeling benefit over an unstructured, uniform, weighting approach. We then illustrate the strength of TAM as an advanced interrogation technique, first by using TMA to prove the existence of (and to quantitatively describe) multiple topologically distinct configurations of a metabolic network that each optimally model a given set of experimental observations. We further show that such alternate topologies are indistinguishable using existing stoichiometric modeling techniques, and we explain the biological significance of the topological variables appearing within our model. By leveraging the manner in which TMA implements metabolite inputs and outputs, we also show that metabolites whose possible metabolic fates are inadequately described by a given network reconstruction can be quickly identified. Lastly, we show how the use of the TMA aggregate objective function (AOF) permits the identification of modeling solutions that can simultaneously consider experimental observations, underlying biological motivations, or even purely engineering- or design-based goals.

  3. Platelet-released supernatant induces osteoblastic differentiation of human mesenchymal stem cells: potential role of BMP-2

    Directory of Open Access Journals (Sweden)

    M Alini

    2010-12-01

    Full Text Available Platelet-rich preparations have recently gained popularity in maxillofacial and dental surgery, but their beneficial effect is still under debate. Furthermore, very little is known about the effect of platelet preparations at the cellular level, and the underlying mechanisms. In this study, we tested the effect of platelet-released supernatant (PRS on human mesenchymal stem cell (MSC differentiation towards an osteoblastic phenotype in vitro. Cultures of MSC were supplemented with PRS and typical osteoblastic markers were assessed at up to 28 days post-confluence. PRS showed an osteoinductive effect on MSC, as shown by an increased expression of typical osteoblastic marker genes such as collagen Ialpha1, bone sialoprotein II, BMP-2 and MMP-13, as well as by increased 45Ca2+ incorporation. Our results suggest that the effect of PRS on human MSC could be at least partially mediated by BMP-2.Activated autologous PRS could therefore provide an alternative to agents like recombinant bone growth factors by increasing osteoblastic differentiation of bone precursor cells at bone repair sites, although further studies are needed to fully support our observations.

  4. Cell-free supernatants from probiotic Lactobacillus casei and Lactobacillus rhamnosus GG decrease colon cancer cell invasion in vitro.

    Science.gov (United States)

    Escamilla, Juanita; Lane, Michelle A; Maitin, Vatsala

    2012-08-01

    Probiotics have been shown to have a preventative role in colorectal carcinogenesis but research concerning their prophylactic potential in the later stages of colorectal cancer, specifically metastasis is limited. This study explored the potential of cell-free supernatants (CFS) from 2 probiotic Lactobacillus sp., Lactobacillus casei and Lactobacillus rhamnosus GG, to inhibit colon cancer cell invasion by influencing matrix metalloproteinase-9 (MMP-9) activity and levels of the tight junction protein zona occludens-1 (ZO-1) in cultured metastatic human colorectal carcinoma cells. HCT-116 cells were treated with CFS from L. casei, L. rhamnosus, or Bacteroides thetaiotaomicron (a gut commensal); or with uninoculated bacterial growth media. Treatment with CFS from both Lactobacillus sp. decreased colorectal cell invasion but treatment with CFS from B. thetaiotaomicron did not. CFS from both Lactobacillus sp. decreased MMP-9 and increased ZO-1 protein levels. L. rhamnosus CFS also lowered MMP-9 activity. To begin elucidating the secreted bacterial factor conveying these responses, Lactobacillus sp. CFS were fractionated into defined molecular weight ranges and cell invasion assessed. Fractionation revealed that the inhibitory activity was contained primarily in the >100 kDa and 50-100 kDa fractions, suggesting the inhibitory compound may be a macromolecule such as a protein, nucleic acid, or a polysaccharide.

  5. Lactobacillus rhamnosus GG supernatant enhance neonatal resistance to systemic Escherichia coli K1 infection by accelerating development of intestinal defense

    Science.gov (United States)

    He, Xiaolong; Zeng, Qing; Puthiyakunnon, Santhosh; Zeng, Zhijie; Yang, Weijun; Qiu, Jiawen; Du, Lei; Boddu, Swapna; Wu, Tongwei; Cai, Danxian; Huang, Sheng-He; Cao, Hong

    2017-01-01

    The objective of this study was to determine whether Lactobacillus rhamnosus GG culture supernatant (LCS) has a preventive effect against gut-derived systemic neonatal Escherichia coli (E. coli) K1 infection. The preventive effects were evaluated in human colonic carcinoma cell line Caco-2 and neonatal rat models. Our in vitro results showed that LCS could block adhesion, invasion and translocation of E. coli K1 to Caco-2 monolayer via up-regulating mucin production and maintaining intestinal integrity. In vivo experiments revealed that pre-treatment with LCS significantly decrease susceptibility of neonatal rats to oral E. coli K1 infection as reflected by reduced bacterial intestinal colonization, translocation, dissemination and systemic infections. Further, we found that LCS treated neonatal rats have higher intestinal expressions of Ki67, MUC2, ZO-1, IgA, mucin and lower barrier permeability than those in untreated rats. These results indicated that LCS could enhance neonatal resistance to systemic E. coli K1 infection via promoting maturation of neonatal intestinal defense. In conclusions, our findings suggested that LCS has a prophylactic effect against systemic E. coli K1 infection in neonates. Future studies aimed at identifying the specific active ingredients in LCS will be helpful in developing effective pharmacological strategies for preventing neonatal E. coli K1 infection. PMID:28262688

  6. An In-Vitro Investigation of the Antibacterial Effects of the Methanol and Aqueous Extracts and the Supernatant of the Algae Chlorella vulgaris CCATM 210-1 on Multiantibiotic-Resistant Staphylococcus aureus Isolates Causing Urinary Tract Infections

    Directory of Open Access Journals (Sweden)

    Senobar Asadi

    2016-04-01

    Full Text Available Algae contain compounds, some of which have antibacterial properties. For example, the antibiotic Chlorellin can be mentioned, which is extracted from Chlorella species. The methanol and aqueous extracts, and the supernatant of the algae Chlorella vulgaris CCATM- 210-1 were used in this study. After culturing the Algae and preparing the supernatant and extracts, the antibacterial effects of the extracts and supernatant of this algae against multidrugresistant (MDR Staphylococcus aureus isolates causing urinary tract infections were determined using the microdilution and checkerboard titration methods. In the microdilution method, first, the aqueous extract of this algae at a concentration of 0.72 ± 0.22 mg/ml could inhibit the growth of the bacteria being tested. Then, the methanol extract and the supernatant of this algae at concentrations of 0.98 ± 0.68 and 1 ± 0.26 mg/ml, respectively, inhibited the growth of the bacteria being studied. The sum of the FIC of the methanol and aqueous extracts of the algae Chlorella vulgaris CCATM 210-1 in this study, showed the indifference of these two extracts towards each other.

  7. An In-Vitro Investigation of the Antibacterial Effects of the Acetone and Ethanol Extracts and the Supernatant of the Algae Chlorella vulgaris CCATM- 210-1 on Some of the Gram-Negative Bacterial Foodborne Pathogens

    Directory of Open Access Journals (Sweden)

    Yasaman Asadi

    2016-09-01

    Full Text Available Algae are rich sources of amino acids, terpenoids, florentines, alkanes, halogenated ketones, steroidal compounds, fatty acids, and phenols that can be used in the pharmaceutical industry. The acetone and ethanol extracts, and the supernatant of the algae Chlorella vulgaris CCATM- 210-1, were used in this study. After culturing the Algae, and preparing the supernatant and extracts, the antibacterial effects of the extracts and supernatant of this algae against some of the gram-negative bacterial foodborne pathogens were determined using the well plate and agar disk diffusion methods. According to the study conducted, the acetone and ethanol extracts and the supernatant of the algae Chlorella vulgaris CCATM- 210-1 showed acceptable antibacterial properties against some of the bacteria being studied, at concentrations of 1.25, 2.5, 5, and 10 mg/ml in the well plate and agar disk diffusion methods. The maximum inhibition zone diameters in the well plate and agar disk diffusion methods, were 36.8 and 35.4 mm, respectively, which were related to the bacterium Proteus mirabilis PTCC 1776 and were observed at a 10 mg/ml concentration of the acetone extract. By comparing the mean inhibition zone diameters among the acetone and ethanol extracts and a mixture of the two extracts using the well plate method, it can be concluded that the acetone and ethanol extracts of Chlorella vulgaris CCATM- 210-1 are indifferent to each other. This analysis was also true of the agar disk diffusion method.

  8. Rheological properties in supernatant of peach gum from almond(Prunus dulcis)

    Institute of Scientific and Technical Information of China (English)

    王森; 谢碧霞; 钟秋平; 杜红岩

    2008-01-01

    The rheological properties in the supernatant of peach gum from Prunnus dulcis were discussed in order to provide more scientific technical parameters and references for developing peach gum as a kind of medicinal gum.The rheological properties in the supernatant of peach gum were comparatively studied in different material ratios,temperatures,shaking times,pH values and salinities.The results show that,1) the mathematical model of shear rate with material ratio and shear stress is Y=0.069X12+0.035X2 -1.174,R2=0.942;2) the mathematical model of shear rate with temperature and shear stress is Y=4.936X12+0.023 2X2-1.688,R2=0.937;3) the mathematical model of shear rate with shaking time and shear stress is Y=0.005 192 X13-0.140 73X12+1.249 045X1+ 0.036 546 X2-3.644 29,R2=0.954 3;4) the effects of pH value on the rheological properties in the supernatant of peach gum are comparatively complicated with a varying range of 3-11 and the shear rate shows a change trend of saddle model;5) the mathematical model of shear rate with the concentration of NaCl and shear stress is Y=-0.037 44X1+0.012 93 X2,R2=0.998;6) the mathematical model of shear rate with the concentration of CaCl2 and shear stress is Y=0.025 789X1+0.016 19X2,R2 =0.999;and 7) the mathematical model of shear rate with the concentration of sorbic acid potassium and shear stress is Y=0.079 5X1+0.017 3X2,R2=0.998.The results show that the material ratio,temperature,shaking time,pH value significantly affect the rheological properties in the supernatant of peach gum,and the concentrations of NaCl and CaCl2 also significantly affect the rheological properties expect the concentration of sorbic acid potassium.

  9. Study of gelatin-agar intermolecular aggregates in the supernatant of its coacervate.

    Science.gov (United States)

    Singh, S Santinath; Bohidar, H B; Bandyopadhyay, S

    2007-05-15

    Intermolecular interaction leading to formation of aggregates between gelatin, a polyampholyte, and agar, a polysaccharide was studied in the supernatant of the complex coacervate formed by these biopolymers. Electrophoresis, laser light scattering and viscometry data were used to determine the interaction and the physical structure of these intermolecular soluble complexes by modeling these to be prolate ellipsoids of revolution (rod-like structures with well defined axial ratio and Perrin's factor). Solution ionic strength was found to reduce the axial ratio of these complexes implying the presence of screened polarization-induced electrostatic interaction between the two biopolymers.

  10. Treatment test of supernatant from sewage sludge by irradiation of high energy electron beams under supersaturation with oxygen

    Energy Technology Data Exchange (ETDEWEB)

    Hosono, Masakazu; Arai, Hidehiko (Japan Atomic Energy Research Inst., Takasaki, Gunma (Japan). Takasaki Radiation Chemistry Research Establishment); Aizawa, Masaki; Shimooka, Toshio; Yamamoto, Ichiro; Shimizu, Ken; Sugiyama, Masashi.

    1993-02-01

    Supernatant comes from dewaterization of sewage sludge, and contains biologically nondegradable organics. Therefore, it is hard to be treated by conventional activated sludge method. The development of a new technology is required to decrease the chemical oxygen demand (COD) effectively below 30 mg/l. Irradiation of high energy electron beams can convert nondegradable organics in water into substances which are biodegradable. However, sufficient dissolved oxygen in water is needed to induce oxidation effectively. In the present study, the treatment of supernatant was studied using an apparatus which can be irradiated by high intensity electron beams in flow system under supersaturation with oxygen by pressurization up to 3 atms. The dependence of oxygen concentration on the reduction in absorbance at 230 nm of azo dye (Acid Red 265) aqueous solution was examined, and it was clarified that sufficient oxygen was supplied in the solution up to about 14 kGy under 3 atms of oxygen. Radiation treatment of supernatant which came from the leather works was carried out using the above apparatus. However, as this supernatant contained high concentration of nitrite, the nitrite was removed by limited aeration activated sludge method. By this pretreatment, COD was reduced from 200 mg/l to 53 mg/l. Then, the biodegradability of supernatant irradiated under supersaturation with oxygen was examined. The final COD of the supernatant was reduced below 30 mg/l by the combined method of irradiation of 7 kGy and biological treatment. (author).

  11. Evolution of composition of dairy manure supernatant in a controlled dung pit.

    Science.gov (United States)

    Rico, C; García, H; Rico, J L; Fernández, J; Renedo, J

    2009-12-01

    Anaerobic conversion of dairy manure into biogas is an attractive way of managing this waste. It is well known that the hydrolysis of large molecules into small, directly biodegradable ones is the rate limiting step of the overall anaerobic process. The present work studies the development of the hydrolytic and acidogenic stages of dairy manure with different solid concentrations (40, 60 and 80 g VS/L) at ambient temperature (20 degrees C). The purpose was to determine the operational conditions that provide a liquid fraction with a high soluble chemical oxygen demand (COD) and a high volatile fatty acids (VFA) content in manure before the methanogenic stage starts up. At 20 degrees C, the evolution of the studied parameters showed that, in a controlled plug-flow dung pit, the hydrolytic and acidogenic stages progressed moderately in a continuous way during the 25 days that the experimentation lasted, whereas no methanization was observed. Supernatant COD and VFA concentrations increased 30% and 107%, respectively, for the 60 g VS/L samples. Manure was also operated at 35 degrees C with a similar increase in supernatant COD but a higher increase in VFA, 154%. For both operational temperatures, the predominant VFAs were, in this order, acetic, propionic and butyric acids. During the operation at 35 degrees C, the methanogenic stage started between days 20 and 25 for the samples with lower solids content, i.e. 40 and 60 g VS/L.

  12. cultural

    Directory of Open Access Journals (Sweden)

    Irene Kreutz

    2006-01-01

    Full Text Available Es un estudio cualitativo que adoptó como referencial teorico-motodológico la antropología y la etnografía. Presenta las experiencias vivenciadas por mujeres de una comunidad en el proceso salud-enfermedad, con el objetivo de comprender los determinantes sócio-culturales e históricos de las prácticas de prevención y tratamiento adoptados por el grupo cultural por medio de la entrevista semi-estructurada. Los temas que emergieron fueron: la relación entre la alimentación y lo proceso salud-enfermedad, las relaciones con el sistema de salud oficial y el proceso salud-enfermedad y lo sobrenatural. Los dados revelaron que los moradores de la comunidad investigada tienen un modo particular de explicar sus procedimientos terapéuticos. Consideramos que es papel de los profesionales de la salud en sus prácticas, la adopción de abordajes o enfoques que consideren al individuo en su dimensión sócio-cultural e histórica, considerando la enorme diversidad cultural en nuestro país.

  13. Peroxynitrite decomposition catalyst prevents apoptotic cell death in a human astrocytoma cell line incubated with supernatants of HIV-infected macrophages

    Directory of Open Access Journals (Sweden)

    Perno Carlo

    2002-09-01

    Full Text Available Abstract Background Oxidative stress has shown to contribute in the mechanisms underlying apoptotic cell death occuring in AIDS-dementia complex. Here we investigated the role of peroxynitrite in apoptosis occurring in astroglial cells incubated with supernatants of HIV-infected human primary macrophages (M/M. Results Flow cytometric analysis (FACS of human cultured astrocytes shortly incubated with HIV-1-infected M/M supernatants showed apoptotic cell death, an effect accompanied by pronounced staining for nitrotyrosine (footprint of peroxynitrite and by abnormal formation of malondialdehyde (MDA. Pretreatment of astrocytes with the peroxynitrite decomposition catalyst FeTMPS antagonized HIV-related astrocytic apoptosis, MDA formation and nitrotyrosine staining. Conclusions Taken together, our results suggest that inibition of peroxynitrite leads to protection against peroxidative stress accompanying HIV-related apoptosis of astrocytes. Overall results support the role of peroxynitrite in HIV-related programmed death of astrocytes and suggest the use of peroxynitrite decomposition catalyst to counteract HIV-1-related neurological disorders.

  14. Cloning and Characterization of a Hybridoma Secreting a 4-(Methylnitrosamino-1-(3-pyridyl-1-butanone (NNK-Specific Monoclonal Antibody and Recombinant F(ab

    Directory of Open Access Journals (Sweden)

    Lawrence K. Silbart

    2013-03-01

    Full Text Available Smokeless tobacco products have been associated with increased risks of oro-pharyngeal cancers, due in part to the presence of tobacco-specific nitrosamines (TSNAs such as 4-(methylnitrosamino-1-(3-pyridyl-1-butanone (NNK. These potent carcinogens are formed during tobacco curing and as a result of direct nitrosation reactions that occur in the oral cavity. In the current work we describe the isolation and characterization of a hybridoma secreting a high-affinity, NNK-specific monoclonal antibody. A structurally-related benzoyl derivative was synthesized to facilitate coupling to NNK-carrier proteins, which were characterized for the presence of the N-nitroso group using the Griess reaction, and used to immunize BALB/c mice. Splenocytes from mice bearing NNK-specific antibodies were used to create hybridomas. Out of four, one was selected for subcloning and characterization. Approximately 99% of the monoclonal antibodies from this clone were competitively displaced from plate-bound NNKB conjugates in the presence of free NNK. The affinity of the monoclonal antibody to the NNKB conjugates was Kd = 2.93 nM as determined by surface plasmon resonance. Free nicotine was a poor competitor for the NNKB binding site. The heavy and light chain antibody F(ab fragments were cloned, sequenced and inserted in tandem into an expression vector, with an FMDV Furin 2A cleavage site between them. Expression in HEK 293 cells revealed a functional F(ab with similar binding features to that of the parent hybridoma. This study lays the groundwork for synthesizing transgenic tobacco that expresses carcinogen-sequestration properties, thereby rendering it less harmful to consumers.

  15. Compressibility of the fouling layer formed by membrane bioreactor sludge and supernatant

    DEFF Research Database (Denmark)

    Jørgensen, Mads Koustrup; Poorasgari, Eskandar; Christensen, Morten Lykkegaard

    the pressure range 0.08 - 0.13 bar. The fouling layer resistance increased while fouling layer compression was reversible. Conversely, the supernatant filtrations showed that the gel layer is compressible within the range 0.1 - 2 bar applied pressure. Calculated resistance of the gel layer indicated....... Compressibility of the gel layer was studied in a dead-end filtration system, whereas the compressibility of a fouling layer formed by MBR sludge was studied in a submerged system hollow sheet membrane by TMP stepping. It was shown that the fouling layer formed by the MBR sludge was highly compressible within...... that higher pressure causes higher change in resistance. The results of the gel compression study showed a lower compressibility of gel layers than the fouling layer formed by sludge flocs. At pressures between 0.1 to 0.5 bar there was no significant change in specific cake resistance with pressure compared...

  16. Pseudolinkage of the duplicate loci for supernatant aspartate aminotransferase in brook trout, Salvelinus fontinalis.

    Science.gov (United States)

    Wright, J E; May, B; Stoneking, M; Lee, G M

    1980-01-01

    Electrophoretic variation involving three alleles is described for the duplicated loci for supernatant aspartate aminotransferase (AAT-1,2), from muscle extracts of brook trout. Both loci exhibit largely disomic inheritance. Exceptional progeny types are proposed to be the result of a form of tetrasomic inheritance. Nonrandom segregation was found among the progeny of males doubly heterozygous for AAT markers; where so-called linkage phase was known, this nonrandom assortment was shown to be pseudolinkage (78.9 percent recombination). Analyses of joint segregation of triply heterozygous males for the AAT-(1,2) loci and for the single alpha glycerophosphate dehydrogenase locus (AGP-1) revealed true linkage of AGP-1 with one AAT locus (mean r = 11 percent), but pseudolinkage with the other AAT locus (r = 74 percent). Intraindividual variation for homoeologous multivalent pairing of two acrocentric with two metacentric chromosomes in males, but with bivalent pairing in females, is proposed to account for pseudolinkage and for the tetrasomically inherited types.

  17. Host markers in Quantiferon supernatants differentiate active TB from latent TB infection: preliminary report

    Directory of Open Access Journals (Sweden)

    Walzl Gerhard

    2009-05-01

    Full Text Available Abstract Background Interferon gamma release assays, including the QuantiFERON® TB Gold In Tube (QFT have been shown to be accurate in diagnosing Mycobacterium tuberculosis infection. These assays however, do not discriminate between latent TB infection (LTBI and active TB disease. Methods We recruited twenty-three pulmonary TB patients and 34 household contacts from Cape Town, South Africa and performed the QFT test. To investigate the ability of new host markers to differentiate between LTBI and active TB, levels of 29 biomarkers in QFT supernatants were evaluated using a Luminex multiplex cytokine assay. Results Eight out of 29 biomarkers distinguished active TB from LTBI in a pilot study. Baseline levels of epidermal growth factor (EGF soluble CD40 ligand (sCD40L, antigen stimulated levels of EGF, and the background corrected antigen stimulated levels of EGF and macrophage inflammatory protein (MIP-1β were the most informative single markers for differentiation between TB disease and LTBI, with AUCs of 0.88, 0.84, 0.87, 0.90 and 0.79 respectively. The combination of EGF and MIP-1β predicted 96% of active TB cases and 92% of LTBIs. Combinations between EGF, sCD40L, VEGF, TGF-α and IL-1α also showed potential to differentiate between TB infection states. EGF, VEGF, TGF-α and sCD40L levels were higher in TB patients. Conclusion These preliminary data suggest that active TB may be accurately differentiated from LTBI utilizing adaptations of the commercial QFT test that includes measurement of EGF, sCD40L, MIP-1β, VEGF, TGF-α or IL-1α in supernatants from QFT assays. This approach holds promise for development as a rapid diagnostic test for active TB.

  18. Supernatant from a cloned helper T cell stimulates resting B cells to express transferrin and IL-2 receptors.

    Science.gov (United States)

    Diu, A; Leclercq, L; Dautry-Varsat, A; Theze, J

    1987-07-01

    We describe the properties of the supernatant from a murine cloned helper T cell (clone 52.3) which is able to polyclonally activate most resting B cells in the absence of any additional stimulus. We hypothesize that an activity which we call BCAF (B-cell-activating factor(s] exists in our supernatant which can activate resting B cells alone or in conjunction with other lymphokines. In the present report, we investigate changes in the surface antigen pattern induced on resting B cells by BCAF-containing supernatant. Analysis of the cells by flow cytometry shows that transferrin receptor and IL-2 receptor expression increase on a large fraction of B cells after 2 days of activation by the T-helper-cell clone supernatant. Monoclonal anti-transferrin receptor antibody inhibits cell division but does not affect blastogenesis, while IL-2 has no effect in our experimental system. Our present results confirm that BCAF-containing supernatants can act on most resting B cells and replace helper T cells in inducing B-cell activation and proliferation.

  19. 抗TPO单克隆抗体杂交瘤细胞系的建立%The preliminary determination of anti-TPO monoclonal antibody hybridoma cell line

    Institute of Scientific and Technical Information of China (English)

    于芳; 朱恒奇; 陈琳; 徐秀英; 黄培堂

    2001-01-01

    Bal b/c mice were immunized with TPO antigen from synthesized peptide, their spleen cells were fused with myeloma cells by polyethyleneglyco (PEG)making for hybridomas cell line. The rate of fusion was 91%. By making ELISA screening and subcloning, one hybridoma cell line was established.%用合成肽TPO作抗原,经腹腔免疫Bal b/c小鼠,鼠抗血清效价为1:1000,ELISA间接法测定的P/N值为3。取小鼠脾细胞与骨髓瘤细胞(SP2/0)在PEG作用下进行融合,细胞融合率达91%。通过ELISA筛选并经过3次亚克隆,获得了1株能分泌抗TPO抗体的单克隆杂交瘤细胞株,P/N值均高于8。

  20. Boildown Study on Supernatant Liquid Retrieved from AW-106 in December 2012

    Energy Technology Data Exchange (ETDEWEB)

    Page, Jason S. [Washington River Protection Solutions, LLC, Richland, WA (United States)

    2013-06-04

    This document reports the results of a boil down study using a composite created from supernatant liquid grab samples retrieved from tank 241-AW-I06 in December of 2012. The composite was made using predetermined volumes of the grab samples which accounted for layering of the supernatant liquid in the tank. The finished composite was a clear, yellow liquid containing no visible solids at hot cell ambient temperatures (24 - 27°C). The density of the test composite was measured in the hot cell immediately before the boildown study and was 1.266 g/mL at 27.1 °C. The boiling temperature of the composite was measured at three different pressures (40, 60, and 80 Torr) throughout the volume reduction, and the results show steadily increasing boiling temperatures with increasing volume reduction and no significant discontinuities. Moderate foaming was observed at the onset of the boildown. The foaming disappeared during the first reduction step, and minimal foaming was observed throughout the rest of the study. The bulk densities at 18.0 °C (D{sub Bulk}{sup 18 °C}) and quantities of settled and centrifuged solids were measured on samples of the boildown concentrates. Estimated values of the bulk densities at the 60-Torr boiling temperatures (D{sub Bulk}{sup 60 Torr}) were also calculated. Solids were first observed at boildown temperatures when the % VWR reached 39.3%. The quantity of solids in the composite quickly increased after this initial formation; the amount of centrifuged solids increased by 22% as the %WVR increased from 39.3 to 44.1 %. A small amount of solids did appear in the samples collected prior to the initial formation during the boildown. These solids precipitated while they sat at hot cell ambient temperature and in the 18. 0 °C water bath. Analysis of boil down test samples indicated that natrophosphate (Na7{sub 3}F(PO{sub 4}){sub 2}{centerdot} 19 H{sub 2}O) and kogarkoite (Na3FS04) accounted for a majority of the initial solids (~80% of the

  1. Experimental prestorage filtration removes antibodies and decreases lipids in RBC supernatants mitigating TRALI in vivo.

    Science.gov (United States)

    Silliman, Christopher C; Kelher, Marguerite R; Khan, Samina Y; LaSarre, Monica; West, F Bernadette; Land, Kevin J; Mish, Barbara; Ceriano, Linda; Sowemimo-Coker, Samuel

    2014-05-29

    Transfusion-related acute lung injury (TRALI) remains a significant cause of transfusion-related mortality with red cell transfusion. We hypothesize that prestorage filtration may reduce proinflammatory activity in the red blood cell (RBC) supernatant and prevent TRALI. Filters were manufactured for both small volumes and RBC units. Plasma containing antibodies to human lymphocyte antigen (HLA)-A2 or human neutrophil antigen (HNA)-3a was filtered, and immunoglobulins and specific HNA-3a and HLA-2a neutrophil (PMN) priming activity were measured. Antibodies to OX27 were added to plasma, and filtration was evaluated in a 2-event animal model of TRALI. RBC units from 31 donors known to have antibodies against HLA antigens and from 16 antibody-negative controls were filtered. Furthermore, 4 RBC units were drawn and underwent standard leukoreduction. Immunoglobulins, HLA antibodies, PMN priming activity, and the ability to induce TRALI in an animal model were measured. Small-volume filtration of plasma removed >96% of IgG, antibodies to HLA-A2 and HNA-3a, and their respective priming activity, as well as mitigating antibody-mediated in vivo TRALI. In RBC units, experimental filtration removed antibodies to HLA antigens and inhibited the accumulation of lipid priming activity and lipid-mediated TRALI. We conclude that filtration removes proinflammatory activity and the ability to induce TRALI from RBCs and may represent a TRALI mitigation step.

  2. Multistage aqueous two-phase extraction of a monoclonal antibody from cell supernatant.

    Science.gov (United States)

    Muendges, Jan; Zalesko, Alexej; Górak, Andrzej; Zeiner, Tim

    2015-01-01

    This article presents results of continuous multistage aqueous two-phase extraction of an immunoglobulin G1 from cell supernatant in a mixer-settler unit. An aqueous two-phase system consisting of polyethylene glycol 2000, phosphate salt, and water was applied without and with sodium chloride (NaCl). Influences of different parameters such as throughput, phase ratio, and stage number on the extraction performance were analyzed. For systems without NaCl, the extraction was carried out as a washing step. An increase of stage number from one to five stages enabled to increase the immunoglobulin G1 purity from 11.8 to 32.6% at a yield of nearly 90%. Furthermore, a reduction of product phase volume due to a higher phase ratio led to an increase of purity from 20.8 to 29.6% in a three-stage countercurrent extraction. For experiments with NaCl moderate partitioning conditions were adjusted by adding 8 wt% NaCl. In that case, the extraction was carried out as a stripping step.

  3. Discovery of Novel Secreted Virulence Factors from Salmonella enterica Serovar Typhimurium by Proteomic Analysis of Culture Supernatants

    Energy Technology Data Exchange (ETDEWEB)

    Niemann, George; Brown, Roslyn N.; Gustin, Jean K.; Stufkens, Afke; Shaikh-Kidwai, Afshan S.; Li, Jie; McDermott, Jason E.; Brewer, Heather M.; Schepmoes, Athena A.; Smith, Richard D.; Adkins, Joshua N.; Heffron, Fred

    2011-01-01

    The intracellular pathogen Salmonella enterica serovar Typhimurium is a leading cause of acute gastroenteritis in the world. This pathogen has two type-III secretion systems (TTSS) necessary for virulence that are encoded in Salmonella pathogenicity islands 1 and 2 (SPI-1 and SPI-2) and are expressed during extracellular or intracellular infectious states, respectively, to deliver virulence factors (effectors) to the host cell cytoplasm. While many have been identified and at least partially characterized, the full repertoire of effectors has not been catalogued. In this mass spectrometry-based proteomics study, we identified effector proteins secreted under minimal acidic medium growth conditions that induced the SPI-2 TTSS and its effectors, and compared the secretome from the parent strain to the secretome from strains missing either essential (SsaK) or regulatory components (SsaL) of the SPI-2 secretion apparatus. We identified 75% of the known TTSS effector repertoire. Excluding translocon components, 95% of the known effectors were biased for identification in the ssaL mutant background, which demonstrated that SsaL regulates SPI-2 type III secretion. To confirm secretion to animal cells, we made translational fusions of several of the best candidates to the calmodulin-dependent adenylate cyclase of Bordetella pertussis and assayed cAMP levels of infected J774 macrophage-like cells. From these infected cells we identified six new TTSS effectors and two others that are secreted independent of TTSS. Our results substantiate reports of additional secretion systems encoded by Salmonella other than TTSS.

  4. Separation of active laccases from Pleurotus sapidus culture supernatant using aqueous two-phase systems in centrifugal partition chromatography.

    Science.gov (United States)

    Schwienheer, C; Prinz, A; Zeiner, T; Merz, J

    2015-10-01

    For the production of bio active compounds, e.g., active enzymes or antibodies, a conserved purification process with a minimum loss of active compounds is necessary. In centrifugal partition chromatography (CPC), the separation effect is based on the different distribution of the components to be separated between two immiscible liquid phases. Thereby, one liquid phase is kept stationary in chambers by a centrifugal field and the mobile phase is pumped through via connecting ducts. Aqueous two phase systems (ATPS) are known to provide benign conditions for biochemical products and seem to be promising when used in CPC for purification tasks. However, it is not known if active biochemical compounds can "survive" the conditions in a CPC where strong shear forces can occur due to the two-phasic flow under centrifugal forces. Therefore, this aspect has been faced within this study by the separation of active laccases from a fermentation broth of Pleurotus sapidus. After selecting a suitable ATPS and operating conditions, the activity yield was calculated and the preservation of the active enzymes could be observed. Therefore, CPC could be shown as potentially suitable for the purification of bio-active compounds.

  5. Discovery of novel secreted virulence factors from Salmonella enterica serovar Typhimurium by proteomic analysis of culture supernatants.

    Science.gov (United States)

    Niemann, George S; Brown, Roslyn N; Gustin, Jean K; Stufkens, Afke; Shaikh-Kidwai, Afshan S; Li, Jie; McDermott, Jason E; Brewer, Heather M; Schepmoes, Athena; Smith, Richard D; Adkins, Joshua N; Heffron, Fred

    2011-01-01

    Salmonella enterica serovar Typhimurium is a leading cause of acute gastroenteritis throughout the world. This pathogen has two type III secretion systems (TTSS) encoded in Salmonella pathogenicity islands 1 and 2 (SPI-1 and SPI-2) that deliver virulence factors (effectors) to the host cell cytoplasm and are required for virulence. While many effectors have been identified and at least partially characterized, the full repertoire of effectors has not been catalogued. In this proteomic study, we identified effector proteins secreted into defined minimal medium designed to induce expression of the SPI-2 TTSS and its effectors. We compared the secretomes of the parent strain to those of strains missing essential (ssaK::cat) or regulatory (ΔssaL) components of the SPI-2 TTSS. We identified 20 known SPI-2 effectors. Excluding the translocon components SseBCD, all SPI-2 effectors were biased for identification in the ΔssaL mutant, substantiating the regulatory role of SsaL in TTS. To identify novel effector proteins, we coupled our secretome data with a machine learning algorithm (SIEVE, SVM-based identification and evaluation of virulence effectors) and selected 12 candidate proteins for further characterization. Using CyaA' reporter fusions, we identified six novel type III effectors and two additional proteins that were secreted into J774 macrophages independently of a TTSS. To assess their roles in virulence, we constructed nonpolar deletions and performed a competitive index analysis from intraperitoneally infected 129/SvJ mice. Six mutants were significantly attenuated for spleen colonization. Our results also suggest that non-type III secretion mechanisms are required for full Salmonella virulence.

  6. Reverse transcriptase activity in chicken embryo fibroblast culture supernatants is associated with particles containing endogenous avian retrovirus EAV-0 RNA.

    OpenAIRE

    Weissmahr, R N; Schüpbach, J; Böni, J

    1997-01-01

    We have recently shown that live attenuated virus vaccines produced on chicken-derived cells contain low levels of particle-associated reverse transcriptase (RT). In both virus and corresponding control harvests produced on chicken embryo fibroblasts, these activities were present at significantly higher concentrations than in the vaccines. In order to identify the putative retrovirus sequence responsible for this activity, a novel method for the selective PCR amplification of particle-associ...

  7. Detecting pM concentrations of prostaglandins in cell culture supernatants by capillary SCX-LC-MS/MS

    DEFF Research Database (Denmark)

    Dahl, Sandra Rinne; Kleiveland, Charlotte Ramstad; Kassem, Moustapha

    2008-01-01

    A highly sensitive, improved online strong cation exchange (SCX)--RP capillary liquid chromatographic (cLC) method with IT mass spectrometric (IT-MS/MS) detection for the simultaneous determination of prostaglandin (PG)A(1), PGD(2), PGE(1), PGE(2), PGF(2alpha), 8-iso-(8i)PGF(2alpha), 6-keto-(6k)P...

  8. Distinct galactofuranose antigens in the cell wall and culture supernatants as a means to differentiate Fusarium from AspergillusspeciesAnnegret

    DEFF Research Database (Denmark)

    Wiedemann, Annegret; Kakoschke, Tamara Katharina; Speth, Cornelia

    2016-01-01

    tDetection of carbohydrate antigens is an important means for diagnosis of invasive fungal infections. Fordiagnosis of systemic Aspergillus infections, galactomannan is commonly used, the core antigenic struc-ture of which consists of chains of several galactofuranose moieties. In this study, we ...

  9. Biological nutrients removal from the supernatant originating from the anaerobic digestion of the organic fraction of municipal solid waste.

    Science.gov (United States)

    Malamis, S; Katsou, E; Di Fabio, S; Bolzonella, D; Fatone, F

    2014-09-01

    This study critically evaluates the biological processes and techniques applied to remove nitrogen and phosphorus from the anaerobic supernatant produced from the treatment of the organic fraction of municipal solid waste (OFMSW) and from its co-digestion with other biodegradable organic waste (BOW) streams. The wide application of anaerobic digestion for the treatment of several organic waste streams results in the production of high quantities of anaerobic effluents. Such effluents are characterized by high nutrient content, because organic and particulate nitrogen and phosphorus are hydrolyzed in the anaerobic digestion process. Consequently, adequate post-treatment is required in order to comply with the existing land application and discharge legislation in the European Union countries. This may include physicochemical and biological processes, with the latter being more advantageous due to their lower cost. Nitrogen removal is accomplished through the conventional nitrification/denitrification, nitritation/denitritation and the complete autotrophic nitrogen removal process; the latter is accomplished by nitritation coupled with the anoxic ammonium oxidation process. As anaerobic digestion effluents are characterized by low COD/TKN ratio, conventional denitrification/nitrification is not an attractive option; short-cut nitrogen removal processes are more promising. Both suspended and attached growth processes have been employed to treat the anaerobic supernatant. Specifically, the sequencing batch reactor, the membrane bioreactor, the conventional activated sludge and the moving bed biofilm reactor processes have been investigated. Physicochemical phosphorus removal via struvite precipitation has been extensively examined. Enhanced biological phosphorus removal from the anaerobic supernatant can take place through the sequencing anaerobic/aerobic process. More recently, denitrifying phosphorus removal via nitrite or nitrate has been explored. The removal of

  10. Upflow anaerobic sludge blanket (UASB) treatment of supernatant of cow manure by thermal pre-treatment.

    Science.gov (United States)

    Yoneyama, Y; Nishii, A; Nishimoto, M; Yamada, N; Suzuki, T

    2006-01-01

    Upflow anaerobic sludge blanket (UASB) methane fermentation treatment of cow manure that was subjected to screw pressing, thermal treatment and subsequent solid-liquid separation was studied. Conducting batch scale tests at temperatures between 140 and 180 degrees C, the optimal temperature for sludge settling and the color suppression was found to be between 160-170 degrees C. UASB treatment was carried out with a supernatant obtained from the thermal treatment at the optimal conditions (170 degrees C for 30 minutes) and polymer-dosed solid-liquid separation. In the UASB treatment with a COD(Cr) loading of 11.7 kg/m3/d and water temperature of 32.2 degrees C, the COD(Cr) level dropped from 16,360 mg/L in raw water to 3,940 mg/L in treated water (COD(Cr), removal rate of 75.9%), and the methane production rate per COD(Cr) was 0.187 Nm3/kg. Using wastewater thermal-treated at the optimal conditions, also a methane fermentation treatment with a continuously stirred tank reactor (CSTR) was conducted (COD(Cr) in raw water: 38,000 mg/L, hydraulic retention time (HRT): 20 days, 35 degrees C). At the COD(Cr) loading of 1.9 kg/m3/d, the methane production rate per COD(Cr), was 0.153 Nm3/kg. This result shows that UASB treatment using thermal pre-treatment provides a COD(Cr), loading of four times or more and a methane production rate of 1.3 times higher than the CSTR treatment.

  11. Boildown Study on Supernatant Liquid Retrieved from AP-107 in May 2010

    Energy Technology Data Exchange (ETDEWEB)

    Callaway, W. S. [Washington River Protection Solutions, LLC, Richland, WA (United States); Page, J. S. [Washington River Protection Solutions, LLC, Richland, WA (United States)

    2013-02-12

    A boildown study was completed on a composite prepared from supernatant liquid grab samples retrieved from tank 241-AP-107 in May of 2010. The composite was a clear, yellow liquid containing no visible solids at hot cell ambient temperatures (25-27 °C). The density of the test composite was 1.216 g/mL at 26.8 °C. The boiling temperature curves generated at three reduced pressures—40-, 60-, and 80 Torr—displayed steadily increasing boiling temperatures with increasing volume reduction with no significant discontinuities. Only minimal foaming was observed after the volume reduction proceeded beyond 50 %WVR (percent waste volume reduction). The bulk densities (D{sub Bulk}{sup 18 °C}) and quantities of settled and centrifuged solids present were measured on samples of the boildown concentrates that were kept at 18 °C for 7-8 days. Estimated values of the bulk densities of the concentrates at 60-Torr boiling temperatures (D{sub Bulk}{sup 60 Torr}) were also calculated. Solids were observed in all boildown concentrates at process temperatures, at hot cell ambient temperatures (25-27 °C), and at 18 °C. The quantity of solids found in the cooled concentrates increased slowly through 50.2 %WVR. The quantity of solids found in concentrates after 54.0 %WVR was noticeably greater. Beyond 54.0 %WVR, the quantity of solids found in cooled concentrates increased dramatically. Analysis of boildown test samples indicated that sodium oxalate and sodium carbonate solids form in cooled concentrates after volume reduction of 8.4 %WVR or less. The major contributors to the large increase in the quantity of solids found in concentrates after 54 %WVR were sodium nitrate and sodium carbonate.

  12. Anti-adherence potential of Enterococcus durans cells and its cell-free supernatant on plastic and stainless steel against foodborne pathogens.

    Science.gov (United States)

    Amel, Ait Meddour; Farida, Bendali; Djamila, Sadoun

    2015-07-01

    It is demonstrated that numerous bacteria are able to attach to surfaces of equipment used for food handling or processing. In this study, a strain of Enterococcus durans, originally isolated from a milking machine surface, was firstly studied for its biofilm formation potential on plastic and stainless steel supports. The strain was found to be a biofilm producer either at 25, 30 or 37 °C on polystyrene microtitre plates, with a best adherence level observed at 25 °C. En. durans showed a strong adhesion to stainless steel AISI-304. Antibacterial and anti-adherence activities of En. durans were tested against four foodborne pathogens (Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27853 and Listeria innocua CLIP 74915) which were shown as biofilm producers on both plastic and stainless steel. En. durans cells and cell-free culture supernatant showed a significant (P < 0.05) inhibition potential of the pathogens either on solid media or in broth co-cultures. Characterization of the antibacterial substances indicated their proteinaceous nature which assigned them most probably to bacteriocins group.

  13. Protocol for Identifying the Presence of and Understanding the Nature of Soluble, Non-pertechnetate Technetium in Hanford Tank Supernatants

    Energy Technology Data Exchange (ETDEWEB)

    Rapko, Brian M.

    2014-02-27

    The objective of this report is to propose a method to evaluate the presence and extent of soluble, non-pertechnetate Tc in Hanford tank supernatants as well as methods that might be used to gain insight as to the nature of the specie(s) that make up this fraction. This study will then provide a recommendation as to the preferred approach for identifying and quantifying the presence of Hanford tank supernatant-soluble, non-pertechnetate, technetium. The recommendation will also describe an approach to address the issue of whether inductively coupled plasma mass spectrometry (ICP-MS) analysis, which is useful as a monitoring tool for Tc, may be confounded by the presence of other mass 99 species.

  14. Laboratory bioassay for assessing the effects of sludge supernatant on plant growth and vesicular-arbuscular mycorrhiza formation

    Energy Technology Data Exchange (ETDEWEB)

    Bohn, K.S.; Liberta, A.E.

    1982-12-01

    A laboratory bioassay is described for assessing the effects of sludge supernatant on juvenile corn growth and the ability of vesicular-arbuscular (VA) mycorrhizal fungi, indigenous to coal spoil, to form mycorrhizae. The bioassay demonstrated that application rates can be identified that have the potential to promote increased plant dry weight without suppressing the formation of VA mycorrhizae in a plant's root system.

  15. Facile purification of Escherichia coli expressed tag-free recombinant human tumor necrosis factor alpha from supernatant.

    Science.gov (United States)

    Zhang, Chun; Liu, Yongdong; Zhao, Dawei; Li, Xiunan; Yu, Rong; Su, Zhiguo

    2014-03-01

    Fusing affinity tag at N-terminus was reported to decrease the biological activity of the recombinant human tumor necrosis factor alpha. Although preparation of tag-free rhTNF-α has already been achieved, the processes were yet laborious, especially in large scale. In this paper, tag-free rhTNF-α was almost equally synthesized by Escherichia coli in both soluble and insoluble forms. A two-step ion exchange chromatography, DEAE-Sepharose combined with CM-Sepharose, was developed to purify the soluble specie from supernatant after cell lysis. Native PAGE and HP-SEC showed the rhTNF-α extracted from supernatant existed in a homogeneous form. HP-SAX and SDS-PAGE analysis demonstrated the purity of the final fraction was over 98% with a very high recovery of 75%. Circular dichroism spectrum demonstrated that β-sheet structure was dominant and fluorescence analysis suggested no dramatic exposure of aromatic amino acid residues on the protein surface. Bioassay indicated that purified rhTNF-α was biologically active with a specific activity of approximately 2.0×10(7)U/mg. All these results suggested that this two-step ion exchange chromatography is efficient for preparation of biologically active tag-free rhTNF-α from supernatant.

  16. Cloning of scFv from hybridomas using a rational strategy: Application as a receptor to sensitive detection microcystin-LR in water.

    Science.gov (United States)

    Zhang, Xiuyuan; He, Kuo; Zhao, Ruiping; Wang, Lixia; Jin, Yandan

    2016-10-01

    Single chain variable fragment (scFv), containing of heavy and light chains (VH and VL) joined by a short peptide linker, has been used widely for immunodetection. Nevertheless, cloning functional variable genes is still a bottle neck for the scFv generation technology. Here, a rational strategy for cloning and selecting variable region genes from an anti-microcystin-LR hybridoma was devised, then the functional VH and VL genes were recloned and assembled to scFv using splicing overlap extension PCR. The resulting scFv gene was recombinantly expressed as a soluble scFv-alkaline phosphatase fusion protein (scFv-AP) by vector PLIP6/GN. Then an indirect competitive chemiluminescent enzyme immunoassay (ic-CLEIA) for detection of microcystin-LR was developed. The half-maximum inhibition concentrations (IC50) and limits of detection (LODs, IC15) were 0.81 ± 0.04 μgL(-1) and 0.13 ± 0.03 μgL(-1), respectively. With the mean coefficient of variation lowing 8%, the mean recovery in intra-assay and inter-assay were 100.06% and 96.46%, The proposed strategy should be useful for generation scFv in a rapid and simple way.

  17. Comparison of indirect immunofluorescence (IF) test with enzyme-linked immunosorbent assay (ELISA) in screening of hybridomas to very virulent Marek's disease virus.

    Science.gov (United States)

    Nakajima, K; Shibayama, T; Naito, M; Kurimura, T; Hirai, K

    1992-01-01

    For identifying virus-specific antigens of Marek's disease virus (MDV), monoclonal antibodies (MAbs) against strain Md5 of serotype 1, which is known to be a very virulent MDV (vvMDV), were isolated. Fifty-eight hybridoma clones that secreted MAbs against vvMDV were obtained. Of these MAbs, 36 gave positive reactions in an immunofluorescence (IF) test, and 22 gave positive reactions on enzyme-linked immunosorbent assay (ELISA). None of these MAbs gave positive reactions in both the IF test and ELISA. Of the MAbs that gave positive reactions in the IF test, 33 clones reacted with MDV1-specific epitopes, the other three reacting with MDV1-HVT intertypic epitopes. None of the clones reacted with MDV1-MDV2 intertypic epitopes. Three virus-specific polypeptides were identified by radioimmunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) or immunoblotting. These polypeptides were recognized by 12 MAbs giving positive reactions by IF, but by none of those giving positive reactions by ELISA. In addition, size heterogeneity of the MDV1-specific phosphorylated polypeptides in the MDV1 strains was shown using the MAbs against Md5.

  18. Diagnosing feline infectious peritonitis using the Sysmex XT-2000iV based on frozen supernatants from cavitary effusions.

    Science.gov (United States)

    Stranieri, Angelica; Paltrinieri, Saverio; Giordano, Alessia

    2017-02-01

    The delta total nucleated cells (ΔTNC) measurement with the Sysmex XT-2000iV (Sysmex Europe, Norderstedt, Germany) has high diagnostic accuracy on effusions in feline infectious peritonitis (FIP) cases, but the test can be performed only on fresh samples. We evaluated whether supernatants from effusions retain the ability to induce cell clumping and assessed the diagnostic accuracy of this modified ΔTNC method. Effusions were collected from FIP cats ( n = 19) and from cats with other diseases ( n = 15). ΔTNC was measured on fresh samples and on frozen-thawed supernatants after the addition of feline blood at 1:10 dilution. Diagnostic accuracy was assessed at the cutoffs of suggestive of FIP (ΔTNC = 1.7) and consistent with FIP (ΔTNC = 3.4). The influence of the protein content, number of added cells, and magnitude of dilution were also investigated. Specificity and positive predictive value were 100% for both the methods. Sensitivity and negative predictive value were higher for the modified ΔTNC (84.2% and 83.3%, respectively, at the cutoff of 1.7; 78.9% and 78.9%, respectively, at the cutoff of 3.4) than for the ΔTNC on fresh samples (78.6% and 81.3%, respectively, at the cutoff of 1.7; 57.1% and 68.4%, respectively, at the cutoff of 3.4). Protein content, total cell count of the added blood, and magnitude of dilutions did not influence the results. Supernatants of frozen effusions from FIP cats retain the ability to induce cell clumping, thus the modified ΔTNC measurement is a reliable tool to diagnose FIP on samples that cannot be analyzed immediately.

  19. Requirement of Simultaneous Assessment of Crystal- and Supernatant-Related Entomotoxic Activities of Bacillus thuringiensis Strains for Biocontrol-Product Development

    Directory of Open Access Journals (Sweden)

    Ronaldo Costa Argôlo-Filho

    2014-05-01

    Full Text Available Bioinsecticides with lower concentrations of endospores/crystals and without loss of efficiency are economically advantageous for pest biocontrol. In addition to Cry proteins, other Bacillus thuringiensis (Bt toxins in culture supernatants (SN have biocontrol potential (e.g., Vip3A, Cry1I, Sip1, whereas others are unwanted (β-exotoxins, as they display widespread toxicity across taxa. A strain simultaneously providing distinct toxin activities in crystals and SN would be desirable for bioinsecticides development; however, strains secreting β-exotoxins should be discarded, independently of other useful entomotoxins. Entomotoxicity of crystals and SN from a Brazilian Bt tolworthi strain (Btt01 was tested against Spodoptera frugiperda to assess the potential for biocontrol-product development based on more than one type of toxin/activity. Tests showed that 107 endospores mL−1 caused >80% of larvae mortality, suggesting Btt01 may be used in similar concentrations as those of other Bt-based biopesticides. When it was applied to cornfields, a significant 60% reduction of larvae infestation was observed. However, bioassays with Btt01 SN revealed a thermostable toxic activity. Physicochemical characterization strongly suggests the presence of unwanted β-exotoxins, with isolate-specific temporal variation in its secretion. Knowledge of the temporal pattern of secretion/activity in culture for all forms of toxins produced by a single strain is required to both detect useful activities and avoid the potential lack of identification of undesirable toxins. These findings are discussed in the contexts of commercial Bt product development, advantages of multiple-activity strains, and care and handling recommended for large-scale fermentation systems.

  20. Requirement of simultaneous assessment of crystal- and supernatant-related entomotoxic activities of Bacillus thuringiensis strains for biocontrol-product development.

    Science.gov (United States)

    Argôlo-Filho, Ronaldo Costa; Costa, Robson Luz; Pinheiro, Daniele Heloisa; Corrêa, Fábio Mathias; Valicente, Fernando Hercos; Pomella, Alan William Vilela; Loguercio, Leandro Lopes

    2014-05-20

    Bioinsecticides with lower concentrations of endospores/crystals and without loss of efficiency are economically advantageous for pest biocontrol. In addition to Cry proteins, other Bacillus thuringiensis (Bt) toxins in culture supernatants (SN) have biocontrol potential (e.g., Vip3A, Cry1I, Sip1), whereas others are unwanted (β-exotoxins), as they display widespread toxicity across taxa. A strain simultaneously providing distinct toxin activities in crystals and SN would be desirable for bioinsecticides development; however, strains secreting β-exotoxins should be discarded, independently of other useful entomotoxins. Entomotoxicity of crystals and SN from a Brazilian Bt tolworthi strain (Btt01) was tested against Spodoptera frugiperda to assess the potential for biocontrol-product development based on more than one type of toxin/activity. Tests showed that 10(7) endospores mL(-1) caused >80% of larvae mortality, suggesting Btt01 may be used in similar concentrations as those of other Bt-based biopesticides. When it was applied to cornfields, a significant 60% reduction of larvae infestation was observed. However, bioassays with Btt01 SN revealed a thermostable toxic activity. Physicochemical characterization strongly suggests the presence of unwanted β-exotoxins, with isolate-specific temporal variation in its secretion. Knowledge of the temporal pattern of secretion/activity in culture for all forms of toxins produced by a single strain is required to both detect useful activities and avoid the potential lack of identification of undesirable toxins. These findings are discussed in the contexts of commercial Bt product development, advantages of multiple-activity strains, and care and handling recommended for large-scale fermentation systems.

  1. Inhibition of biofilm development and spoilage potential of Shewanella baltica by quorum sensing signal in cell-free supernatant from Pseudomonas fluorescens.

    Science.gov (United States)

    Zhao, Aifei; Zhu, Junli; Ye, Xiaofeng; Ge, Yangyang; Li, Jianrong

    2016-08-02

    The objective of this study was to in vitro evaluate the effect of a cell-free supernatant (CFS) containing quorum sensing (QS) signal of Pseudomonas fluorescens on the growth, biofilm development and spoilage potential of Shewanella baltica, and preliminarily assess the interactive influences of various chemically synthesized autoinducers on spoilage phenotypes of S. baltica. PF01 strain isolated from spoiled Pseudosciaen crocea was identified P. fluorescens. The addition of 25% and 50% CFS to S. baltica culture had no effect on the growth rate during the lag and exponential phase, however, caused cell decline during the stationary phase. The presence of CFS from P. fluorescens significantly inhibited biofilm development, and greatly decreased the production of trimethylamine (TMA) and biogenic amino in S. baltica. Various signal molecules of QS in the CFS of P. fluorescens culture were detected, including seven N-acyl-l-homoserine lactones (AHLs), autoinducer-2 (AI-2) and two diketopiperazines (DKPs). Exogenous supplement of synthesized seven AHLs containing in the CFS decreased biofilm formation and TMA production in S. baltica, while exposure to exogenous cyclo-(l-Pro-l-Leu) was showed to promote spoilage potential, which revealed that S. baltica also sense the two QS molecules. Furthermore, the stimulating effect of cyclo-(l-Pro-l-Leu) was affected when AHL was simultaneously added, suggesting that the inhibitory activity of spoilage phenotypes in S. baltica might be attributed to a competitive effect of these QS compounds in the CFS of P. fluorescens. The present studies provide a good basis for future research on the role of QS in the regulation of spoilage microbial flora.

  2. Requirement of Simultaneous Assessment of Crystal- and Supernatant-Related Entomotoxic Activities of Bacillus thuringiensis Strains for Biocontrol-Product Development

    Science.gov (United States)

    Argôlo-Filho, Ronaldo Costa; Costa, Robson Luz; Pinheiro, Daniele Heloisa; Corrêa, Fábio Mathias; Valicente, Fernando Hercos; Pomella, Alan William Vilela; Loguercio, Leandro Lopes

    2014-01-01

    Bioinsecticides with lower concentrations of endospores/crystals and without loss of efficiency are economically advantageous for pest biocontrol. In addition to Cry proteins, other Bacillus thuringiensis (Bt) toxins in culture supernatants (SN) have biocontrol potential (e.g., Vip3A, Cry1I, Sip1), whereas others are unwanted (β-exotoxins), as they display widespread toxicity across taxa. A strain simultaneously providing distinct toxin activities in crystals and SN would be desirable for bioinsecticides development; however, strains secreting β-exotoxins should be discarded, independently of other useful entomotoxins. Entomotoxicity of crystals and SN from a Brazilian Bt tolworthi strain (Btt01) was tested against Spodoptera frugiperda to assess the potential for biocontrol-product development based on more than one type of toxin/activity. Tests showed that 107 endospores mL−1 caused >80% of larvae mortality, suggesting Btt01 may be used in similar concentrations as those of other Bt-based biopesticides. When it was applied to cornfields, a significant 60% reduction of larvae infestation was observed. However, bioassays with Btt01 SN revealed a thermostable toxic activity. Physicochemical characterization strongly suggests the presence of unwanted β-exotoxins, with isolate-specific temporal variation in its secretion. Knowledge of the temporal pattern of secretion/activity in culture for all forms of toxins produced by a single strain is required to both detect useful activities and avoid the potential lack of identification of undesirable toxins. These findings are discussed in the contexts of commercial Bt product development, advantages of multiple-activity strains, and care and handling recommended for large-scale fermentation systems. PMID:24854738

  3. Analysis of tank 4 (FTF-4-15-22, 23) surface and subsurface supernatant samples in support of enrichment control, corrosion control and evaporator feed qualification programs

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-09-09

    This report provides the results of analyses on Savannah River Site Tank 4 surface and subsurface supernatant liquid samples in support of the Enrichment Control Program (ECP), the Corrosion Control Program (CCP) and the Evaporator Feed Qualification (EFQ) Program. The purpose of the ECP sample taken from Tank 4 in August 2015 was to determine if the supernatant liquid would be “acceptable feed” to the 2H and 3H evaporator systems.

  4. Use of Bacillus thuringiensis supernatant from a fermentation process to improve bioremediation of chlorpyrifos in contaminated soils.

    Science.gov (United States)

    Aceves-Diez, Angel E; Estrada-Castañeda, Kelly J; Castañeda-Sandoval, Laura M

    2015-07-01

    The aim of this research was to investigate the potential of a nutrient-rich organic waste, namely the cell-free supernatant of Bacillus thuringiensis (BtS) gathered from fermentation, as a biostimulating agent to improve and sustain microbial populations and their enzymatic activities, thereby assisting in the bioremediation of chlorpyrifos-contaminated soil at a high dose (70 mg kg(-1)). Experiments were performed for up to 80 d. Chlorpyrifos degradation and its major metabolic product, 3,5,6-trichloro-2-pyridinol (TCP), were quantified by high-performance liquid chromatography (HPLC); total microbial populations were enumerated by direct counts in specific medium; and fluorescein diacetate (FDA) hydrolysis was measured as an index of soil microbial activity. Throughout the experiment, there was higher chlorpyrifos degradation in soil supplemented with BtS (83.1%) as compared to non-supplemented soil. TCP formation and degradation occurred in all soils, but the greatest degradation (30.34%) was observed in soil supplemented with BtS. The total microbial populations were significantly improved by supplementation with BtS. The application of chlorpyrifos to soil inhibited the enzymatic activity; however, this negative effect was counteracted by BtS, inducing an increase of approximately 16% in FDA hydrolysis. These results demonstrate the potential of B. thuringiensis supernatant as a suitable biostimulation agent for enhancing chlorpyrifos and TCP biodegradation in chlorpyrifos-contaminated soils.

  5. A combination turbidity and supernatant microplate assay to rank-order the supersaturation limits of early drug candidates.

    Science.gov (United States)

    Morrison, John S; Nophsker, Michelle J; Haskell, Roy J

    2014-10-01

    A unique opportunity exists at the drug discovery stage to overcome inherently poor solubility by selecting drug candidates with superior supersaturation propensity. Existing supersaturation assays compare either precipitation-resistant or precipitation-inhibiting excipients, or higher-energy polymorphic forms, but not multiple compounds or multiple concentrations. Furthermore, these assays lack sufficient throughput and compound conservation necessary for implementation in the discovery environment. A microplate-based combination turbidity and supernatant concentration assay was therefore developed to determine the extent to which different compounds remain in solution as a function of applied concentration in biorelevant media over a specific period of time. Dimethyl sulfoxide stock solutions at multiple concentrations of four poorly soluble, weak base compounds (Dipyridamole, Ketoconazole, Albendazole, and Cinnarizine) were diluted with pH 6.5 buffer as well as FaSSIF. All samples were monitored for precipitation by turbidity at 600 nm over 1 h and the final supernatant concentrations were measured. The maximum supersaturation ratio was calculated from the supersaturation limit and the equilibrium solubility in each media. Compounds were rank-ordered by supersaturation ratio: Ketoconazole > Dipyridamole > Cinnarizine ∼ Albendazole. These in vitro results correlated well with oral AUC ratios from published in vivo pH effect studies, thereby confirming the validity of this approach. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

  6. Lipopolysaccharide-Induced Profiles of Cytokine, Chemokine, and Growth Factors Produced by Human Decidual Cells Are Altered by Lactobacillus rhamnosus GR-1 Supernatant.

    Science.gov (United States)

    Li, Wei; Yang, Siwen; Kim, Sung O; Reid, Gregor; Challis, John R G; Bocking, Alan D

    2014-07-01

    The aim of this study was to assess the effects of bacterial lipopolysaccharide (LPS) and Lactobacillus rhamnosus GR-1 supernatant (GR-1SN) on secretion profiles of cytokines, chemokines, and growth factors from primary cultures of human decidual cells. Lipopolysaccharide significantly increased the output of proinflammatory cytokines (interleukin [IL]-1B, IL-2, IL-6, IL-12p70, IL-15, IL-17A, interferon gamma [IFN-γ], and tumor necrosis factor [TNF]); anti-inflammatory cytokines (IL-1RN, IL-4, IL-9, and IL-10); chemokines (IL-8, eotaxin, IFN-inducible protein 10 [IP-10], monocyte chemoattractant protein 1 [MCP-1], macrophage inflammatory protein-1α [MIP-1α], macrophage inflammatory protein-1β [MIP-1β], and regulated on activation normal T cell expressed and secreted [RANTES]); and growth factors (granulocyte colony-stimulating factor [CSF] 3, CSF-2, and vascular endothelial growth factor A [VEGFA]). Lactobacillus rhamnosus GR-1SN alone significantly increased CSF-3, MIP-1α MIP-1β, and RANTES but decreased IL-15 and IP-10 output. The GR-1SN also significantly or partially reduced LPS-induced proinflammatory cytokines TNF, IFN-γ, IL-1β, IL-2 IL-6, IL-12p70, IL-15, IL-17, and IP-10; partially reduced LPS-induced anti-inflammatory cytokines IL-1RN, IL-4 and IL-10, and LPS-induced VEGFA output but did not affect CSF-3, MIP-1α, MIP-1β, MCP-1, IL-8, and IL-9. Our results demonstrate that GR-1SN attenuates the inflammatory responses to LPS by human decidual cells, suggesting its potential role in ameliorating intrauterine infection.

  7. Analysis of tank 7 surface supernatant sample (FTF-7-15-26) in support of corrosion control program

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-10-01

    This report provides the results of analyses on Savannah River Site Tank 7 surface supernatant liquid sample in support of the Corrosion Control Program (CCP). The measured nitrate, nitrite and free-hydroxide concentrations for the Tank 7 surface sample averaged, 3.74E-01 ± 1.88E-03, 4.17E-01 ± 9.01E-03 and 0.602 ± 0.005 M, respectively. The Tank 7 surface cesium-137, sodium and silicon concentrations were, respectively, 3.99E+08, ± 3.25E+06 dpm/mL, 2.78 M and <3.10 mg/L. The measured aluminum concentration in the Tank 7 surface sample averaged 0.11 M.

  8. Mescaline-induced changes of brain-cortex ribosomes. Mescaline demethylase activity of brain-cortex soluble supernatant.

    Science.gov (United States)

    Datta, R K; Ghosh, J J

    1977-02-01

    Brain-cortex slices demethylate mescaline and p-methoxyacetanilide, a reference O-demethylating substrate, though the rate of demethylation of mescaline is about one third that of the reference substrate. The demethylase activity is localized mostly in the soluble supernatant (105 000 x g). It is purified 47-fold with respect to the demethylation of mescaline by ammonium sulfate precipitation and DEAE cellulose chromatography. The partially purified demethylase, which is stable for 3-5 days at -5 degrees C in the presence of dithiothreitol and glutathione and is inhibited by p-chloromercuribenzoate, has maximal activity at pH between 7.2 and 8.0. It demethylates mescaline into 3,4-dimethoxy-5-hydroxyphenethylamine and 3,5-dimethoxy-4-hydroxyphenethylamine and some unidentified derivatives.

  9. Sponge Hybridomas: Applications and Implications

    NARCIS (Netherlands)

    Pomponi, S.A.; Jevitt, A.; Patel, J.; Diaz, M.C.

    2013-01-01

    Many sponge-derived natural products with applications to human health have been discovered over the past three decades. In vitro production has been proposed as one biological alternative to ensure adequate supply of marine natural products for preclinical and clinical development of drugs. Althoug

  10. Hydrofocusing Bioreactor for Three-Dimensional Cell Culture

    Science.gov (United States)

    Gonda, Steve R.; Spaulding, Glenn F.; Tsao, Yow-Min D.; Flechsig, Scott; Jones, Leslie; Soehnge, Holly

    2003-01-01

    The hydrodynamic focusing bioreactor (HFB) is a bioreactor system designed for three-dimensional cell culture and tissue-engineering investigations on orbiting spacecraft and in laboratories on Earth. The HFB offers a unique hydrofocusing capability that enables the creation of a low-shear culture environment simultaneously with the "herding" of suspended cells, tissue assemblies, and air bubbles. Under development for use in the Biotechnology Facility on the International Space Station, the HFB has successfully grown large three-dimensional, tissuelike assemblies from anchorage-dependent cells and grown suspension hybridoma cells to high densities. The HFB, based on the principle of hydrodynamic focusing, provides the capability to control the movement of air bubbles and removes them from the bioreactor without degrading the low-shear culture environment or the suspended three-dimensional tissue assemblies. The HFB also provides unparalleled control over the locations of cells and tissues within its bioreactor vessel during operation and sampling.

  11. Determining antioxidant activities of lactobacilli cell-free supernatants by cellular antioxidant assay: a comparison with traditional methods.

    Directory of Open Access Journals (Sweden)

    Jiali Xing

    Full Text Available Antioxidant activity of lactic acid bacteria is associated with multiple health-protective effects. Traditional indexes of chemical antioxidant activities poorly reflect the antioxidant effects of these bacteria in vivo. Cellular antioxidant activity (CAA assay was used in this study to determine the antioxidant activity of cell-free supernatants (CFSs of 10 Lactobacillus strains. The performance of the CAA assay was compared with that of four chemical antioxidant activity assays, namely, DPPH radical scavenging, hydroxyl radical scavenging (HRS, reducing power (RP, and inhibition of linoleic acid peroxidation (ILAP. Results of the CAA assay were associated with those of DPPH and ILAP assays, but not with those of RP and HRS assays. The inter- and intra-specific antioxidant activities of CFS were characterized by chemical and CAA assays. L. rhamnosus CCFM 1107 displayed a high antioxidative effect similar to positive control L. rhamnosus GG ATCC 53103 in all of the assays. The CAA assay is a potential method for the detection of antioxidant activities of lactobacilli CFSs.

  12. MicroRNA Expression in Salivary Supernatant of Patients with Pancreatic Cancer and Its Relationship with ZHENG

    Directory of Open Access Journals (Sweden)

    Song Gao

    2014-01-01

    Full Text Available In traditional Chinese medicine (TCM, diagnosis and prescriptions are based on the signs and symptoms which are recognized as ZHENG. The cornerstone of TCM is to differentially treat one ZHENG from others, which is also known as syndrome differentiation, and this relies on the gathering of clinical information through inspection, auscultation and olfaction, inquiry, and palpation. However, the biomolecular basis of the ZHENG remains unclear. In this study, the expressions of 384 cancer-related miRNAs in salivary supernatant of patients with pancreatic cancer were assessed by miRNA polymerase chain reaction (PCR array, and the different expression patterns of miRNA in three different groups of ZHENG were studied with use of real-time quantitative PCR (qRT-PCR. Some miRNAs were found to be specifically expressed in some ZHENGs, for instance, miR-17, miR-21, and miR-181b in Shi-Re ZHENG and miR-196a in Pi-Xu ZHENG. This indicates that these miRNAs may play important roles in different ZHENG condition. Therefore, this study to some extent revealed the molecular basis of TCM ZHENG in pancreatic cancer.

  13. Analysis of tank 39H (HTF-39-15-61, 62) surface and subsurface supernatant samples in support of corrosion control program

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-08-19

    This report provides the results of analyses on Tanks 39H surface and subsurface supernatant liquid samples in support of the Corrosion Control Program. Analyses included warm acid strike preparation followed by analysis for silicon, aluminum, and sodium and water dilution preparation followed by analysis for anions. Other reported analytical results include analyses results for uranium, Pu-241 and Pu-239.

  14. Evaluation of the Antioxidative, Antibacterial, and Anti-Inflammatory Effects of the Aloe Fermentation Supernatant Containing Lactobacillus plantarum HM218749.1

    Directory of Open Access Journals (Sweden)

    Meixiu Jiang

    2016-01-01

    Full Text Available Little work is done to develop Aloe vera (AV using probiotics. To explore the potential benefits, the antioxidant effects and the antibacterial effects on foodborne pathogens of Aloe fermentation supernatant were evaluated in vitro. Our results indicated that the Aloe fermentation supernatant fermented by Lactobacillus plantarum HM218749.1 had very strong scavenging capacities of the DPPH (86%, O2•- (85%, OH• (76%, and Fe2+ chelation (82% and reducing powers (242.5 mg/L, and the inhibition zones for Salmonella typhimurium, Salmonella enteritidis, Shigella flexneri, Escherichia coli, Listeria monocytogenes, S. dysenteriae 301, Staphylococcus aureus Cowan1, and Propionibacterium acnes were 16, 15, 19, 20, 21, 20, and 27 mm. Moreover, the low concentration of Aloe fermentation supernatant had significantly reduced the production of IL-1β, TNF-α, and IL-6 in both mRNA and protein levels (P<0.01. Therefore, the Aloe fermentation supernatant can be used as functional beverage or cosmetic ingredients to guard human intestinal health, delaying senescence, and prevent chronic diseases.

  15. Evaluation of the Antioxidative, Antibacterial, and Anti-Inflammatory Effects of the Aloe Fermentation Supernatant Containing Lactobacillus plantarum HM218749.1

    Science.gov (United States)

    Deng, Kan; Jiang, Chunling; Fu, Mingui; Guo, Chunlan; Wang, Xiaolei; Wang, Xin; Meng, Fanjing; Yang, Shaoguo; Deng, Keyu

    2016-01-01

    Little work is done to develop Aloe vera (AV) using probiotics. To explore the potential benefits, the antioxidant effects and the antibacterial effects on foodborne pathogens of Aloe fermentation supernatant were evaluated in vitro. Our results indicated that the Aloe fermentation supernatant fermented by Lactobacillus plantarum HM218749.1 had very strong scavenging capacities of the DPPH (86%), O2•− (85%), •OH (76%), and Fe2+ chelation (82%) and reducing powers (242.5 mg/L), and the inhibition zones for Salmonella typhimurium, Salmonella enteritidis, Shigella flexneri, Escherichia coli, Listeria monocytogenes, S. dysenteriae 301, Staphylococcus aureus Cowan1, and Propionibacterium acnes were 16, 15, 19, 20, 21, 20, and 27 mm. Moreover, the low concentration of Aloe fermentation supernatant had significantly reduced the production of IL-1β, TNF-α, and IL-6 in both mRNA and protein levels (P < 0.01). Therefore, the Aloe fermentation supernatant can be used as functional beverage or cosmetic ingredients to guard human intestinal health, delaying senescence, and prevent chronic diseases. PMID:27493450

  16. [The comparative characteristics of antibacterial properties of the peptides of the active site of GM-CSF, and substances delivered from supernatants of hematopoietic progenitor CD34+45- cells].

    Science.gov (United States)

    Zurochka, A V; Zurochka, V A; Kostolomova, E G; Dobrynina, M A; Sykhoveĭ, Iu G; Gritsenko, V A

    2012-01-01

    The antibacterial activity of synthetic peptides of the active site of GM-CSF and supernatants of CD34+45- hematopoietic progenitor cells has been investigated GM-CSF peptides and cell supernatants were found to possess pronounced antibacterial activity, at that a combination of these substances has a more pronounced activity in comparison with the single substances. Possible mechanisms of the identified effects of synthetic peptides and substances from the supernatants of CD34+5- cells are discussed.

  17. Fermentanomics informed amino acid supplementation of an antibody producing mammalian cell culture.

    Science.gov (United States)

    Read, Erik K; Bradley, Scott A; Smitka, Tim A; Agarabi, Cyrus D; Lute, Scott C; Brorson, Kurt A

    2013-01-01

    Fermentanomics, or a global understanding of a culture state on the molecular level empowered by advanced techniques like NMR, was employed to show that a model hybridoma culture supplied with glutamine and glucose depletes aspartate, cysteine, methionine, tryptophan, and tyrosine during antibody production. Supplementation with these amino acids prevents depletion and improves culture performance. Furthermore, no significant changes were observed in the distribution of glycans attached to the IgG3 in cultures supplemented with specific amino acids, arguing that this strategy can be implemented without fear of impact on important product quality attributes. In summary, a targeted strategy of quantifying media components and designing a supplementation strategy can improve bioprocess cell cultures when enpowered by fermentanomics tools.

  18. Culture medium for amylase production by toxigenic fungi

    Directory of Open Access Journals (Sweden)

    Figueira Edson Luiz Zangrando

    2000-01-01

    Full Text Available Mycelial growth and amylase production by a mycotoxigenic strain of Fusarium moniliforme and Aspergillus flavus were evaluated in a culture medium containing starch, glycerol, wheat bran or corn. With emphasis on corn, different fractions composed by germ, degermed seed, starch, milky stage corn and the respective starch or supernatant fraction were analyzed for F. moniliforme growth . The medium composed of milky stage corn supernatant promoted the best mycelial growth (p<0.05, and it was used to prepare amylase production medium in the next step. The medium composed with 2% ground corn in milky stage corn supernatant (350g of milky stage corn blended with 250mL water and centrifuged promoted the highest amylase production, which was at the 10th day of fermentation, both for F. moniliforme (42.32U/mL and A. flavus (4,745.54U/mL.

  19. Analysis of tank 51H (HTF-51-15-77) subsurface supernatant sample in support of enrichment and corrosion control programs

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-08-18

    This report provides the results of analyses on Tank 51H subsurface supernatant liquid sample in support of the Enrichment Control Program (ECP) and the Corrosion Control Program (CCP). The purpose of the ECP sample taken from Tank 51H in early June was to determine if the later decants would be “acceptable feed” to the 2H and 3H evaporator systems.

  20. Analysis of tank 51H (HTF-51-15-77) subsurface supernatant sample in support of enrichment and corrosion control programs

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-08-18

    This report provides the results of analyses on Tank 51H subsurface supernatant liquid sample in support of the Enrichment Control Program (ECP) and the Corrosion Control Program (CCP).The purpose of the ECP sample taken from Tank 51H in early June was to determine if the later decants would be “acceptable feed” to the 2H and 3H evaporator systems.

  1. 热损伤角质形成细胞培养上清液对成纤维细胞生物学行为的影响%Effect of heat injured keratinocytes supernatant on biological behavior of fibroblasts

    Institute of Scientific and Technical Information of China (English)

    白晓智; 胡大海; 张万福; 张战凤; 石继红; 蔡维霞; 朱华宇; 朱雄翔; 汤朝武

    2010-01-01

    Objective To observe the effect of the supernatant of heat injured keratinocytes (KC) on biological behavior of the dermal fibroblasts ( Fb). Methods Human dermal Fb were isolated and cultured. A model of heat injured KC ( HaCaT) was reproduced in vitro. Supernatant of normal KC and the supernatant of KC culture 12 hours after heat injury were collected and diluted with non-serum DMEM in 1:1 volume ratio to make normal KC conditioned medium ( NKCM ) and heat injury KC conditioned medium ( HKCM) respectively. Fb was respectively treated with non-serum DMEM and 2 kinds of conditioned medium. (1) The proliferation of Fb was detected with MTT method at post culture hour (PCH) 12, 24, 36, 48. (2) The apoptosis of Fb was determined by flow cytometry at PCH 12 ( Fb were heat injured in advance; Fb without heat treatment was used as control). (3) At PCH 24, expression of a-SMA in Fb cytoplasm was determined with immunofluorescence method; expression of a-SMA mRNA in Fb was determined with real-time quantified PCR. Data were processed with one-way analysis of variance, and pairwise comparison among groups with LSD-t test. Results (1) The proliferation of Fb: the absorbance value of Fb cultured with HKCM at PCH 12, 24, 36, 48 wa9 respectively higher than that of Fb cultured with non-serum DMEM ( with t value respectively 1. 89, 2. 35 , 2.02, 1.94, and P values all below 0. 01). There were significant statistical differences between the absorbance values of Fb cultured with HKCM and those of Fb cultured with 1MKCM at PCH 12, 24, and 48 (at PCH 12, t = 1. 83, P <0. 01; at PCH 24,t =2.91, P < 0. 05 ; at PCH 48 , t = 1. 83 , P < 0.05). (2 ) Apoptosis of Fb cultured with HKCM was diminished as compared with that of Fb cultured with NKCM and of Fb without treatment ( t =3.31 , P <0.05; t = 1.47, P < 0.01). (3) The expression of α-SMA (red fluorescence) in Fb cultured with non-serum DMEM or NKCM was less as seen under fluorescence scope, and it was obviously increased in Fb

  2. Activation of resting human B cells by helper T-cell clone supernatant: characterization of a human B-cell-activating factor.

    Science.gov (United States)

    Diu, A; Gougeon, M L; Moreau, J L; Reinherz, E L; Thèze, J

    1987-12-01

    The effects of helper T-cell clone supernatants on resting human B cells were investigated. Four different helper T-cell clones (two T4+ and two T8+) were stimulated by anti-T3 monoclonal antibodies on Sepharose beads or anti-T11(2) plus anti-T11(3) monoclonal antibodies. The supernatants from these activated clones induced the proliferation of highly purified resting B lymphocytes from the peripheral blood. The B cells exhibited a cell size and a surface-antigen pattern (4F2 antigen and transferrin receptor) of phase G0 B cells, and they were functionally resting. In response to T-cell supernatants a large fraction of the B cells enlarged and expressed 4F2 antigens and transferrin receptors. In gel filtration, the corresponding activity migrated with an apparent Mr of 12,000-15,000. Our findings strongly support the existence of a human B-cell-activating factor acting on resting B cells and causing them to enter phase G1 of the cell cycle.

  3. [Induced sputum supernatant prostaglandin E2 during oral aspirin challenge of asthmatic patients with and without aspirin hypersensitivity and healthy controls--pilot study].

    Science.gov (United States)

    Ignacak, Maria; Celejewska-Wójcik, Natalia; Wójcik, Krzysztof; Sałapa, Kinga; Konduracka, Ewa; Sanak, Marek; Tyrak, Katarzyna; Sładek, Krzysztof; Musiał, Jacek; Mastalerz, Lucyna

    2016-01-01

    The aim of this pilot study was to evaluate changes in the concentration of prostaglandin E2 (PGE2) in induced sputum supernatant in 3 groups: sub- jects with NSAID-exacerbated respira- tory disease (NERD), aspirin tolerant asthma (ATA) and healthy controls (HC), before and after oral aspirin chal- lenge test. The study was conducted in the years 2014-2015 at the Clinical Department of the Pulmonology Clinic at the University Hospital in Cracow. 43 patients were enrolled in the study (NERD - n = 15, ATA - n = 15 and HC - n = 13). All of them underwent a placebo-controlled oral aspirin challenge. Sputum was induced 24 hours before the challenge and immediately after the test. Induced sputum was processed in order to obtain cystospin slides to depict inflammatory cell patterns and supernatants, in which PGE2 was measured. The concentration of PGE2 was determined using mass spectrometry coupled with gas chromatography (gas chromatography/mass spectrometry - GC/MS). After aspirin challenge, the concentration of PGE2 in induced sputum supernatant decreased in both asthmatics hypersensitive to aspirin (p = 0.01) and those who tolerated aspirin well (p = 0.17). The change in the healthy control group was not statistically significant. These results support the cyclooxygenase theory of PGE2 inhibition by aspirin. However, the mechanism of bronchoconstriction after aspirin administration alone in patients with NSAID-exacerbated respiratory disease remains unclear.

  4. Fast and simple online sample preparation coupled with capillary LC-MS/MS for determination of prostaglandins in cell culture supernatants

    DEFF Research Database (Denmark)

    Rinne, Sandra; Ramstad Kleiveland, Charlotte; Kassem, Moustapha

    2007-01-01

    An online 2-D strong cation exchange (SCX)-RP capillary liquid chromatographic (cLC) method with IT mass spectrometric (IT-MS/MS) detection for the simultaneous determination of prostaglandin (PG) A(1), PGD(2), PGE(1), PGE(2), PGF(2a), 6-keto-(6k)PGF(1a), and 15-Delta(12,14)-deoxy-PGJ(2) (15dPGJ(...

  5. High-throughput screening techniques for rapid PEG-based precipitation of IgG4 mAb from clarified cell culture supernatant.

    Science.gov (United States)

    Knevelman, Carol; Davies, Jim; Allen, Lee; Titchener-Hooker, Nigel J

    2010-01-01

    Locating optimal protein precipitation conditions for complex biological feed materials is problematic. This article describes the application of a series of high-throughput platforms for the rapid identification and selection of conditions for the precipitation of an IgG(4) monoclonal antibody (mAb) from a complex feedstock using only microliter quantities of material. The approach uses 96-microwell filter plates combined with high-throughput analytical methods and a method for well volume determination for product quantification. The low material, time and resource requirements facilitated the use of a full factorial Design of Experiments (DoE) for the rapid investigation into how critical parameters impact the IgG(4) precipitation. To aid the DoE, a set of preliminary range-finding studies were conducted first. Data collected through this approach describing Polyethylene Glycol (PEG) precipitation of the IgG(4) as a function of mAb concentration, precipitant concentration, and pH are presented. Response surface diagrams were used to explore interactions between parameters and to inform selection of the most favorable conditions for maximum yield and purification. PEG concentrations required for maximum yield and purity were dependant on the IgG(4) concentration; however, concentrations of 14 to 20% w/v, pH 6.5, gave optimal levels of yield and purity. Application of the high-throughput approach enabled 1,155 conditions to be examined with less than 1 g of material. The level of insights gained over such a short time frame is indicative of the power of microwell experimentation in allowing the rapid identification of appropriate processing conditions for key bioprocess operations. Copyright 2009 American Institute of Chemical Engineers

  6. Tetracycline and Glutathione Inhibit Matrix Metalloproteinase Activity: An In Vitro Study Using Culture Supernatants of L929 and Dalton Lymphoma Cell Lines

    Directory of Open Access Journals (Sweden)

    Gajanan Kendre

    2013-01-01

    Full Text Available Tetracycline and glutathione inhibited the protease activities of matrix metalloproteinase-2 and matrix metalloproteinase-9 expressed by mouse fibrosarcoma cells (L929 and Dalton lymphoma cells, respectively. The inhibitory activity of the tetracycline may be due to its ability to chelate metal ions such as calcium and zinc. Gelatin-zymography technique was used to demonstrate the inhibitory activity of both tetracycline and glutathione. The intensity of the bands corresponding to metalloproteinase activity in zymography gel was reduced in the presence of 50–100 μg/mL of tetracycline. The presence of 10–100 μg/mL of tetracycline in the medium increased the adherence of L929 cancer cells. These results clearly indicate the antimetastatic property of tetracycline. Reduced glutathione, a compound which is produced endogenously by the cells to maintain the redox status, was shown to inhibit the matrix metalloproteinase activity (in vitro. Therefore, it is assumed that decreased glutathione levels in synovial fluids or plasma might increase the activity of MMP. Reduced glutathione at 100 μg/mL inhibited the metalloproteinase activity in gelatin-zymographic gel. As both tetracycline and glutathione exhibited an inhibitory effect on matrix metalloproteinase activity, it was of great interest to check their clinical effects on various MMP associated pathological conditions such as cancer metastasis and arthritis. Here we report that tetracycline and reduced glutathione inhibited the activity of MMP2 completely and activity of MMP9 partly.

  7. Screening for epitope specificity directly on culture supernatants in the early phase of monoclonal antibody production by an ELISA with biotin-labeled antigen

    DEFF Research Database (Denmark)

    Andersen, Ditte C; Jensen, Charlotte H; Gregersen, Annemette

    2004-01-01

    This report describes an assay for comparison of epitope specificity in groups of monoclonal antibodies against a given antigen. The only prerequisite is the biotin-labeled antigen. One of the monoclonal antibodies is captured onto a plastic surface via a rabbit anti-mouse Ig, and the other...... preincubated with biotinylated antigen. When the two antibodies react with the same epitope subsequent binding of the biotin-labeled antigen is abolished (inhibition). In the cases where no inhibition was observed, the two antibodies were considered to react with distinct, independent epitopes. The obvious...

  8. Toxigenicity testing of clinical isolates of non-typhoidal salmonellae in Vero cell culture & Caenorhabditis elegans model.

    Science.gov (United States)

    Jesudason, Mary V; V Balaji, V; Densibai, Shoba

    2006-06-01

    The non-typhoidal salmonellae (NTS) are recognized agents of gastroenteritis worldwide. Some of the NTS do not produces cytotoxic changes in tissue culture and not much is known about the endotoxicity of the clinical isolates of NTS (mostly Salmonella enterica serotype Typhimurium and Salmonella enterica serotype Enteritidis). We examined the exotoxic (cytotoxin) and endotoxic activity of clinical isolates of NTS in two assay models namely Vero cell culture and the nematode, Caenorhabditis elegans. Bacteria-free culture supernatants of 40 isolates NTS were tested in 96 well microtitre plate containing confluent monolayers of Vero cells. For the effects on C. elegans, the worms were exposed to bacteria free culture supernatants in 24 well microtitre plate for 24 h and then transferred to OP50 Escherichia coli lawn culture. The endotoxic activity of the live bacterium was studied by feeding the worms in the lawn culture of NTS separately. No cytopathic effect was observed with NTS tested in Vero cell culture assay. Likewise, the worms exposed to the bacteria-free culture supernatants were found active up to 7 days. In the co-culture killing assay, worms were found dead with characteristic stiff and straight appearance by 16(th) day. The worms were alive up to 21 days in OP50 E. coli. Bacteria-free culture supernatants did not have any deleterious effect on worms or in Vero cell culture, suggesting that there is no soluble toxic factor (diffusible toxin) in the culture supernatants. However, live NTS were found to be lethal to the worms; indicating that direct interaction between viable NTS and C. elegans is necessary for killing.

  9. Effects of human promyelocytic leukemia HL-60 cel supernatant on differetiation of mononulear cel s from human umbilical cord blood%急性早幼粒白血病细胞培养上清液对人脐带血单个核细胞分化的影响

    Institute of Scientific and Technical Information of China (English)

    郑毅; 方宁; 陈代雄

    2013-01-01

    目的白血病细胞能够通过自分泌方式分泌细胞因子而作用于自身,使细胞大量的增殖而不分化;白血病细胞所分泌的细胞因子也能作用于人脐带血(UCB)单个核细胞(MNCs)中的CD34+/CD133+细胞亚群使之增殖而抑制其分化。关于急性早幼粒白血病细胞(HL-60细胞)培养上清液能否使UCB- MNCs分化尚未见报道,本实验主要目的是观察HL-60细胞培养上清液对人UCB-MNCs分化的影响。方法实验分为①有HL-60细胞培养上清液+StemSpan® SFEM组;②StemSpan® SFEM组;③HL-60细胞培养上清液+高糖(HG)-DMEM组;④HG-DMEM组;⑤HL-60细胞培养上清液+低糖(LG)-DMEM组;⑥LG-DMEM组。收分离的人UCB-MNCs按2.5×106/瓶分别接种于6组培养液中,37℃,5%CO2,饱和湿度培养,于培养前和培养12天用流式细胞仪(FCM)分析表型CD34,CD90和CD166的变化。结果各实验组CD34,CD34/CD90和CD166阳性细胞百分比均较培养前升高。结论%Objective:Leukemic cel s can excrete cytokines by an autocrine way, which contribute to its large-scale proliferation, and the cytokines may also contribute to the proliferations of CD34+/CD133+ subpopulation of UCB-MNCs too. To the effect of human promyelocytic leukemia HL-60 cel supernatant on the differentiation of UCB-MNCs, there is not related report so far. The aim of this study is to investigate effects of HL-60 cel supernatant on differentiations of mononuclear cel s from human umbilical cord blood. Methods:Six various cultural system are listed as fol ows: ① HL-60 cel supernatant+ StemSpan–SFEM; ② StemSpan–SFEM; ③ HL-60 cel supernatant+ HG-DMEM; ④ HG-DMEM; ⑤HL-60 cel supernatant LG-DMEM; ⑥ LG-DMEM. 2.50×106 of UCB-MNCs per bottle with the above each cultural system is incubated at 37℃ in a 5%CO2, saturation humidity of incubator, the changes of phenotype of CD34,CD90 and CD166 analyzed by FCM after 12 days. Results: Positive cel s with CD34+, CD166+ and CD34+/CD

  10. Primary culture of Rhodnius prolixus (Hemiptera: Reduviidae salivary gland cells

    Directory of Open Access Journals (Sweden)

    Fernanda F Rocha

    2010-03-01

    Full Text Available In the present paper, we developed a primary culture of Rhodnius prolixus salivary gland and main salivary canal cells. Cells remained viable in culture for 30 days. Three types of cells were indentified in the salivary gland cultures, with binuclear cells being the most abundant. The supernatants of salivary cultures contained mainly 16-24 kDa proteins and presented anticoagulant and apyrase activities. Secretion vesicles were observed budding from the cellular monolayer of the main salivary canal cells. These results indicate that R. prolixus salivary proteins may be produced in vitro and suggest that the main salivary canal may have a possible secretory role.

  11. Determination of Sr{sup 90}, Sb{sup 125}, Cs{sup 137} and absolute beta activities in First Cycle Supernatent

    Energy Technology Data Exchange (ETDEWEB)

    Helmholz, H.R.

    1954-03-15

    This report discusses procedures have been established for determining the absolute activities of Sr{sup 90}, Sb{sup 125}, Cs{sup 137}, and gross beta in First Cycle Supernatant. These procedures are also valid for other samples such as scavenged Radioactive Waste (RAW), the primary restriction being that the age of the material be at least 250 days so that any Sr{sup 89} activity (53 day half-life) is negligible. With the exception of the gross beta analysis the results are believed to be accurate to within 10%.

  12. Rapid method for identification of group B streptococci in neonatal blood cultures.

    OpenAIRE

    Holmes, R. L.; Harada, W A

    1981-01-01

    A rapid technique used for the identification of Streptococcus agalactiae, Lancefield group B, from the blood cultures of two neonatal infants is reported. The method utilized the Phadebact Streptococcus Test System (Pharmacia Diagnostics, Piscataway, N.J.) and the supernatant from 13- and 14-h blood cultures. Additional studies with simulated neonatal blood cultures revealed that this method was reproducible. Additional studies also revealed that some non-specific agglutination did occur, wh...

  13. Analysis of Tank 38H (HTF-38-15-47, 49) and Tank 43H (HTF-43-15-51, 53) surface and subsurface supernatant samples in support of enrichment and corrosion control programs

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-06-30

    This report provides the results of analyses on Tanks 38H and 43H surface and subsurface supernatant liquid samples in support of the Enrichment Control Program (ECP) and the Corrosion Control Program (CCP).

  14. Analysis of Tank 38H (HTF-38-14-150, 151) and Tank 43H (HTF- 43-14-152, 53) Surface and Subsurface Supernatant Samples in Support of Enrichment Control, Corrosion Control and Sodium Aluminosilicate Formation Potential Programs

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-01-14

    This report provides the results of analyses on Tanks 38H and 43H surface and subsurface supernatant liquid samples in support of the Enrichment Control Program (ECP), the Corrosion Control Program and Sodium Aluminosilicate Formation Potential in the Evaporator.

  15. Differentiation efficiency of human umbilical cord mesenchymal stem cells into hepatocytes under two kinds of liver homogenate supernatants: a comparative study%两种肝组织匀浆上清液诱导人脐带间充质干细胞向肝细胞分化的比较

    Institute of Scientific and Technical Information of China (English)

    闫成; 薛改; 吴丽颖; 刘建芳; 侯艳宁

    2015-01-01

    BACKGROUND:Previous studies have demonstrated that normal rat liver homogenate supernatant can induce human umbilical cord mesenchymal stem cels to differentiate into hepatocyte-like cels with partial hepatocyte functions. However, whether fibrotic liver homogenate supernatant can work or how the inducing effect is remains unclear. OBJECTIVE:To investigate the differentiation potential of human umbilical cord mesenchymal stem cels into hepatocytes under the normal liver and fibrotic liver microenvironment in vitro. METHODS:Liver fibrosis was induced in the SD rats by repeated intraperitoneal injections of 3% thioacetamide at a dose of 200 mg/kg body mass, twice a week for 4 weeks, and then fibrotic liver tissues and normal liver tissues were used to prepare liver homogenate supernatants. Passage 3 human umbilical cord mesenchymal stem cels were used and divided into standard control group (cels were cultured in DMEM/F12 with 10% fetal bovine serum), fibrotic liver homogenate supernatants group (cels were cultured in DMEM/F12 with 10% fetal bovine serum and 50 g/L fibrotic liver homogenate supernatants), normal liver homogenate supernatants group (cels were cultured in DMEM/F12 with 10% fetal bovine serum and 100 g/L normal liver homogenate supernatants). The morphological changes of the cels in each group were recorded under inverted microscope; the protein levels of CK18, AFP, CYP3A4, CYP2E1, CYP2D6 and TPH2 were evaluated using western blot assay. Furthermore, the concentration of albumin in the cels was measured. RESULTS AND CONCLUSION:After a 7-day inducement, the stem cels in liver homogenate supernatants groups lost their fusiform shape and changed into hepatocyte-like cels with the morphous of round shape. Compared with the standard control group, the hepatocyte-like cels in the two liver homogenate supernatants groups exhibited human hepatocyte biomarkers, CK18 and AFP. The standard control group cels could express a little amount of CYP2E1, while cels in

  16. Impact of Cell-free Supernatant of Lactic Acid Bacteria on Putrescine and Other Polyamine Formation by Foodborne Pathogens in Ornithine Decarboxylase Broth.

    Science.gov (United States)

    Ozogul, Fatih; Tabanelli, Giulia; Toy, Nurten; Gardini, Fausto

    2015-06-24

    Conversion of ornithine to putrescine by Salmonella Paratyphi A, Listeria monocytogenes, Staphylococcus aureus, and Escherichia coli was investigated in ornithine decarboxylase broth (ODB) using cell-free supernatants (CFSs) obtained from Leuconostoc mesenterodies subsp. cremoris, Pediococcus acidilactici, Lactococcus lactis subsp. lactis, Streptococcus thermophilus. Two groups of cell-free supernatants (25 or 50%) and control (only ODB) were prepared to investigate putrescine (PUT) and other polyamine formation by foodborne pathogens (FBPs). Significant differences (p < 0.05) were observed among the species for each amine. All of the CFSs reduced the formation of PUT by ≥65%. The production of cadaverine (CAD) was scarcely affected by the presence of CFSs, with the exception of the samples inoculated with L. monocytogenes. The variation in polyamine was found with respect to the control samples. Spermidine (SPD) was produced in lower amount in many samples, especially in Gram-negative FBPs, whereas spermine (SPN) increased drastically in the major part of the samples concerning the control. Histamine (HIS) was characterized by a marked concentration decrease in all of the samples, and tyramine (TYR) was accumulated in very low concentrations in the controls. Therefore, the ability of bacteria to produce certain biogenic amines such as HIS, TYR, PUT, and CAD has been studied to assess their risk and prevent their formation in food products. The results obtained from this study concluded that the lactic acid bacteria (LAB) strains with non-decarboxylase activity are capable of avoiding or limiting biogenic amine formation by FBP.

  17. Effect of phenobarbital pretreatment on benzene biotransformation in the rat. Pt. 2. 9. 000 g supernatant and isolated perfused liver versus living rat

    Energy Technology Data Exchange (ETDEWEB)

    Gut, I.; Hatle, K.; Zizkova, L.

    1981-03-01

    Factors responsible for different quantitative effect of phenobarbital (PB) pretreatment on benzene metabolism to phenol in vivo and in vitro were studied in male Wistar rats. A more than 4-fold increase of benzene metabolism was observed with 9,000 g supernatant of liver homogenate, 2.8- to 4-fold increase with isolated perfused liver; phenol formation in vivo after oral benzene was increased by PB 2-fold, but only shortly following benzene administration and the enhancement rapidly diminished to 1.15-fold increase in the total excreted phenol. Benzene concentrations in 9,000 g supernatant incubations were 2 mM, those with isolated perfused livers were up to 4 mM, but those in blood in vivo were below 0.3 mM; the effect of PB induction in vivo disappeared along with decreasing benzene and increasing phenol blood concentrations which surpassed benzene 2-3 h after oral benzene administration. The effect of benzene concentration on the manifestation of PB induction is also supported by almost a 2-fold increased phenol formation in PB rats over controls in vivo after repeated administration of benzene. The elimination of radioactive metabolites of orally administered benzene-/sup 14/C, in urine was markedly inhibited by intraperitoneal administration of phenol, but not by pyrocatechol, resorcinol or hydroquinol suggesting that phenol might inhibit benzene metabolism in vivo especially when its concentration exceeds that of benzene.

  18. Microwell engineering characterization for mammalian cell culture process development.

    Science.gov (United States)

    Barrett, Timothy A; Wu, Andrew; Zhang, Hu; Levy, M Susana; Lye, Gary J

    2010-02-01

    Experimentation in shaken microplate formats offers a potential platform technology for the rapid evaluation and optimization of cell culture conditions. Provided that cell growth and antibody production kinetics are comparable to those found in currently used shake flask systems then the microwell approach offers the possibility to obtain early process design data more cost effectively and with reduced material requirements. This work describes a detailed engineering characterization of liquid mixing and gas-liquid mass transfer in microwell systems and their impact on suspension cell cultures. For growth of murine hybridoma cells producing IgG1, 24-well plates have been characterized in terms of energy dissipation (P/V) (via Computational Fluid Dynamics, CFD), fluid flow, mixing and oxygen transfer rate as a function of shaking frequency and liquid fill volume. Predicted k(L)a values varied between 1.3 and 29 h(-1); liquid-phase mixing time, quantified using iodine decolorization experiments, varied from 1.7 s to 3.5 h; while the predicted P/V ranged from 5 to 35 W m(-3). CFD simulations of the shear rate predicted hydrodynamic forces will not be detrimental to cells. For hybridoma cultures however, high shaking speeds (>250 rpm) were shown to have a negative impact on cell growth, while a combination of low shaking speed and high well fill volume (120 rpm, 2,000 microL) resulted in oxygen limited conditions. Based on these findings a first engineering comparison of cell culture kinetics in microwell and shake flask formats was made at matched average energy dissipation rates. Cell growth kinetics and antibody titer were found to be similar in 24-well microtiter plates and 250 mL shake flasks. Overall this work has demonstrated that cell culture performed in shaken microwell plates can provide data that is both reproducible and comparable to currently used shake flask systems while offering at least a 30-fold decrease in scale of operation and material

  19. Near infrared spectroscopy combined with multivariate analysis for monitoring the ethanol precipitation process of fraction I+II+III supernatant in human albumin separation.

    Science.gov (United States)

    Li, Can; Wang, Fei; Zang, Lixuan; Zang, Hengchang; Alcalà, Manel; Nie, Lei; Wang, Mingyu; Li, Lian

    2017-03-15

    Nowadays, as a powerful process analytical tool, near infrared spectroscopy (NIRS) has been widely applied in process monitoring. In present work, NIRS combined with multivariate analysis was used to monitor the ethanol precipitation process of fraction I+II+III (FI+II+III) supernatant in human albumin (HA) separation to achieve qualitative and quantitative monitoring at the same time and assure the product's quality. First, a qualitative model was established by using principal component analysis (PCA) with 6 of 8 normal batches samples, and evaluated by the remaining 2 normal batches and 3 abnormal batches. The results showed that the first principal component (PC1) score chart could be successfully used for fault detection and diagnosis. Then, two quantitative models were built with 6 of 8 normal batches to determine the content of the total protein (TP) and HA separately by using partial least squares regression (PLS-R) strategy, and the models were validated by 2 remaining normal batches. The determination coefficient of validation (Rp(2)), root mean square error of cross validation (RMSECV), root mean square error of prediction (RMSEP) and ratio of performance deviation (RPD) were 0.975, 0.501g/L, 0.465g/L and 5.57 for TP, and 0.969, 0.530g/L, 0.341g/L and 5.47 for HA, respectively. The results showed that the established models could give a rapid and accurate measurement of the content of TP and HA. The results of this study indicated that NIRS is an effective tool and could be successfully used for qualitative and quantitative monitoring the ethanol precipitation process of FI+II+III supernatant simultaneously. This research has significant reference value for assuring the quality and improving the recovery ratio of HA in industrialization scale by using NIRS.

  20. Near infrared spectroscopy combined with multivariate analysis for monitoring the ethanol precipitation process of fraction I + II + III supernatant in human albumin separation

    Science.gov (United States)

    Li, Can; Wang, Fei; Zang, Lixuan; Zang, Hengchang; Alcalà, Manel; Nie, Lei; Wang, Mingyu; Li, Lian

    2017-03-01

    Nowadays, as a powerful process analytical tool, near infrared spectroscopy (NIRS) has been widely applied in process monitoring. In present work, NIRS combined with multivariate analysis was used to monitor the ethanol precipitation process of fraction I + II + III (FI + II + III) supernatant in human albumin (HA) separation to achieve qualitative and quantitative monitoring at the same time and assure the product's quality. First, a qualitative model was established by using principal component analysis (PCA) with 6 of 8 normal batches samples, and evaluated by the remaining 2 normal batches and 3 abnormal batches. The results showed that the first principal component (PC1) score chart could be successfully used for fault detection and diagnosis. Then, two quantitative models were built with 6 of 8 normal batches to determine the content of the total protein (TP) and HA separately by using partial least squares regression (PLS-R) strategy, and the models were validated by 2 remaining normal batches. The determination coefficient of validation (Rp2), root mean square error of cross validation (RMSECV), root mean square error of prediction (RMSEP) and ratio of performance deviation (RPD) were 0.975, 0.501 g/L, 0.465 g/L and 5.57 for TP, and 0.969, 0.530 g/L, 0.341 g/L and 5.47 for HA, respectively. The results showed that the established models could give a rapid and accurate measurement of the content of TP and HA. The results of this study indicated that NIRS is an effective tool and could be successfully used for qualitative and quantitative monitoring the ethanol precipitation process of FI + II + III supernatant simultaneously. This research has significant reference value for assuring the quality and improving the recovery ratio of HA in industrialization scale by using NIRS.

  1. Astrocytes Enhance Streptococcus suis-Glial Cell Interaction in Primary Astrocyte-Microglial Cell Co-Cultures.

    Science.gov (United States)

    Seele, Jana; Nau, Roland; Prajeeth, Chittappen K; Stangel, Martin; Valentin-Weigand, Peter; Seitz, Maren

    2016-06-13

    Streptococcus (S.) suis infections are the most common cause of meningitis in pigs. Moreover, S. suis is a zoonotic pathogen, which can lead to meningitis in humans, mainly in adults. We assume that glial cells may play a crucial role in host-pathogen interactions during S. suis infection of the central nervous system. Glial cells are considered to possess important functions during inflammation and injury of the brain in bacterial meningitis. In the present study, we established primary astrocyte-microglial cell co-cultures to investigate interactions of S. suis with glial cells. For this purpose, microglial cells and astrocytes were isolated from new-born mouse brains and characterized by flow cytometry, followed by the establishment of astrocyte and microglial cell mono-cultures as well as astrocyte-microglial cell co-cultures. In addition, we prepared microglial cell mono-cultures co-incubated with uninfected astrocyte mono-culture supernatants and astrocyte mono-cultures co-incubated with uninfected microglial cell mono-culture supernatants. After infection of the different cell cultures with S. suis, bacteria-cell association was mainly observed with microglial cells and most prominently with a non-encapsulated mutant of S. suis. A time-dependent induction of NO release was found only in the co-cultures and after co-incubation of microglial cells with uninfected supernatants of astrocyte mono-cultures mainly after infection with the capsular mutant. Only moderate cytotoxic effects were found in co-cultured glial cells after infection with S. suis. Taken together, astrocytes and astrocyte supernatants increased interaction of microglial cells with S. suis. Astrocyte-microglial cell co-cultures are suitable to study S. suis infections and bacteria-cell association as well as NO release by microglial cells was enhanced in the presence of astrocytes.

  2. Enhancing the culturability of bacteria from the gastrointestinal tract of farmed adult turbot Scophthalmus maximus

    Science.gov (United States)

    Xing, Mengxin; Hou, Zhanhui; Qu, Yanmei; Liu, Bin

    2014-03-01

    Eighteen agar media were tested for the culture of gut-associated bacteria from farmed adult turbot ( Scophthalmus maximus), including 16 agar media with or without 1% gastrointestinal (GI) supernatant, or with 2% or 4% GI supernatant. A total of 1 711 colonies were analyzed and 24 operational taxonomic units (OTUs) were identified. The greatest bacterial diversity was isolated on Zobell 2216E/Zobell 2216E+ agar media, whereas MRS/MRS+ agar media produced a low diversity of colonies. Agar media with GI supernatant (1%, 2%, or 4%) showed increased diversity and yielded different profiles of OTUs from the corresponding original media, suggesting that GI supernatant provides substances that enhance the culture efficiency of bacteria from the turbot GI tract. The large majority of the colonies (82%) were γ-Proteobacteria, whereas 15.6% and 2.4% of colonies were Firmicutes and Actinobacteria, respectively. At the genus level, 49.4% of all colonies were assigned to Vibrio. Other potential pathogens, including Pseudomonas, Photobacterium, and Enterobacter, and potential probiotics, including Bacillus, Paenibacillus, and Pseudomonas, were also isolated on agar media. Most cultured bacteria belonged to species that were first described in the turbot GI tract. The impact of these species on turbot physiology and health should be investigated further.

  3. A self-feeding roller bottle for continuous cell culture.

    Science.gov (United States)

    Berson, R Eric; Friederichs, Goetz

    2008-01-01

    The concept of a self-feeding roller bottle that delivers a continuous supply of fresh media to cells in culture, which is mechanically simplistic and works with existing roller apparatuses, is presented here. A conventional roller bottle is partitioned into two chambers; one chamber contains the fresh culture media reservoir, and the other contains the cell culture chamber. A spiroid of tubing inside the fresh media reservoir acts as a pump when the bottle rotates on its horizontal axis, continuously delivering fresh media through an opening in the partition to the cell culture chamber. The modified bottle proved capable of maintaining steady-state cell densities of a hybridoma cell line over the 10-day period tested, although at lower densities than reached during batch operation due to the continuous volume dilution. Steady-state density proved to be controllable by adjusting the perfusion rate, which changes with the rotation rate of the bottle. Specific antibody production rate is as much as 3.7 times the rate in conventional roller bottles operating with intermittent batch feeding.

  4. Characteristics of HCV replication and expression in a cultured human liver carcinoma cell line in vitro

    Institute of Scientific and Technical Information of China (English)

    SONG Zhi-qing; HAO Fei; MIN Feng; LIU Dao-jian

    2001-01-01

    Objective: To establish a cell culture system to support HCV long-term replication in vitro. Methods: A human hepatoma cell line 7721 was tested for its susceptibility to HCV by incubating with a serum from chronic hepatitis C patient. Cells and supernatant of the culture medium were harvested at various time-phases during the culturing periods. The presence of HCV RNA, the expression of HCV antigens in cells and/or supematant were examined with RT-PCR, in situ hybridization and immunohistochemistry respectively. Results: It was found that the intracellular HCV RNA was first detected on the 2nd day after culture, and then could be intermittently detected in both cells and supernatant over a period of at least 3 months after culture. HCV NS3, CP10 antigens were expressed in the cells. The fresh cells could be infected with the supernatant from cultured infected cells and the transmission of viral genome from HCV-infected 7721 cells to peripheral blood mononuclear cells (PBMCs) was also observed. Conclusion: Our findings suggest that the human liver carcinoma cell line7721 is not only susceptible to HCV but also can support its long replication in vitro. This cell line with HCV infection in vitro can serve as a useful tool for the study of the mechanism of HCV infection and replication, the evaluation of antiviral agents, and the primary selection of neutralization assays and HCV vaccine development.

  5. Catalase expression is modulated by vancomycin and ciprofloxacin and influences the formation of free radicals in Staphylococcus aureus cultures

    DEFF Research Database (Denmark)

    Wang, Ying; Hougaard, Anni Bygvrå; Paulander, Wilhelm Erik Axel

    2015-01-01

    formation in the human pathogen Staphylococcus aureus treated with a bactericidal antibiotic, vancomycin or ciprofloxacin. While we were unable to detect ESR signals in bacterial cells, hydroxyl radicals were observed in the supernatant of bacterial cell cultures. Surprisingly, the strongest signal...

  6. Identification and localization of a soluble antigen, Ag2, of 136 kDa from Plasmodium falciparum in vitro cultures

    DEFF Research Database (Denmark)

    Jakobsen, P H; Grellier, P; Theander, T G

    1991-01-01

    The soluble antigens, antigen 2 (Ag2) and antigen 6 (Ag6), were copurified from supernatants of P. falciparum in vitro cultures by affinity chromatography and Fast Protein Liquid Chromatography. Rabbit antibodies to Ag2 were raised and characterized by crossed immunoelectrophoresis. Ag2 appeared ...... vacuole and in clefts in the infected erythrocyte cytoplasma as shown by immunogold electron microscopy....

  7. Inflammatory cytokine detection in adenotonsill and peripheral blood mononuclear cells- culture in adenotonsillectomy patients: a comparative study

    Directory of Open Access Journals (Sweden)

    Farhadi M

    2013-04-01

    Full Text Available Background: Tonsils and adenoid hypertrophy is a major respiratory symptom in children which is partly due to recruitment of inflammatory cells in upper airway lymph nodes as a result of the effects of synthesis and release of different inflammatory cytokines. It seems that infections play role in concert with these cytokines leading to tonsilar hypertrophy and other pathologic consequences. It is proposed that cellular infiltrate of tonsils and adenoids may secrete different quantities of these cytokines compared with peripheral blood mononuclear cells (PBMC cultures.Methods: Among patients who were admitted for adenotonsillectomy to the ENT ward, 37 patients, under 1-12 years old patients with fulfill criteria selected to include the study. Excised adenoid and tonsils cultured and inflammatory cytokines Interferon-γ (INF-γ, Interlukine-1 (IL-1, IL-6, IL-8 and tumor necrosis factor-α (TNF-α measured in cellular culture supernatant. The same cytokines measured in PBMC cultures.Results: The data shows that there is a significant difference between IFN-γ and IL-8 amounts in adenoid tissue culture supernatant and PBMC culture of our patients. Furth-ermore, the amounts of IFN-γ, IL-1 and IL-8 showed considerable difference between tonsilar tissue culture supernatant and PBMC culture of these patients. Although there is a significant correlation between IL-6 amounts in tissue culture supernatant and PBMC culture (P=0.02, the respective data for TNF is only almost significant.Conclusion: Inflammatory cytokines may have significant role in the early provoke of inflammation occurred in hypertrophied tonsils and adenoid. The majority of these cyt-okines increase the expression of adhesion molecules on epithelial cells and influence the recruitment of leucocytes and inflamed tonsils. On the other hand lack of sufficient cytokine release may lead to persistent infections and may cause chronic inflammation and hypertrophied tissue.

  8. Diminished exoproteome of Frankia spp. in culture and symbiosis.

    Science.gov (United States)

    Mastronunzio, J E; Huang, Y; Benson, D R

    2009-11-01

    Frankia species are the most geographically widespread gram-positive plant symbionts, carrying out N(2) fixation in root nodules of trees and woody shrubs called actinorhizal plants. Taking advantage of the sequencing of three Frankia genomes, proteomics techniques were used to investigate the population of extracellular proteins (the exoproteome) from Frankia, some of which potentially mediate host-microbe interactions. Initial two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of culture supernatants indicated that cytoplasmic proteins appeared in supernatants as cells aged, likely because older hyphae lyse in this slow-growing filamentous actinomycete. Using liquid chromatography coupled to tandem mass spectrometry to identify peptides, 38 proteins were identified in the culture supernatant of Frankia sp. strain CcI3, but only three had predicted export signal peptides. In symbiotic cells, 42 signal peptide-containing proteins were detected from strain CcI3 in Casuarina cunninghamiana and Casuarina glauca root nodules, while 73 and 53 putative secreted proteins containing signal peptides were identified from Frankia strains in field-collected root nodules of Alnus incana and Elaeagnus angustifolia, respectively. Solute-binding proteins were the most commonly identified secreted proteins in symbiosis, particularly those predicted to bind branched-chain amino acids and peptides. These direct proteomics results complement a previous bioinformatics study that predicted few secreted hydrolytic enzymes in the Frankia proteome and provide direct evidence that the symbiosis succeeds partly, if not largely, because of a benign relationship.

  9. Analysis of tank 39H (HTF-39-15-61, 62) surface and subsurface supernatant samples in support of corrosion control program

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2015-08-01

    This report provides the results of analyses on Tanks 39H surface and subsurface supernatant liquid samples in support of the Corrosion Control Program. Analyses included warm acid strike preparation followed by analysis for silicon, aluminum, and sodium and water dilution preparation followed by analysis for anions. Other reported analytical results include analyses results for uranium, Pu-241 and Pu-239. The measured sodium concentration averaged, respectively, 4.28E+00 ± 9.30E-02 M and 4.32E+00 ± 1.076E-01 M in the Tank 39H surface sample and Tank 39H subsurface sample. In general, the nitrate, nitrite, free-OH and specific gravity of the Tank 39H surface and subsurface samples were all about the same in magnitude, respectively, averaging 1.98 M, 0.314 M, 1.26 M and 1.24. The measured silicon concentration for the Tank 39H surface and subsurface samples were, respectively, 3.84E+01± 5.51E+00 and 4.14E+01± 1.17E+00 mg/L. Based on the uranium, Pu-241 and Pu-239 concentrations, the calculated U-235 equivalent is 21.41 wt% for the surface sample and 21.32 wt% for the subsurface sample.

  10. Bio-chemical process for nitrogen and phosphorus removal by draining out anaerobic rich phosphate supernatant in ERP-SBR system

    Institute of Scientific and Technical Information of China (English)

    JIFangying; XUXiaoyi; LUOGuyuan

    2003-01-01

    The method of fixed phosphate coming from anaerobic reactor by the auxiliary chemical process is applied in External Recycle Process-SBR (ERP-SBR). This process changes the model of draining out activated sludge in the traditional biological phosphorus removal system to discharge anaerobic poly-phosphate supernatant. This process eliminates the contradiction of control for Solid Removal Time (SRT) in process of biological nitrogen and phosphorus removal. It can obtain high removal efficiency of nitrogen(N) and phosphorus(P) in longer SRT. Experiment results show that: when SRT=50 ~ 80 d, TN=28.6~ 58.3 mg/L, TP=5.5~ 13.5 mg/L in influent, COD≤ 34mg/L, TN≤ 6.02 mg/L, PO43-≤0.23 mg/L in effluent. The amount of lime is only 5% of traditional methods. The phosphorus content in the chemical sludge is 12 %~15 % and the recycle of phosphorus can be realized easily.

  11. Isolation of monoclonal antibody from a Chinese hamster ovary supernatant. II: dynamics of the integrated separation on ion exchange and hydrophobic interaction chromatography media.

    Science.gov (United States)

    Marek, Wojciech; Muca, Renata; Woś, Sylwia; Piątkowski, Wojciech; Antos, Dorota

    2013-08-30

    Dynamics of the purification process of a CHO derived monoclonal antibody by ion exchange chromatography (IEC), hydrophobic interaction chromatography (HIC) and their integration has been investigated. To quantify the adsorption behavior of the target protein (IgG1) and impurities contained in the supernatant, their elution course on IEC and HIC columns has been analyzed versus pH and/or the salt concentration in the mobile phase. A short-cut method has been proposed for mathematical modeling and determining underlying kinetic and thermodynamic parameters. The accuracy of the model predictions has been verified by comparing the simulated and experimental band profiles recorded in both chromatographic processes. After verification, the model was used to optimize operating conditions for the column loading and chromatographic elution in the integrated process IEC/HIC. Two alternative loading techniques based on the upstream and downstream feed dilution were taken into account in the optimization routine. In the first one the feed stream was diluted with the loading buffer prior to the column loading, while in the latter one the feed dilution was realized inside the column using the multiple-injection technique. It was shown that the downstream dilution allowed significant reduction of the contact time between the protein and the loading buffer.

  12. Use of an adaptable cell culture kit for performing lymphocyte and monocyte cell cultures in microgravity

    Science.gov (United States)

    Hatton, J. P.; Lewis, M. L.; Roquefeuil, S. B.; Chaput, D.; Cazenave, J. P.; Schmitt, D. A.

    1998-01-01

    The results of experiments performed in recent years on board facilities such as the Space Shuttle/Spacelab have demonstrated that many cell systems, ranging from simple bacteria to mammalian cells, are sensitive to the microgravity environment, suggesting gravity affects fundamental cellular processes. However, performing well-controlled experiments aboard spacecraft offers unique challenges to the cell biologist. Although systems such as the European 'Biorack' provide generic experiment facilities including an incubator, on-board 1-g reference centrifuge, and contained area for manipulations, the experimenter must still establish a system for performing cell culture experiments that is compatible with the constraints of spaceflight. Two different cell culture kits developed by the French Space Agency, CNES, were recently used to perform a series of experiments during four flights of the 'Biorack' facility aboard the Space Shuttle. The first unit, Generic Cell Activation Kit 1 (GCAK-1), contains six separate culture units per cassette, each consisting of a culture chamber, activator chamber, filtration system (permitting separation of cells from supernatant in-flight), injection port, and supernatant collection chamber. The second unit (GCAK-2) also contains six separate culture units, including a culture, activator, and fixation chambers. Both hardware units permit relatively complex cell culture manipulations without extensive use of spacecraft resources (crew time, volume, mass, power), or the need for excessive safety measures. Possible operations include stimulation of cultures with activators, separation of cells from supernatant, fixation/lysis, manipulation of radiolabelled reagents, and medium exchange. Investigations performed aboard the Space Shuttle in six different experiments used Jurkat, purified T-cells or U937 cells, the results of which are reported separately. We report here the behaviour of Jurkat and U937 cells in the GCAK hardware in ground

  13. Organ culture system as a means to detect celiac disease.

    Science.gov (United States)

    Picarelli, Antonio; Libanori, Valerio; De Nitto, Daniela; Saponara, Annarita; Di Tola, Marco; Donato, Giuseppe

    2010-01-01

    Anti-endomysial and anti-transglutaminase antibodies can be produced in vitro by the intestinal mucosa of celiac disease (CD) patients in clinical remission, when the culture is performed in the presence of gliadin peptides. Our aim was to use this organ culture system as a means to detect the pathognomonic antibodies of celiac disease (CD) in the culture supernatants. Organ culture was performed in the presence of three different activators to evaluate which one induced the strongest antibody response in intestinal mucosa from patients in clinical remission of CD. Our data confirm the high efficiency of synthetic peptide 31-43 as a specific immunological activator in CD and demonstrate its capability to stimulate production/secretion of CD-specific antibodies. We envision that this organ culture system may prove to be useful as a new technique for CD diagnosis.

  14. Concentration Dependent Influence of Lipopolysaccharides on Separation of Hoof Explants and Supernatant Lactic Acid Concentration in an Ex Vivo/In Vitro Laminitis Model.

    Science.gov (United States)

    Reisinger, Nicole; Schaumberger, Simone; Nagl, Veronika; Hessenberger, Sabine; Schatzmayr, Gerd

    2015-01-01

    Laminitis is one of the most common diseases in horses. It is not only painful for the animal, but also has a significant financial impact on the equine industry. This multifactorial disease affects the connective tissue of the hoof. However, the pathogenesis of laminitis is still not fully understood. Endotoxins, also known as lipopolysaccharides (LPS), and bacterial exotoxins seem to play an important role during the development of laminitis. The aim of our study was to investigate the effect of increasing LPS concentrations (0, 2.5, 5, 10, and 100 μg/mL) on cell viability of isolated epidermal and dermal hoof cells as well as on the tissue integrity of hoof explants. Furthermore, glucose, acetic acid, lactic acid, and propionic acid concentrations in explant supernatants were measured to evaluate the energy metabolism in the hoof tissue. LPS did not exhibit cytotoxic effects on epidermal or dermal cells. Force required to separate LPS treated hoof explants decreased in a concentration dependent manner. Specifically, explants incubated with 10 and 100 μg/mL needed significantly less force to separate compared to control explants. Lactic acid concentrations were significantly decreased in explants incubated with 5, 10, or 100 μg/mL LPS, while glucose, acetic acid and propionic acid concentrations were unaffected by LPS treatment. Our study indicates that LPS has no cytotoxic effect on epidermal and dermal cells isolated from hoof tissue, but impairs integrity of hoof explants. In addition, LPS led to an alteration of the lactic acid production in the lamellar tissue. Since our data highlight that LPS can affect the integrity of the equine hoof tissue in vitro, endotoxins should be further explored for their contribution to facilitate the development of laminitis.

  15. Concentration Dependent Influence of Lipopolysaccharides on Separation of Hoof Explants and Supernatant Lactic Acid Concentration in an Ex Vivo/In Vitro Laminitis Model.

    Directory of Open Access Journals (Sweden)

    Nicole Reisinger

    Full Text Available Laminitis is one of the most common diseases in horses. It is not only painful for the animal, but also has a significant financial impact on the equine industry. This multifactorial disease affects the connective tissue of the hoof. However, the pathogenesis of laminitis is still not fully understood. Endotoxins, also known as lipopolysaccharides (LPS, and bacterial exotoxins seem to play an important role during the development of laminitis. The aim of our study was to investigate the effect of increasing LPS concentrations (0, 2.5, 5, 10, and 100 μg/mL on cell viability of isolated epidermal and dermal hoof cells as well as on the tissue integrity of hoof explants. Furthermore, glucose, acetic acid, lactic acid, and propionic acid concentrations in explant supernatants were measured to evaluate the energy metabolism in the hoof tissue. LPS did not exhibit cytotoxic effects on epidermal or dermal cells. Force required to separate LPS treated hoof explants decreased in a concentration dependent manner. Specifically, explants incubated with 10 and 100 μg/mL needed significantly less force to separate compared to control explants. Lactic acid concentrations were significantly decreased in explants incubated with 5, 10, or 100 μg/mL LPS, while glucose, acetic acid and propionic acid concentrations were unaffected by LPS treatment. Our study indicates that LPS has no cytotoxic effect on epidermal and dermal cells isolated from hoof tissue, but impairs integrity of hoof explants. In addition, LPS led to an alteration of the lactic acid production in the lamellar tissue. Since our data highlight that LPS can affect the integrity of the equine hoof tissue in vitro, endotoxins should be further explored for their contribution to facilitate the development of laminitis.

  16. Lactobacillus rhamnosus GG supernatant promotes intestinal barrier function, balances Treg and TH17 cells and ameliorates hepatic injury in a mouse model of chronic-binge alcohol feeding.

    Science.gov (United States)

    Chen, Rui-Cong; Xu, Lan-Man; Du, Shan-Jie; Huang, Si-Si; Wu, He; Dong, Jia-Jia; Huang, Jian-Rong; Wang, Xiao-Dong; Feng, Wen-Ke; Chen, Yong-Ping

    2016-01-22

    Impaired intestinal barrier function plays a critical role in alcohol-induced hepatic injury, and the subsequent excessive absorbed endotoxin and bacterial translocation activate the immune response that aggravates the liver injury. Lactobacillus rhamnosus GG supernatant (LGG-s) has been suggested to improve intestinal barrier function and alleviate the liver injury induced by chronic and binge alcohol consumption, but the underlying mechanisms are still not clear. In this study, chronic-binge alcohol fed model was used to determine the effects of LGG-s on the prevention of alcoholic liver disease in C57BL/6 mice and investigate underlying mechanisms. Mice were fed Lieber-DeCarli diet containing 5% alcohol for 10 days, and one dose of alcohol was gavaged on Day 11. In one group, LGG-s was supplemented along with alcohol. Control mice were fed isocaloric diet. Nine hours later the mice were sacrificed for analysis. Chronic-binge alcohol exposure induced an elevation in liver enzymes, steatosis and morphology changes, while LGG-s supplementation attenuated these changes. Treatment with LGG-s significantly improved intestinal barrier function reflected by increased mRNA expression of tight junction (TJ) proteins and villus-crypt histology in ileum, and decreased Escherichia coli (E. coli) protein level in liver. Importantly, flow cytometry analysis showed that alcohol reduced Treg cell population while increased TH17 cell population as well as IL-17 secretion, which was reversed by LGG-s administration. In conclusion, our findings indicate that LGG-s is effective in preventing chronic-binge alcohol exposure-induced liver injury and shed a light on the importance of the balance of Treg and TH17 cells in the role of LGG-s application.

  17. Urine culture

    Science.gov (United States)

    Culture and sensitivity - urine ... when urinating. You also may have a urine culture after you have been treated for an infection. ... when bacteria or yeast are found in the culture. This likely means that you have a urinary ...

  18. An integrated system for synchronous culture of animal cells under controlled conditions.

    Science.gov (United States)

    Mendoza-Pérez, Elena; Hernández, Vanessa; Palomares, Laura A; Serrato, José A

    2016-01-01

    The cell cycle has fundamental effects on cell cultures and their products. Tools to synchronize cultured cells allow the study of cellular physiology and metabolism at particular cell cycle phases. However, cells are most often arrested by methods that alter their homeostasis and are then cultivated in poorly controlled environments. Cell behavior could then be affected by the synchronization method and culture conditions used, and not just by the particular cell cycle phase under study. Moreover, only a few viable cells are recovered. Here, we designed an integrated system where a large number of cells from a controlled bioreactor culture is separated by centrifugal elutriation at high viabilities. In contrast to current elutriation methods, cells are injected directly from a bioreactor into an injection loop, allowing the introduction of a large number of cells into the separation chamber without stressful centrifugation. A low pulsation peristaltic pump increases the stability of the elutriation chamber. Using this approach, a large number of healthy cells at each cell cycle phase were obtained, allowing their direct inoculation into fully instrumented bioreactors. Hybridoma cells synchronized and cultured in this system behaved as expected for a synchronous culture.

  19. Safeguards Culture

    Energy Technology Data Exchange (ETDEWEB)

    Frazar, Sarah L.; Mladineo, Stephen V.

    2012-07-01

    The concepts of nuclear safety and security culture are well established; however, a common understanding of safeguards culture is not internationally recognized. Supported by the National Nuclear Security Administration, the authors prepared this report, an analysis of the concept of safeguards culture, and gauged its value to the safeguards community. The authors explored distinctions between safeguards culture, safeguards compliance, and safeguards performance, and evaluated synergies and differences between safeguards culture and safety/security culture. The report concludes with suggested next steps.

  20. Organizational Culture

    Directory of Open Access Journals (Sweden)

    Adrian HUDREA

    2006-02-01

    Full Text Available Cultural orientations of an organization can be its greatest strength, providing the basis for problem solving, cooperation, and communication. Culture, however, can also inhibit needed changes. Cultural changes typically happen slowly – but without cultural change, many other organizational changes are doomed to fail. The dominant culture of an organization is a major contributor to its success. But, of course, no organizational culture is purely one type or another. And the existence of secondary cultures can provide the basis for change. Therefore, organizations need to understand the cultural environments and values.

  1. The method used to culture host cells (Sf9 cells) can affect the qualities of baculovirus budding particles expressing recombinant proteins.

    Science.gov (United States)

    Hattori, Tomomi; Nakanishi, Kohei; Mori, Takaaki; Tomita, Masahiro; Tsumoto, Kanta

    2016-01-01

    Budded virus (BV) particles of baculovirus (Autographa californica nucleopolyhedrovirus, AcNPV) are harvested from the supernatant of liquid culture of Sf9 host cells by ultracentrifugation. Using polyacrylamide gel electrophoresis, Western blot and transmission electron microscopy (TEM) of BV samples fractionated closely by sucrose density gradient centrifugation, we observed that BVs exhibited different qualities depending on whether they had been harvested from the supernatant from a standing (static), shaking (suspension), or standing/shaking (pre-/post-infection) culture of Sf9 cells. The amount of BV protein apparently increased in the order of standing, standing/shaking, and shaking procedure, and the yield of intact particles showed an opposite trend. TEM observation clearly showed that appropriate fractions of the standing and standing/shaking cultures contained more intact BV particles than those from the shaking culture. These results suggest that the qualities of recombinant BV particles may be related to the culture conditions of the host cells.

  2. Identification and localization of a soluble antigen, Ag2, of 136 kDa from Plasmodium falciparum in vitro cultures

    DEFF Research Database (Denmark)

    Jakobsen, P H; Grellier, P; Theander, T G

    1991-01-01

    The soluble antigens, antigen 2 (Ag2) and antigen 6 (Ag6), were copurified from supernatants of P. falciparum in vitro cultures by affinity chromatography and Fast Protein Liquid Chromatography. Rabbit antibodies to Ag2 were raised and characterized by crossed immunoelectrophoresis. Ag2 appeared ...... vacuole and in clefts in the infected erythrocyte cytoplasma as shown by immunogold electron microscopy....

  3. Industrial cultures

    DEFF Research Database (Denmark)

    Rasmussen, Lauge Baungaard

    1996-01-01

    The chapter deals with different paradigms andtheories of cultural development. The problem toexplain change and methods to analyse developmentin different cultures are presented and discussed.......The chapter deals with different paradigms andtheories of cultural development. The problem toexplain change and methods to analyse developmentin different cultures are presented and discussed....

  4. Human cerebrospinal fluid fatty acid levels differ between supernatant fluid and brain-derived nanoparticle fractions, and are altered in Alzheimer's disease.

    Directory of Open Access Journals (Sweden)

    Alfred N Fonteh

    Full Text Available Although saturated (SAFA, monounsaturated (MUFA, and polyunsaturated (PUFA fatty acids are important structural components of neuronal membranes and precursors of signaling molecules, knowledge of their metabolism in Alzheimer's disease (AD is limited. Based on recent discovery that lipids in cerebrospinal fluid (CSF are distributed in both brain-derived nanoparticles (NP and supernatant fluid (SF, we hypothesized that fatty acid (FA abundance and distribution into these compartments is altered in early AD pathology.We assayed the FA composition and abundance in CSF fractions from cognitively healthy (CH, mild cognitive impairment (MCI, and AD study participants using gas chromatography-mass spectrometry. In the SF fraction, concentration of docosahexaenoic acid [DHA, (C22:6n-3] was less in AD compared with CH, while alpha linolenic acid [α-LNA, (C18:3n-3] was lower in MCI compared with CH. In the NP fraction, levels of SAFAs (C15:0, C16:0 and a MUFA (C15:1 differentiated CH from MCI, while two MUFAs (C15:1, C19:1 and four PUFAs (C20:2n-6, C20:3n-3, C22:4n-6, C22:5n-3 were higher in AD compared with CH. Levels of even-chain free SAFA and total free FA levels were higher in AD, levels of odd-chain free SAFAs, MUFAs, n-3 PUFAs, and total PUFA, were lower in AD compared with CH. Free n-6 PUFA levels were similar in all three groups.FA metabolism is compartmentalized differently in NP versus SF fractions of CSF, and altered FA levels reflect the importance of abnormal metabolism and oxidative pathways in AD. Depleted DHA in CSF fractions in AD is consistent with the importance of n-3 PUFAs in cognitive function, and suggests that disturbed PUFA metabolism contributes to AD pathology. This study of FA levels in CSF fractions from different cognitive stages shows potential AD biomarkers, and provides further insight into cell membrane dysfunctions, including mechanisms leading to amyloid production.

  5. Combined Analysis of IFN-γ, IL-2, IL-5, IL-10, IL-1RA and MCP-1 in QFT Supernatant Is Useful for Distinguishing Active Tuberculosis from Latent Infection.

    Directory of Open Access Journals (Sweden)

    Maho Suzukawa

    Full Text Available The QuantiFERON®-TB Gold In-Tube test (QFT, an interferon-γ release assay, is used to diagnose Mycobacterium tuberculosis, but its inaccuracy in distinguishing active tuberculosis from latent infection is a major concern. There is thus a need for an easy and accurate tool for achieving that goal in daily clinical settings. This study aimed to identify candidate cytokines for specifically differentiating active tuberculosis from latent infection. Our study population consisted of 31 active TB (tuberculosis patients, 29 LTBI (latent tuberculosis infection patients and 10 healthy control subjects. We assayed for 27 cytokines in QFT supernatants of both specific antigen-stimulated blood samples (TBAg and negative-control samples (Nil. We analyzed their specificities and sensitivities by creating receiver operating characteristic (ROC curves and measuring the area under those curves (AUCs. In TBAg-Nil supernatants, IL-10, IFN-γ, MCP-1 and IL-1RA showed high AUCs of 0.8120, 0.7842, 0.7419 and 0.7375, respectively. Compared with each cytokine alone, combined assay for these top four cytokines showed positive rates in diagnosing active TB, and GDA analysis revealed that MCP-1 and IL-5 are potent in distinguishing active TB from LTBI, with Wilk's lambda = 0.718 (p < 0.001. Furthermore, utilizing the unique characteristic of IL-2 that its TBAg-Nil supernatant levels are higher in LTBI compared to active TB, the difference between IFN-γ and IL-2 showed a large AUC of 0.8910. In summary, besides IFN-γ, IL-2, IL-5, IL-10, IL-1RA and MCP-1 in QFT supernatants may be useful for distinguishing active TB from LTBI. Those cytokines may also help us understand the difference in pathogenesis between active TB and LTBI.

  6. Cervical Cancer Cell Supernatants Induce a Phenotypic Switch from U937-Derived Macrophage-Activated M1 State into M2-Like Suppressor Phenotype with Change in Toll-Like Receptor Profile

    Science.gov (United States)

    Sánchez-Reyes, Karina; Bravo-Cuellar, Alejandro; Hernández-Flores, Georgina; Lerma-Díaz, José Manuel; Jave-Suárez, Luis Felipe; Gómez-Lomelí, Paulina; de Celis, Ruth; Aguilar-Lemarroy, Adriana; Domínguez-Rodríguez, Jorge Ramiro; Ortiz-Lazareno, Pablo Cesar

    2014-01-01

    Cervical cancer (CC) is the second most common cancer among women worldwide. Infection with human papillomavirus (HPV) is the main risk factor for developing CC. Macrophages are important immune effector cells; they can be differentiated into two phenotypes, identified as M1 (classically activated) and M2 (alternatively activated). Macrophage polarization exerts profound effects on the Toll-like receptor (TLR) profile. In this study, we evaluated whether the supernatant of human CC cells HeLa, SiHa, and C-33A induces a shift of M1 macrophage toward M2 macrophage in U937-derived macrophages. Results. The results showed that soluble factors secreted by CC cells induce a change in the immunophenotype of macrophages from macrophage M1 into macrophage M2. U937-derived macrophages M1 released proinflammatory cytokines and nitric oxide; however, when these cells were treated with the supernatant of CC cell lines, we observed a turnover of M1 toward M2. These cells increased CD163 and IL-10 expression. The expression of TLR-3, -7, and -9 is increased when the macrophages were treated with the supernatant of CC cells. Conclusions. Our result strongly suggests that CC cells may, through the secretion of soluble factors, induce a change of immunophenotype M1 into M2 macrophages. PMID:25309919

  7. Cervical Cancer Cell Supernatants Induce a Phenotypic Switch from U937-Derived Macrophage-Activated M1 State into M2-Like Suppressor Phenotype with Change in Toll-Like Receptor Profile

    Directory of Open Access Journals (Sweden)

    Karina Sánchez-Reyes

    2014-01-01

    Full Text Available Cervical cancer (CC is the second most common cancer among women worldwide. Infection with human papillomavirus (HPV is the main risk factor for developing CC. Macrophages are important immune effector cells; they can be differentiated into two phenotypes, identified as M1 (classically activated and M2 (alternatively activated. Macrophage polarization exerts profound effects on the Toll-like receptor (TLR profile. In this study, we evaluated whether the supernatant of human CC cells HeLa, SiHa, and C-33A induces a shift of M1 macrophage toward M2 macrophage in U937-derived macrophages. Results. The results showed that soluble factors secreted by CC cells induce a change in the immunophenotype of macrophages from macrophage M1 into macrophage M2. U937-derived macrophages M1 released proinflammatory cytokines and nitric oxide; however, when these cells were treated with the supernatant of CC cell lines, we observed a turnover of M1 toward M2. These cells increased CD163 and IL-10 expression. The expression of TLR-3, -7, and -9 is increased when the macrophages were treated with the supernatant of CC cells. Conclusions. Our result strongly suggests that CC cells may, through the secretion of soluble factors, induce a change of immunophenotype M1 into M2 macrophages.

  8. Glycerol production by Oenococcus oeni during sequential and simultaneous cultures with wine yeast strains.

    Science.gov (United States)

    Ale, Cesar E; Farías, Marta E; Strasser de Saad, Ana M; Pasteris, Sergio E

    2014-07-01

    Growth and fermentation patterns of Saccharomyces cerevisiae, Kloeckera apiculata, and Oenococcus oeni strains cultured in grape juice medium were studied. In pure, sequential and simultaneous cultures, the strains reached the stationary growth phase between 2 and 3 days. Pure and mixed K. apiculata and S. cerevisiae cultures used mainly glucose, producing ethanol, organic acids, and 4.0 and 0.1 mM glycerol, respectively. In sequential cultures, O. oeni achieved about 1 log unit at 3 days using mainly fructose and L-malic acid. Highest sugars consumption was detected in K. apiculata supernatants, lactic acid being the major end-product. 8.0 mM glycerol was found in 6-day culture supernatants. In simultaneous cultures, total sugars and L-malic acid were used at 3 days and 98% of ethanol and glycerol were detected. This study represents the first report of the population dynamics and metabolic behavior of yeasts and O. oeni in sequential and simultaneous cultures and contributes to the selection of indigenous strains to design starter cultures for winemaking, also considering the inclusion of K. apiculata. The sequential inoculation of yeasts and O. oeni would enhance glycerol production, which confers desirable organoleptic characteristics to wines, while organic acids levels would not affect their sensory profile.

  9. Cultural psychology.

    Science.gov (United States)

    Heine, Steven J; Ruby, Matthew B

    2010-03-01

    Humans are a cultural species, constantly navigating a complex web of culturally bound practices, norms, and worldviews. This article provides a brief overview of the relatively young field of cultural psychology, which investigates the many ways psychology and culture interweave with one another. Highlighting the cultural nature of the human species, it draws upon research on cultural evolution, enculturation, and developmental processes. This review further summarizes a number of cultural differences in how people perceive the self, and the behavioral consequences that follow from these differences, in the domains of internal and external attribution styles, motivations for self-enhancement, approach/avoidance, primary and secondary control, as well as motivations for distinctiveness and conformity. Additionally, the review discusses research on the intersection of culture and emotion, as well as cultural differences in cognition, perception, and reasoning. Copyright © 2010 John Wiley & Sons, Ltd. For further resources related to this article, please visit the WIREs website. Copyright © 2010 John Wiley & Sons, Ltd.

  10. Cultural commons and cultural evolution

    OpenAIRE

    Giangiacomo Bravo

    2010-01-01

    Culture evolves following a process that is akin to biological evolution, although with some significant differences. At the same time culture has often a collective good value for human groups. This paper studies culture in an evolutionary perspective, with a focus on the implications of group definition for the coexistence of different cultures. A model of cultural evolution is presented where agents interacts in an artificial environment. The belonging to a specific memetic group is a majo...

  11. Repellent Culture.

    Science.gov (United States)

    Carroll, Jeffrey

    2001-01-01

    Considers defining "culture," noting how it is difficult to define because those individuals defining it cannot separate themselves from it. Relates these issues to student writing and their writing improvement. Addresses violence in relation to culture. (SG)

  12. Culturing Protozoa.

    Science.gov (United States)

    Stevenson, Paul

    1980-01-01

    Compares various nutrient media, growth conditions, and stock solutions used in culturing protozoa. A hay infusion in Chalkey's solution maintained at a stable temperature is recommended for producing the most dense and diverse cultures. (WB)

  13. Throat Culture

    Science.gov (United States)

    ... products and services. Advertising & Sponsorship: Policy | Opportunities Throat Culture Share this page: Was this page helpful? Collecting | ... treatment | Getting results | see BLOOD SAMPLE Collecting A culture is a test that is often used to ...

  14. Culturing Protozoa.

    Science.gov (United States)

    Stevenson, Paul

    1980-01-01

    Compares various nutrient media, growth conditions, and stock solutions used in culturing protozoa. A hay infusion in Chalkey's solution maintained at a stable temperature is recommended for producing the most dense and diverse cultures. (WB)

  15. Safety culture

    Energy Technology Data Exchange (ETDEWEB)

    Keen, L.J. [Canadian Nuclear Safety Commission, Ottawa, Ontario (Canada)

    2003-07-01

    Safety culture has become a topic of increasing interest for industry and regulators as issues are raised on safety problems around the world. The keys to safety culture are organizational effectiveness, effective communications, organizational learning, and a culture that encourages the identification and resolution of safety issues. The necessity of a strong safety culture places an onus on all of us to continually question whether the safety measures already in place are sufficient, and are being applied. (author)

  16. Cultural clashes

    National Research Council Canada - National Science Library

    2001-01-01

    ... is essentially a cultural, not religious, practice. Siddiq Bazarwala Singapore From Georgina Dubourcq Sir: While I don't disagree with Dr Dalrymple's comments about the way Muslim culture can treat women, I wonder how a culture which condones lap-dancing, pornographic videos and often turns a blind eye to under-age prostitution can really feel superior. I...

  17. Handling Culture

    NARCIS (Netherlands)

    Pieter van Nispen tot Pannerden

    2011-01-01

    The article indicates how companies may prepare for and deal with cultural differences. Because the research base is still rather limited an overall perspective may not be realised. After discussing definitions and concepts of culture, as well as values, cultural differences between states are discu

  18. Handling Culture

    NARCIS (Netherlands)

    Nispen tot Pannerden, P.J.M. van

    2011-01-01

    The article indicates how companies may prepare for and deal with cultural differences. Because the research base is still rather limited an overall perspective may not be realised. After discussing definitions and concepts of culture, as well as values, cultural differences between states are discu

  19. Beyond Culture.

    Science.gov (United States)

    Barron, Daniel D.

    1993-01-01

    Discusses the lack of literature relating to cultural differences and school library media programs and reviews the book "Beyond Culture" by Edward T. Hall. Highlights include the population/environment crisis, cultural literacy, the use of technology, and Marshall McLuhan's idea of the global village. (LRW)

  20. Cultural Rights and Cultural Diversity

    Institute of Scientific and Technical Information of China (English)

    WANG SIXIN

    2011-01-01

    @@ Culture is a very big concept, big enough almost to comprise all the activities of human beings and the tangible and intangible results caused by human activities.Therefore, it is very difficult to define culture in a few words.

  1. Culture evolves.

    Science.gov (United States)

    Whiten, Andrew; Hinde, Robert A; Laland, Kevin N; Stringer, Christopher B

    2011-04-12

    Culture pervades human lives and has allowed our species to create niches all around the world and its oceans, in ways quite unlike any other primate. Indeed, our cultural nature appears so distinctive that it is often thought to separate humanity from the rest of nature and the Darwinian forces that shape it. A contrary view arises through the recent discoveries of a diverse range of disciplines, here brought together to illustrate the scope of a burgeoning field of cultural evolution and to facilitate cross-disciplinary fertilization. Each approach emphasizes important linkages between culture and evolutionary biology rather than quarantining one from the other. Recent studies reveal that processes important in cultural transmission are more widespread and significant across the animal kingdom than earlier recognized, with important implications for evolutionary theory. Recent archaeological discoveries have pushed back the origins of human culture to much more ancient times than traditionally thought. These developments suggest previously unidentified continuities between animal and human culture. A third new array of discoveries concerns the later diversification of human cultures, where the operations of Darwinian-like processes are identified, in part, through scientific methods borrowed from biology. Finally, surprising discoveries have been made about the imprint of cultural evolution in the predispositions of human minds for cultural transmission.

  2. Spatial Culture

    DEFF Research Database (Denmark)

    2012-01-01

    – 2006. The essays published here allow us to subdivide the field of spatial culture into five major domains, summarized in the titles of chapters in the book: ”Perception and Strategies: Architecture”, ”Politics and Poetics: Urban Spaces”, ”Movements and Cityscape: Textuality”, ”Crisis and Construction......Spatial Culture – A Humanities Perspective Abstract of introductory essay by Henrik Reeh Secured by alliances between socio-political development and cultural practices, a new field of humanistic studies in spatial culture has developed since the 1990s. To focus on links between urban culture...... and modern society is, however, an intellectual practice which has a much longer history. Already in the 1980s, the debate on the modern and the postmodern cited Paris and Los Angeles as spatio-cultural illustrations of these major philosophical concepts. Earlier, in the history of critical studies, the work...

  3. Culture Shock

    Institute of Scientific and Technical Information of China (English)

    宋文玲

    2004-01-01

    Specialists say that it is not easy to get used to life in a new culture.“Culture shock”is the term these specialists use when talking about the feelings that people have in a new environment.There are three stages of culture shock,say the specialists.In the first stage,the newcomers like their new environment,Then when the fresh experience

  4. Cultural diversity

    OpenAIRE

    Raghavan, Raghu

    2011-01-01

    The concept of cultural diversity has emerged as an influential one having impact on multiple policy and legal instruments especially following the adoption of the UNESCO Convention on the Protection and Promotion of the Diversity of Cultural Expressions in 2005. The discussions on its appropriate implementation are however profoundly fragmented and often laden with political considerations. The present brief paper offers some thoughts on the meaning of cultural diversity and its implementati...

  5. Skin or nail culture

    Science.gov (United States)

    Mucosal culture; Culture - skin; Culture - mucosal; Nail culture; Culture - fingernail; Fingernail culture ... There, it is placed in a special dish (culture). It is then watched to see if bacteria, ...

  6. Suppression of MMP activity in bovine cartilage explants cultures has little if any effect on the release of aggrecanase-derived aggrecan fragments

    DEFF Research Database (Denmark)

    Wang, Bijue; Chen, Pingping; Jensen, Anne-Christine Bay

    2009-01-01

    - and aggrecanase-derived fragments of aggrecan and type II collagen into the supernatant of bovine cartilage explants cultures using neo-epitope specific immunoassays, and to associate the release of these fragments with the activity of proteolytic enzymes using inhibitors. FINDINGS: Bovine cartilage explants were...... cultured in the presence or absence of the catabolic cytokines oncostatin M (OSM) and tumor necrosis factor alpha (TNFalpha). In parallel, explants were co-cultured with protease inhibitors such as GM6001, TIMP1, TIMP2 and TIMP3. Fragments released into the supernatant were determined using a range of neo......-epitope specific immunoassays; (1) sandwich (342)FFGVG-G2 ELISA, (2) competition NITEGE(373)ELISA (3) sandwich G1-NITEGE(373 )ELISA (4) competition (374)ARGSV ELISA, and (5) sandwich (374)ARGSV-G2 ELISA all detecting aggrecan fragments, and (6) sandwich CTX-II ELISA, detecting C-telopeptides of type II collagen...

  7. Development of murine monoclonal antibodies for the immunohistochemical diagnosis of systemic bovine aspergillosis

    DEFF Research Database (Denmark)

    Jensen, H.E.; Aalbaek, B.; Lind, Peter

    1996-01-01

    Murine monoclonal antibodies (MAbs) against water-soluble somatic antigens (WSSA) and the wall fraction (WF) from Aspergillus fumigatus were produced by fusion of splenocytes from immunized BALB/c mice with mouse myeloma X63-Ag 8.653 cells. The supernatants of in vitro cultured hybridomas were in.......e., aspergillosis was diagnosed. The characteristics of the 2 MAbs (MAb-WF-AF-1 and-2) raised against the WF of A. fumigatus in ELISA and immunoblotting and immunohistochemical assays justify their application for the in situ diagnosis of systemic aspergillosis of cattle....

  8. Studies on the Primary Toxic Factor of Bt9816C Culture Supernatant%Bt9816C培养上清中杀虫活性成分的研究

    Institute of Scientific and Technical Information of China (English)

    蔡峻; 关彬; 陈月华; 任改新

    2004-01-01

    Bt9816C的培养上清对棉铃虫和甜菜夜蛾具有很高的杀虫活性.热敏实验表明β-外毒素和Zwittermicin A不是其主要杀虫活性成分.Bt9816C的无晶体株保留有卵磷酯酶C和溶血素活性,但是不再具有vip3A基因,其培养上清也丧失了杀虫活性.SDS-PAGE结果显示Bt9816C培养上清中有Vip3A蛋白特征大小的89 kD的蛋白,而其无晶体株正好缺失了该蛋白.以上结果表明Bt9816C培养上清中主要的杀虫活性成分是Vip3A蛋白.

  9. Cultural probes

    DEFF Research Database (Denmark)

    Madsen, Jacob Østergaard

    2016-01-01

    The aim of this study was thus to explore cultural probes (Gaver, Boucher et al. 2004), as a possible methodical approach, supporting knowledge production on situated and contextual aspects of occupation.......The aim of this study was thus to explore cultural probes (Gaver, Boucher et al. 2004), as a possible methodical approach, supporting knowledge production on situated and contextual aspects of occupation....

  10. Manuscript Cultures

    DEFF Research Database (Denmark)

    What do Mesoamerica, Greece, Byzantium, Island, Chad, Ethiopia, India, Tibet, China and Japan have in common? Like many other cultures of the world, they share a particular form of cultural heritage: ancient handwritten documents. In 2007, scholars from some20 countries around the world gathered...

  11. Culture Stories

    DEFF Research Database (Denmark)

    Jensen, Ole B.

    2007-01-01

    by certain representations and embedded in certain norms and values. The analytical framework is applied on a case of cultural urban branding. The case is the harbour front in Aalborg, Denmark where a number of flagship architecture projects and cultural institutions are being planned. It is shown how...

  12. Cultural Communications.

    Science.gov (United States)

    Armas, Jose

    It is too often taken for granted that the communication process with culturally different children takes place as readily as it might with children from Anglo cultures. Most teachers receive training in verbal and formal communication skills; children come to school with nonverbal and informal communication skills. This initially can create…

  13. CULTURAL TOURISM

    Directory of Open Access Journals (Sweden)

    Dana POP

    2016-07-01

    Full Text Available In this paper we will try to analyse the cultural tourism. We will start by referring to the complex concepts of tourism and culture and to the synergies existing between them. We will define cultural tourism and present its appearance and evolution as well as its importance as a modern form of tourism. We will present the various types of cultural tourism with their characteristics and the specific features of cultural tourists according to their interests. We will also mention that there are advantages and disadvantages for any kind of tourism depending on the position – local communities, companies or tourists. For the future we will refer to the new partnership between UNWTO and UNESCO.

  14. Cultural History and Cultural Materialism.

    Science.gov (United States)

    Berman, Ronald

    1990-01-01

    Historicism critiques cultural history and cultural materialism as a methodology for literary analysis. Questions the finality of interpretation, how original values change, and whether dramatic history implies actual history. Using Shakespearean plays, analyzes the power and politics of a play in relation to its audience; posits that cultural…

  15. Cultural History and Cultural Materialism.

    Science.gov (United States)

    Berman, Ronald

    1990-01-01

    Historicism critiques cultural history and cultural materialism as a methodology for literary analysis. Questions the finality of interpretation, how original values change, and whether dramatic history implies actual history. Using Shakespearean plays, analyzes the power and politics of a play in relation to its audience; posits that cultural…

  16. Digging culture and doing culture

    NARCIS (Netherlands)

    Andries van den Broek; Jos de Haan; Frank Huysmans

    2009-01-01

    Original title: Cultuurbewonderaars en cultuurbeoefenaars. There are people who love art and culture and there are people who practise it; people who enjoy it and people who are themselves creative in their leisure time. Who are these culture-lovers and practitioners? How has participation in

  17. Culture Matters

    Directory of Open Access Journals (Sweden)

    Gillian Warner-Søderholm

    2012-12-01

    Full Text Available Whether managers are concerned with financial issues, marketing, or human resource management (HRM, cultural values and practices do matter. The purpose of this article is to understand Norwegian managers’ cultural values within the cross-cultural landscape of her neighbors in the “Scandinavian cluster.” Clearly, subtle but disturbing differences may surface even when representatives from similar cultures work together. As a follow on from the GLOBE project, data based on the GLOBE instrument were collected on culture and communication values in Norway from 710 Norwegian middle managers for this present study. Although the Scandinavian cultures appear ostensibly similar, the results illustrate that research can reveal subtle but important cultural differences in nations that are similar yet dissimilar. All three Scandinavian societies appear intrinsically egalitarian; they appear to value low Power Distance, directness, and consensus in decision making and to promote Gender Egalitarianism. Nevertheless, there are significant differences in the degrees of commitment to these values by each individual Scandinavian partner. These differences need to be understood and appreciated to avoid misunderstandings.

  18. Fermentanomics: Relating quality attributes of a monoclonal antibody to cell culture process variables and raw materials using multivariate data analysis.

    Science.gov (United States)

    Rathore, Anurag S; Kumar Singh, Sumit; Pathak, Mili; Read, Erik K; Brorson, Kurt A; Agarabi, Cyrus D; Khan, Mansoor

    2015-01-01

    Fermentanomics is an emerging field of research and involves understanding the underlying controlled process variables and their effect on process yield and product quality. Although major advancements have occurred in process analytics over the past two decades, accurate real-time measurement of significant quality attributes for a biotech product during production culture is still not feasible. Researchers have used an amalgam of process models and analytical measurements for monitoring and process control during production. This article focuses on using multivariate data analysis as a tool for monitoring the internal bioreactor dynamics, the metabolic state of the cell, and interactions among them during culture. Quality attributes of the monoclonal antibody product that were monitored include glycosylation profile of the final product along with process attributes, such as viable cell density and level of antibody expression. These were related to process variables, raw materials components of the chemically defined hybridoma media, concentration of metabolites formed during the course of the culture, aeration-related parameters, and supplemented raw materials such as glucose, methionine, threonine, tryptophan, and tyrosine. This article demonstrates the utility of multivariate data analysis for correlating the product quality attributes (especially glycosylation) to process variables and raw materials (especially amino acid supplements in cell culture media). The proposed approach can be applied for process optimization to increase product expression, improve consistency of product quality, and target the desired quality attribute profile.

  19. Cultural Spectrum

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Exorbitant Performance Fees ProhibitedThe Ministry of Culture has taken measures todiscourage performance artists from charging excessively for their stage appearances. According to the ministry, payment for performers and ticket prices must be in line with average con-

  20. Blood culture

    Science.gov (United States)

    Culture - blood ... A blood sample is needed . The site where blood will be drawn is first cleaned with an antiseptic such ... organism from the skin getting into (contaminating) the blood sample and causing a false-positive result (see ...

  1. Rectal culture

    Science.gov (United States)

    ... 2016:chap 283. Haines CF, Sears CL. Infectious enteritis and proctocolitis. In: Feldman M, Friedman LS, Brandt ... PA: Elsevier; 2017:chap 22. Read More Cryptosporidium enteritis Fecal culture Proctitis Review Date 5/11/2016 ...

  2. callus culture

    African Journals Online (AJOL)

    steve

    and KH2PO4 -NH4NO3 depletion was fatal to grape cells and resulted in formation ... Individual phenolic compounds found in the grape cell cultures were ...... Characterization of in vitro anthocyanin-producing sour cherry ... Process Biochem.

  3. Culture School

    Institute of Scientific and Technical Information of China (English)

    MATTHEW LIM

    2008-01-01

    @@ Until recently, employees posted abroad would, if they were lucky, receive some limited language training before they relocated. How they would cope with living in a completely new culture when they arrived was left up to them to figure out.

  4. Yangshao Culture

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Yangshao culture was born in the Neolithic Age. In 1921, archaeologists unearthed a number of chipped stone implements such as knives and axes; bone objects and everyday ceramic items. Thus Yangshao Village and its matriarchal society have

  5. Culture perspectives.

    Science.gov (United States)

    Locsin, Rozzano C

    2002-10-01

    All cultures have had means and techniques that express their immediate aims. The thing that interests me is that today, painters do not have to go to a subject matter outside of themselves. They work from a different source. They work from within. It seems to me that the modern artist cannot express this age, the airplane, the atom bomb, the radio, in the old form of the Renaissance or of any of the old cultures.

  6. Cultural Imports

    Institute of Scientific and Technical Information of China (English)

    1996-01-01

    FOREIGN commodities have flowed into China since the country opened its doors to the outside world. China is an expansive territory with a huge population offering a vast potential consumer market. There are absolutely no limits to the world of culture in China, with Chinese people having access to foreign films, dramas, music and books, all of which have helped to strengthen exchanges between Chinese and Western cultures.

  7. Cell culture plastics with immobilized interleukin-4 for monocyte differentiation

    DEFF Research Database (Denmark)

    Hansen, Morten; Hjortø, Gertrud Malene; Met, Özcan;

    2011-01-01

    Standard cell culture plastic was surface modified by passive adsorption or covalent attachment of interleukin (IL)-4 and investigated for its ability to induce differentiation of human monocytes into mature dendritic cells, a process dose-dependently regulated by IL-4. Covalent attachment of IL-4...... proceeded via anthraquinone photochemistry to introduce amine functionalities at the surface followed by coupling of IL-4 through a bifunctional amine-reactive linker. X-ray photoelectron spectroscopy showed that undesirable multilayer formation of the photoactive compound could be avoided by reaction...... in water instead of phosphate-buffered saline. Passively adsorbed IL-4 was observed to induce differentiation to dendritic cells, but analysis of cell culture supernatants revealed that leakage of IL-4 into solution could account for the differentiation observed. Covalent attachment resulted in bound IL-4...

  8. Urine culture - catheterized specimen

    Science.gov (United States)

    Culture - urine - catheterized specimen; Urine culture - catheterization; Catheterized urine specimen culture ... urinary tract infections may be found in the culture. This is called a contaminant. You may not ...

  9. Development of myiasis vaccine: In vitro detection of immunoprotective responses of peritrophic membrane protein, first instar larva Ll supernatant and pellet antigen of fly Chrysomyia bezziana in sheep

    Directory of Open Access Journals (Sweden)

    Sukarsih

    1999-10-01

    Full Text Available Myiasis control by means of individual treatment of animals which are mainly rised extensively is time consumed and expensive. The alternative way to control this disease by vaccination is considered effective and economically accepted. However the expected vaccine is now still being developed under a collaborative project between CSIRO, Inter-University Centre on Biotechnology-ITB and Research Institute for Veterinary Science and funded by ACIAR. There are several antigens have been identified as vaccine candidates and an in vitro bioassay technique has been developed for assessing the immunoresponses of vaccine in sheep. Three antigens were used for vaccines in this study, these included protein peritrophic membrane (PM, soluble extract (SE and pellet extract (PE of 1st instar larvae of Chrysomya bezziana. Twenty four experimental sheep were divided into 4 groups of 6 animals, 3 groups of animals were injected with PM, SE and PE vaccines with the dose rate of 0.5 g PM/head, 0.8 g PE/head and 4.2 ml LE/head respectively, and the other one group was injected with 4 ml PBS/head as a control group. Vaccination with the same dose was repeated 4 weeks after the 1st vaccination as a booster, and 2 weeks after the booster the sheep were challenged with live larvae, 3 days after challenge animals were killed. Sera were collected at the day of vaccination, 4 weeks after vaccination, 2 weeks after booster, and 3 days after challenge. An in vitro bioassay technique was conducted by culturing 1st instar larvae on five media containing sera collected from each experimental animal. The effects of sera on cultivated larvae were assessed by means of larval weight and larval mortality rate. The results indicated that the growth rate and survival of cultivated larvae in media containing anti-PM sera were significantly lower (P<0.01 compared to the larvae cultivated on media with sera on the day of vaccination. The larval weight depression by anti- PM sera

  10. Cultural tourism and tourism cultures

    DEFF Research Database (Denmark)

    Ooi, Can-Seng

    Presenting a comprehensive and dynamic understanding of cultural tourism, this volume examines cultural mediators and how they help tourists appreciate foreign cultures. It also shows how tourism experiences are strategically crafted by mediators, the complexity of the mediation process, and how...... various products are mediated differently. A number of different products are investigated, including destination brand identities, "living" cultures and everyday life, art and history. The author illustrates his arguments by comparing the tourism strategies of Copenhagen and Singapore, and demonstrates...... how tourism is an agent for social change. The author also offers an original and refreshing way of understanding tourist behaviour through the concept of the "versatile tourist". The book's empirical cases and dialogic framework provide new and deep insights into tourism activities. In his...

  11. Preparation and use of lipid microemulsions as nutritional supplements for culturing mammalian cells.

    Science.gov (United States)

    Darfler, F J

    1990-08-01

    Cells grown in vitro generally have a requirement for an exogenous source of lipid. This requirement is often met by the addition of serum, lipoproteins, or lipids complexed to albumin. To overcome the disadvantages of using lipoproteins or albumin for culturing cells in serum-free media, a method has been devised to provide necessary lipids. This report describes the preparation and use of protein-free lipid microemulsions suitable for use in tissue culture. The microemulsions are prepared from purified, synthetic lipids to produce a homogeneous, water-soluble, stable suspension that can be sterile-filtered. The best results were obtained using a sonicate of cholesterol oleate, dipalmitoyl phosphatidylcholine, dilinoleoyl phosphatidylcholine, dimyristoyl phosphatidylglycerol, cholesterol, sphingomyelin, alpha-tocopherol, alpha-tocopherol acetate, and Tween 80. Using Chinese hamster ovary (CHO) cells in a protein-free medium, cell growth was 222% vs. control (no microemulsion) in a 5-d assay. Inclusion of the microemulsion to protein-free media also increased the growth rate of murine hybridomas, H9 transformed T lymphoblasts, and human skin keratinocytes.

  12. A Cultural Sexuality or a Sexual Culture?

    NARCIS (Netherlands)

    Vandermeersch, Patrick

    1990-01-01

    P. Vandermeersch, A Cultural Sexuality or a Sexual Culture? In: F. VAN DE VIJVER & G. HUTSCHEMAEKERS (ed.), The Investigation of Culture. Current Issues in Cultural Psychology, Tilburg, Tilburg University Press, 1990, 43-58.

  13. Usefulness of whole blood, plasma, peritoneal fluid, and peritoneal fluid supernatant glucose concentrations obtained by a veterinary point-of-care glucometer to identify septic peritonitis in dogs with peritoneal effusion.

    Science.gov (United States)

    Koenig, Amie; Verlander, Lindsey Lane

    2015-11-01

    To evaluate the usefulness of a veterinary point-of-care glucometer for identification of septic peritonitis in dogs with peritoneal effusion (PE). Prospective clinical evaluation. 39 dogs with PE. Blood and peritoneal fluid convenience samples were collected concurrently in all dogs at the time of initial evaluation. A veterinary point-of-care glucometer was used to measure glucose concentration in heparinized whole blood, plasma, peritoneal fluid, and peritoneal fluid supernatant samples. Seventeen dogs had confirmed septic peritonitis, and 22 dogs had nonseptic PE. Sensitivity, specificity, positive and negative predictive values, and accuracy of identification of dogs with septic peritonitis were calculated for glucose concentration differences for whole blood versus peritoneal fluid (WB-PF), plasma versus peritoneal fluid (P-PF), and plasma versus peritoneal fluid supernatant (P-PFS). With a cutoff of > 20 mg/dL, the glucose concentration difference for WB-PF was an insensitive indicator of septic peritonitis (sensitivity, 41.2%; specificity, 100%). In comparison, the glucose concentration differences for P-PF and P-PFS had a higher sensitivity for septic peritonitis (88.2% and 82.4%, respectively) but a lower specificity (80% and 77.8%, respectively). With a glucose concentration difference cutoff of ≥ 38 mg/dL, specificity, positive predictive value, and accuracy of P-PF and P-PFS improved. Determination of the glucose concentration difference for WB-PF with the veterinary point-of-care glucometer was not useful in identifying all dogs with septic peritonitis. A glucose concentration difference of ≥ 38 mg/dL for P-PF or P-PFS, however, supported an accurate diagnosis of septic peritonitis in dogs with PE.

  14. Disinfection effects of ultraviolet on supernatants of flocculated digestate from swine farm%猪场沼液絮凝上清液的紫外线杀菌效果

    Institute of Scientific and Technical Information of China (English)

    李同; 董红敏; 陶秀萍

    2014-01-01

    There are a large number of microorganisms in the liquid digestate from a swine farm. The liquid digestate should be disinfected prior to discharge or utilization for sanitary and environmental safety, but there is a lack of literature on the disinfection of liquid digestate from animal feeding operations. In view of the high turbidity and chromaticity, the transmittance (T254) of liquid digestate from swine farm is nearly zero, and liquid digestate must be treated prior to ultraviolet disinfection. Flocculation was applied to pre-treat liquid digestate in this study in order to obtain supernatants with different transmittances (T254) for the following disinfecting trials. In terms of total bacteria count, total coliforms, and fecal coliforms, experiments were conducted to investigate the disinfecting feasibility and the effectiveness of ultraviolet on the supernatants of flocculated digestate under 60 experimental conditions, which were formed by 4×3×5 factorial combinations of transmittances (T254) (0.01%, 0.69%, 3.78%and 8.54%), water depth (WD) (1, 2 and 3 cm), and hydraulic retention times (HRT) (1, 5, 10, 15 and 20 min). Results showed that T254, WD, and HRT all significantly affected (P<0.01) the disinfection effects of ultraviolet on the supernatants of flocculated digestate, and significant interactions (P<0.01) were also detected among the experimental factors. The disinfection rates of ultraviolet on total bacteria count, total coliforms, and fecal coliforms were (99.99±1.20)%, (99.99±1.43)%, and (100.00±0.01)%respectively, under the condition of 0.69%T254, 2 cm WD, and 15 min HRT. In such cases, the fecal coliforms in the supernatant of flocculated digestate decreased from 3.9×106 count/L to less than 3 count/L. Therefore, ultraviolet could be applied for the disinfection of the supernatants of flocculated digestate from swine farms, and the disinfected effluent could meet the sanitary requirements of the current national standards. It is

  15. Cultural Robotics: The Culture of Robotics and Robotics in Culture

    OpenAIRE

    2013-01-01

    In this paper, we have investigated the concept of "Cultural Robotics" with regard to the evolution of social into cultural robots in the 21st Century. By defining the concept of culture, the potential development of a culture between humans and robots is explored. Based on the cultural values of the robotics developers, and the learning ability of current robots, cultural attributes in this regard are in the process of being formed, which would define the new concept of cultural robotics. Ac...

  16. Cultural Resurrection

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    "Who are we?Where are we from?"Humans have been pondering these questions since the day they first came into being.One of the ways we preserve memories of the past is through our cul- tural heritage that has been passed on from generation to genera- tion.Intangible cultural heritage,as well as tangible cultural her- itage,is essential to the continuity of human civilization. Since the United Nations Educational,Scientific and Cultural Organization(UNESCO)unveiled the Masterpieces of the Oral and Intangible Heritage of Humanity in 2001,China has had Kunqu opera,Guqin and its music,the art of Uygur Muqam of Xinjiang and the traditional Mongolian folk song Long Song added to UNESCO’s protection list.It is now one of the coun-

  17. Japanese Shame Culture and American Guilt Culture

    Institute of Scientific and Technical Information of China (English)

    Lu Weijie

    2016-01-01

    Culture is an important factor contributing to the success of intercultural communication. In the east and west, there are many different cultures, among which Japanese shame culture and American guilt culture are two typical ones. Influenced by different cultures, these two countries have different characteristics, which reminds us that in intercultural communication culture should be paid much attention to.

  18. Culturally Responsive Teaching: Understanding Disability Culture

    Science.gov (United States)

    Darrow, Alice-Ann

    2013-01-01

    To be culturally responsive teachers, we must first have an understanding of other cultures and how students from these cultures differ from one another. As we consider the many cultures represented in our classrooms, we might also consider students with disabilities as a cultural group. Within any main culture are subgroups differentiated by…

  19. Hepatitis B virus infection and replication in primarily cultured human fetal hepatocytes

    Institute of Scientific and Technical Information of China (English)

    Min Lin; Qun Chen; Li-Ye Yang; Wen-Yu Li; Xi-Biao Cao; Jiao-Ren Wu; You-Peng Peng; Mo-Rui Chen

    2007-01-01

    AIM:To investigate the infection and replication of hepatitis B virus(HBV)in primarily cultured human fetal hepatocytes(HFHs).METHODS:The human fetal hepatocytes were cultured in serum-free medium,HBV-positive serum was added into the medium to study the susceptibility of hepatocytes to HBV infection.The supernatant was collected for ELISA assay of HBsAg and HBeAg,and quantitative fluorescence PCR for HBV-DNA assay daily.Albumin and HBcAg,CK8 and CK18 expressions were detected by immunohistochemistry in cultured hepatocytes.Content of lactate dehydrogenate(LDH)was measured to find out the integrity of the cell membrane.RESULTS:A stable hepatocyte culture system was established.HBV could infect the hepatocytes and replicate,and HBcAg expression could be detected by immunohistochemistry in hepatocyte-like cells.HBV-DNA in the supernatant could be detected from d 2 to d 18 and HBsAg and HBeAg were positive on d 3-d 18 after HBV infection.HBV in medium increased from d 0 to d 6 and subsequently decreased as the cells were progressively loosing their hepatocyte phenotypes.CONCLUSION:HBV could infect human fetal hepatocytes and replicate.This in vitro model allowed a detailed Study on early events associated with human HBV entry into cells and subsequent replication.

  20. Biocompatibility of three bioabsorbable membranes assessed in FGH fibroblasts and human osteoblast like cells culture.

    Science.gov (United States)

    Soares, Michelle Pereira Costa Mundim; Soares, Paulo Vinícius; Pereira, Analice Giovani; Moura, Camilla Christian Gomes; Soares, Priscila Barbosa Ferreira; Naves, Lucas Zago; de Magalhães, Denildo

    2014-08-06

    Specific physical and chemical features of the membranes may influence the healing of periodontal tissues after guided tissue regeneration (GTR). The aim of the present investigation was to analyze the biological effects of three bioabsorbable membranes. The hypothesis is that all tested membranes present similar biological effects. Human osteoblast like-cells (SaOs-2) and gingival fibroblasts FGH (BCRJ -RJ) were cultured in DMEM medium. The viability of the cells cultured on the membranes was assesses using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Quantitative determination of activated human Transforming Growth Factor beta 1 (TGF-β1) on the supernatants of the cell culture was observed. Samples were examined using scanning electron microscope (SEM). SaOs2, in 24 hours, PLA group showed higher values when compared to other groups (P statistical significance values when compared two times. In 4 h and 24 h, for the fibroblasts group, significantly difference was found to PLA membrane, when compared with the other groups (p statistically significant difference (p analysis of culture supernatants of fibroblasts, in 24 hours, only PLA group presented significant difference (p = 0,008). The biomaterials analyzed did not show cytotoxicity, since no membrane presented lower results than the control group. PLA membrane presented the best performance due to its higher cell viability and absorbance levels of proliferation. Both collagen membranes showed similar results either when compared to each other or to the control group.

  1. Information cultures

    DEFF Research Database (Denmark)

    2017-01-01

    The purpose of this article is to suggest a genealogy of the concept of information beyond the 20th century. The article discusses how the concept of information culture might provide a way of formulating such a genealogic strategy. The article approaches this purpose by providing a general...... narrative of premodern information cultures, examining works on early-modern scholars and 18th century savants and discussion of what seems to be a Foucauldian rupture in the conceptualization of information in 19th century England. The findings of the article are situated in the thinking that a genealogy...... of information would reveal that information had specific purposes in specific settings....

  2. Mayan Culture

    DEFF Research Database (Denmark)

    Hervik, Peter Bent

    1992-01-01

    The social categories « Maya » and « mestizo » habe been applied to denote the Yucatec Mayan people in Mexico. The A. examines the cluster of perceived attributes (schemata) evoked by the terms and how they relate to each other. He shows that there is an incongruency between them along the lines ...... of local and academic categorization, which is an implication of the different social spaces in which they arise. In spite of the incongruency and the cultural plurality evoked by their usage, the A. argues that the people of Yucatec share a single culture....

  3. Culture Consciousness

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    This year, June 10 marked China's first Cultural Heritage Day. The designation by the Chinese Government aims to raise awareness of the need to protect and understand the value of the nation's abundant cultural treasures. In future the second Saturday in June each year will be set aside for this purpose. Recently, the State Council published the sixth group of major relics under state protection. On the list are 1,080 historic relics such as the Grand Canal from Beijing to Hangzhou and the

  4. Boosting Culture

    Institute of Scientific and Technical Information of China (English)

    ZHOU JIANXIONG

    2011-01-01

    Culture makes up an indispensable part of our lives,just like material comfort.It is thought of as an important source of a nation's vitality and creativity,and constitutes a key factor uniting the nation,while making it distinctive from other countries.It is also said culture is a productive power that not only shapes human concepts and impacts their behavior,but also contributes in no small measure to the betterment of our material as well as spiritual world.

  5. MARKETING CULTURAL

    OpenAIRE

    Ramírez, Claudia Gómez

    2013-01-01

    Este artículo analiza la definición de "Marketing" Cultural y la adaptación y beneficios del "marketing" tradicional respecto al conjunto de manifestaciones artísticas de las diversas industrias involucradas en el sector cultural o artístico; asimismo, se desagregan los conceptos básicos que lo componen como factor de éxito en dichas empresas. Se hace uso de la exposición de casos específicos para ilustrar la articulación de estos dos conceptos aparentemente contrapuestos, cultura y "marketin...

  6. Morphology of primary human venous endothelial cell cultures before and after culture medium exchange.

    Science.gov (United States)

    Krüger-Genge, A; Fuhrmann, R; Jung, F; Franke, R P

    2015-01-01

    The evaluation of the interaction of human, venous endothelial cells (HUVEC) with body foreign materials on the cellular level cannot be performed in vivo, but is investigated in vitro under standard culture conditions. To maintain the vitality, proliferation and morphology of HUVEC seeded on body foreign substrates over days, the cell culture medium is usually exchanged every second day. It is well known, that alterations in the microenvironment of cells bear the risk of influencing cell morphology and function. In the current study the influence of cell culture medium exchange on HUVEC cytoskeletal microfilament structure and function was investigated. HUVEC in the third passage were seeded on extracellular matrix (ECM) - which was secreted from bovine corneal endothelial cells on glass- until functional confluence was reached. The experiment started 11 days after HUVEC seeding with an exchange of the cell culture medium followed by a staining of the actin microfilaments with phalloidin-rhodamin 1.5 and 5 minutes after medium exchange. The microfilaments were documented by use of an Olympus microscope (IMT-2) equipped with a UV lamp and online connected to a TV chain (Sony XC 50 ST/monochrome) implying an OPTIMAS - Image analysis system. Prostacyclin was analysed in the cell culture supernatant. 1.5 min after culture medium exchange in the functionally confluent cultures a slight disturbance of the actin microfilament structure with a broadening of the marginal filament band, a partial disconnection of cell-cell contacts and the appearance of intercellular fenestrations were observed. 5 minutes after medium exchange a redevelopment of the slightly disturbed microfilament structure with a condensation and narrowing of the marginal filament band was seen. 12 h later a further consolidation of the microfilament structure occurred. In addition, a perturbation of the cultured HUVEC occurred after cell culture medium exchange. The prostacyclin concentration in the

  7. Whole-blood culture is a valid low-cost method to measure monocytic cytokines - A comparison of cytokine production in cultures of human whole-blood, mononuclear cells and monocytes

    DEFF Research Database (Denmark)

    Damsgaard, Camilla T.; Lauritzen, Lotte; Calder, Philip C.

    2009-01-01

    assessed the intra- and inter-individual variation in cytokine production. In 64 healthy men (age 19-40 years) IL-6, TNF and IL-10 were measured by enzyme-linked immunosorbent assay in supernatants from whole-blood, PBMC and monocytes cultured 24 h with lipopolysaccharide (LPS) or UV-killed L acidophilus......Whole-blood and peripheral blood mononuclear cell (PBMC) cultures are used as non-validated surrogate measures of monocytic cytokine production. The aim of this investigation was to compare ex vivo cytokine production from human whole-blood and PBMC with that from isolated monocytes. We also...

  8. Cultural Usability

    DEFF Research Database (Denmark)

    Shi, Qingxin

    2007-01-01

    relationship and communicate effectively with the user in order to find relevant usability problems in culturally localized applications. It includes three parts, pilot study, field study and experiments, to get both qualitative data and quantitative data. From this project, we hope to find an effective way...

  9. Hydroponic Culture

    Science.gov (United States)

    Steucek, G. L.; Yurkiewicz, W. J.

    1973-01-01

    Describes a hydroponic culture technique suitable for student exercises in biology. This technique of growing plants in nutrient solutions enhances plant growth, and is an excellent way to obtain intact plants with root systems free of soil or other particulate matter. (JR)

  10. Culture Shock

    Science.gov (United States)

    Brown, Angela Khristin

    2013-01-01

    The migration of blacks in North America through slavery became united. The population of blacks past downs a tradition of artist through art to native born citizens. The art tradition involved telling stories to each generation in black families. The black culture elevated by tradition created hope to determine their personal freedom to escape…

  11. Hydroponic Culture

    Science.gov (United States)

    Steucek, G. L.; Yurkiewicz, W. J.

    1973-01-01

    Describes a hydroponic culture technique suitable for student exercises in biology. This technique of growing plants in nutrient solutions enhances plant growth, and is an excellent way to obtain intact plants with root systems free of soil or other particulate matter. (JR)

  12. Viable cell recycle with an inclined settler in the perfusion culture of suspended recombinant Chinese hamster ovary cells.

    Science.gov (United States)

    Searles, J A; Todd, P; Kompala, D S

    1994-01-01

    The perfusion culture of suspended mammalian cells requires a cell retention device, the best of which will retain all viable cells and reject all nonviable cells and debris. The inclined settler is a passive, simple, inexpensive, and easy-to-maintain device that has been shown in the past to selectively remove single nonviable cells of hybridoma cultures. In this work, we have demonstrated the preferential return of viable recombinant Chinese hamster ovary (CHO) cells through the use of a three-port settler maintained at lower temperatures and vibrated to reduce cell attachment and enhance cell return to the bioreactor. The residence time of CHO cells in the cooled, vibrated settler was determined by flow-cytometric discrimination of tracer recombinant CHO cells. Cells returning to the bioreactor through the underflow had an average residence time of 1.46 h in the settler. During perfusion cultures with cell densities above 10(6) cells/mL, cells seen to be stalled within the settler were easily dislodged by periodic air bubbling using a simple back-flushing procedure in which headspace gas was brought through the settler underflow port. The resuspended cells were returned to the bioreactor within an average of 32 min after bubbling. This study demonstrates that inclined sedimentation technology can be utilized to selectively recycle viable recombinant CHO cells with only a short retention time in an inclined settler.

  13. Primary targets in photochemical inactivation of cells in culture

    Science.gov (United States)

    Berg, Kristian; Jones, Stuart G.; Prydz, Kristian; Moan, Johan

    1995-01-01

    The mechanisms of photoinactivation of NHIK 3025 cells in culture sensitized by tetrasulfonated phenylporphines (TPPS4) are described). Ultracentrifugation studies on postnuclear supernatants indicated that the intracellular distribution of TPPS4 resembles that of (beta) -N-acetyl-D-glucosaminidase ((beta) -AGA), a lysosomal marker enzyme, and that the cytosolic content of TPPS4 is below the detection limit of the ultracentrifugation method. Upon light exposure more than 90% of TPPS4 was lost from the lysosomal fractions, due to lysosomal rupture. The content of TPPS4 in the postnuclear supernatants was reduced by 30 - 40% upon exposure to light. This is most likely due to binding of TPPS4 to the nuclei, which were removed from the cell extracts before ultracentrifugation, after photochemical treatment. The unpolymerized form of tubulin seems to be an important target for the photochemical inactivation of NHIK 3025 cells. Since TPPS4 is mainly localized in lysosomes it was assumed that a dose of light disrupting a substantial number of lysosomes followed by microtubule depolymerization by nocodazole would enhance the sensitivity of the cells to photoinactivation. This was confirmed by using a colony-forming assay. The increased phototoxic effect exerted by such a treatment regime could be explained by an enhanced sensitivity of tubulin to light. Another cytosolic constituent, lactate dehydrogenase, was not photoinactivated by TPPS4 and light.

  14. In vitro differentiation of MSCs into retina-like cells by the supernatant fluid of light-injured neurosensory retina%光损伤鼠视网膜片培养上清液诱导 MSCs 分化为视网膜样细胞的研究

    Institute of Scientific and Technical Information of China (English)

    白月; 徐国兴

    2014-01-01

    AIM: To explore the possibility of inducing rat mesenchymal stem cells ( MSCs) into retina-like cells by the supernatant fluid of light-injured neurosensory retina in vitro. METHODS: MSCs were isolated and attached to the wall of culture dishes by their specific adherent ability. Then the cells were characterized by flow cytometry.The neurosensory retina was isolated from retina of SD rat and it was tested by hematoxylin-eosin ( HE ) staining.The pathological changes of light-injured neurosensory retina was observed under transmission electron microscope. Three kinds of supernatant fluid of light -injured neurosensory retina of SD rats were prepared.The third passage of MSCs were cultured with these mixed medium for 7-8d, we used RT-PCR to see whether they could express rhodopsin, neuron-specific enolase (NSE), and glial fibrillary acidic protein ( GFAP ) , and positive cells were counted and analyzed. RESULTS: HE staining showed the retinal sheets included full-thickness neural retina.Neurosensory retina developed ultrastructural destructions by light injury.RT-PCR showed that the medium of mixed I expressed higher positive rate of rhodopsin (0.3915±0.00644), NSE (0.2019± 0.00682), GFAP (0.1972 ±0.00211) than the medium of mixed Ⅱ rhodopsin (0.0983 ±0.00319), NSE (0.1048 ± 0.00323), GFAP (0.1040±0.00254) and medium of mixedⅢrhodopsin(0.0044±0.00126), NSE (0.0498±0.00149), GFAP (0.0467±0.00333).The difference of intergroup has statistical significance. CONCLUTION:The supernatant fluid of light-injured neurosensory retina of SD rats can induce MSCs to differentiate into retina-like cells and provide new insights of stem cell therapy for retinopathy.%目的:应用大鼠视网膜片光损伤后的培养上清液,在体外诱导大鼠骨髓间充质干细胞( mesenchymal stem cells , MSCs)成为视网膜样细胞的可能性。  方法:贴壁筛选法分离、培养大鼠MSCs ,流式细胞仪对其细胞纯度鉴定。取材大鼠视网

  15. Serum-free culture of H pylori intensifies cytotoxicity

    Institute of Scientific and Technical Information of China (English)

    Hiroyuki Ohno; Akiyuki Murano

    2007-01-01

    AIM: To perform a long culture passage of H pylori without serum, taking into account its cytotoxicity and the presence of the probable new cytotoxic factor.METHODS: One sample of H pylori 60190 (ATCC49503) was grown on Brain Heart Infusion (BHI) agar containing 0.5% 2,6-di-O-methyl-β-cyclodextrin without any serum, being passaged 70-100 times every 3-4 d for approximately 2 h, while another sample of H pylori contained 70 mL/L fetal calf serum without 2,6-di-Omethyl-β-cyclodextrin. Their supernatant and extract after 16 h in culture were evaluated for changes in cell morphology and for cell viability using HeLa cells. Furthermore, the characteristics of the probable cytotoxic factor in the extract were examined on partial purification studies and its cytotoxicity was evaluated in various human cells.RESULTS: The supernatant and the extract of the bacterium grown on serum-free medium had strong cytotoxicity compared with those grown on serumcontaining medium. They irreversibly damaged HeLa cells without vacuolation that was altogether different from that of the bacterium when grown with serum.Their cytotoxicity was easily measured by cell viability assay. The probable cytotoxic factor partially purified and detected by chromatography had characteristics difference from that of vacuolating toxin and a broad cytotoxicity toward various cell lines.CONCLUSION: Serum-free long culture method of H pylori makes its supematant and its extract cytotoxic enough to be easily measured by cell viability assay. The probable cytotoxic factor has a unique characteristic and might be a new cytotoxin.

  16. Regulation of Interleukin—6 Production by Cultured Human Thyrocytes

    Institute of Scientific and Technical Information of China (English)

    段宇; 刘超; 等

    2002-01-01

    Objective To study the modulation of interleukin-6(IL-6) generation from human thyroid cells.Methods Thryoid tissues were obtained at surgery from patients with Graves disease.IL-6 in the supernatant of cultured thyrocytes was detected with ultra-sensitive ELISA.mRNA was extracted respectively from primany human cultured thyroidal cells under stimulated and unstimulated conditions,and semiquantitative reverse transcriptase-polymerase chain reaction(RTPCR) technique was used for IL-6 mRNA detection.Results:(1) In the basic conditions,thyroid cells can produce high concentration IL-6.Thyrotropin(TSH,103mU/L),tumor necrosis factorα(TNF-α,10-1U/ml),interleukin-1(IL-1,10 U/ml) and adrenaline(10-5 mol/L) significantly increased Il-6 levels in the supernatant of cultured thyroid cells,whereas dexamethasone(10 mmol/L) markedly suppressed IL-6 release from thryocytes.Nal of 10-4 mol/L exerted no effect on the production of Il-6.(2)Compared with the basal results,at a concentration of 10-103 mmol/L,dexamethasone could gradually suppress IL-6 gene expression on human thryocytes,while TNF-α could obviously stimulate the production of IL-6 mRNA in the range of 10-1-102 U/ml.On the level of 102 U/ml,IL-1 increased the expression of IL-6,but the same effect could not be shown when IL-1 was 10-1 U/ml.Sodium iodine(NaI) could not affect Il-6 gene from the level of 10-8 to 10-4 mol/L.Conclusion IL-6 produced by thyrocytes might be regulated by many factors that modulate thyroid functions in vitro as well as in vivo.

  17. Talking Culture

    DEFF Research Database (Denmark)

    Holmgreen, Lise-Lotte

    When Danish businesses move production abroad, ‘culture’ is often seen as a huge challenge to the successful outcome of cross-border collaboration. Therefore, business leaders often seek information and guidelines of how to cope in the vast amount of literature on culture and intercultural...... communication. Much of this literature is based on functionalist approaches providing the dos and don’ts of intercultural encounters. This involves inter alia conceptualising ‘culture’ as a relatively fixed, homogeneous entity of values, attitudes and norms shared by members of a group, often leading readers...... to adopt dichotomised understandings and discourses about other cultures (see e.g. Hofstede 2001; Jandt 1998; Trompenaars & Hampden-Turner 1997). However, experience shows that the world in which intercultural encounters take place is not as simple and easy to categorise as these approaches may suggest...

  18. MARKETING CULTURAL

    Directory of Open Access Journals (Sweden)

    Claudia Gómez Ramírez

    2013-07-01

    Full Text Available Este artículo analiza la definición de "Marketing" Cultural y la adaptación y beneficios del "marketing" tradicional respecto al conjunto de manifestaciones artísticas de las diversas industrias involucradas en el sector cultural o artístico; asimismo, se desagregan los conceptos básicos que lo componen como factor de éxito en dichas empresas. Se hace uso de la exposición de casos específicos para ilustrar la articulación de estos dos conceptos aparentemente contrapuestos, cultura y "marketing", y registra algunas reflexiones para que el lector se involucre en la construcción del concepto aquí presentado.

  19. Studies of protein oxidation as a product quality attribute on a scale-down model for cell culture process development.

    Science.gov (United States)

    Lee, Nacole D; Kondragunta, Bhargavi; Uplekar, Shaunak; Vallejos, Jose; Moreira, Antonio; Rao, Govind

    2015-01-01

    Of importance to the biological properties of proteins produced in cell culture systems are the complex post-translational modifications that are affected by variations in process conditions. Protein oxidation, oxidative modification to intracellular proteins that involves cleavage of the polypeptide chain, and modifications of the amino acid side chains can be affected by such process variations. Dissolved oxygen is a parameter of increasing interest since studies have shown that despite the necessity of oxygen for respiration, there may also be some detrimental effects of oxygen to the cell. Production and accumulation of reactive oxygen species can cause damage to proteins as a result of oxidation of the cell and cellular components. Variation, or changes to cell culture products, can affect function, clearance rate, immunogenicity, and specific activity, which translates into clinical implications. The effect of increasing dissolved oxygen on protein oxidation in immunoglobulin G3-producing mouse hybridoma cells was studied using 50 mL high-throughput mini-bioreactors that employ non-invasive optical sensor technology for monitoring and closed feedback control of pH and dissolved oxygen. Relative protein carbonyl concentration of proteins produced under varying levels of dissolved oxygen was measured by enzyme-linked immunosorbent assay and used as an indicator of oxidative damage. A trend of increasing protein carbonyl content in response to increasing dissolved oxygen levels under controlled conditions was observed. Protein oxidation, oxidative modification to intracellular proteins that involves cleavage of the polypeptide chain, and modifications of the amino acid side chains can be affected by variations in dissolved oxygen levels in cell culture systems. Studies have shown that despite the necessity of oxygen for respiration, there may be detrimental effects of oxygen to the cell. Production and accumulation of reactive oxygen species can cause damage to

  20. Cultural neurolinguistics.

    Science.gov (United States)

    Chen, Chuansheng; Xue, Gui; Mei, Leilei; Chen, Chunhui; Dong, Qi

    2009-01-01

    As the only species that evolved to possess a language faculty, humans have been surprisingly generative in creating a diverse array of language systems. These systems vary in phonology, morphology, syntax, and written forms. Before the advent of modern brain-imaging techniques, little was known about how differences across languages are reflected in the brain. This chapter aims to provide an overview of an emerging area of research - cultural neurolinguistics - that examines systematic cross-cultural/crosslinguistic variations in the neural networks of languages. We first briefly describe general brain networks for written and spoken languages. We then discuss language-specific brain regions by highlighting differences in neural bases of different scripts (logographic vs. alphabetic scripts), orthographies (transparent vs. nontransparent orthographies), and tonality (tonal vs. atonal languages). We also discuss neural basis of second language and the role of native language experience in second-language acquisition. In the last section, we outline a general model that integrates culture and neural bases of language and discuss future directions of research in this area.

  1. Dialysis cultures.

    Science.gov (United States)

    Pörtner, R; Märkl, H

    1998-10-01

    Dialysis techniques are discussed as a means for effective removal of low-molecular-mass components from fermentation broth to reach high cell density. Reactor systems and process strategies, the relevant properties of membranes and examples for high-density fermentation with dialysis, and problems related to scale-up are addressed. The dialysis technique has turned out to be very efficient and reliable for obtaining high cell densities. As in dialysis processes the membranes are not perfused, membrane clogging is not a problem as it is for micro- and ultrafiltration. By applying a "nutrient-split" feeding strategy, the loss of nutrients can be avoided and the medium is used very efficiently. The potential of dialysis cultures is demonstrated on the laboratory scale in a membrane dialysis reactor with an integrated membrane and in reactor systems with an external dialysis loop. In dialysis cultures with different microorganisms (Staphylococci, Escherichia coli, extremophilic microorganisms, Lactobacilli) the cell densities achieved were up to 30 times higher than those of other fermentation methods. The technique enables high cell densities to be attained without time-consuming medium optimization. For animal cell cultures the concept of a fixed bed coupled with dialysis proved to be very effective.

  2. Cultural Robotics: The Culture of Robotics and Robotics in Culture

    Directory of Open Access Journals (Sweden)

    Hooman Samani

    2013-12-01

    Full Text Available In this paper, we have investigated the concept of "Cultural Robotics" with regard to the evolution of social into cultural robots in the 21st Century. By defining the concept of culture, the potential development of a culture between humans and robots is explored. Based on the cultural values of the robotics developers, and the learning ability of current robots, cultural attributes in this regard are in the process of being formed, which would define the new concept of cultural robotics. According to the importance of the embodiment of robots in the sense of presence, the influence of robots in communication culture is anticipated. The sustainability of robotics culture based on diversity for cultural communities for various acceptance modalities is explored in order to anticipate the creation of different attributes of culture between robots and humans in the future.

  3. Increased production of recombinant prourokinase with porous microcarrier cell culture by periodic pressure oscillation in a stirred tank reactor

    Institute of Scientific and Technical Information of China (English)

    Hu Xianwen; Gao Lihua; Li Zuohu; Xiao Chengzu; Xu Zhaoping

    2006-01-01

    An rCHO cell line expressing recombinant human prourokinase (pro-UK at the level of 5μg/106cells/d was cultivated on Cytopore cellulose porous microcarriers in a 7.5L Biostat CT stirred tank reactor. A periodic pressure oscillation of 0.04 MPa and 0.04 Hz was adopted to introduce a physical stimulus on the rCHO cells and to improve mass transfer characteristic between cells and medium in the process of porous microcarrier CHO cell culture. Compared to constant pressure culture, the oscillation culture didn't influence specific cell growth rate significantly, but could enhance the pro-UK specific production by 10%~40%, and reduce production of lactate by 10%~30%. In the perfusion culture of recombinant CHO cell with serum-free medium for 67 days, cell density could reach 2.64×107/ml, the maximal prourokinase concentration in harvested supernatant was about 118mg/L, a total of 21.1 grams of prourokinase was produced in 313 liters of supernatant. In conclusion, the perfusion cell culture with periodic pressure oscillation can enhance the production of recombinant protein and increase the reactor specific productivity.

  4. Responsive hydrogels produced via organic sol-gel chemistry for cell culture applications.

    Science.gov (United States)

    Patil, Smruti; Chaudhury, Pulkit; Clarizia, Lisa; McDonald, Melisenda; Reynaud, Emmanuelle; Gaines, Peter; Schmidt, Daniel F

    2012-08-01

    In this study, we report the synthesis of novel environmentally responsive polyurea hydrogel networks prepared via organic sol-gel chemistry and demonstrate that the networks can stabilize pH while releasing glucose both in simple aqueous media and in mammalian cell culture settings. Hydrogel formulations have been developed based on the combination of an aliphatic triisocyanate with pH-insensitive amine functional polyether and pH-sensitive poly(ethyleneimine) segments in a minimally toxic solvent suitable for the sol-gel reaction. The polyether component of the polyurea network is sufficiently hydrophilic to give rise to some level of swelling independent of environmental pH, while the poly(ethyleneimine) component contains tertiary amine groups providing pH sensitivity to the network in the form of enhanced swelling and release under acidic conditions. The reaction of these materials to form a network is rapid and requires no catalyst. The resultant material exhibits the desired pH-responsive swelling behavior and demonstrates its ability to simultaneously neutralize lactic acid and release glucose in both cell-free culture media and mammalian cell culture, with no detectable evidence of cytotoxicity or changes in cell behavior, in the case of either SA-13 human hybridomas or mouse embryonic stem cells. Furthermore, pH is observed to have a clear effect on the rate at which glucose is released from the hydrogel network. Such characteristics promise to maintain a favorable cell culture environment in the absence of human intervention. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  5. Hepatitis E Virus Produced from Cell Culture Has a Lipid Envelope.

    Directory of Open Access Journals (Sweden)

    Ying Qi

    Full Text Available The absence of a productive cell culture system hampered detailed analysis of the structure and protein composition of the hepatitis E virion. In this study, hepatitis E virus from a robust HEV cell culture system and from the feces of infected monkeys at the peak of virus excretion was purified by ultra-centrifugation. The common feature of the two samples after ultracentrifugation was that the ORF2 protein mainly remained in the top fractions. The ORF2 protein from cell culture system was glycosylated, with an apparent molecular weight of 88 kDa, and was not infectious in PLC/PRF/5 cells. The ORF2 protein in this fraction can bind to and protect HEV RNA from digestion by RNase A. The RNA-ORF2 product has a similar sedimentation coefficient to the virus from feces. The viral RNA in the cell culture supernatant was mainly in the fraction of 1.15 g/cm3 but that from the feces was mainly in the fraction of 1.21 g/cm3. Both were infectious in PLC/PRF/5 cells. And the fraction in the middle of the gradient (1.06 g/cm3 from the cell culture supernatant,but not that from the feces, also has ORF2 protein and HEV RNA but was not infectious in PLC/PRF/5.The infectious RNA-rich fraction from the cell culture contained ORF3 protein and lipid but the corresponding fraction from feces had no lipid and little ORF3 protein. The lipid on the surface of the virus has no effect on its binding to cells but the ORF3 protein interferes with binding. The result suggests that most of the secreted ORF2 protein is not associated with HEV RNA and that hepatitis E virus produced in cell culture differs in structure from the virus found in feces in that it has a lipid envelope.

  6. Challenges of culturing human norovirus in three-dimensional organoid intestinal cell culture models.

    Directory of Open Access Journals (Sweden)

    Efstathia Papafragkou

    Full Text Available Human noroviruses are the most common cause of acute gastroenteritis worldwide. Recently, cell culture systems have been described using either human embryonic intestinal epithelial cells (Int-407 or human epithelial colorectal adenocarcinoma cells (Caco-2 growing on collagen-I porous micro carrier beads in a rotating bioreactor under conditions of physiological fluid shear. Here, we describe the efforts from two independent laboratories to implement this three dimensional (3D cell culture system for the replication of norovirus. Int-407 and Caco-2 were grown in a rotating bioreactor for up to 28 days. Prior to infection, cells were screened for the presence of microvilli by electron microscopy and stained for junction proteins (zonula occludens-1, claudin-1, and β-catenin. Differentiated 3D cells were transferred to 24-well plates and infected with bacteria-free filtrates of various norovirus genotypes (GI.1, GI.3, GI.8, GII.2, GII.4, GII.7, and GII.8. At 12 h, 24 h, and 48 h post inoculation, viral RNA from both cells and supernatants were collected and analyzed for norovirus RNA by real-time reverse transcription PCR. Despite observations of high expression of junction proteins and microvilli development in stained thin sections, our data suggest no significant increase in viral titer based on norovirus RNA copy number during the first 48 h after inoculation for the different samples and virus culture conditions tested. Our combined efforts demonstrate that 3D cell culture models using Int-407 or Caco-2 cells do not support norovirus replication and highlight the complexity and difficulty of developing a reproducible in vitro cell culture system for human norovirus.

  7. In vitro culture and characterization of alveolar bone osteoblasts isolated from type 2 diabetics

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Dao-Cai [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China); Department of Stomatology, The 291st Hospital of P.L.A, Baotou (China); Li, De-Hua [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China); Ji, Hui-Cang [Military Sanatorium of Retired Cadres, Baotou (China); Rao, Guo-Zhou [Center of Laboratory, School of Stomatology, Xi' an Jiaotong University, Xi' an (China); Liang, Li-Hua [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China); Ma, Ai-Jie [Xi' an Technology University, Xi' an (China); Xie, Chao; Zou, Gui-Ke; Song, Ying-Liang [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China)

    2012-04-05

    In order to understand the mechanisms of poor osseointegration following dental implants in type 2 diabetics, it is important to study the biological properties of alveolar bone osteoblasts isolated from these patients. We collected alveolar bone chips under aseptic conditions and cultured them in vitro using the tissue explants adherent method. The biological properties of these cells were characterized using the following methods: alkaline phosphatase (ALP) chemical staining for cell viability, Alizarin red staining for osteogenic characteristics, MTT test for cell proliferation, enzyme dynamics for ALP contents, radio-immunoassay for bone gla protein (BGP) concentration, and ELISA for the concentration of type I collagen (COL-I) in the supernatant. Furthermore, we detected the adhesion ability of two types of cells from titanium slices using non-specific immunofluorescence staining and cell count. The two cell forms showed no significant difference in morphology under the same culture conditions. However, the alveolar bone osteoblasts received from type 2 diabetic patients had slower growth, lower cell activity and calcium nodule formation than the normal ones. The concentration of ALP, BGP and COL-I was lower in the supernatant of alveolar bone osteoblasts received from type 2 diabetic patients than in that received from normal subjects (P < 0.05). The alveolar bone osteoblasts obtained from type 2 diabetic patients can be successfully cultured in vitro with the same morphology and biological characteristics as those from normal patients, but with slower growth and lower concentration of specific secretion and lower combining ability with titanium than normal ones.

  8. xMAP-based analysis of three most prevalent staphylococcal toxins in Staphylococcus aureus cultures.

    Science.gov (United States)

    Simonova, Maria A; Petrova, Elena E; Dmitrenko, Olga A; Komaleva, Ravilya L; Shoshina, Natalia S; Samokhvalova, Larisa V; Valyakina, Tatiana I; Grishin, Eugene V

    2014-10-01

    Detection of staphylococcal toxins presents a great interest for medical diagnostics. Screening of clinical samples for the presence of several types of staphylococcal toxins using traditional methods-biological tests on animals or cell cultures as well as ELISA-is laborious. Multiplex detection methods would simplify testing. We have designed an xMAP-based assay to detect three staphylococcal toxins-enterotoxins A and B (SEA and SEB) and toxic shock syndrome toxin (TSST)-in cultural supernatants obtained from different strains of Staphylococcus aureus. The limits of detection of SEA, SEB, and TSST multiplex detection in S. aureus growth medium were 10, 1,000, and 5 pg/mL, respectively. Fifty-nine samples of S. aureus cultural supernatants were tested with the xMAP assay. The developed assay has proved highly effective detection of the natural toxins in the samples obtained due to bacterial cells cultivation. In prospect, the developed test system can be used in clinical diagnostics and in monitoring of foodstuffs and environmental objects.

  9. IL-10 release by bovine epithelial cells cultured with Trichomonas vaginalis and Tritrichomonas foetus

    Science.gov (United States)

    Vilela, Ricardo Chaves; Benchimol, Marlene

    2013-01-01

    Trichomonas vaginalis and Tritrichomonas foetus are parasitic protists of the human and bovine urogenital tracts, respectively. Several studies have described the cytotoxic effects of trichomonads on urogenital tract epithelial cells. However, little is known about the host cell response against trichomonads. The aim of this study was to determine whether T. foetus and T. vaginalis stimulated the release of the cytokine interleukin (IL)-10 from cultured bovine epithelial cells. To characterise the inflammatory response induced by these parasites, primary cultures of bovine oviduct epithelial cells were exposed to either T. vaginalis or T. foetus. Within 12 h after parasite challenge, supernatants were collected and cytokine production was analysed. Large amounts of IL-10 were detected in the supernatants of cultures that had been stimulated with T. foetus. Interestingly, T. vaginalis induced only a small increase in the release of IL-10 upon exposure to the same bovine cells. Thus, the inflammatory response of the host cell is species-specific. Only T. foetus and not T. vaginalis induced the release of IL-10 by bovine oviduct epithelial cells. PMID:23440124

  10. IL-10 release by bovine epithelial cells cultured with Trichomonas vaginalis and Tritrichomonas foetus

    Directory of Open Access Journals (Sweden)

    Ricardo Chaves Vilela

    2013-02-01

    Full Text Available Trichomonas vaginalis and Tritrichomonas foetus are parasitic protists of the human and bovine urogenital tracts, respectively. Several studies have described the cytotoxic effects of trichomonads on urogenital tract epithelial cells. However, little is known about the host cell response against trichomonads. The aim of this study was to determine whether T. foetus and T. vaginalis stimulated the release of the cytokine interleukin (IL-10 from cultured bovine epithelial cells. To characterise the inflammatory response induced by these parasites, primary cultures of bovine oviduct epithelial cells were exposed to either T. vaginalis or T. foetus. Within 12 h after parasite challenge, supernatants were collected and cytokine production was analysed. Large amounts of IL-10 were detected in the supernatants of cultures that had been stimulated with T. foetus. Interestingly, T. vaginalis induced only a small increase in the release of IL-10 upon exposure to the same bovine cells. Thus, the inflammatory response of the host cell is species-specific. Only T. foetus and not T. vaginalis induced the release of IL-10 by bovine oviduct epithelial cells.

  11. Selective crystallization of tank supernatant liquid

    Energy Technology Data Exchange (ETDEWEB)

    Herting, D.

    1996-10-01

    The objective of this task is to demonstrate the feasibility of selectively removing sodium nitrate (NaNO{sub 3}) from Hanford Site tank waste by a large-scale fractional crystallization process. Two thirds of all the nuclear waste stored in Hanford`s underground storage tanks is sodium nitrate (mass basis, excluding water). Fractional crystallization can remove essentially nonradioactive NaNO{sub 3} and other sodium salts from the waste, thereby reducing the volume of low-level waste glass by as much as 90%.

  12. La arquitectura cultural. / Cultural architecture.

    Directory of Open Access Journals (Sweden)

    Lobos, Jorge

    2004-12-01

    Full Text Available Este artículo releva la Arquitectura Cultural, que es plural y diversa en términos estéticos y conceptuales. Sugiere la apertura a otras dinámicas de comprensión de lo arquitectónico, a otras lógicas de construcción de las ciudades. Para aclarar el concepto se expone un breve ejemplo práctico./This article points out the "cultural architecture", which is plural and diverse in concept and aesthetic terms. It suggests the opening to other dinamics of comprehention of the architectural issue and the building of cities. The article presents a brief empirical example that clarifies the conceptual approach.

  13. Cultural Robotics: The Culture of Robotics and Robotics in Culture

    OpenAIRE

    Hooman Samani; Elham Saadatian; Natalie Pang; Doros Polydorou; Owen Noel Newton Fernando; Ryohei Nakatsu; Jeffrey Tzu Kwan Valino Koh

    2013-01-01

    In this paper, we have investigated the concept of “Cultural Robotics” with regard to the evolution of social into cultural robots in the 21st Century. By defining the concept of culture, the potential development of a culture between humans and robots is explored. Based on the cultural values of the robotics developers, and the learning ability of current robots, cultural attributes in this regard are in the process of being formed, which would define the new concept of cultural robotics. Ac...

  14. Reinventing Culture

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Singer puts a modern musical face on ancient cultural classics In traditional loose sleeved Chinese costume and a hairstyle straight from ancient China, a slim female singer stepped onto the stage, singing a wistful song called Li. "Li is respect, Li is elegance, Li is purity, Li is tranquility; Li comes from a smile, Li comes from love, Li comes from the heart," she sings, accompanied by a melody of haunting court music. The singer is appearing on CCTV, China’s National TV station, in a music program aired during the 2008 Olympic Games.

  15. Black Culture

    Directory of Open Access Journals (Sweden)

    Angela Khristin Brown

    2013-07-01

    Full Text Available The migration of blacks in North America through slavery became united.  The population of blacks past downs a tradition of artist through art to native born citizens. The art tradition involved telling stories to each generation in black families. The black culture elevated by tradition created hope to determine their personal freedom to escape from poverty of enslavement and to establish a way of life through tradition. A way of personal freedoms was through getting a good education that lead to a better foundation and a better way of life.

  16. Culture Jamming Versus Popular Culture

    Directory of Open Access Journals (Sweden)

    Leonardia Acynthia Putri

    2013-11-01

    Full Text Available Abstract: This literature study researched Adbusters, the anti-commercial organization, and described the organization’s activities and media usage, mainly in the period of 2007-2010, which critized the populer culture. Adbusters is an organization which performs “Culture Jamming”; a rebellious act reacting towards commercialism domination in many aspects including popular culture. Compared to other similar organizations, Adbusters has been executing more various activisms using several media which other organizations do not use. This study used the Adbusters’ official website and blogs as main data sources. The data of Adbusters’ activities and media usage were categorized and analyzed, thus the tendency of its development can be described. This study also analyzed Adbusters’ activity using Media Hegemony Theory and Political Economy Media Theory. The media has been dominated by a certain group that owns politic and economic power, so the information flow has been dominated by them. Media and its contents have been commercialized, thus capitalism and commercialism have been considered as a common system that should run the world. Adbusters has been trying to stop the domination and change the society’s way of thinking into a more critical way of thinking.   Abstrak: Studi literatur ini meneliti tentang Adbusters, sebuah organisasi anti komersial, dengan mendeskripsikan aktivitas serta penggunaan media organisasi tersebut dari tahun 2007-2010 dalam mengkritisi budaya populer. Adbusters adalah organisasi yang melakukan Culture Jamming, aksi perlawanan terhadap dominasi komersialisme di segala aspek termasuk popular culture. Dibandingkan dengan organisasi lain yang serupa, aktivitas Adbusters lebih bervariasi dan menggunakan media-media yang tidak biasa digunakan organisasi lain. Penelitian ini menggunakan situs online resmi Adbusters sebagai sumber data utama. Data mengenai aktivitas dan

  17. Comparison of procedures for the extraction of supernatants and cytotoxicity tests in Vero cells, applied to assess the toxigenic potential of Bacillus spp. and Lactobacillus spp., intended for use as probiotic strains.

    Science.gov (United States)

    Blanch, Anicet R; Méndez, Javier; Castel, Susana; Reina, Manuel

    2014-08-01

    Interest in using Bacillus strains as probiotic components of animal feeds has grown in recent years. However, some of these strains, especially those taxonomically related to the Bacillus cereus group, may have enterotoxigenic activity. Assessment of their toxigenic potential by well-established and robust protocols is required before authorizing their use in animal nutrition. Three methods of extraction and concentration of supernatants of Bacillus and Lactobacillus strains (methanol extraction, ammonium sulphate and ultrafiltration concentration) and three cytotoxic tests in Vero cells (WST-1, LDH and protein synthesis inhibition assays) for the assessment of the cytotoxicity activity of Lactobacillus strains (as probiotic strains in human and animal nutrition) and Bacillus toyonensis BCT-7112(T) (as animal probiotic strain in animal nutrition-Toyocerin®-) were evaluated in this study. Methanol extraction was not useful under any circumstances. The other two concentration methods (ammonium sulphate and ultrafiltration) were feasible, with slightly greater sensitivity achieved by ultrafiltration. The probiotic strain B. toyonensis BCT-7112(T) proved to be a non-cytotoxic strain in all the protocols tested. However, some Lactobacillus strains showed cytotoxicity activity, regardless of the protocols applied. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Culture collections.

    Science.gov (United States)

    Smith, David

    2012-01-01

    Culture collections no matter their size, form, or institutional objectives play a role in underpinning microbiology, supplying the resources for study, innovation, and discovery. Their basic roles include providing a mechanism for ex situ conservation of organisms; they are repositories for strains subject to publication, taking in safe, confidential, and patent deposits from researchers. They supply strains for use; therefore, the microorganisms provided must be authentic and preserved well, and any associated information must be valid and sufficient to facilitate the confirmation of their identity and to facilitate their use. The organisms must be collected in compliance with international conventions, international and national legislation and distributed to users indicating clearly the terms and conditions under which they are received and can be used. Collections are harmonizing approaches and characterizing strains to meet user needs. No one single collection can carry out this task alone, and therefore, it is important that output and strategy are coordinated to ensure culture collections deliver the basic resources and services microbiological innovation requires. This chapter describes the types of collection and how they can implement quality management systems and operate to deliver their basic functions. The links to information sources given not only provide support for the practitioners within collections but also provide guidance to users on accessing the huge resource available and how they can help ensure microbiology has the resources and a solid platform for future development.

  19. Interferon Response in Hepatitis C Virus (HCV) Infection: Lessons from Cell Culture Systems of HCV Infection.

    Science.gov (United States)

    Sung, Pil Soo; Shin, Eui-Cheol; Yoon, Seung Kew

    2015-01-01

    Hepatitis C virus (HCV) is a positive-stranded RNA virus that infects approximately 130-170 million people worldwide. In 2005, the first HCV infection system in cell culture was established using clone JFH-1, which was isolated from a Japanese patient with fulminant HCV infection. JFH-1 replicates efficiently in hepatoma cells and infectious virion particles are released into the culture supernatant. The development of cell culture-derived HCV (HCVcc) systems has allowed us to understand how hosts respond to HCV infection and how HCV evades host responses. Although the mechanisms underlying the different outcomes of HCV infection are not fully understood, innate immune responses seem to have a critical impact on the outcome of HCV infection, as demonstrated by the prognostic value of IFN-λ gene polymorphisms among patients with chronic HCV infection. Herein, we review recent research on interferon response in HCV infection, particularly studies using HCVcc infection systems.

  20. Bacterial Wound Culture

    Science.gov (United States)

    ... and services. Advertising & Sponsorship: Policy | Opportunities Bacterial Wound Culture Share this page: Was this page helpful? Also known as: Aerobic Wound Culture; Anaerobic Wound Culture Formal name: Culture, wound Related ...

  1. Culture Wars in Denmark

    DEFF Research Database (Denmark)

    Ørum, Tania

    2016-01-01

    In the 1960s high and low culture were brought into sharp conflict, as the Ministry of Culture was established to support cultural enlightenment and democratic access to art and culture, while popular demands for more entertainment were raised.......In the 1960s high and low culture were brought into sharp conflict, as the Ministry of Culture was established to support cultural enlightenment and democratic access to art and culture, while popular demands for more entertainment were raised....

  2. Polyamines in relation to growth in carrot cell cultures.

    Science.gov (United States)

    Fallon, K M; Phillips, R

    1988-09-01

    Changes in polyamine metabolism were investigated in relation to growth of cell suspension cultures of carrot (Daucus carota, cv Chantenay). Changes in levels of the major amines putrescine and spermidine throughout the culture period correlated poorly with changes in fresh weight, but a closer correlation with the minor component spermine was observed. The arginine decarboxylase (ADC) inhibitor difluoromethylarginine (DFMA) strongly and specifically inhibited ADC activity in the supernatant, reduced the major amine (putrescine) by 95% and the total amine content by 80%. It had no effect on cell number and stimulated fresh weight by over 25% through increased cell expansion. Spermine content, in contrast, increased with DFMA concentration in parallel with fresh weight increases. Difluoromethylornithine strongly inhibited ornithine decarboxylase activity in the pellet, but had little effect on either polyamine levels or culture growth. It was concluded that little evidence for a correlation between free polyamines and cell number in carrot cultures could be detected, but that a possible correlation between spermine content and cell expansion was observed.

  3. Polyamines in Relation to Growth in Carrot Cell Cultures 1

    Science.gov (United States)

    Fallon, Kevin M.; Phillips, Richard

    1988-01-01

    Changes in polyamine metabolism were investigated in relation to growth of cell suspension cultures of carrot (Daucus carota, cv Chantenay). Changes in levels of the major amines putrescine and spermidine throughout the culture period correlated poorly with changes in fresh weight, but a closer correlation with the minor component spermine was observed. The arginine decarboxylase (ADC) inhibitor difluoromethylarginine (DFMA) strongly and specifically inhibited ADC activity in the supernatant, reduced the major amine (putrescine) by 95% and the total amine content by 80%. It had no effect on cell number and stimulated fresh weight by over 25% through increased cell expansion. Spermine content, in contrast, increased with DFMA concentration in parallel with fresh weight increases. Difluoromethylornithine strongly inhibited ornithine decarboxylase activity in the pellet, but had little effect on either polyamine levels or culture growth. It was concluded that little evidence for a correlation between free polyamines and cell number in carrot cultures could be detected, but that a possible correlation between spermine content and cell expansion was observed. PMID:16666271

  4. Study of HLA-DR synthesis in cultured human keratinocytes.

    Science.gov (United States)

    Wikner, N E; Huff, J C; Norris, D A; Boyce, S T; Cary, M; Kissinger, M; Weston, W L

    1986-11-01

    Within the normal human epidermis only Langerhans and indeterminate cells express HLA-DR. Human keratinocytes (HK), however, may also express HLA-DR in certain disease states characterized by mononuclear cell infiltrates. Previous studies have shown that HK synthesize HLA-DR in response to stimulation by interferon gamma (INF-gamma). The purposes of this study were to define conditions under which cultured HK might express HLA-DR and to compare the HLA-DR synthesis of HK with that of monocytes. HLA-DR expression by HK as determined by indirect immunofluorescence of HK cultures was absent under standard low calcium conditions and remained absent with the addition of calcium, serum, mitogens, and supernatants from Pam-212 cells containing epidermal thymocyte-activating factor. HLA-DR expression in HK was induced by cocultivation with concanavalin A-stimulated peripheral blood mononuclear cells (PBMC), but not unstimulated PBMC. This effect was time-dependent and directly related to the number of PBMC. HLA-DR expression was also induced in a time- and dose-dependent manner by addition of supernatant from stimulated PBMC (SS) or by addition of recombinant INF-gamma but not by addition of interleukin (IL)-1 or IL-2. Induction by either SS or INF-gamma was blocked by an antiserum to INF-gamma. As determined by a semiquantitative immunoprecipitation technique, HLA-DR synthesis by HK was directly related to INF-gamma concentration. The pattern of HLA-DR peptides produced by HK was similar to that of monocytes, but the relative quantity synthesized was far less than that of monocytes.

  5. Determination of amino acids in cell culture and fermentation broth media using anion-exchange chromatography with integrated pulsed amperometric detection.

    Science.gov (United States)

    Hanko, Valoran P; Rohrer, Jeffrey S

    2004-01-01

    Cell culture and fermentation broth media are used in the manufacture of biotherapeutics and many other biological materials. Characterizing the amino acid composition in cell culture and fermentation broth media is important because deficiencies in these nutrients can reduce desired yields or alter final product quality. Anion-exchange (AE) chromatography using sodium hydroxide (NaOH) and sodium acetate gradients, coupled with integrated pulsed amperometric detection (IPAD), determines amino acids without sample derivatization. AE-IPAD also detects carbohydrates, glycols, and sugar alcohols. The presence of these compounds, often at high concentrations in cell culture and fermentation broth media, can complicate amino acid determinations. To determine whether these samples can be analyzed without sample preparation, we studied the effects of altering and extending the initial NaOH eluent concentration on the retention of 42 different carbohydrates and related compounds, 30 amino acids and related compounds, and 3 additional compounds. We found that carbohydrate retention is impacted in a manner different from that of amino acid retention by a change in [NaOH]. We used this selectivity difference to design amino acid determinations of diluted cell culture and fermentation broth media, including Bacto yeast extract-peptone-dextrose (yeast culture medium) broth, Luria-Bertani (bacterial culture medium) broth, and minimal essential medium and serum-free protein-free hybridoma medium (mammalian cell culture media). These media were selected as representatives for both prokaryotic and eukaryotic culture systems capable of challenging the analytical technique presented in this paper. Glucose up to 10mM (0.2%, w/w) did not interfere with the chromatography, or decrease recovery greater than 20%, for the common amino acids arginine, lysine, alanine, threonine, glycine, valine, serine, proline, isoleucine, leucine, methionine, histidine, phenylalanine, glutamate, aspartate

  6. Cultural Misreading in ELT

    Institute of Scientific and Technical Information of China (English)

    SunCuilan; RenHuaiping; MaDaoshan

    2004-01-01

    In inter-linguistic-cultural communication, cultural misreading is unavoidable. The same is true in foreign language teaching and learning owning to the cultural dissimilarities, for the influence exerted by cultural components upon languages constitutes the major barriers. Language can not exist without culture as its component. Culture consists of all the shared

  7. Advertising cultures

    DEFF Research Database (Denmark)

    Malefyt, Timothy de Waal; Moeran, Brian

    The growth, success and secrets of advertizing are legendary. Advertizing agencies ceaselessly churn out evermore sophisticated campaigns that, when successful, manage to capture the every essence of consumer desire. The secrets of advertizing are perhaps best understood by turning......, but exposes, through in-depth accounts based on personal experience, the inner workings of the advertizing industry. How do adverts manage to capture "real" life? What issues do agencies have to consider when using an advert in a range of different countries? What specific methods are used to persuade us...... to the relationship between advertizing and anthropology. The link between them may come as a surprise to those who consider advertizing to be firmly rooted in commerce and anthropology in culture. Through the lens of anthropologists, this book not only shows how anthropology and advertizing are connected...

  8. patrimonio cultural

    Directory of Open Access Journals (Sweden)

    Esther Fernández de Paz

    2006-01-01

    Full Text Available Desde el momento en que Europa sacralizó un determinado conjunto de objetos y los convirtió en referentes patrimoniales activados y protegidos por los representantes de la cultura oficial, hasta el presente, mucho se han ensanchado los estrechos límites patrimoniales; se ha superado la concepción objetual, historicista y esteticista para abarcar todo el conjunto de bienes de valor cultural. El patrimonio deja así de ser contemplado exclusivamente como un tesoro histórico-artístico para pasar a convertirse en algo mucho más valioso: en elementos -materiales e inmateriales- fundamentales para comprender nuestra identidad. No obstante, la creciente demanda turística de supuestas autenticidades está hoy provocando que este patrimonio se oferte, en no pocas ocasiones, como la expresión de un pasado idealizado.

  9. Academic Culture and Campus Culture of Universities

    Science.gov (United States)

    Shen, Xi; Tian, Xianghong

    2012-01-01

    Academic culture of universities mainly consists of academic outlooks, academic spirits, academic ethics and academic environments. Campus culture in a university is characterized by individuality, academic feature, opening, leading, variety and creativity. The academic culture enhances the construction of campus culture. The campus culture…

  10. Culture-lovers and Culture-leavers

    NARCIS (Netherlands)

    Frank Huysmans; Andries van den Broek; Jos de Haan

    2005-01-01

    Who are the people in the Netherlands with an active interest in cultural heritage and the performing arts, and who prefer to leave these forms of culture alone? Have the size and composition of the groups of 'culture-lovers' and 'culture-leavers' changed since the end of the 1970s? These are the ce

  11. Isolation and characterization of in vitro culture of hair follicle cells differentiated from umbilical cord blood mesenchymal stem cells.

    Science.gov (United States)

    Bu, Zhang-Yu; Wu, Li-Min; Yu, Xiao-Hong; Zhong, Jian-Bo; Yang, Ping; Chen, Jian

    2017-07-01

    The present investigation explored the in vitro culture, isolation and characterization of hair follicle cell differentiation from umbilical cord blood mesenchymal stem cells (MSCs). Flow cytometry was used to obtain MSCs from the isolation and purification of human umbilical cord blood MSCs. Culture suspension of hair follicle organ was centrifuged and the supernatant used in the culture medium of MSCs, and the entire process of induced differentiation was recorded by photomicroscopy. The expression level of surface marker CK15 of hair follicle cells obtained from induced differentiation was detected with immunofluorescence. RT-PCR method was used to further detect the difference in expression of CK15 between hair follicle cells and umbilical cord blood MSCs, and statistical analysis was carried out. CD44(+)CD29(+) double-labeled cells accounted for 50.8% of all the samples of umbilical cord blood MSCs in this study. The diameter of hair follicle cells differentiated from umbilical cord blood stem cells reached 800×10(-3) mm after 3 weeks of cell culture. Based on the detection and colocalization of CK15 expression in induced hair follicle cells, the overlap ratio between CK15 and nuclei reached 83% in hair follicle cells, which was obviously higher than that in umbilical cord blood stem cells. The difference had statistical significance (Pumbilical cord blood stem cells by using the supernatant from hair follicle cells. This method can be used for high-speed induced differentiation with high purity, which is promising for clinical application.

  12. Expression of Cyt C and Caspase-3 of bone marrow supernatant in patients with chronic mountain sickness%慢性高原病患者骨髓CytC和Caspase-3表达及意义研究

    Institute of Scientific and Technical Information of China (English)

    戴昕; 李建平; 李文倩; 冯建明

    2014-01-01

    目的:探讨细胞凋亡在慢性高原病(CMS)发生发展过程中的作用,从而寻求新的诊断和治疗途径。方法选取 CMS 患者20例为研究对象,健康人群14例为对照组,采用固相双抗体夹心 ELISA 方法测定 CMS 组及对照组骨髓上清液 Cyt C 和 Caspase-3水平。结果 CMS 患者骨髓上清液中 Cyt C 水平[(21.43±8.92)ng/ml]水平与对照组[(10.36±3.55)ng/ml]比较差异有统计学意义(P<0.05);CMS 患者骨髓上清液中 Caspase-3水平[(15.45±4.73)ng/ml]水平与对照组[(7.14±1.93)ng/ml]比较差异有统计学意义(P<0.05);CMS 患者骨髓上清液中 Cyt C 与 Caspase-3水平呈正相关(r=0.706,P<0.01);CMS 患者骨髓上清液中 Cyt C 和 Caspase-3表达水平均与血红蛋白(Hb)浓度呈正相关(r=0.524,P<0.01;r=0.577,P<0.01),与血氧饱和度(SaO2)呈负相关(r=-0.505,P<0.05;r=-0.554,P<0.01)。结论推测细胞凋亡可能在 CMS 病理生理过程中发挥着重要作用。%Objective To investigate the apoptosis occurred during the development of function in CMS, so as to seek a new means of diagnosis and treatment. Methods The bone marrow supernatant levels of Cyt C and Caspase-3 in 20 CMS pa-tients and 14 healthy people were determined by quantitative sandwich enzyme immunoassay. Results The bone marrow su-pernatant levels of Cyt C[(21.43±8.92)ng/ml] and Caspase-3[(10.36±3.55)ng/ml] in CMS were significantly higher than those in control group [(15.45±4.73)ng/ml] and [(7.14±1.93)ng/ml] respectively. And there were positive relationship between the levels of Cyt C and Caspase-3 (r=0.706,P<0.01),and between the levels of Cyt C and Hb (r=0.524,P<0.01),and also between the levels of Caspase-3 and Hb (r=0.577,P<0.01). There were negative relationship between the levels of Cyt C and SaO2(r=-0.554, P<0.05),and between the levels of Caspase-3 and SaO2 (r=-0.505,P<0.05). Conclusion Speculate that apoptosis may play an

  13. Oral Platelet Gel Supernatant Plus Supportive Medical Treatment Versus Supportive Medical Treatment in the Management of Radiation-induced Oral Mucositis: A Matched Explorative Active Control Trial by Propensity Analysis.

    Science.gov (United States)

    Bonfili, Pierluigi; Gravina, Giovanni L; Marampon, Francesco; Rughetti, Anna; Di Staso, Mario; Dell'Orso, Luigi; Vittorini, Francesca; Moro, Roberto; La Verghetta, Maria E; Parente, Silvia; Reale, Marilisa; Ruggieri, Valeria; Franzese, Pietro; Tombolini, Vincenzo; Masciocchi, Carlo; Di Cesare, Ernesto

    2017-08-01

    In this active control trial, the rate of radio-induced WHO grade 3/4 oral mucositis and the change in quality of life, assessed by OMWQ-HN, were measured in subjects with head and neck cancer treated by platelet gel supernatant (PGS) and supportive medical treatment versus subjects treated by supportive medical treatment alone. Eighty patients with nonmetastatic head and neck cancer underwent curative or adjuvant radiotherapy. All patients underwent supportive medical treatment and/or PGS at the beginning and during radiotherapy. Sixteen patients received PGS in association with supportive medical treatment. To obtain 2 groups virtually randomized for important clinical characteristics subjects were matched, by propensity analysis, with a group of subjects (64 patients) treated with supportive medical treatment alone. Subjects treated with standard supportive treatment experienced significant higher WHO grade 3/4 toxicity (55%; 35/64) than subjects treated by PGS (13%; 3/16). The reduced toxicity found in PGS group paralleled with the evidence that they developed later symptoms with respect to controls. The Cox proportional hazard model indicated that patients treated with standard supportive medical treatment experienced 2.7-fold increase (hazard ratio=2.7; 95% confidence interval, 1.3-5.7) in the occurrence of WHO grade 3/4 toxicity. PGS group significantly experienced higher quality of life than control groups as measured by OMWQ-HN. A significant decrease in the opioid analgesics usage was found in the PGS group. These preliminary data should be interpreted with caution and could serve as a framework around which to design future trials.

  14. Analysis of Tank 13H (HTF-13-14-156, 157) Surface and Subsurface Supernatant Samples in Support of Enrichment Control, Corrosion Control and Sodium Aluminosilicate Formation Potential Programs

    Energy Technology Data Exchange (ETDEWEB)

    Oji, L. N. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL). Advanced Characterization and Processing

    2015-02-18

    The 2H Evaporator system includes mainly Tank 43H (feed tank) and Tank 38H (drop tank) with Tank 22H acting as the DWPF recycle receipt tank. The Tank 13H is being characterized to ensure that it can be transferred to the 2H evaporator. This report provides the results of analyses on Tanks 13H surface and subsurface supernatant liquid samples to ensure compliance with the Enrichment Control Program (ECP), the Corrosion Control Program and Sodium Aluminosilicate Formation Potential in the Evaporator. The U-235 mass divided by the total uranium averaged 0.00799 (0.799 % uranium enrichment) for both the surface and subsurface Tank 13H samples. This enrichment is slightly above the enrichment for Tanks 38H and 43H, where the enrichment normally ranges from 0.59 to 0.7 wt%. The U-235 concentration in Tank 13H samples ranged from 2.01E-02 to 2.63E-02 mg/L, while the U-238 concentration in Tank 13H ranged from 2.47E+00 to 3.21E+00 mg/L. Thus, the U-235/total uranium ratio is in line with the prior 2H-evaporator ECP samples. Measured sodium and silicon concentrations averaged, respectively, 2.46 M and 1.42E-04 M (3.98 mg/L) in the Tank 13H subsurface sample. The measured aluminum concentration in Tanks 13H subsurface samples averaged 2.01E-01 M.

  15. Fermentation culture of Cryptococcus neoformans benefiting for purification of mannoprotein%利于甘露糖蛋白纯化的新生隐球菌培养方法研究

    Institute of Scientific and Technical Information of China (English)

    李平; 谭宏月; 钟彬; 陈丽华; 胡婵; 朱红梅; 温海

    2012-01-01

    Objective To study the possibility of fermentation culture of Cryptococcus neoformans and the advantages of fermentation culture supernatant used for purificating mannoprotein. Methods ConA Sepharose 4B affinity column was used to purificate mannoprotein from fermentation culture supernatant of CAP67. Results Up to milligram of mannoprotein was obtained from fermentation supernatants of Cryptococcus neoformans acapsular strain CAP67. Conclusion Fermentation method can be used to culture Cryptococcus neoformans , and the fermentation supernatants can be used for the purification of mannoprotein, which possessed the superiority of shorter time-consuming and higher yields compared to the traditional method.%目的 研究发酵法培养新生隐球菌的可能性,探索发酵法较传统方法培养上清用于纯化甘露糖蛋白的优势.方法 采用发酵法培养新生隐球菌CAP67,收集上清使用ConA琼脂糖凝胶4B亲和柱亲和纯化隐球菌甘露糖蛋白.结果 从新生隐球菌无荚膜株CAP67的发酵培养上清中获得了毫克级的甘露糖蛋白.结论 发酵法可以成功培养新生隐球菌,上清可用于纯化隐球菌甘露糖蛋白,与传统方法相比具有耗时少,产量高等优点.

  16. CONNECTION BETWEEN ECONOMICS, CULTURE AND CULTURAL DIPLOMACY

    Directory of Open Access Journals (Sweden)

    Agil Valiyev

    2017-09-01

    Full Text Available Today, culture is one of the main feeble factors of economic development.  The leading role of culture in economic development should be argued as multiplied: so, on firstly, as domestic value, on secondly, as a main factor of regional economic development advanced to raised gravity of different regions for residents, tourists and investors, on thirdly, as major parameters of social development based on tolerance, creativity and knowledge. To the different international experiences, culture is main part of economic development in our life. Cultural diversities are combined into a main reason economic development model. The article consist of explainations about the understanding of culture, cultural diplomacy and economics, approach on conflicts between culture and economics, to find how affecting of culture to economic development, the role of culture in economic development of Azerbaijan. The article can be considered as a useful resource  for experts and researchers conducting research in this field.

  17. Porphyromonas gingivalis displays a competitive advantage over Aggregatibacter actinomycetemcomitans in co-cultured biofilm.

    Science.gov (United States)

    Takasaki, K; Fujise, O; Miura, M; Hamachi, T; Maeda, K

    2013-06-01

    Biofilm formation occurs through the events of cooperative growth and competitive survival among multiple species. Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans are important periodontal pathogens. The aim of this study was to demonstrate competitive or cooperative interactions between these two species in co-cultured biofilm. P. gingivalis strains and gingipain mutants were cultured with or without A. actinomycetemcomitans. Biofilms formed on glass surfaces were analyzed by crystal violet staining and colony counting. Preformed A. actinomycetemcomitans biofilms were treated with P. gingivalis culture supernatants. Growth and proteolytic activities of gingipains were also determined. Monocultured P. gingivalis strains exhibited a range of biofilm-formation abilities and proteolytic activities. The ATCC33277 strain, noted for its high biofilm-formation ability and proteolytic activity, was found to be dominant in biofilm co-cultured with A. actinomycetemcomitans. In a time-resolved assay, A. actinomycetemcomitans was primarily the dominant colonizer on a glass surface and subsequently detached in the presence of increasing numbers of ATCC33277. Detachment of preformed A. actinomycetemcomitans biofilm was observed by incubation with culture supernatants from highly proteolytic strains. These results suggest that P. gingivalis possesses a competitive advantage over A. actinomycetemcomitans. As the required biofilm-formation abilities and proteolytic activities vary among P. gingivalis strains, the diversity of the competitive advantage is likely to affect disease recurrence during periodontal maintenance. © 2012 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Teaching Culture Through Films

    Institute of Scientific and Technical Information of China (English)

    徐婷

    2016-01-01

    Cultural teaching is an issue which is associated with complexity and paradox and also it is a big challenge for faculty. Teaching culture through films has become an important way of cross-cultural teaching This paper focuses on the reasons for teaching culture through films, the value and how it works. And finally it leads out the prospects of cultural teaching through films.

  19. Culture and Translation

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-hui; REN Pei-hong

    2015-01-01

    Language, as a kind of symbol system of social culture, is strongly connected with culture. Language is a carrier of cul⁃ture and a form of culture. Just because the strong connection between culture and language, so we can believe that language is a mirror of culture.

  20. Stable lentiviral transformation of CHO cells for the expression of the hemagglutinin H5 of avian influenza virus in suspension culture

    Directory of Open Access Journals (Sweden)

    Alaín González Pose

    2014-09-01

    Full Text Available Avian influenza virus H5N1 has caused extensive damage worldwide among poultry and humans. Effective expression systems are needed for the production of viral proteins required for monitoring this devastating disease. The present study deals with the establishment of a stable expression system for the hemagglutinin H5 (HAH5 of avian influenza virus using CHO cells in suspension culture transduced with a recombinant lentiviral vector. The synthetic gene coding the HAH5 protein was inserted in a lentiviral vector with the aim of performing a stable transduction of CHO cells. After the selection of recombinant clones, the one with the highest expression level was adapted to suspension culture and the HAH5 protein was purified by immunoaffinity chromatography from the culture supernatant. There were no significant differences when this protein, purified or direct from the culture supernatant of CHO or SiHa cells, was utilized in an immunologic assay using positive and negative sera as reference. It was also demonstrated that the HAH5 protein in its purified form is able to bind anti-HAH5 antibodies generated with proper and non-proper folded proteins. The results demonstrate that the CHO cell line stably transduced with a lentiviral vector coding the sequence of the HAH5 protein and cultured in suspension can be a suitable expression system to obtain this protein for diagnostic purpose in a consistent and reliable manner.

  1. Dynamics of the Content of Lactobacilli, Microbial Metabolites and Antimicrobial Activity of Growing Culture of Lactobacillus Plantarum 8P-A3

    Directory of Open Access Journals (Sweden)

    I. Yu. Chicherin

    2013-01-01

    Full Text Available The dynamics of the content of lactobacilli, microbial metabolites and antimicrobial activity of growing cultures of Lactobacillus plantarum 8Р-А3 was studied. Lactobacilli L. plantarum 8Р-А3 and test microorganisms isolated from the intestinal contents of patients with dysbacteriosis were used in experiments. Study of the component composition of growing culture supernatant of lactobacilli was carried out by gas liquid chromatography with mass selective detection. By 54 h of cultivation the content of viable microbial cells in the native culture of Lactobacillus achieves 3,0·109 in 1 mL without further increase during the cultivation. The principal component of lactobacilli culture medium possessing antibacterial activity is lactic acid. In addition to lactic acid, which accounts for 70% of the total metabolites, the culture medium and the supernatant contain salts of phosphoric acid (14% as well as amino acids, carboxylic acids, fatty acids, sugars and polyhydric alcohol constituting of up to 16% of the total metabolites. It is found that during the cultivation in liquid medium lactobacilli produce metabolites which possess antibacterial activity against pathogenic bacteria that cause intestinal infections.

  2. Exploring Culture : Exercises, Stories and Synthetic Cultures

    NARCIS (Netherlands)

    Hofstede, G.J.

    2002-01-01

    A unique training book containing over 100 culture awareness exercises, dialogues, stories incidents and simulations that bring to life Geert Hofstede's five dimensions of culture. These dimensions are: power distance, collectivism versus individualism, femininity versus masculinity, uncertainly avo

  3. Culture ou Intercultures (Culture or Intercultural).

    Science.gov (United States)

    Steele, Ross

    1996-01-01

    While planet Earth endeavors to transmit information instantaneously, cultural misunderstanding interferes with communication more than any language barrier. The article urges teachers of French to be cognizant of their role as cultural mediators. (Author/CK)

  4. Exploring Culture : Exercises, Stories and Synthetic Cultures

    NARCIS (Netherlands)

    Hofstede, G.J.

    2002-01-01

    A unique training book containing over 100 culture awareness exercises, dialogues, stories incidents and simulations that bring to life Geert Hofstede's five dimensions of culture. These dimensions are: power distance, collectivism versus individualism, femininity versus masculinity, uncertainly avo

  5. Exploring Culture : Exercises, Stories and Synthetic Cultures

    NARCIS (Netherlands)

    Hofstede, G.J.

    2002-01-01

    A unique training book containing over 100 culture awareness exercises, dialogues, stories incidents and simulations that bring to life Geert Hofstede's five dimensions of culture. These dimensions are: power distance, collectivism versus individualism, femininity versus masculinity, uncertainly

  6. Complejidad Cultural

    Directory of Open Access Journals (Sweden)

    Juan Soto Ramírez

    2010-07-01

    Full Text Available A inicios del 2007, un llamativo suceso se convirtió en el ‘foco de atención' de la opinión pública: 600 internas de La Villa de las niñas de Chalco, presentaron síntomas como: mareo, náuseas, vómitos y problemas musculares. Una vez descartados los factores orgánicos y con el aval de la ‘ciencia médica' se procedió a construir una versión oficial respaldada por la ‘ciencia' y los ‘sistemas de expertos'. En las entrevistas televisivas aparecían ‘académicos' de distintas instituciones del país opinando al respecto y lo sorprendente es que su ‘punto de vista' sobre el caso, resultaba ser tan inverosímil como el de las instancias de salud. Incluso, como se verá, las ‘versiones académicas' terminaron otorgándole, quizá sin quererlo, verosimilitud a la ‘versión oficial' de la Secretaría de Salud. Lo interesante del caso es que las explicaciones que se produjeron para justificar la existencia de los ‘hechos' se apegaron con estricto fervor romántico a las suposiciones hipnótico-epidemiológicas desarrolladas por la ‘psicología de masas' de finales del siglo XIX y principios del XX. Sirva el presente ensayo para hacer una revisión no sólo del curioso caso de la villa de las niñas sino de la forma en que la construcción de versiones, descripciones y explicaciones, circulan de modos particulares y que el entendimiento de la forma en cómo circulan dichas versiones, descripciones y explicaciones, permite entender la complejidad cultural de cualquier entorno social. 

  7. Neurotoxicity evaluation of three root canal sealers on cultured rat trigeminal ganglion neurons

    Science.gov (United States)

    Ayar, Ahmet; Kalkan, Omer-Faruk; Canpolat, Sinan; Tasdemir, Tamer; Ozan, Ulku

    2017-01-01

    Background The aim of this study was to investigate the possible neurotoxic effects of 3 root canal sealers (RCSs) (AH Plus, GuttaFlow, iRoot SP) on cultured rat trigeminal ganglion (TG) neurons. Material and Methods Primary cultures of TG neurons were obtained from 1 to 2-day old rats. Freshly mixed RCSs were incubated in sterile phosphate buffered saline and cells were incubated with supernatants of the RCSs for different time intervals (1-, 3-, 6- and 24-h; 1 or 1/10 diluted) and viability/cytotoxicity was tested by counting the number of live cells. Pair of dishes with cells from the same culture incubated with only culture medium was considered as negative controls. Cell images were captured and acquired at x200 magnification using a microscope equipped with a camera using special image program. The viable cells were manually counted assigned from the images for each dose and incubation duration. Data was analysed by using 1-way analysis of variance with Tukey post hoc tests. Results There was no significant change in cell viability after short duration of incubation (1- and 3-h) with the supernatant of any of RCSs, except for undiluted-AH Plus at 3-h. When AH Plus was compared with other RCSs, for diluted supernatants, there was only significant difference between iRoot SP and AH Plus at 24-h (P<0.05). Whereas undiluted-AH Plus was significantly more cytotoxic for 3-, 6- and 24-h periods as compared to respective incubation periods of undiluted other groups (P<0.05). GuttaFlow groups had similar neurotoxic effect on cells for all test periods. Conclusions All tested RCSs exhibited a variable degree of neurotoxicity on these primary sensory neurons of orofacial tissues, depending on their chemical compositions. GuttaFlow and iRoot SP evoked a less toxic response to TG cells than AH Plus. Key words:Neurotoxicity, trigeminal ganglia, cell culture, root canal sealer, AH Plus, GuttaFlow, iRoot SP. PMID:28149460

  8. Monitoring of intracellular ribonucleotide pools is a powerful tool in the development and characterization of mammalian cell culture processes.

    Science.gov (United States)

    Grammatikos, S I; Tobien, K; No, W; Werner, R G

    1999-08-05

    Efficient cell culture process development for the industrial production of recombinant therapeutics is characterized by constraints which pertain to issues such as costs, competitiveness and the meeting of project timelines. These constraints require tools which can help the developer learn as much as possible as quickly as possible about the cell at hand and identify features of a particular culture which are amenable to improvement. Current on- and off-line monitoring parameters, however useful, provide only late indications (cell concentration, viability) and circumstantial evidence (lactate, ammonia, etc.) with regard to the physiologic status of cells at the time of sampling. The relative intracellular content of purine to pyrimidine nucleotide triphosphates as well as the ratio of UTP to UDP-N-acetylhexosamines have been previously described as sensitive indicators of a cell's metabolic status, growth potential, and overall physiological condition. The sensitivity of such nucleotide ratios and their usefulness in commercially relevant process development and characterization were tested at Boehringer Ingelheim Pharma KG in a large number of fermentations (>80) with a variety of culture modes, cells, and products in scales up to 10,000 litres. Monitoring of these intracellular parameters allows a timely and reliable assessment of cell state and growth potential, which is possible neither by classical cell number and viability measurements nor by a variety of fermentation data typically monitored. The view inside the cell afforded by nucleotide monitoring enables prediction of the behavior of a culture up to 2 days before any hint of physiological changes is given by cell number and viability estimation. In this paper, data relating the growth behavior of CHO and hybridoma cell lines to their nucleotide pools are shown. Two very different processes for the production of recombinant tPA in 10,000-litre bioreactors are compared and characterized with respect to

  9. Cultural Understanding Through Cross-Cultural Analysis.

    Science.gov (United States)

    Briere, Jean-Francois

    1986-01-01

    A college course used an explicit intercultural approach and collective research activities to compare French and American cultures and to examine the reasons for cultural attitudes and culture conflict. Class assignments dealt with contrastive analyses of American and French institutions like advertising, cinema, feminism, etc. (MSE)

  10. Cultural Understanding Through Cross-Cultural Analysis.

    Science.gov (United States)

    Briere, Jean-Francois

    1986-01-01

    A college course used an explicit intercultural approach and collective research activities to compare French and American cultures and to examine the reasons for cultural attitudes and culture conflict. Class assignments dealt with contrastive analyses of American and French institutions like advertising, cinema, feminism, etc. (MSE)

  11. LL-37, HNP-1, and HBD2/3 modulate the secretion of cytokines TNF-α, IL-6, IFN-γ, IL-10 and MMP1 in human primary cell cultures.

    Science.gov (United States)

    Medina Santos, Carlos Erik; López Hurtado, Carmen Nathaly; Rivas Santiago, Bruno; Gonzalez-Amaro, Roberto; Cataño Cañizales, Yolanda Guadalupe; Martínez Fierro, Margarita de la Luz; Enciso-Moreno, José Antonio; García Hernández, Mariana Haydee

    2016-09-01

    The aim of this study was to evaluate the effects of the LL-37, HNP-1 and HBD2/3 peptides on cytokine and MMP production in human polymorphonuclear cells, mononuclear cells and chondrocytes. The levels of cytokines in supernatants from mononuclear and polymorphonuclear cell cultures were measured with a cytometric bead array by flow cytometry. Likewise, the levels of metalloproteinase/MMP-1, 3, and 13 were measured in supernatants from chondrocyte cultures using an ELISA. The expression of RANKL on lymphocytes was analyzed by flow cytometry. We observed increased levels of TNF-α, IL-6 and IL-10 in mononuclear and polymorphonuclear cell cultures stimulated with HBD-2/3. We also observed increased levels of IFN-γ, IL-10, and IL-6 in mononuclear cell cultures stimulated with HNP-1, and increased IL-6 levels were observed in polymorphonuclear cell cultures exposed to HNP-1. We also found that the MMP-1 level increased in the chondrocyte cultures stimulated with HBD-3, whereas the MMP-1 level was decreased in cultures exposed to LL-37. The present report is the first study to determine that HNP-1and HBD2/3 promote the secretion of pro-inflammatory cytokines by polymorphonuclear and mononuclear cells and the secretion of MMP by chondrocytes, whereas LL-37 diminishes MMP1 secretion. Our results suggest that HBD-2/3 and HNP1 might play a pathological role in rheumatoid arthritis, while LL-37 might have a protective role.

  12. (Catharanthus roseus) tissue culture

    African Journals Online (AJOL)

    STORAGESEVER

    2008-08-18

    Aug 18, 2008 ... indole alkaloids in plant tissue cultures of C. roseus have. *Corresponding ... alkaloids from C. roseus cell cultures have failed (review- ed by Van der ..... that vinblastine occur in callus culture with differentiated roots. Dimeric ...

  13. Microalgal Culture Collection Transfers

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Milford Microalgal culture Collection holds over 200 live cultures representing 13 classes of of algae. The cultures are maintained in three different growing...

  14. Culture - joint fluid

    Science.gov (United States)

    Joint fluid culture ... fungi, or viruses grow. This is called a culture. If these germs are detected, other tests may ... is no special preparation needed for the lab culture. How to prepare for the removal of joint ...

  15. Routine sputum culture

    Science.gov (United States)

    Sputum culture ... There, it is placed in a special dish (culture). It is then watched to see if bacteria ... Chernecky CC, Berger BJ. Culture, routine. In: Chernecky CC, Berger BJ, ... . 6th ed. Philadelphia, PA: Elsevier Saunders; 2013:409- ...

  16. Lymph node culture

    Science.gov (United States)

    Culture - lymph node ... or viruses grow. This process is called a culture. Sometimes, special stains are also used to identify specific cells or microorganisms before culture results are available. If needle aspiration does not ...

  17. Blood Culture (For Parents)

    Science.gov (United States)

    ... Kids to Be Smart About Social Media Blood Culture KidsHealth > For Parents > Blood Culture Print A A ... adjust the treatment choice. Why Do a Blood Culture? During some illnesses, certain infection-causing bacteria and ...

  18. Cross-cultural awareness

    OpenAIRE

    2016-01-01

    The article deals with the importance of cultural awareness for businesspeople when they go abroad. It also gives some cultural advice and factors which are thought to be the most important in creating a culture.

  19. Animal culture: chimpanzee conformity?

    Science.gov (United States)

    van Schaik, Carel P

    2012-05-22

    Culture-like phenomena in wild animals have received much attention, but how good is the evidence and how similar are they to human culture? New data on chimpanzees suggest their culture may even have an element of conformity.

  20. Isolation and characterization of a native strain of Aspergillus niger ZRS14 with capability of high resistance to zinc and its supernatant application towards extracellular synthesis of zinc oxide nanoparticles

    Directory of Open Access Journals (Sweden)

    Morahem Ashengroph

    2013-01-01

    Full Text Available Introduction: Zinc oxide nanoparticles have quite a few applications in the fields of biology, optics, mechanics, magnetism, energy, hygiene and medicine. Due to serious problems associated with physiochemical synthesis of ZnO nanoparticles, including environmental pollution, complicated and costly processes, there is a growing need to develop a simple biological procedure for synthesis of nanoparticles to achieve the monodisperse-sized particles with a higher purity, low energy consumption and a cleaner environment. We conducted this investigation to screen and isolate native fungi strains capable of high zinc metal tolerance ability and a potential for extracellular synthesis of ZnO nanoparticles using fungal secretions as biological catalysts.Materials and methods: 15 different strains of fungi were isolated from soil samples collected from lead and zinc mines of Angoran-Zanjan using conventional enrichment process and characterized initially based on macroscopic and microscopic characteristics and colony morphology. The intrinsic tolerance of the isolated strains to zinc toxic metal was measured in the synthetic and complex media using the agar dilution method. The supernatants of isolated fungi were incubated with zinc acetate solution in a shaker incubator for 72h; then, the strain that was able to synthesis ZnO nanoparticle was identified. The ZnO nanoparticles formation was investigated by using spectroscopic techniques and microscopic observations.Results: Among the 15 isolated strains, the strain ZRS14 had highest zinc metal tolerance ability and was selected and identified as Aspergillus niger strain ZRS14 (GenBank accession number KF414527 based on morphological and molecular phylogenetic analysis. For synthesis of ZnO nanoparticles by isolated A. niger ZRS14, fungal cell-free filtrate of the strain was collected and incubated in the presence of zinc acetate solution at a final concentration of 250 mg/l zinc metal ion at 28º C for

  1. Cultural Context and Translation

    Institute of Scientific and Technical Information of China (English)

    张敏

    2009-01-01

    cultural context plays an important role in translation. Because translation is a cross-culture activity, the culture context that influ-ences translating is consisted of both the culture contexts of source language and target language. This article firstly analyzes the concept of context and cultural context, then according to the procedure of translating classifies cultural context into two stages and talks about how they respectively influence translating.

  2. Cultural Analysis - towards cross-cultural understanding

    DEFF Research Database (Denmark)

    Gullestrup, Hans

    The book considers intercultural understanding and co-action, partly by means of general insight into concept of culture and the dimensions which bring about cultural differences, and partly as a methodology to analyse a certain culture - whether one's own or others'. This leads towards an unders......The book considers intercultural understanding and co-action, partly by means of general insight into concept of culture and the dimensions which bring about cultural differences, and partly as a methodology to analyse a certain culture - whether one's own or others'. This leads towards...... a theoretical/abstract proposal for cultural understanding. The second part presents a theoretical/abstract proposal for under-standing intercultural plurality and complexity. The third part provides an empirical model for the analysis of intercultural co-action. Finally, the fourth part present and discusses...

  3. FROM CULTURAL IMPOTENCE TO CULTURAL AMPUTATION

    Directory of Open Access Journals (Sweden)

    Вячеслав Владимирович Суханов

    2013-04-01

    Full Text Available Cultural space of any state is formed by a population that is within its borders. In this article, the author introduces a new cultural definitions «cultural impotence» and «cultural amputation», justifying their use, both in terms of population of the Russian Federation and the European Union and America. The article analyzes the state of society and the cultural factors that influence the development of society in Russia, there are options to bring the country out of a deep cultural crisis. Also established a close relationship between the domestic policy of the state and development of culture.DOI: http://dx.doi.org/10.12731/2218-7405-2013-2-1

  4. Eye extract improves cell migration out of lymphoid organ explants of L. vannamei and viability of the primary cell cultures.

    Science.gov (United States)

    Li, Wenfeng; Van Tuan, Vo; Van Thuong, Khuong; Bossier, Peter; Nauwynck, Hans

    2015-08-01

    Since no cell line from shrimp has been established up till now, an optimization of the primary cell culture protocol is necessary. In this context, the effect of extracts (supernatant of a 1:50 (w/v) suspension) from different shrimp organs (muscle, brain, ganglia, eyestalk, ovary, and eye) on the performance of primary lymphoid cell cultures was evaluated. Ten percent of eye extract and 3% of ovary extract enhanced maximally the migration and survival of cells of the lymphoid organ of Litopenaeus vannamei significantly at 48, 96, and 144 h post seeding. Extracts from the eyestalk (10%), muscle (10%), and brain (1%) significantly promoted the migration and survival of cells at 48 and 96 h post seeding but not anymore at 144 h post seeding. In conclusion, it may be advised to add 10% of eye extract or 3% of ovary extract to cells for the maximal health of primary cell cultures from the lymphoid organ of L. vannamei.

  5. Isolation and risk assessment of Geotrichum spp. in the white shrimp (Litopenaeus vannamei Boone, 1931 from culture ponds

    Directory of Open Access Journals (Sweden)

    José Luis Ochoa

    2015-09-01

    Full Text Available The present study was done in order to identify the fungus invading some of the supralittoral ponds used for shrimp aquaculture in the CIBNOR facilities in La Paz, Baja California Sur (BCS, México during the summer season. From the walls and bottoms of the ponds, two strains of Geotrichum spp. were isolated and morphologically identified. Fungal adhesion towards hemocytes and primary cultures of various white shrimp (Litopeneaus vannamei tissues (gill, tegument, and gut was analyzed to determine infectivity. Extracellular protease, lipase, and amylase activity were evaluated as virulence factors. Survival of shrimp post-larvae (PL8 exposed to fungal culture supernatant or to their filaments was also investigated. The results showed that shrimp tegument cells and hemocytes were very susceptible to Geotrichum spp. invasion, and that this fungus provokes great mortality of post-larvae. Hence, Geotrichum spp. could be considered an opportunistic pathogen that might represent a serious health risk to shrimp in culture.

  6. Trypanosoma acomys (Wenyon, 1909): continuous culturing with a mouse fibroblast cell-line (A9).

    Science.gov (United States)

    Abdallah, M A; Abdel-Hafez, S K; al-Yaman, F M

    1990-01-01

    The continuous culturing of Trypanosoma acomys in the presence of a murine areolar-adipose cell line (A9) was possible for the 1st time. The trypanosomes were cultured at 37 degrees C with A9 in DMEM supplemented with 20% heat inactivated fetal bovine serum, using an initial inoculum from primary cultures of lung or blood clots from infected spiny mice. The cultures were maintained for 115 days and underwent 15 passages before termination and cryopreservation. Using this culture system T. acomys subcultures were initiated from 3 different initial inocula (3 x 10(4), 1.5 x 10(5) and 7.4 x 10(5) parasites/ml) and growth curves revealed that the lowest inoculum gave the best growth pattern. This inoculum yielded a population doubling time of less than 12 h for 4 days, a high peak density of 7 x 10(6) parasites/ml and the most gradual decline compared to the other 2 inocula. Rosetting epimastigotes and nests of amastigotes were observed in close association with the feeder layer cells. Epimastigotes were the most predominant form in culture supernatants but other morphological forms observed included trypomastigotes and sphaeromastigotes.

  7. In vitro culture and characterization of human umbilical cord blood-derived plasmacytoid dendritic cell subsets

    Directory of Open Access Journals (Sweden)

    PENG Jianping

    2015-11-01

    Full Text Available ObjectiveTo establish a method for in vitro culture of plasmacytoid dendritic cell (pDC. MethodsUmbilical cord blood (40 ml was collected from healthy parturients in the First Affiliated Hospital of Hunan University of Chinese Medicine, and cord blood mononuclear cell (CBMC were isolated. The CBMC were cultured for 7 days with RPMI 1640 complete medium containing rh Flt3-ligand (Flt3-L (100 ng/ml and rh interleukin (IL-3 (10 ng/ml, and the medium was half changed every 2 days. On the eighth day, CpG ODN (2 μg/ml was added to the cells, and the attached cells and supernatant were collected 24 h later for flow cytometry and interferon (IFNα measurement, respectively. On days 1, 3, 5, 7, and 8 of cell culture, the morphological changes of pDC were observed. Results After 2 h of culture, the CBMC showed circular, flat morphology. Twenty-four hours later, the cells began to adhere to the wall, with extended cytoplasm and increased volumes, and they became round and translucent, with scattered small colonies. On days 3-4 of culture, the cell volume continued increasing; most cells were round, and some had small protrusions; few cells were spindle-, tadpole-, star- or irregularly shaped; the number and volumes of colonies increased substantially. On days 5-8 of culture, the number of colonies and the number of cells in colonies gradually decreased, and suspended cells that were round or had small protrusions gradually increased in the medium. The cells expressing CD123, BDCA-2, and BDCA-4, which were considered pDC, were detected by flow cytometry. Flow cytometry revealed that the proportion of pDC in CBMC increased during the culture: increasing from 1.08% at the beginning of culture to 5.32% on day 4, and finally reaching a peak of 19.8% on day 8. On day 8, the level of IFNα in pDC culture supernatant was(11 302.61±1745.31 pg/ml. ConclusionpDC can be successfully induced in vitro by rh Flt3-L combined with IL-3 from human umbilical CBMC.

  8. Information and Corporate Cultures.

    Science.gov (United States)

    Drake, Miriam A.

    1984-01-01

    This paper defines "corporate culture" (set of values and beliefs shared by people working in an organization which represents employees' collective judgments about future) and discusses importance of corporate culture, nature of corporate cultures in business and academia, and role of information in shaping present and future corporate cultures.…

  9. Culture and English Teaching

    Institute of Scientific and Technical Information of China (English)

    李莹

    2008-01-01

    There is a natural relationship between culture and language. Language reflects how the people of a nation form the unique way of life and the way of thinking. Therefore, English teaching necessarily involves cultural education as well. This paper analyzes the influence of social culture in English teaching and tries to set up a principle of teaching English culture.

  10. Culture Wars in Denmark

    DEFF Research Database (Denmark)

    Ørum, Tania

    2016-01-01

    In the 1960s high and low culture were brought into sharp conflict i Denmark. In 1961 a Ministry of Culture was established for the first time. The first minister of culture, the social democrat Julius Bomholt, saw art and culture as an important part of education for democracy that should be made...

  11. Many Forms of Culture

    Science.gov (United States)

    Cohen, Adam B.

    2009-01-01

    Psychologists interested in culture have focused primarily on East-West differences in individualism-collectivism, or independent-interdependent self-construal. As important as this dimension is, there are many other forms of culture with many dimensions of cultural variability. Selecting from among the many understudied cultures in psychology,…

  12. Cultural Identity Through CLIL

    OpenAIRE

    Oprescu Monica

    2015-01-01

    The CLIL approach is a modern manner of teaching English, which has been adapted in Romanian schools and universities. An interesting aspect of learning a foreign language is the contact with its culture/s and the changes it produces in terms of identity. Therefore, a challenging question to be answered is whether a CLIL approach focusing on culture influences students' cultural identity.

  13. Screening of Cytotoxic B. cereus on Differentiated Caco-2 Cells and in Co-Culture with Mucus-Secreting (HT29-MTX) Cells.

    Science.gov (United States)

    Castiaux, Virginie; Laloux, Laurie; Schneider, Yves-Jacques; Mahillon, Jacques

    2016-11-05

    B. cereus is an opportunistic foodborne pathogen able to cause diarrhoea. However, the diarrhoeal potential of a B. cereus strain remains difficult to predict, because no simple correlation has yet been identified between the symptoms and a unique or a specific combination of virulence factors. In this study, 70 B. cereus strains with different origins (food poisonings, foods and environment) have been selected to assess their enterotoxicity. The B. cereus cell-free supernatants have been tested for their toxicity in vitro, on differentiated (21 day-old) Caco-2 cells, using their ATP content, LDH release and NR accumulation. The genetic determinants of the main potential enterotoxins and virulence factors (ces, cytK, entFM, entS, hbl, nhe, nprA, piplC and sph) have also been screened by PCR. This analysis showed that none of these genes was able to fully explain the enterotoxicity of B. cereus strains. Additionally, in order to assess a possible effect of the mucus layer in vitro, a cytotoxicity comparison between a monoculture (Caco-2 cells) and a co-culture (Caco-2 and HT29-MTX mucus-secreting cells) model has been performed with selected B. cereus supernatants. It appeared that, in these conditions, the mucus layer had no notable influence on the cytotoxicity of B. cereus supernatants.

  14. Language and Culture

    Institute of Scientific and Technical Information of China (English)

    徐君

    2011-01-01

    As the carrier of culture,language is considered as the main expressional form of culture which develops with nation,country and society’s development.Language is a part of a nation’s culture.The different nations own their unique cultures,his-tory,manners and customs and so on.However,various cultural characteristics can be displayed in the form of language.This ar-ticle,by analyzing the influence and the difference of historical culture,regional culture and custom culture,mainly reveals the relationship between language and culture which is interdependent and interactive.What’s more,a better comprehension of this relationship prevents us from misunderstanding in cross-culture communication.

  15. Cultural Capital in Context:

    DEFF Research Database (Denmark)

    Andersen, Ida Gran; Jæger, Mads Meier

    This paper analyzes the extent to which the effect of cultural capital on academic achievement varies across high- and low-achieving schooling environments. We distinguish three competing theoretical models: Cultural reproduction (cultural capital yields higher returns in high-achieving schooling...... environments than in low-achieving ones), cultural mobility (cultural capital yields higher returns in low-achieving environments), and cultural resources (cultural capital yields the same returns in different environments). We analyze PISA data from six countries and find that returns to cultural capital tend...... to be higher in low-achieving schooling environments than in high-achieving ones. These results support the cultural mobility explanation and are in line with previous research suggesting that children from low-SES families benefit more from cultural capital than children from high-SES families....

  16. Absorbing the Culture Shock

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Participants at a forum on communication between cultures generate ideas on how China can deal with its growing cultural deficit Five months ago, Ding Wei, Assistant Minister of Culture, described China's deficit in international cultural trade as "huge" at a press conference held by the State Council Information Office. "Our statistics years ago showed that the ratio of imports of cultural products to exports was 10 to 1," he

  17. KEEPING CULTURAL GENES ALIVE

    Institute of Scientific and Technical Information of China (English)

    Bai Shi

    2012-01-01

    China's contemporary culture and the protection of its diverse cultural heritage have become some of the most talked about issues today.Cultural prosperity was put forward as an important objective of the Central Government's national development strategy last year.However,the industrialization and commercialization of China's culture have been both criticized and celebrated.Many scholars believe industrialization and enormous government investment may not be the best means to protect intangible cultural heritage (ICH).

  18. DIAGNOSIS OF CULTURAL ORGANIZATIONS

    Directory of Open Access Journals (Sweden)

    ALBU MĂDĂLINA

    2015-04-01

    Full Text Available Cultural institution management is to direct the organization to a specific cultural profile purpose, namely production values esthetic sense, artistic, moral, spiritual, distribution, promotion of these values, protection and circulation of cultural heritage. In this regard, an analysis in the diagnosis cultural organizations aims to determine the main strengths and weaknesses, assess the potential and making recommendations focused on the root causes of failures and positive aspects. This paper presents considerations diligence activity Culture House "IL Caragiale "in Ploiesti. The mission of this organization is to contribute to the cultural development of the community by initiating projects and cultural programs, offer development programs and services to meet cultural needs, increase public access to diverse cultural life, providing a constant presence institution circuit local, national, European and international level. Conclusions drawn from the analysis shows that in a world of economic globalization, information and culture in a company in constant change, in a competitive market where there is information readily available means and leisure, but not cultural consistency in a social environment where interest in culture of people is declining, the situation of the population is impaired, the remuneration of staff working in the field of cultural education is demotivating, the funds allocated to culture have grown lately effectively lead a cultural institution is a challenge.

  19. Bovine lactotroph cultures for the study of prolactin synthesis functions.

    Science.gov (United States)

    Wang, Jianfa; Yang, Zhanqing; Fu, Shoupeng; Liu, Bingrun; Wu, Dianjun; Wang, Wei; Sun, Dongbo; Wu, Rui; Liu, Juxiong

    2016-03-01

    The aim of this study was to establish a bovine anterior pituitary-derived lactotroph (BAPDL) line that expresses prolactin (PRL) in vitro to study the mechanisms of bovine PRL synthesis and secretion. Immunohistochemistry assay of PRL in the newborn calves' anterior pituitary glands showed that most lactotrophs were located within the superior border of the lateral wings of the anterior pituitary. Tissues of the superior border of the lateral wings of the anterior pituitary were dispersed and cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS). The limiting dilution method was used to establish BAPDL from single cell clone. BAPDL cells constantly expressed mRNAs for PRL and pituitary-specific transcription factor 1 (Pit-1) gene and grew steadily and rapidly in the DMEM supplemented with 10% FBS. PRL immunoreactivity was present in BAPDL at passage 20. The concentration of bovine PRL in BAPDL at passage 20 culture supernatant was decreased to below 35% compared with that in BAPDL at passage 1. The effects of human epidermal growth factor (hEGF) and dopamine (DA) on the expression and secretion of PRL in BAPDL at passage 4 were also investigated. The results are consistent with those of previous studies. Thus, it can be used successfully for studying the mechanisms of stimuli regulating PRL synthesis and release.

  20. Corneal organ cultures in tyrosinemia release chemotactic factors.

    Science.gov (United States)

    Lohr, K M; Hyndiuk, R A; Hatchell, D L; Kurth, C E

    1985-05-01

    Corneal inflammation with subsequent scarring and blindness occurs in the inherited human metabolic disease tyrosinemia type II, yet putative inflammatory mediators in this disorder and in the avascular cornea in general are poorly defined. In a Tyr-fed rat model of tyrosinemia type II, intracellular crystals, presumably Tyr, are hypothesized to be responsible for the increased lysosomal activity observed in corneal epithelial lesions. Because polymorphonuclear leukocytes (PMNs) are seen only at the site of these lesions, we used this model to study humoral mediators released from Tyr-fed rat corneal organ cultures. Only Tyr-fed rats developed stromal edema and linear granular opacities in gray edematous corneal epithelium, compatible with a noninfectious keratitis. Electron micrographs confirmed epithelial edema and showed focal epithelial necrosis with PMN invasion of the stroma. Only Tyr-fed rat corneal culture supernatants contained chemotactic activity that was heat labile and moderately trypsin sensitive. Four peaks with varying amounts of chemotactic activity were found on Sephadex G-75 chromatography. Although the identity of these peaks of activity has not yet been established, we suggest that they may be responsible for the PMN infiltration observed in this model of corneal inflammation.

  1. Evaluation of predictive tools for cell culture clarification performance.

    Science.gov (United States)

    Senczuk, Anna; Petty, Krista; Thomas, Anne; McNerney, Thomas; Moscariello, John; Yigzaw, Yinges

    2016-03-01

    Recent advances in the productivity of industrial mammalian cell culture processes have resulted in part in increased cell density. This increase and the associated increase in cellular debris are known to challenge harvest operations, however this understanding is limited and largely qualitative. Part of the issue arises from the heterogeneous size and composition of cellular debris, which makes harvest feed stream extremely difficult to characterize. Improved characterization methods would facilitate the development of clarification approaches that are consistent and scalable. This work describes how both particle size and cholesterol analysis can be used to characterize the feed stream. Particle size analysis by focused beam reflectance and dynamic light scattering are shown to be predictive of centrate filterability under certain harvest conditions. Because of the particle size range limitations of each detector, their applicability is limited to a particular stage or method of clarification. The measurement of cholesterol present in the cell culture supernatant or centrate was successfully used in providing relative amount of lysed cellular debris and enabled us to predict clarification performance of acid precipitated harvest regardless of particle size distribution profile. © 2015 Wiley Periodicals, Inc.

  2. DEVELOPMENT OF PRIMARY CELL CULTURE FROM TAIL EPIDERMAL TISSUE OF KOI CARP (Cyprinus carpio koi

    Directory of Open Access Journals (Sweden)

    Lila Gardenia

    2014-06-01

    Full Text Available Primary cell culture from tail epidermal tissue of koi carp (Cyprinus carpio koi was developed. Cells were grown in Leibovits-15 medium supplemented with 20% fetal bovine serum and antibiotics (Penicillin/Streptomycin and Kanamycin. Cell growth was observed in a range of incubation temperature (17oC±2oC, 22oC±2oC, 27oC±2oC, and 32oC±2oC in order to determine the optimum temperature. The cells were able to grow at a range of temperature between 17oC to 32oC with optimal growth at 22oC. Primary cells infected with koi herpes virus produced typical cytopathic effects characterized by severe vacuolation and deformation of nuclei, which is consistent with those of previous reports. Artificial injection experiment by using supernatant koi herpes virus SKBM-1 isolate revealed that it could cause 90% mortality in infected fish within two weeks. PCR test with Sph I-5 specific primers carried out with DNA template from supernatant virus, pellet cell, and gills of infected fish showed positive results in all samples (molecular weight of DNA target 290 bp. The cells were found to be susceptible to koi herpes virus and can be used for virus propagation.

  3. Ambroxol inhibits rhinovirus infection in primary cultures of human tracheal epithelial cells.

    Science.gov (United States)

    Yamaya, Mutsuo; Nishimura, Hidekazu; Nadine, Lusamba Kalonji; Ota, Chiharu; Kubo, Hiroshi; Nagatomi, Ryoichi

    2014-04-01

    The mucolytic drug ambroxol hydrochloride reduces the production of pro-inflammatory cytokines and the frequency of exacerbation in patients with chronic obstructive pulmonary disease (COPD). However, the inhibitory effects of ambroxol on rhinovirus infection, the major cause of COPD exacerbations, have not been studied. We examined the effects of ambroxol on type 14 rhinovirus (RV14) infection, a major RV group, in primary cultures of human tracheal epithelial cells. RV14 infection increased virus titers and cytokine content in the supernatants and RV14 RNA in the cells. Ambroxol (100 nM) reduced RV14 titers and cytokine concentrations of interleukin (IL)-1β, IL-6 and IL-8 in the supernatants and RV14 RNA in the cells after RV14 infection, in addition to reducing susceptibility to RV14 infection. Ambroxol also reduced the expression of intercellular adhesion molecule-1 (ICAM-1), the receptor for RV14, and the number of acidic endosomes from which RV14 RNA enters the cytoplasm. In addition, ambroxol reduced the activation of the transcription factor nuclear factor kappa B (NF-κB) in the nucleus. These results suggest that ambroxol inhibits RV14 infection partly by reducing ICAM-1 and acidic endosomes via the inhibition of NF-κB activation. Ambroxol may modulate airway inflammation by reducing the production of cytokines in rhinovirus infection.

  4. Characterizing European cultural landscapes

    DEFF Research Database (Denmark)

    Tieskens, Koen F.; Schulp, Catharina J E; Levers, Christian

    2017-01-01

    European cultural landscapes are mostly characterized by only one of the dimensions. Our paper can help to identify pressures to cultural landscapes and thus to target measures for the conservation of these landscapes, to link similar landscapes in different regions, and to inform policy design on the most......Almost all rural areas in Europe have been shaped or altered by humans and can be considered cultural landscapes, many of which now are considered to entail valuable cultural heritage. Current dynamics in land management have put cultural landscapes under a huge pressure of agricultural...... intensification and land abandonment. To prevent the loss of cultural landscapes, knowledge on the location of different types of cultural landscapes is needed. In this paper, we present a characterization of European cultural landscapes based on the prevalence of three key dimensions of cultural landscapes...

  5. Cultural Molding: A Modular Approach. Cultural Anthropology.

    Science.gov (United States)

    Kassebaum, Peter

    Designed for use as supplementary instructional material in a cultural anthropology course, this learning module introduces the student to cultural molding, the idea that most human behavior can be traced to enculturation and exposure rather than to a socio-biological explanation of human behavior. Following a brief description of socialization,…

  6. Robotic art, culture and cultural imagination

    DEFF Research Database (Denmark)

    Romic, Bojana

    2015-01-01

    In the recent years there has been an ongoing debate about the notion and (possible) function of robotic culture; with the development of the new generation of drones, as well as the advancement in social and health robotics, questions about robot culture seem to open a variety of discussions...... – beginning with the inquiry of how can we grasp the notion of culture, as an umbrella term for the range of habitual and technological practices, in relation to equally heterogeneous field of robotics. This article aims to accentuate the importance of cultural imagination of robots in situating the robotic...... research, observing it 'as a mixed register of fantasy and an actual practice' (Kakoudaki, 2007). The emphasis will be put on the robotic art which, I argue, is in the fluid state of exchange with other areas of robotic research, equally benefiting from the larger context of cultural imagination of robots...

  7. Developing Cultural Awareness

    OpenAIRE

    2005-01-01

    This paper aims at emphasizing the issue of teaching of culture in foreign language teaching.  In this respect, the reasons of teaching culture in foreign language classes are focused on initially.  So, the justifications of teaching culture are considered and explained and by the help of a dialogue.  Right after this, ways of developing cultural awareness is taken into account.  At this step, types of courses to develop cultural awareness are dealt with.  Developing cultural awareness in cla...

  8. Cultural Activation of Consumers.

    Science.gov (United States)

    Siegel, Carole E; Reid-Rose, Lenora; Joseph, Adriana M; Hernandez, Jennifer C; Haugland, Gary

    2016-02-01

    This column discusses "cultural activation," defined as a consumer's recognition of the importance of providing cultural information to providers about cultural affiliations, challenges, views about, and attitudes toward behavioral health and general medical health care, as well as the consumer's confidence in his or her ability to provide this information. An aid to activation, "Cultural Activation Prompts," and a scale that measures a consumer's level of activation, the Cultural Activation Measurement Scale, are described. Suggestions are made about ways to introduce cultural activation as a component of usual care.

  9. Cultural Exchange Strengthens Ties

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    The Year of Chinese Culture in Australia is poised to cement bonds between the Chinese and Australians As the Year of Australian Culture in China drew to a close in June,the Year of Chinese Culture in Australia,titled Experience China,came ons tage to showcase the rich variety of China’s traditional and contemporary culture to the Australians.The opening ceremony of the event was held on June 24 at the Sydney State Theater.It featured famous Chinese dancer Yang Liping’s original dance drama The Legend of Shangri-La.The Year of Chinese Culture in Australiais by far the largest Chinese cultural festival

  10. On value and culture

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Value stands for the significance of things,and concepts of value are ideas,opinions and attitudes about what kind of things are significant.Studies on the concept of value cannot be separated from culture.Every society has its own distinct culture and concept of value shared by its people.The relationship between concept of value and culture shows that the building of the concept of value must be based on culture.Developing culture,providing excellent products of culture and better humanitarian environment are the premise for people's possession of the correct concept of value.

  11. Dexamethasone inhibition of IL-1-induced collagen degradation by corneal fibroblasts in three-dimensional culture.

    Science.gov (United States)

    Lu, Ying; Fukuda, Ken; Liu, Yang; Kumagai, Naoki; Nishida, Teruo

    2004-09-01

    Corticosteroids regulate the functions of inflammatory cells. The purpose of the present study was to investigate the effect of dexamethasone on collagen degradation by corneal fibroblasts, an underlying cause of corneal ulceration. Rabbit corneal fibroblasts were cultured in three-dimensional gels of type I collagen and in the absence or presence of IL-1beta or dexamethasone. The extent of collagen degradation was determined by measurement of the amount of hydroxyproline generated by acid-heat hydrolysis of culture supernatants. The expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) was evaluated by immunoblot analysis, gelatin zymography, and reverse transcription and real-time polymerase chain reaction. The phosphorylation of mitogen-activated protein kinases (MAPKs) in corneal fibroblasts was assessed by immunoblot analysis. Dexamethasone inhibited IL-1beta-induced collagen degradation by corneal fibroblasts in a dose-dependent manner. Both the synthesis and activation of MMPs and the expression of TIMPs were inhibited by dexamethasone, as was the activity of plasmin in culture supernatants. Dexamethasone also inhibited the IL-1beta-induced phosphorylation of the MAPKs extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), but not that of p38. Dexamethasone exerted multiple effects on the MMP-TIMP system in corneal fibroblasts and thereby inhibited IL-1beta-induced collagen degradation by these cells. Inhibition of the IL-1beta-induced activation of ERK and JNK may contribute to these effects of dexamethasone. Copyright Association for Research in Vision and Ophthalmology

  12. In vitro culture and characterization of alveolar bone osteoblasts isolated from type 2 diabetics

    Directory of Open Access Journals (Sweden)

    Dao-Cai Sun

    2012-06-01

    Full Text Available In order to understand the mechanisms of poor osseointegration following dental implants in type 2 diabetics, it is important to study the biological properties of alveolar bone osteoblasts isolated from these patients. We collected alveolar bone chips under aseptic conditions and cultured them in vitro using the tissue explants adherent method. The biological properties of these cells were characterized using the following methods: alkaline phosphatase (ALP chemical staining for cell viability, Alizarin red staining for osteogenic characteristics, MTT test for cell proliferation, enzyme dynamics for ALP contents, radio-immunoassay for bone gla protein (BGP concentration, and ELISA for the concentration of type I collagen (COL-I in the supernatant. Furthermore, we detected the adhesion ability of two types of cells from titanium slices using non-specific immunofluorescence staining and cell count. The two cell forms showed no significant difference in morphology under the same culture conditions. However, the alveolar bone osteoblasts received from type 2 diabetic patients had slower growth, lower cell activity and calcium nodule formation than the normal ones. The concentration of ALP, BGP and COL-I was lower in the supernatant of alveolar bone osteoblasts received from type 2 diabetic patients than in that received from normal subjects (P < 0.05. The alveolar bone osteoblasts obtained from type 2 diabetic patients can be successfully cultured in vitro with the same morphology and biological characteristics as those from normal patients, but with slower growth and lower concentration of specific secretion and lower combining ability with titanium than normal ones.

  13. Cultural Dyphemisms in the Turkish Culture Region

    Directory of Open Access Journals (Sweden)

    İlhami Durmuş

    2014-12-01

    Full Text Available Turkish steppe culture constituted a typical structure with regard to its creation and development. It emerged from steppe geography. The geography affected social, politic, economical, military and religious structure of the society substantially. The Turkish culture was shaped up under these influences. The agents of the culture appeared as a shepherd generally. Shepherding provided a basic to the animal husbandry. The shepherds headed to search the fertile grass. Shepherding obliged people to became seminomadic. Thus the life continued in a manner of searching new pastures and shuttling between plateau and winter quarters . The horse that is one of the important ingredient of the culture provided mobility and speed to the culture. In social, politic, and military life of society the horse came into prominence. The nomadic houses or the cars contributed to Turks’ life very much. These nomad houses were shelter at the nights and cold days for them. Going from the plateau to winter quarters and from pasture to another pasture was made easy thanks to these. The finding which took out from graves reflects to all features of the culture.The ruins of nomad houses and horses were arised from the graves . These support the informations which was given in the written sources. Among the findings, there were also The animal struggle scenes which reflect art concept of theseminomadism between tomb finds. This art concept named as turkish animal style. It’s tried to determine The scope of the Turkish culture by emphasizing steppe, shepherding, semi-nomadism, horse, nomad house, tomb. It is also emphasized that aforementioned notions can be used for Turkish culture. Thus, cultural dysphemisms in the turkish culture based on that idea.

  14. Effect of Bifidobacterium upon Clostridium difficile growth and toxicity when co-cultured in different prebiotic substrates

    Directory of Open Access Journals (Sweden)

    Lorena Valdés Varela

    2016-05-01

    Full Text Available The intestinal overgrowth of Clostridium difficile, often after disturbance of the gut microbiota by antibiotic treatment, leads to C. difficile infection (CDI which manifestation ranges from mild diarrhoea to life-threatening conditions. The increasing CDI incidence, not only in compromised subjects but also in traditionally considered low-risk populations, together with the frequent relapses of the disease, has attracted the interest for prevention/therapeutic options. Among these, probiotics, prebiotics or synbiotics constitute a promising approach. In this study we determined the potential of selected Bifidobacterium strains for the inhibition of C. difficile growth and toxicity in different carbon sources. We conducted co-cultures of the toxigenic strain C. difficile LMG21717 with four Bifidobacterium strains (Bifidobacterium longum IPLA20022, Bifidobacterium breve IPLA20006, Bifidobacterium bifidum IPLA20015, and Bifidobacterium animalis subsp. lactis Bb12 in the presence of various prebiotic substrates (Inulin, Synergy and Actilight or glucose, and compared the results with those obtained for the corresponding mono-cultures. C. difficile and bifidobacteria levels were quantified by qPCR; the pH and the production of short chain fatty acids was also determined. Moreover, supernatants of the cultures were collected to evaluate their toxicity using a recently developed model. Results showed that co-culture with B. longum IPLA20022 and B. breve IPLA20006 in the presence of short-chain fructooligosaccharides, but not of Inulin, as carbon source significantly reduced the growth of the pathogen. With the sole exception of B. animalis Bb12, whose growth was enhanced, the presence of C. difficile did not show major effects upon the growth of the bifidobacteria. In accordance with the growth data, B. longum and B. breve were the strains showing higher reduction in the toxicity of the co-culture supernatants.

  15. Clarification of interactions among microorganisms and development of co-culture system; Biseibutsukan sokosayo no kaiseki to kongo baiyo shisutemu no kaihatsu

    Energy Technology Data Exchange (ETDEWEB)

    Taniguchi, Masayuki [Niigata University, Niigata (Japan). Dept. of Materials Science and Technology

    1999-03-10

    Co-culture systems containing two microorganisms for production of useful substances are described. The co-culture of Bifidobacterium longum and Propionibacterium freudenreichii, where lactic acid produced once from lactose by B. longum is converted to acetic and propionic acids by P. freudenreichii, was carried out. Thought the sequential conversion of lactose using the two microorganisms, the culture supernatant containing a mixture of acetic and propionic acids without lactic acid is produced. The antimicrobial activity of the mixture is higher than that obtained in the cultivation of B. longum alone. We developed a novel co-culture system composed of two fermentors and two micro filtration modules for efficient ethanol production from a mixture of glucose and xylose by co-culture of Pichia stipitis and Saccharomyces cerevisiae. The proposed co-culture system allowed regulation of the dissolved oxygen concentration at a level suitable for an individual yeast in each fermentor, as well as the successful exchange of culture medium between two fermentors. When P. stipitis and S. cerevisiae are cultivated individually under different oxygen supply conditions in the new co-culture system, the yield and productivity of ethanol from a glucose and xylose mixture are higher than in single culture of P. stipitis alone. By clarifying the interactions among microorganisms, new bioprocesses in which similar performance to co-culture systems is expressed even using a single microorganism, are expected to be developed for improvement of biochemical reaction systems. (author)

  16. Production of polyclonal and monoclonal antibodies against the Bacillus thuringiensis vegetative insecticidal protein Vip3Aa16.

    Science.gov (United States)

    Ben Hamadou-Charfi, Dorra; Sauer, Annette Juliane; Abdelkafi-Mesrati, Lobna; Jaoua, Samir; Stephan, Dietrich

    2015-03-01

    The aim of this study is to establish a quantitative determination of the vegetative insecticidal protein Vip3A from the culture supernatant of Bacillus thuringiensis either by ELISA or by the conventional quantification method of the Western blot band. The Vip3A protein was produced by fermentation of the B. thuringiensis reference strain BUPM95 in 3 L. By Western blot, the Vip3Aa16 toxin was detected in the culture supernatant during the exponential growth phase of B. thuringiensis BUPM95. However, the detection of Vip3Aa16 on Western blot showed in addition to the toxin two other strips (62 and 180 kDa) recognized by the anti-Vip3Aa16 polyclonal antibodies prepared at the Centre of Biotechnology of Sfax Tunisia. For that reason and in order to develop a technique for reliable quantification of the toxin, we have considered the production of polyclonal antibodies at the Julius Kühn Institute, Germany. These antibodies were the basis for the production of monoclonal antibodies directed against the protein produced by the Vip3Aa16 recombinant strain Escherichia coli BL21 (DE3). These monoclonal antibodies were tested by plate-trapped antigen (PTA) and triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA). The selection of hybridoma supernatants gave us four positive clones producing monoclonal antibodies.

  17. Human colon tissue in organ culture: calcium and multi-mineral-induced mucosal differentiation.

    Science.gov (United States)

    Dame, Michael K; Veerapaneni, Indiradevi; Bhagavathula, Narasimharao; Naik, Madhav; Varani, James

    2011-01-01

    We have recently shown that a multi-mineral extract from the marine red algae, Lithothamnion calcareum, suppresses colon polyp formation and inflammation in mice. In the present study, we used intact human colon tissue in organ culture to compare responses initiated by Ca(2+) supplementation versus the multi-mineral extract. Normal human colon tissue was treated for 2 d in culture with various concentrations of calcium or the mineral-rich extract. The tissue was then prepared for histology/immunohistochemistry, and the culture supernatants were assayed for levels of type I procollagen and type I collagen. At higher Ca(2+) concentrations or with the mineral-rich extract, proliferation of epithelial cells at the base and walls of the mucosal crypts was suppressed, as visualized by reduced Ki67 staining. E-cadherin, a marker of differentiation, was more strongly expressed at the upper third of the crypt and at the luminal surface. Treatment with Ca(2+) or with the multi-mineral extract influenced collagen turnover, with decreased procollagen and increased type I collagen. These data suggest that calcium or mineral-rich extract has the capacity to (1) promote differentiation in human colon tissue in organ culture and (2) modulate stromal function as assessed by increased levels of type I collagen. Taken together, these data suggest that human colon tissue in organ culture (supporting in vivo finding in mice) will provide a valuable model for the preclinical assessment of agents that regulate growth and differentiation in the colonic mucosa.

  18. Yokukansan, a Traditional Japanese Medicine, Adjusts Glutamate Signaling in Cultured Keratinocytes

    Directory of Open Access Journals (Sweden)

    Maki Wakabayashi

    2014-01-01

    Full Text Available Glutamate plays an important role in skin barrier signaling. In our previous study, Yokukansan (YKS affected glutamate receptors in NC/Nga mice and was ameliorated in atopic dermatitis lesions. The aim of this study was to assess the effect of YKS on skin and cultured human keratinocytes. Glutamate concentrations in skin of YKS-treated and nontreated NC/Nga mice were measured. Then, glutamate release from cultured keratinocytes was measured, and extracellular glutamate concentrations in YKS-stimulated cultured human keratinocytes were determined. The mRNA expression levels of NMDA receptor 2D (NMDAR2D and glutamate aspartate transporter (GLAST were also determined in YKS-stimulated cultured keratinocytes. The glutamate concentrations and dermatitis scores increased in conventional mice, whereas they decreased in YKS-treated mice. Glutamate concentrations in cell supernatants of cultured keratinocytes increased proportionally to the cell density. However, they decreased dose-dependently with YKS. YKS stimulation increased NMDAR2D in a concentration-dependent manner. Conversely, GLAST decreased in response to YKS. Our findings indicate that YKS affects peripheral glutamate signaling in keratinocytes. Glutamine is essential as a transmitter, and dermatitis lesions might produce and release excess glutamate. This study suggests that, in keratinocytes, YKS controls extracellular glutamate concentrations, suppresses N-methyl-D-aspartate (NMDA receptors, and activates glutamate transport.

  19. Modulation of mAb quality attributes using microliter scale fed-batch cultures.

    Science.gov (United States)

    Rouiller, Yolande; Périlleux, Arnaud; Vesin, Marie-Noëlle; Stettler, Matthieu; Jordan, Martin; Broly, Hervé

    2014-01-01

    A high-throughput DoE approach performed in a 96-deepwell plate system was used to explore the impact of media and feed components on main quality attributes of a monoclonal antibody. Six CHO-S derived clonal cell lines expressing the same monoclonal antibody were tested in two different cell culture media with six components added at three different levels. The resulting 384 culture conditions including controls were simultaneously tested in fed-batch conditions, and process performance such as viable cell density, viability, and product titer were monitored. At the end of the culture, supernatants from each condition were purified and the product was analyzed for N-glycan profiles, charge variant distribution, aggregates, and low molecular weight forms. The screening described here provided highly valuable insights into the factors and combination of factors that can be used to modulate the quality attributes of a molecule. The approach also revealed specific intrinsic differences of the selected clonal cell lines - some cell lines were very responsive in terms of changes in performance or quality attributes, whereas others were less affected by the factors tested in this study. Moreover, it indicated to what extent the attributes can be impacted within the selected experimental design space. The outcome correlated well with confirmations performed in larger cell culture volumes such as small-scale bioreactors. Being fast and resource effective, this integrated high-throughput approach can provide information which is particularly useful during early stage cell culture development. © 2014 American Institute of Chemical Engineers.

  20. CULTURE, CULTURE LEARNING AND NEW TECHNOLOGIES: TOWARDS A PEDAGOGICAL FRAMEWORK

    OpenAIRE

    Mike Levy

    2007-01-01

    This paper seeks to improve approaches to the learning and teaching of culture using new technologies by relating the key qualities and dimensions of the culture concept to elements within a pedagogical framework. In Part One, five facets of the culture concept are developed: culture as elemental; culture as relative; culture as group membership; culture as contested; and culture as individual (variable and multiple). Each perspective aims to provide a focus for thinking about culture, and th...

  1. Cross-Cultural Nongeneralizations.

    Science.gov (United States)

    Patton, Michael Quinn

    1985-01-01

    This synthesis of the previous articles concludes that cultural considerations are important for effective evaluation practice. Culturally sensitive and situationally responsive evaluation practices can contribute to international understanding. (BS)

  2. Cultural changes in aerospace

    Science.gov (United States)

    Strobl, Bill

    1991-01-01

    Cultural changes; people and jobs; examples of cultural changes required; advanced launch system (ALS) philosophy; ALS operability capabilities; and ALS operability in design are outlined. This presentation is represented by viewgraphs.

  3. Plant tissue culture techniques

    Directory of Open Access Journals (Sweden)

    Rolf Dieter Illg

    1991-01-01

    Full Text Available Plant cell and tissue culture in a simple fashion refers to techniques which utilize either single plant cells, groups of unorganized cells (callus or organized tissues or organs put in culture, under controlled sterile conditions.

  4. Nordic cultural policies

    DEFF Research Database (Denmark)

    Duelund, Peter

    2008-01-01

    A critical view on Nordic Cultural Policy 1961-2008 - Aims, measures, forms of organisation, state og national identity......A critical view on Nordic Cultural Policy 1961-2008 - Aims, measures, forms of organisation, state og national identity...

  5. Culture in the Classroom

    Science.gov (United States)

    Medin, Douglas L.; Bang, Megan

    2014-01-01

    Culture plays a large but often unnoticeable role in what we teach and how we teach children. We are a country of immense diversity, but in classrooms the dominant European-American culture has become the language of learning.

  6. Cultural Communication and Transmission

    Science.gov (United States)

    Tschumi, R.

    1973-01-01

    Part of a larger work, of which the French version, Theorie de la Culture'' (Theory of Culture), is to be published first; shorter version read at the International Federation for Modern Languages and Literatures Congress, Cambridge, England, 1972. (RS)

  7. Plant tissue culture techniques

    OpenAIRE

    Rolf Dieter Illg

    1991-01-01

    Plant cell and tissue culture in a simple fashion refers to techniques which utilize either single plant cells, groups of unorganized cells (callus) or organized tissues or organs put in culture, under controlled sterile conditions.

  8. Culture in the Classroom

    Science.gov (United States)

    Medin, Douglas L.; Bang, Megan

    2014-01-01

    Culture plays a large but often unnoticeable role in what we teach and how we teach children. We are a country of immense diversity, but in classrooms the dominant European-American culture has become the language of learning.

  9. Culture and social class.

    Science.gov (United States)

    Miyamoto, Yuri

    2017-08-08

    A large body of research in Western cultures has demonstrated the psychological and health effects of social class. This review outlines a cultural psychological approach to social stratification by comparing psychological and health manifestations of social class across Western and East Asian cultures. These comparisons suggest that cultural meaning systems shape how people make meaning and respond to material/structural conditions associated with social class, thereby leading to culturally divergent manifestations of social class. Specifically, unlike their counterparts in Western cultures, individuals of high social class in East Asian cultures tend to show high conformity and other-orientated psychological attributes. In addition, cultures differ in how social class impacts health (i.e. on which bases, through which pathways, and to what extent). Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Rectal culture (image)

    Science.gov (United States)

    A rectal culture test is performed by inserting a cotton swab in the rectum. The swab is rotated gently, and withdrawn. A smear of the swab is placed in culture media to encourage the growth of microorganisms. The ...

  11. Armenian Cultural Astronomy

    Science.gov (United States)

    Farmanyan, S. V.; Mickaelian, A. M.

    2015-07-01

    Cultural Astronomy is the reflection of sky events in various fields of nations' culture. In foreign literature this field is also called "Astronomy in Culture" or "Astronomy and Culture". Cultural astronomy is the set of interdisciplinary fields studying the astronomical systems of current or ancient societies and cultures. It is manifested in Religion, Mythology, Folklore, Poetry, Art, Linguistics and other fields. In recent years, considerable attention has been paid to this sphere, particularly international organizations were established, conferences are held and journals are published. Armenia is also rich in cultural astronomy. The present paper focuses on Armenian archaeoastronomy and cultural astronomy, including many creations related to astronomical knowledge; calendars, rock art, mythology, etc. On the other hand, this subject is rather poorly developed in Armenia; there are only individual studies on various related issues (especially many studies related to Anania Shirakatsi) but not coordinated actions to manage this important field of investigation.

  12. Native Culture Issues in Cross-cultural Teaching

    Institute of Scientific and Technical Information of China (English)

    万欣

    2012-01-01

      The bi-direction of cross-cultural communication determines culture teaching should include both target culture and native culture. Currently, however, mere emphasis of target culture with ignoring native culture has resulted in“two-skin”phenomenon and“aphasia of Chinese culture”. Therefore, this paper aims to underline native culture teaching, to explore proper techniques for native culture teaching, to achieve integration of target culture and native culture, to enhance students’expressive competence in native culture and finally to carry out effective cross-cultural communication.

  13. Construction of Hong-dae cultural district : cultural place, cultural policy and cultural politics

    OpenAIRE

    Cho, Mihye

    2007-01-01

    This dissertation examines how the process of creating the "Hong-dae cultural district" in Seoul has involved the mobilisation of various social groups and triggered the (re)institutionalisation of the meaning of "the cultural". It seeks to explicate how a cultural policy project can stimulate the emergence of social groups, which challenge existing policy provisions and laws and lead to the (re)institutionalisation of "Hong-dae culture". In so doing, the author will be able to simultaneously...

  14. The Politics of Culture

    Directory of Open Access Journals (Sweden)

    John Storey

    2017-01-01

    Full Text Available This article provides an overview over the evolution of thinking about "culture" in the work of Raymond Williams. With the introduction of Antonio Gramsci's concept of hegemony culture came to be understood as consisting of not only shared, but contested meanings as well. On the basis of this redefinition by Williams, cultural studies was able to delineate culture as the production, circulation, and consumption of meanings that become embodied and embedded in social practice.

  15. Euphemism and Social Culture

    Institute of Scientific and Technical Information of China (English)

    Yue Nan

    2008-01-01

    Euphemism,as a unique form in language expression,conveys a lot about the society and culture in which it exists.The study of euphemism is one method to understand the relation between language,society and culture.This paper analyzes the application of euphemism in social-cultural activities,studies the features of euphemisms in such application with the purpose to reveal connections between language and culture.

  16. Urine, faeces and culture

    DEFF Research Database (Denmark)

    Quitzau, M.

    This article looks upon the importance of considering cultural aspects in relation to toilet technologies. It is outlined how culture theoretically can be seen as an integrated part of every day actions and technology.......This article looks upon the importance of considering cultural aspects in relation to toilet technologies. It is outlined how culture theoretically can be seen as an integrated part of every day actions and technology....

  17. Research on audit culture

    Institute of Scientific and Technical Information of China (English)

    王仪

    2016-01-01

    It is the basis of promoting the scientific development of the audit business to strengthen the cultural construction of the audit staff, and also can improve the comprehensive quality of the audit staff. Aiming at the shortcomings of the current audit culture, this paper analyzes the reasons, and then puts forward some countermeasures for the construction of the audit culture, which is the goal of accelerating the construction of the audit culture and enhancing the strength of the audit team.

  18. Diamantina: World Cultural Heritage

    Directory of Open Access Journals (Sweden)

    Fernanda de Alencar Machado Albuquerque

    2012-11-01

    Full Text Available Awareness on preservation of cultural values in Brazil has been characterized as a current trend, and local communities play an important role in this process. The country’s preservationist policy has emerged with the Institute of National Historical and Artistic Heritage that aims at identifying and preserving the historical, cultural and artistic heritage. In the Brazilian scene the city of Diamantina/MG stands out for its remarkable cultural heritage, considered by UNESCO a World Cultural Heritage.

  19. Rupestrian culture in Italy

    OpenAIRE

    Crescenzi, C

    2012-01-01

    Rupestrian culture in Italy. L'articolo descrive sinteticamente le aree di studio, di alcune regioni italiane interessate dal fenomeno dell’architettura rupestre, che sono state oggetto dei workshop realizzati nell'ambito del progetto di ricerca internazionale Cultural Rupestrian Heritage in the Circum-Mediterraneam Area-cinp. Programme Culture 2007-2013, Budget 2010, Strand 1.1 Multi-annual cooperation project, Strand 1.2.1- Cooperation measures. estrian culture in Italy

  20. Cultural Identity Through CLIL

    Directory of Open Access Journals (Sweden)

    Oprescu Monica

    2015-11-01

    Full Text Available The CLIL approach is a modern manner of teaching English, which has been adapted in Romanian schools and universities. An interesting aspect of learning a foreign language is the contact with its culture/s and the changes it produces in terms of identity. Therefore, a challenging question to be answered is whether a CLIL approach focusing on culture influences students' cultural identity.

  1. Culture Differences and English Teaching

    Science.gov (United States)

    Wang, Jin

    2011-01-01

    Language is a part of culture, and plays a very important role in the development of the culture. Some sociologists consider it as the keystone of culture. They believe, without language, culture would not be available. At the same time, language is influenced and shaped by culture, it reflects culture. Therefore, culture plays a very important…

  2. Insect Cell Culture

    NARCIS (Netherlands)

    Oers, van M.M.; Lynn, D.E.

    2010-01-01

    Insect cell cultures are widely used in studies on insect cell physiology, developmental biology and microbial pathology. In particular, insect cell culture is an indispensable tool for the study of insect viruses. The first continuously growing insect cell cultures were established from lepidoptera

  3. Culture and Development

    NARCIS (Netherlands)

    D.R. Gasper (Des)

    2006-01-01

    textabstractDiscourses on culture and development vary according to their conceptions of culture and of development and according to their standpoint. The ‘culture and development’ problematic has typically: (1) arisen from a conception of ‘culture’ as a relatively fixed, homogeneous set of mental

  4. The University Culture

    Science.gov (United States)

    Simplicio, Joseph

    2012-01-01

    In this article the author discusses the role university culture can play on a campus and how it can impact policy and practice. The article explores how a university's history, values, and vision form its culture and how this culture in turn affects its stability and continuity. The article discusses how newcomers within the university are…

  5. Insect Cell Culture

    NARCIS (Netherlands)

    Oers, van M.M.; Lynn, D.E.

    2010-01-01

    Insect cell cultures are widely used in studies on insect cell physiology, developmental biology and microbial pathology. In particular, insect cell culture is an indispensable tool for the study of insect viruses. The first continuously growing insect cell cultures were established from lepidoptera

  6. Restoring Cultural Heritage Sites

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    Most of the post-quake cultural heritage rescue and protection projects in Sichuan have been completed The State Administration of Cultural Heritage recently rewarded 132 individuals and organizations for their work in rescuing and protecting cultural relics damaged by the Wenchuan earthquake on May12,2008.

  7. Culture and Language Teaching

    Institute of Scientific and Technical Information of China (English)

    李琳

    2008-01-01

    @@ Ⅰ.What Is Culture In 1871,in his classic book Primitive Culture,British anthropologist Edward Tylor first gave the definition of culture which is widely quoted: "Culture…is that complex whole which includes knowledge,beliefs,arts,morals,law,custom and any other capacities and habits acquired by man as a member of society".

  8. TESOL and Culture.

    Science.gov (United States)

    Atkinson, Dwight

    1999-01-01

    Looks at the question of how culture is understood in the Teaching English as a Second/Other-Language (TESOL) profession. Examines the perspectives toward culture implicitly or explicitly expressed in recent "TESOL Quarterly" articles, and concludes that different views of culture exist in the field. (Author/VWL)

  9. Cultural Industries Bloom

    Institute of Scientific and Technical Information of China (English)

    ZAN JIFANG

    2010-01-01

    @@ The market scale of China's cultural industries reached around 800 billion yuan($118 billion)in 2009,said a report on China's cultural industry development jointly released by a cultural research center under the Chinese Academy of Social Sciences and the academy's Social Sciences Academic Press on May 6.

  10. Why Teach Visual Culture?

    Science.gov (United States)

    Passmore, Kaye

    2007-01-01

    Visual culture is a hot topic in art education right now as some teachers are dedicated to teaching it and others are adamant that it has no place in a traditional art class. Visual culture, the author asserts, can include just about anything that is visually represented. Although people often think of visual culture as contemporary visuals such…

  11. Language, Perception, Culture & Communication

    Institute of Scientific and Technical Information of China (English)

    DU Man-li

    2015-01-01

    The paper explores the prospect of introducing language, perception, culture and communication. Starting with some definitions of language, perception, culture and communication, the paper argues for the internal connection among them. It pro⁃vides better understanding of these factors in foreign language learning and encourages learners to achieve the better learning re⁃sult to communicate effectively through language, culture etc.

  12. The Value of Culture

    NARCIS (Netherlands)

    Klamer, Arjo

    1997-01-01

    Culture manifests itself in everything human, including the ordinary business of everyday life. Culture and art have their own value, but economic values are also constrained. Art sponsorships and subsidies suggest a value that exceeds market price. So what is the real value of culture? Unlike the u

  13. Peritoneal fluid culture

    Science.gov (United States)

    Culture - peritoneal fluid ... sent to the laboratory for Gram stain and culture. The sample is checked to see if bacteria ... The peritoneal fluid culture may be negative, even if you have ... diagnosis of peritonitis is based on other factors, in addition ...

  14. Culture Difference and Translation

    Institute of Scientific and Technical Information of China (English)

    何冬兰

    2012-01-01

    Culture difference is necessary to be paid attention to during the process of translating.Culture difference is caused by different history,regions,customs,religions and the modes of thinking.Having the awareness of the culture difference will make translation more accurate and successful.

  15. Organizational culture, Anthropology of

    DEFF Research Database (Denmark)

    Krause-Jensen, Jakob; Wright, Susan

    2015-01-01

    cultures’ into transnational corporations and organizations concerned with international governance. In such organizations, anthropology graduates are increasingly employed as ‘cultural experts.’ We track the anthropological research on organizational culture and argue that the sensibilities and analytical...... skills acquired and the concepts developed through the ethnographic encounter gives anthropology a unique voice in the study of cultural matters in organizations....

  16. Cultur(ally) Jammed: Culture Jams as a Form of Culturally Responsive Teaching

    Science.gov (United States)

    Martinez, Ulyssa

    2012-01-01

    Does the person become the name or does the name become the person? This question was asked by a participant of my culture jam entitled, "What's my name?" In this culture jam, I asked people to discern the name of a person based solely on their appearance and a list of possible names below their picture. This article aims to show how culture jams…

  17. A Cultural Classroom Library

    Science.gov (United States)

    Lawrence, Maria

    2007-01-01

    Native American and other cultural stories provide students with a broader perspective on the world. In addition, cultural stories connect science content and knowledge about the world to cultural interpretations and people's life ways. By implementing the ideas suggested in this article, you can select books that both enrich your science library…

  18. Teaching Language, Teaching Culture.

    Science.gov (United States)

    Liddicoat, Anthony J., Ed.; Crozet, Chantal, Ed.

    1997-01-01

    Essays and research reports on the relationship between teaching second languages and teaching culture include: "Teaching Culture as an Integrated Part of Language Teaching: An Introduction" (Chantal Crozet, Anthony J. Liddicoat); "Primary Socialization and Cultural Factors in Second Language Learning: Wending Our Way through Semi-Charted…

  19. Managing culture in IJVs

    DEFF Research Database (Denmark)

    Dao, Li

    2012-01-01

    The purpose of this paper is to extend a cultural sense-making perspective to the context of international joint ventures. The dominant literature on cultural issues in this inter-firm setting has been criticized for relying on a narrow view of culture mainly as a country-level construct. The paper...

  20. Enriched cultures of lactic acid bacteria from selected Zimbabwean fermented food and medicinal products with potential as therapy or prophylaxis against yeast infections

    Directory of Open Access Journals (Sweden)

    Alec Chabwinja

    2017-10-01

    Full Text Available Objective: To investigate the antifungal activity of crude cultures of putative strains of lactic acid bacteria (LAB from a selection of Zimbabwean traditional and commercial food/ medicinal products against yeasts (strains of environmental isolates of Candida albicans and Rhodotorula spp.. Methods: Cultures of putative LAB from our selection of fermented products were enriched in de Man, Rogosa and Sharpe and isolated on de Man, Rogosa and Sharpe agar. Results: The crude microbial cultures from the products that showed high antifungal activities (zone of inhibition, mm were as follows: supernatant-free microbial pellet (SFMP from an extract of Melia azedarach leaves [(27.0 ± 2.5 mm] > cell-free culture supernatants (CFCS from Maaz Dairy sour milk and Mnandi sour milk [approximately (26.0 ± 1.8/2.5 mm] > CFCS and SFMP from Amansi hodzeko [(25.0 ± 1.5 mm] > CFCS from Parinari curatellifolia fruit [(24.0 ± 1.5 mm], SFMP from Parinari curatellifolia fruit [(24.0 ± 1.4 mm] and SFMP from mahewu [(20.0 ± 1.5 mm]. These cultures also showed high tolerance to acidic conditions (pH 4.0 and pH 5.0. However, culture from WAYA LGG (shown elsewhere to harbour antimicrobial activities showed no antifungal activity. The LAB could have inhibited yeasts by either competitive exclusion or the release of antimicrobial metabolites. Conclusions: Our cultures of LAB from a selection of Zimbabwean fermented products, especially Ziziphus mauritiana and fermented milk products have great potential for use as antifungal probiotics against yeast infections. Studies are ongoing to determine the exact mechanisms that are employed by the putative LAB to inhibit Candida albicans.

  1. Heterogeneous profiles of a factor that renders neutrophils cytotoxic obtained from a concanavalin A-stimulated spleen cell culture in partial purification process

    Energy Technology Data Exchange (ETDEWEB)

    Fujii, Y.; Inoue, T.; Ito, M.; Kimura, S.; Fuyama, S.; Arai, S.; Naiki, M.; Sendo, F.

    1986-05-15

    Concanavalin A (Con A)-stimulated rat spleen cells were cultured in a serum-free conditioned medium. This culture supernatant contained a certain factor(s) that renders neutrophil cytotoxic for various tumor cells. The factor was tentatively termed neutrophil-activating factor (NAF). NAF activity was eluted in broad fractions by the ion exchange chromatography and the gel filtration. Moreover, on the Con A column, some NAF activities were bound to the column, but other activities passed through the column. These results showed the heterogeneity or polydispersity of NAF activity in both molecular size and charge-based separation properties. Monoclonal antibodies were produced by fusing BALB/c myeloma cells (P3-X63 Ag8.653) with spleen cells from syngeneic mice immunized with partially purified NAF (pNAF) obtained from the gel filtration. Absorbent beads which were linked with one monoclonal antibody (ANAF-10) partially absorbed NAF activity from supernatants of a Con A-stimulated spleen cell culture. By further purification of pNAF the NAF activity was concentrated about 10,000-fold. Heterogeneity of NAF activity, however, did not disappear in even this affinity chromatography. On the other hand, /sup 125/I-labeled material of the final product migrated to one major band corresponding with an m.w. of about 20,000 as determined by SDS-PAGE analysis, and NAF activity was detected in the same band.

  2. Culture-sensitive psychotraumatology

    Directory of Open Access Journals (Sweden)

    Ulrich Schnyder

    2016-07-01

    Full Text Available Background: Although there is some evidence of the posttraumatic stress disorder (PTSD construct's cross cultural validity, trauma-related disorders may vary across cultures, and the same may be true for treatments that address such conditions. Experienced therapists tailor psychotherapy to each patient's particular situation, to the nature of the patient's psychopathology, to the stage of therapy, and so on. In addition, culture-sensitive psychotherapists try to understand how culture enhances the meaning of their patient's life history, the cultural components of their illness and help-seeking behaviors, as well as their expectations with regard to treatment. We cannot take for granted that all treatment-seeking trauma survivors speak our language or share our cultural values. Therefore, we need to increase our cultural competencies. Methods: The authors of this article are clinicians and/or researchers from across the globe, working with trauma survivors in various settings. Each author focused on one or more specific cultural aspects of working with trauma survivors and highlighted the following aspects. Results: As a result of culture-specific individual and collective meanings linked to trauma and trauma-related disorders survivors may be exposed to (self-stigma in the aftermath of trauma. Patients who are reluctant to talk about their traumatic experiences may instead be willing to write or use other ways of accessing the painful memories such as drawing. In other cultures, community and family cohesion are crucial elements of recovery. While awareness of culture-specific aspects is important, we also need to beware of premature cultural stereotyping. When disseminating empirically supported psychotherapies for PTSD across cultures, a number of additional challenges need to be taken into account: many low and middle income countries have very limited resources available and suffer from a poor health infrastructure. Conclusions: In summary

  3. Theories of Culture

    Institute of Scientific and Technical Information of China (English)

    王燕

    2012-01-01

      “Culture” is such a broad concept which is understood and defined differently by different people and has been remaining a focus for research. Some view culture as skills, values, understandings, knowledge or ways of being achieved as members of society and it is acquired and transmitted over generations; some regard culture as meaning which is established and constructed in practice and it is the context of production of new meaning and constraint of action. In this article it will focus on two theories of culture, namely, Cultural Relativism and Cultural Structuralism, and will illustrate the general ideas, main representatives and their arguments of these two theories.

  4. Culture shock and travelers.

    Science.gov (United States)

    Stewart, L; Leggat, P A

    1998-06-01

    As travel has become easier and more affordable, the number of people traveling has risen sharply. People travel for many and varied reasons, from the business person on an overseas assignment to backpackers seeking new and exotic destinations. Others may take up residence in different regions, states or countries for family, business or political reasons. Other people are fleeing religious or political persecution. Wherever they go and for whatever reason they go, people take their culture with them. Culture, like language, is acquired innately in early childhood and is then reinforced through formal and complex informal social education into adulthood. Culture provides a framework for interpersonal and social interactions. Therefore, the contact with a new culture is often not the exciting or pleasurable experience anticipated. When immersed in a different culture, people no longer know how to act when faced with disparate value systems. Contact with the unfamiliar culture can lead to anxiety, stress, mental illness and, in extreme cases, physical illness and suicide. "Culture shock" is a term coined by the anthropologist Oberg. It is the shock of the new. It implies that the experience of the new culture is an unpleasant surprise or shock, partly because it is unexpected and partly because it can lead to a negative evaluation of one's own culture. It is also known as cross-cultural adjustment, being that period of anxiety and confusion experienced when entering a new culture. It affects people intellectually, emotionally, behaviorally and physically and is characterized by symptoms of psychological distress. Culture shock affects both adults and children. In travelers or workers who have prolonged sojourns in foreign countries, culture shock may occur not only as they enter the new culture, but also may occur on their return to their original culture. Children may also experience readjustment problems after returning from leading sheltered lives in expatriate

  5. Cultural Capital in Context:

    DEFF Research Database (Denmark)

    Andersen, Ida Gran; Jæger, Mads Meier

    This paper analyzes the extent to which the effect of cultural capital on academic achievement varies across high- and low-achieving schooling environments. We distinguish three competing theoretical models: Cultural reproduction (cultural capital yields higher returns in high-achieving schooling...... to be higher in low-achieving schooling environments than in high-achieving ones. These results support the cultural mobility explanation and are in line with previous research suggesting that children from low-SES families benefit more from cultural capital than children from high-SES families....

  6. INCREASING LEARNERS’ CULTURAL AWARENESS

    Institute of Scientific and Technical Information of China (English)

    1996-01-01

    IntroductionCommunicative competence is now widely recognised as the goal of language teaching.A studentcannot obtain this competence in the target language without knowledge of the target culture sincelanguage and culture are closely interrelated.From this,it follows that EFL teaching involvesteaching of two languages and two cultures,in our case,English and Chinese.Then what is culture?Culture is‘the customs,beliefs,and music,and all the other products of human thought made by aparticular group of people at a particular time.’(Longman Dictionary of Contemporary English,1990:251)

  7. Leadership and Organizational Culture

    Institute of Scientific and Technical Information of China (English)

    宋丽娜

    2015-01-01

    This essay attempts to explore the relationship between leaders, organizational culture, and national culture. Leaders cre⁃ate“climate of the organization”with six mechanisms. Furthermore, leaders style of management is considerably influenced by their national culture based on Hofstede’s organizational culture theory. Varieties of examples and cases are analyzed to illustrate that leadership beliefs and practices have direct relationship with organizational culture and shape their individualistic communica⁃tion styles and goals that influence to a significant degree in establishing shared values, beliefs and practices among employees within an organization.

  8. Culture and Negotiation

    DEFF Research Database (Denmark)

    Bülow, Anne Marie; Kumar, Rajesh

    2011-01-01

    The literature on cross-cultural negotiation has expanded considerably over the past few decades, but the findings are often ambiguous and sometimes even contradictory. This introduction highlights the critical areas where objections are commonly raised about the relevance of national culture......, the applicability of typologies that treat cultures as static, and the problem of ambiguous terminology. It may not be surprising that studies contradict each other given the ambiguity of the national cultural construct and variations in the context of the negotiating situations that are studied. The articles...... in this issue contribute to deepening our understanding about cross-cultural negotiation processes....

  9. Field-Portable Immunoassay Instruments and Reagents to Measure Chelators and Mobile Forms of Uranium

    Energy Technology Data Exchange (ETDEWEB)

    Blake, Diane A.

    2006-01-23

    Progress Report Date: 01/23/06 (report delayed due to Hurricane Katrina) Report of results to date: The goals of this 3-year project are to: (1) update and successfully deploy our present immunosensors at DOE sites; (2) devise immunosensor-based assays for Pb(II), Hg(II), chelators, and/or Cr(III) in surface and groundwater; and (3) develop new technologies in antibody engineering that will enhance this immunosensor program. Note: Work on this project was temporarily disrupted when Hurricane Katrina shut down the University on August 29, 2005. While most of the reagents stored in our refrigerators and freezers were destroyed, all of our hybridoma cell lines were saved because they had been stored in liquid nitrogen. We set up new tissue culture reactors with the hybridomas that synthesize the anti-uranium antibodies, and are purifying new monoclonal antibodies from these culture supernatants. Both the in-line and the field-portable sensor were rescued from our labs in New Orleans in early October, and we continued experiments with these sensors in the temporary laboratory we set up in Hammond, LA at Southeastern Louisiana University.

  10. Sperm-immobilizing monoclonal antibody to human seminal plasma antigens.

    Science.gov (United States)

    Shigeta, M; Watanabe, T; Maruyama, S; Koyama, K; Isojima, S

    1980-01-01

    Rat spleen cells immunized to human azoospermic semen (a mixture of seminal plasma components) and mouse myeloma cells (P3/X63 Ag8U1; P3U1) (Marguilies et al., 1976) were successfully fused with polyethylene glycol (PEG 1500) and 19 of 89 fused cell cultures were found to produce sperm-immobilizing antibody. The cells that produced antibody indicating the highest sperm-immobilizing activity were distributed into wells for further recloning and 10 clones producing sperm-immobilizing antibody were established. The clone (1C4) producing the highest antibody titre was found to produce a large amount of IgG in culture supernatants and to contain a mixture of rat and mouse chromosomes. It was proved by immunodiffusion test that the monoclonal antibody was produced to the human seminal plasma antigen No. 7 which is common to human milk protein. Using this hybridoma which produced a large amount of monoclonal sperm-immobilizing antibody, a new method could be developed for purifying human seminal plasma antigen by immunoaffinity chromatography with bound antibody from the hybridoma. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:6783353

  11. Effect of operating conditions in production of diagnostic Salmonella Enteritidis O-antigen-specific monoclonal antibody in different bioreactor systems.

    Science.gov (United States)

    Ayyildiz-Tamis, Duygu; Nalbantsoy, Ayse; Elibol, Murat; Deliloglu-Gurhan, Saime Ismet

    2014-01-01

    In this study, different cultivation systems such as roller bottles (RB), 5-L stirred-tank bioreactor (STR), and disposable bioreactors were used to cultivate hybridoma for lab-scale production of Salmonella Enteritidis O-antigen-specific monoclonal antibody (MAb). Hybridoma cell line was cultivated in either serum-containing or serum-free medium (SFM) culture conditions. In STR, MAb production scaled up to 4 L, and production capabilities of the cells were also evaluated in different featured production systems. Moreover, the growth parameters of the cells in all production systems such as glucose consumption, lactate and ammonia production, and also MAb productivities were determined. Collected supernatants from the reactors were concentrated by a cross-flow filtration system. In conclusion, cells were not adapted to SFM in RB and STR. Therefore, less MAb titer in both STR and RB systems with SFM was observed compared to the cultures containing fetal bovine serum-supplemented medium. A higher MAb titer was gained in the membrane-aerated system compared to those in STR and RB. Although the highest MAb titer was obtained in the static membrane bioreactor system, the highest productivity was obtained in STR operated in semicontinuous mode with overlay aeration.

  12. Cultural aspects of suicide.

    Science.gov (United States)

    Maharajh, Hari D; Abdool, Petal S

    2005-09-08

    Undefined cultural factors cannot be dismissed and significantly contribute to the worldwide incidence of death by suicide. Culture is an all embracing term and defines the relationship of an individual to his environment. This study seeks to investigate the effect of culture on suicide both regionally and internationally. Culture-bound syndrome with suicidal behaviours specific to a particular culture or geographical region are discussed. Opinions are divided as to the status of religious martyrs. The law itself is silent on many aspects of suicidal behaviour and despite decriminalization of suicide as self-murder, the latter remains on the statutes of many developing countries. The Caribbean region is of concern due to its steady rise in mean suicide rate, especially in Trinidad and Tobago where socio-cultural factors are instrumental in influencing suicidal behaviour. These include transgenerational cultural conflicts, psycho-social problems, media exposure, unemployment, social distress, religion and family structure. The methods used are attributed to accessibility and lethality. Ingestion of poisonous substances is most popular followed by hanging. The gender differences seen with regard to suicidality can also be attributed to gender related psychopathology and psychosocial differences in help-seeking behaviour. These are influenced by the cultural environment to which the individual is exposed. Culture provides coping strategies to individuals; as civilization advances many of these coping mechanisms are lost unclothing the genetic predisposition of vulnerable groups. In the management of suicidal behaviour, a system of therapeutic re-culturation is needed with an emphasis on relevant culture- based therapies.

  13. Developing cultural sensitivity

    DEFF Research Database (Denmark)

    Ruddock, Heidi; Turner, deSalle

    2007-01-01

    . Background. Many countries are becoming culturally diverse, but healthcare systems and nursing education often remain mono-cultural and focused on the norms and needs of the majority culture. To meet the needs of all members of multicultural societies, nurses need to develop cultural sensitivity......Title. Developing cultural sensitivity: nursing students’ experiences of a study abroad programme Aim. This paper is a report of a study to explore whether having an international learning experience as part of a nursing education programme promoted cultural sensitivity in nursing students...... and incorporate this into caregiving. Method. A Gadamerian hermeneutic phenomenological approach was adopted. Data were collected in 2004 by using in-depth conversational interviews and analysed using the Turner method. Findings. Developing cultural sensitivity involves a complex interplay between becoming...

  14. Cultural effects on mindreading.

    Science.gov (United States)

    Perez-Zapata, Daniel; Slaughter, Virginia; Henry, Julie D

    2016-01-01

    People from other cultural backgrounds sometimes seem inscrutable. We identified a potential cause of this phenomenon in two experiments demonstrating that adults' mental state inferences are influenced by the cultural identity of the target. We adapted White, Hill, Happé, and Frith's (2009) Strange Stories to create matched intra-cultural and cross-cultural mindreading and control conditions. Experiment 1 showed that Australian participants were faster to respond and received higher scores in the intra-cultural mindreading condition relative to the cross-cultural mindreading condition, but performance in the control conditions was equivalent. Experiment 2 replicated this pattern in independent samples of Australian and Chilean participants. These findings have important implications for cross-cultural communication and understanding.

  15. Cultural Stress Revisited

    Institute of Scientific and Technical Information of China (English)

    Hastings; K; Shula; Aizhong; Liu

    2011-01-01

    Cultural stress is no longer a rare phenomenon because the world has been reduced to the size of a village due to modern technology and advancements. It is a concept that grows in magnitude each year. More and more people are affected. In this paper, we discuss the assessment of cultural stress by combining some instruments like the Perceived Stress Scale, the Depression Anxiety, and Stress Scale with the Cultural Stress Scale. They appear to be valid and can be used across different cultures. We discuss the need to come up with a standard instrument for measuring cultural stress as opposed to having so many. We also outline ways of coping with cultural stress as it occurs at different stages. There is need for more research to counter the negative effects of cultural stress.

  16. AccessCulture

    DEFF Research Database (Denmark)

    Valtysson, Bjarki

    in cultural production and consumption. The first part of this works looks at how these changes respond to the field of cultural policy, as well as suggesting a possible culturepolitical reaction in a model which I refer to as access culture. In terms of theoretical approach, the notion of digital cultural...... on the system, the lifeworld, and the inter-mediating public sphere, and in order to adapt his theory better to the network society, I make much use of Manuel Castells' theories on the global network of new media and the culture of realvirtuality. Finally, the third main theoretician which I make use of, is Lev...... and the Audiovisual Media Services Directive from 2007. In order to exemplify the functions of digital cultural public spheres adequately, I therefore take a thorough look at EU's interventions within the cultural, media and communication sectors. Finally, I also analyse projects and programmes that the European...

  17. Cultural Goods Production, Cultural Capital Formation and the Provision of Cultural Services

    OpenAIRE

    Cheng, Sao-Wen

    2005-01-01

    Cultural capital is assumed to benefit all members of society. It is accumulated through the consumption of cultural services and is diminished through depreciation. Using the stock of cultural goods, cultural services are provided by the cultural services industry; the stock of cultural goods is enlarged by the flow of new cultural goods created by individuals who are both consumers and creators of culture and whose utility is positively affected by the cultural goods they created. In the no...

  18. Characterization of chemical-induced sterile inflammation in vitro: application of the model compound ketoconazole in a human hepatic co-culture system.

    Science.gov (United States)

    Wewering, Franziska; Jouy, Florent; Wissenbach, Dirk K; Gebauer, Scarlett; Blüher, Matthias; Gebhardt, Rolf; Pirow, Ralph; von Bergen, Martin; Kalkhof, Stefan; Luch, Andreas; Zellmer, Sebastian

    2017-02-01

    Liver injury as a result of a sterile inflammation is closely linked to the activation of immune cells, including macrophages, by damaged hepatocytes. This interaction between immune cells and hepatocytes is as yet not considered in any of the in vitro test systems applied during the generation of new drugs. Here, we established and characterized a novel in vitro co-culture model with two human cell lines, HepG2 and differentiated THP-1. Ketoconazole, an antifungal drug known for its hepatotoxicity, was used as a model compound in the testing of the co-culture. Single cultures of HepG2 and THP-1 cells were studied as controls. Different metabolism patterns of ketoconazole were observed for the single and co-culture incubations as well as for the different cell types. The main metabolite N-deacetyl ketoconazole was found in cell pellets, but not in supernatants of cell cultures. Global proteome analysis showed that the NRF2-mediated stress response and the CXCL8 (IL-8) pathway were induced by ketoconazole treatment under co-culture conditions. The upregulation and ketoconazole-induced secretion of several pro-inflammatory cytokines, including CXCL8, TNF-α and CCL3, was observed in the co-culture system only, but not in single cell cultures. Taking together, we provide evidence that the co-culture model applied might be suitable to serve as tool for the prediction of chemical-induced sterile inflammation in liver tissue in vivo.

  19. Tick cell culture isolation and growth of Rickettsia raoultii from Dutch Dermacentor reticulatus ticks.

    Science.gov (United States)

    Alberdi, M Pilar; Nijhof, Ard M; Jongejan, Frans; Bell-Sakyi, Lesley

    2012-12-01

    Tick cell lines play an important role in research on ticks and tick-borne pathogenic and symbiotic microorganisms. In an attempt to derive continuous Dermacentor reticulatus cell lines, embryo-derived primary cell cultures were set up from eggs laid by field ticks originally collected as unfed adults in The Netherlands and maintained for up to 16 months. After several months, it became evident that cells in the primary cultures were infected with a Rickettsia-like intracellular organism. Supernatant medium containing some D. reticulatus cells was inoculated into cultures of 2 Rhipicephalus (Boophilus) microplus cell lines, BME/CTVM2 and BME/CTVM23, where abundant growth of the bacteria occurred intracellularly on transfer to both cell lines. Bacterial growth was monitored by light (live, inverted microscope, Giemsa-stained cytocentrifuge smears) and transmission electron microscopy revealing heavy infection with typical intracytoplasmic Rickettsia-like bacteria, not present in uninfected cultures. DNA was extracted from bacteria-infected and uninfected control cultures, and primers specific for Rickettsia 16S rRNA, ompB, and sca4 genes were used to generate PCR products that were subsequently sequenced. D. reticulatus primary cultures and both infected tick cell lines were positive for all 3 Rickettsia genes. Sequencing of PCR products revealed 99-100% identity with published Rickettsia raoultii sequences. The R. raoultii also grew abundantly in the D. nitens cell line ANE58, poorly in the D. albipictus cell line DALBE3, and not at all in the D. andersoni cell line DAE15. In conclusion, primary tick cell cultures and cell lines are useful systems for isolation and propagation of fastidious tick-borne microorganisms. In vitro isolation of R. raoultii from Dutch D. reticulatus confirms previous PCR-based detection in field ticks, and presence of the bacteria in the tick eggs used to initiate the primary cultures confirms that transovarial transmission of this

  20. Production of functional killer protein in batch cultures upon a shift from aerobic to anaerobic conditions

    Directory of Open Access Journals (Sweden)

    Gildo Almeida da Silva

    2011-06-01

    Full Text Available The aim of this work was to study the production of functional protein in yeast culture. The cells of Saccharomyces cerevisiae Embrapa 1B (K+R+ killed a strain of Saccharomyces cerevisiae Embrapa 26B (K-R-in grape must and YEPD media. The lethal effect of toxin-containing supernatant and the effect of aeration upon functional killer production and the correlation between the products of anaerobic metabolism and the functional toxin formation were evaluated. The results showed that at low sugar concentration, the toxin of the killer strain of Sacch. cerevisiae was only produced under anaerobic conditions . The system of killer protein production showed to be regulated by Pasteur and Crabtree effects. As soon as the ethanol was formed, the functional killer toxin was produced. The synthesis of the active killer toxin seemed to be somewhat associated with the switch to fermentation process and with concomitant alcohol dehydrogenase (ADH activity.