WorldWideScience

Sample records for cell culture techniques

  1. Aseptic technique for cell culture.

    Science.gov (United States)

    Coté, R J

    2001-05-01

    This unit describes some of the ways that a laboratory can deal with the constant threat of microbial contamination in cell cultures. A protocol on aseptic technique is described first. This catch-all term universally appears in any set of instructions pertaining to procedures in which noncontaminating conditions must be maintained. In reality, aseptic technique encompasses all aspects of environmental control, personal hygiene, equipment and media sterilization, and associated quality control procedures needed to ensure that a procedure is, indeed, performed with aseptic, noncontaminating technique. Although cell culture can theoretically be carried out on an open bench in a low-traffic area, most cell culture work is carried out using a horizontal laminar-flow clean bench or a vertical laminar-flow biosafety cabinet. Both are described here. PMID:18228291

  2. Techniques for mammalian cell tissue culture.

    Science.gov (United States)

    Phelan, Mary C

    2006-05-01

    This unit opens with detailed discussions on the latest principles of sterile technique and preparation of culture media. Step-by-step protocols describe trypsinizing and subculturing monolayer cultures, passaging suspension cultures, freezing and thawing cells, counting cells using a hemacytometer, and preparing cells for transport. PMID:18770828

  3. An Introductory Undergraduate Course Covering Animal Cell Culture Techniques

    Science.gov (United States)

    Mozdziak, Paul E.; Petitte, James N.; Carson, Susan D.

    2004-01-01

    Animal cell culture is a core laboratory technique in many molecular biology, developmental biology, and biotechnology laboratories. Cell culture is a relatively old technique that has been sparingly taught at the undergraduate level. The traditional methodology for acquiring cell culture training has been through trial and error, instruction when…

  4. 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.

  5. 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.

  6. A simple technique for preparation of chicken-embryo-skin cell cultures.

    Science.gov (United States)

    Silim, A; El Azhary, M A; Roy, R S

    1982-01-01

    A simple, rapid technique was developed for preparing chicken-embryo-skin cell cultures utilizing trypsinization of the skin of intact 12-day-old chicken embryos. When cell cultures were inoculated with fowl pox virus, those that consisted of at least 80% epithelial cells yielded a higher virus titer than fibroblast cell cultures. PMID:6284112

  7. Biotechnology Apprenticeship for Secondary-Level Students: Teaching Advanced Cell Culture Techniques for Research.

    Science.gov (United States)

    Lewis, Jennifer R.; Kotur, Mark S.; Butt, Omar; Kulcarni, Sumant; Riley, Alyssa A.; Ferrell, Nick; Sullivan, Kathryn D.; Ferrari, Mauro

    2002-01-01

    Discusses small-group apprenticeships (SGAs) as a method for introducing cell culture techniques to high school participants. Teaches cell culture practices and introduces advance imaging techniques to solve various biomedical engineering problems. Clarifies and illuminates the value of small-group laboratory apprenticeships. (Author/KHR)

  8. Biotechnology Apprenticeship for Secondary-Level Students: Teaching Advanced Cell Culture Techniques for Research

    OpenAIRE

    Lewis, Jennifer R.; Kotur, Mark S.; Butt, Omar; Kulcarni, Sumant; Riley, Alyssa A.; Ferrell, Nick; Sullivan, Kathryn D.; Ferrari, Mauro

    2002-01-01

    The purpose of this article is to discuss small-group apprenticeships (SGAs) as a method to instruct cell culture techniques to high school participants. The study aimed to teach cell culture practices and to introduce advanced imaging techniques to solve various biomedical engineering problems. Participants designed and completed experiments using both flow cytometry and laser scanning cytometry during the 1-month summer apprenticeship. In addition to effectively and efficiently teaching cel...

  9. Isolation, culturing and characterization of rat adipose tissue-derived mesenchymal stem cells: a simple technique

    OpenAIRE

    NİYAZ, Mehmet; Özer Aylin GÜRPINAR; GÜNAYDIN, Serdar; Onur, Mehmet Ali

    2012-01-01

    In this study, our aim was to develop a new simple technique for isolation of mesenchymal stem cells from adipose tissue. For this purpose, mesenchymal stem cells were isolated from rat adipose tissue by using the primary explant culture technique. When the cells became confluent, they were passaged 4 times by using the standard trypsinization method with trypsin/EDTA solution. Cells at second passage were characterized by using immunofluorescence staining against CD13 and CD29 markers. The r...

  10. Developments in techniques for the isolation, enrichment, main culture conditions and identification of spermatogonial stem cells.

    Science.gov (United States)

    He, Yanan; Chen, Xiaoli; Zhu, Huabin; Wang, Dong

    2015-12-01

    The in vitro culture system of spermatogonial stem cells (SSCs) provides a basis for studies on spermatogenesis, and also contributes to the development of new methods for the preservation of livestock and animal genetic modification. In vitro culture systems have mainly been established for mouse SSCs, but are lacking for farm animals. We reviewed and analyzed the current progress in SSC techniques such as isolation, purification, cultivation and identification. Based on the published studies, we concluded that two-step enzyme digestion and magnetic-activated cell sorting are fast becoming the main methods for isolation and enrichment of SSCs. With regard to the culture systems, serum and feeders were earlier thought to play an important role in the self-renewal and proliferation of SSCs, but serum- and feeder-free culture systems as a means of overcoming the limitations of SSC differentiation in long-term SSC culture are being explored. However, there is still a need to establish more efficient and ideal culture systems that can also be used for SSC culture in larger mammals. Although the lack of SSC-specific surface markers has seriously affected the efficiency of purification and identification, the transgenic study is helpful for our identification of SSCs. Therefore, future studies on SSC techniques should focus on improving serum- and feeder-free culture techniques, and discovering and identifying specific surface markers of SSCs, which will provide new ideas for the optimization of SSC culture systems for mice and promote related studies in farm animals. PMID:25749914

  11. Study on production of useful metabolites by development of advanced cell culture techniques using radiation

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Byung Yeoup; Lee, Seung Sik; Bai, Hyounwoo; Singh, Sudhir; Lee, Eun Mi; Hong, Sung Hyun; Park, Chul Hong; Srilatha, B.; Kim, Mi Ja; Lee, Ohchul

    2012-01-15

    The purpose of this project is improvement of investigation, materialization and evaluation techniques on effectiveness for functional natural compounds throughout development of tissue/cell culture techniques for mass production of useful metabolites using radiation. Research scope includes Development of a technique for radiation tissue and cell culture, Database construction for radiation response in plants and radiation effects, Construction of general-purpose national based techniques of cell culture technique using radiation. Main results are as follow: Development of a technique for radiation tissue and cell culture for Erigeron breviscapus (Vant.) Hand. Mazz.; Identification and functional analysis of AtTDX (chaperone and peroxidase activities); Functional analysis of radiation(gamma ray, electron beam, and proton beam) induced chaperon protein activities (AtTDX); Determine the action mechanism of yPrx2; Development of transgenic plant with bas I gene from Arabidopsis; Development of transgenic plant with EoP gene from centipedegrass; Identification of radiation induced multi functional compounds from Aloe; Determination of the effects of radiation on removing undesirable color and physiological activities (Schizandra chinensis baillon, centipedegrass); Determine the action mechanism of transgenic plant with 2-Cys Prx for heat stress resistance; Determination of the effects of centipedegrass extracts on anti-cancer activities; Functional analysis of centipedegrass extracts (anti-virus effects)

  12. Study on production of useful metabolites by development of advanced cell culture techniques using radiation

    International Nuclear Information System (INIS)

    The purpose of this project is improvement of investigation, materialization and evaluation techniques on effectiveness for functional natural compounds throughout development of tissue/cell culture techniques for mass production of useful metabolites using radiation. Research scope includes Development of a technique for radiation tissue and cell culture, Database construction for radiation response in plants and radiation effects, Construction of general-purpose national based techniques of cell culture technique using radiation. Main results are as follow: Development of a technique for radiation tissue and cell culture for Erigeron breviscapus (Vant.) Hand. Mazz.; Identification and functional analysis of AtTDX (chaperone and peroxidase activities); Functional analysis of radiation(gamma ray, electron beam, and proton beam) induced chaperon protein activities (AtTDX); Determine the action mechanism of yPrx2; Development of transgenic plant with bas I gene from Arabidopsis; Development of transgenic plant with EoP gene from centipedegrass; Identification of radiation induced multi functional compounds from Aloe; Determination of the effects of radiation on removing undesirable color and physiological activities (Schizandra chinensis baillon, centipedegrass); Determine the action mechanism of transgenic plant with 2-Cys Prx for heat stress resistance; Determination of the effects of centipedegrass extracts on anti-cancer activities; Functional analysis of centipedegrass extracts (anti-virus effects)

  13. Study on production of useful metabolites by development of advanced cell culture techniques using radiation

    International Nuclear Information System (INIS)

    The purpose of this project is improvement of investigation, materialization and evaluation techniques on effectiveness for functional natural compounds throughout development of tissue/cell culture techniques for mass production of useful metabolites using radiation. Research scope includes 1) Development of a technique for radiation tissue and cell culture, 2) Database construction for radiation response in plants and radiation effects, 3) Construction of general-purpose national based techniques of cell culture technique using radiation. Main results are as follow: mass culture of the adventitious roots of mountain ginseng (Panax ginseng C. A. Meyer) roots using rare earth elements in bioreactor: characterization of a transcription factor EoP gene from centipedegrass and the transcription regulation of LexA from Synechocystis sp PCC6803 and E. coli: identification of gamma-ray induced hydrogenase synthesis in hox gene transformed E. coli: transformation and the selection of the EoP transgene from Arabidopsis, rice and lettuce: Identification of the maysin and maysin derivatives in centipedegrass: characterization of gamma-ray induced color change in Taxus cuspidata: verification of the expression of antioxidant proteins (POD, APX and CAT) to gamma-ray in Arabidopsis: comparison of the response of the expression level to gamma-ray or H2O2 in Arabidopsis; verification of the responses and effects to gamma-ray from plants (analysis of NPQ and ROS levels): the development method for rapidly enhancing maysin content of centipede grass; establishment of mass culture system for red beet

  14. NucleoCounter—An efficient technique for the determination of cell number and viability in animal cell culture processes

    OpenAIRE

    Shah, Dimpalkumar; Naciri, Mariam; Clee, Paul; Al-Rubeai, Mohamed

    2006-01-01

    The NucleoCounter is a novel, portable cell counting device based on the principle of fluorescence microscopy. The present work establishes its use with animal cells and checks its reliability, consistency and accuracy in comparison with other cytometric techniques. The main advantages of this technique are its ability to handle a large number of samples with a high degree of precision and its simplicity and specificity in detecting viable cells quantitatively in a heterogeneous culture. The ...

  15. Study on production of useful metabolites by development of advanced cell culture techniques using radiation

    International Nuclear Information System (INIS)

    The purpose of this project is improvement of investigation, materialization and evaluation techniques on effectiveness for functional natural compounds throughout development of tissue/cell culture techniques for mass production of useful metabolites using radiation. Research scope includes Development of a technique for radiation tissue and cell culture, Database construction for radiation response in plants and radiation effects, Construction of general-purpose national based techniques of cell culture technique using radiation. Main results are as follow: Isolation and identification of radiation induced basI gene; Determination of stresses sensitivities by transformating basI gene into arabidopsis; Isolation and identification of radiation induced chaperon proteins (PaAhpC and yPrxII) from Pseudomonas and yeast, and structural and functional analysis of the proteins; Determination of oxidative and heat resistance by transformating PaAhpC; Isolation and identification of maysin and its derivatives from centipedgrass; Investigation of enhancement technique for improving maysin and its derivatives production using radiation; Investigation of removing undesirable color in maysin and its derivatives using radiation; Determination of the effect of radiation on physiological functions of centipedgrass extracts; Identification of H2O2 removing enzyme in radiation irradiated plant (Spinach); Determination of the effects of centipedgrass extracts on anti-obesity and anti-cancer activities

  16. Study on production of useful metabolites by development of advanced cell culture techniques using radiation

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Byung Yeoup; Kim, Jinhong; Lee, Seung Sik; Bai, Hyounwoo; An, Byung Chull; Lee, Eun Mi; Lee, Jae Taek; Kim, Mi Ja

    2010-12-15

    The purpose of this project is improvement of investigation, materialization and evaluation techniques on effectiveness for functional natural compounds throughout development of tissue/cell culture techniques for mass production of useful metabolites using radiation. Research scope includes Development of a technique for radiation tissue and cell culture, Database construction for radiation response in plants and radiation effects, Construction of general-purpose national based techniques of cell culture technique using radiation. Main results are as follow: Isolation and identification of radiation induced basI gene; Determination of stresses sensitivities by transformating basI gene into arabidopsis; Isolation and identification of radiation induced chaperon proteins (PaAhpC and yPrxII) from Pseudomonas and yeast, and structural and functional analysis of the proteins; Determination of oxidative and heat resistance by transformating PaAhpC; Isolation and identification of maysin and its derivatives from centipedgrass; Investigation of enhancement technique for improving maysin and its derivatives production using radiation; Investigation of removing undesirable color in maysin and its derivatives using radiation; Determination of the effect of radiation on physiological functions of centipedgrass extracts; Identification of H{sub 2}O{sub 2} removing enzyme in radiation irradiated plant (Spinach); Determination of the effects of centipedgrass extracts on anti-obesity and anti-cancer activities.

  17. A Rapid Culture Technique Produces Functional Dendritic-Like Cells from Human Acute Myeloid Leukemia Cell Lines

    Directory of Open Access Journals (Sweden)

    Jian Ning

    2011-01-01

    Full Text Available Most anti-cancer immunotherapeutic strategies involving dendritic cells (DC as vaccines rely upon the adoptive transfer of DC loaded with exogenous tumour-peptides. This study utilized human acute myeloid leukemia (AML cells as progenitors from which functional dendritic-like antigen presenting cells (DLC were generated, that constitutively express tumour antigens for recognition by CD8+ T cells. DLC were generated from AML cell lines KG-1 and MUTZ-3 using rapid culture techniques and appropriate cytokines. DLC were evaluated for their cell-surface phenotype, antigen uptake and ability to stimulate allogeneic responder cell proliferation, and production of IFN-γ; compared with DC derived from normal human PBMC donors. KG-1 and MUTZ-3 DLC increased expression of CD80, CD83, CD86, and HLA-DR, and MUTZ-3 DLC downregulated CD14 and expressed CD1a. Importantly, both KG-1 and MUTZ-3-derived DLC promoted proliferation of allogeneic responder cells more efficiently than unmodified cells; neither cells incorporated FITC-labeled dextran, but both stimulated IFN-γ production from responding allogeneic CD8+ T cells. Control DC produced from PBMC using the FastDC culture also expressed high levels of critical cell surface ligands and demonstrated good APC function. This paper indicates that functional DLC can be cultured from the AML cell lines KG-1 and MUTZ-3, and FastDC culture generates functional KG-1 DLC.

  18. Development of a cell culture surface conversion technique using alginate thin film for evaluating effect upon cellular differentiation

    International Nuclear Information System (INIS)

    Here, we sought to develop a cell culture surface conversion technique that would not damage living cells. An alginate thin film, formed on a glass plate by spin coating of sodium alginate solution and dipping into calcium chloride solution, was used to inhibit adhesion of cells. The film could be removed by ethylenediaminetetraacetate (EDTA) at any time during cell culture, permitting observation of cellular responses to conversion of the culture surface in real time. Additionally, we demonstrated the validity of the alginate thin film coating method and the performance of the film. The thickness of the alginate thin film was controlled by varying the rotation speed during spin coating. Moreover, the alginate thin film completely inhibited the adhesion of cultured cells to the culture surface, irrespective of the thickness of the film. When the alginate thin film was removed from the culture surface by EDTA, the cultured cells adhered to the culture surface, and their morphology changed. Finally, we achieved effective differentiation of C2C12 myoblasts into myotube cells by cell culture on the convertible culture surface, demonstrating the utility of our novel technique

  19. Biocompatibility of microplates for culturing epithelial renal cells evaluated by a microcalorimetric technique.

    Science.gov (United States)

    Xie, Y; DePierre, J W; Nässberger, L

    2000-09-01

    In the present study we have developed a microcalorimetric procedure which allows convenient investigation of biocompatibility in a microsystem. We examined the biocompatibility of a porcine renal epithelial tubule cell line LLC-PK1 and a human primary renal epithelial tubule cell (RPTEC) with microplates composed of three different materials, i.e. Thermanox, transparent film and titanium. All three materials showed equal biocompatibility with LLC-PK1 cells, judging from the attainment of steady-state power curves and the same rate of heat production per cell (2.5 microW / microg DNA). The human renal cells were poorly biocompatible with the Thermanox and transparent film. However, on titanium the RPTEC cell did adhere, as demonstrated by a steady-state power curve. The human cells also showed a higher metabolic activity (3.0 microW / microg DNA), than did LLC-PK1 cells cultured on the same type of microplates. In research on biocompatibility there is a need for alternatives to experimental animal investigations. The present technique allows studies of cellular interactions with different biomaterials in a rapid and standardized manner and may therefore prove to be a useful screening procedure. PMID:15348389

  20. Light and electron microscopic localization of GABAA-receptors on cultured cerebellar granule cells and astrocytes using immunohistochemical techniques

    DEFF Research Database (Denmark)

    Hansen, G H; Hösli, E; Belhage, B;

    1991-01-01

    GABAA-receptors were localized in explant cultures of rat cerebellum and in dissociated primary cultures of rat cerebellar granule cells and rat cerebellar astrocytes using the monoclonal antibody bd-17 directed against the beta-subunit of the GABAA/benzodiazepine/chloride channel complex. At the...... light microscope level specific staining of GABAA-receptors was localized in various types of neurones in explant cultures of rat cerebellum using the indirect peroxidase-antiperoxidase (PAP) technique, whereas no specific staining was found in astrocytes. At the electron microscope level labeling of...... in dissociated primary cultures of cerebellar astrocytes....

  1. NucleoCounter-An efficient technique for the determination of cell number and viability in animal cell culture processes.

    Science.gov (United States)

    Shah, Dimpalkumar; Naciri, Mariam; Clee, Paul; Al-Rubeai, Mohamed

    2006-05-01

    The NucleoCounter is a novel, portable cell counting device based on the principle of fluorescence microscopy. The present work establishes its use with animal cells and checks its reliability, consistency and accuracy in comparison with other cytometric techniques. The main advantages of this technique are its ability to handle a large number of samples with a high degree of precision and its simplicity and specificity in detecting viable cells quantitatively in a heterogeneous culture. The work addresses and overcomes the problems of subjectivity, and some of the inherent sampling errors associated with using the traditional haemocytometer and Trypan Blue exclusion method. NucleoCounter offers reduced intra- and inter-observer variation as well as consistency in repetitive analysis that establishes it as an efficient and highly potential device for at-line monitoring of animal cell processes. Furthermore, since the only manual steps required are sample aspiration and mixing with two reagents, it is feasible that the whole method could be automated and brought on-line for process monitoring and control. PMID:19002893

  2. OPTIMIZATION OF COMPOSITION CULTURE MEDIA, TECHNIQUE OF CONCENTRATING AND FREEZE DRYING CONDITIONS FOR CELL OF LACTOBACILLUS ACIDOPHILUS

    OpenAIRE

    Lysenko Y. A.; Luneva A. V.; Volkova S. A.; Nikolaenko S. N.; Petrova V. V.

    2014-01-01

    This study has investigated the relationship between culture media, technique of concentrating and freeze drying conditions for the cells of Lactobacillus aci-dophilus. The study has demonstrated that milk whey tomato juice-enriched medium has the high-growth properties. This medium allows ensuring high freeze drying survival of Lactobacillus acidophilus

  3. Characterization of cultured epithelial cells using a novel technique not requiring enzymatic digestion for subculturing.

    Science.gov (United States)

    Peramo, Antonio; Feinberg, Stephen E; Marcelo, Cynthia L

    2013-09-01

    Our laboratory had developed a methodology to expand epithelial cells in culture by growing keratinocyte monolayers, under large volumes of medium that produces large numbers of keratinocytes that leave the monolayer and move into suspension. The cells have been defined as epithelial Pop Up Keratinocytes or ePUKs cells and appear to be highly suitable for clinical applications. In this publication we extend the characterization of the cells with a detailed analysis of the capabilities of the monolayer of a single culture flask to produce, over time, ePUK cells. The cells were characterized using standard epithelial markers for proliferation and differentiation. Analysis of morphology of the monolayer formed and total number of cells produced is presented for a variety of human epithelial cell strains. These keratinocytes provide an additional controlled human cell system for investigation of the mechanisms regulating epithelia cell growth and differentiation and since they are produced in large numbers, they are highly suitable for use in epithelial cell banking. PMID:23149549

  4. Light and electron microscopic localization of GABAA-receptors on cultured cerebellar granule cells and astrocytes using immunohistochemical techniques

    DEFF Research Database (Denmark)

    Hansen, Gert Helge; Hösli, E; Belhage, B;

    1991-01-01

    light microscope level specific staining of GABAA-receptors was localized in various types of neurones in explant cultures of rat cerebellum using the indirect peroxidase-antiperoxidase (PAP) technique, whereas no specific staining was found in astrocytes. At the electron microscope level labeling of...... GABAA-receptors was observed in the plasma membrane of both the cell bodies and processes in dissociated primary cultures of cerebellar granule cells using an indirect preembedding immunogold staining technique which in contrast to the classical PAP technique allows quantitative estimations to be...... performed. Quantification of the labeling intensity revealed a higher concentration of GABAA-receptors per microns plasma membrane in the cell bodies than in the processes. In discrete areas an extremely high density of the GABAA-receptors was observed. No specific labeling of GABAA-receptors was observed...

  5. Mammalian Cell Culture Simplified.

    Science.gov (United States)

    Moss, Robert; Solomon, Sondra

    1991-01-01

    A tissue culture experiment that does not require elaborate equipment and that can be used to teach sterile technique, the principles of animal cell line maintenance, and the concept of cell growth curves is described. The differences between cancerous and normal cells can be highlighted. The procedure is included. (KR)

  6. Digital Microfluidic Cell Culture.

    Science.gov (United States)

    Ng, Alphonsus H C; Li, Bingyu Betty; Chamberlain, M Dean; Wheeler, Aaron R

    2015-01-01

    Digital microfluidics (DMF) is a droplet-based liquid-handling technology that has recently become popular for cell culture and analysis. In DMF, picoliter- to microliter-sized droplets are manipulated on a planar surface using electric fields, thus enabling software-reconfigurable operations on individual droplets, such as move, merge, split, and dispense from reservoirs. Using this technique, multistep cell-based processes can be carried out using simple and compact instrumentation, making DMF an attractive platform for eventual integration into routine biology workflows. In this review, we summarize the state-of-the-art in DMF cell culture, and describe design considerations, types of DMF cell culture, and cell-based applications of DMF. PMID:26643019

  7. Spatial and temporal single-cell volume estimation by a fluorescence imaging technique with application to astrocytes in primary culture

    Science.gov (United States)

    Khatibi, Siamak; Allansson, Louise; Gustavsson, Tomas; Blomstrand, Fredrik; Hansson, Elisabeth; Olsson, Torsten

    1999-05-01

    Cell volume changes are often associated with important physiological and pathological processes in the cell. These changes may be the means by which the cell interacts with its surrounding. Astroglial cells change their volume and shape under several circumstances that affect the central nervous system. Following an incidence of brain damage, such as a stroke or a traumatic brain injury, one of the first events seen is swelling of the astroglial cells. In order to study this and other similar phenomena, it is desirable to develop technical instrumentation and analysis methods capable of detecting and characterizing dynamic cell shape changes in a quantitative and robust way. We have developed a technique to monitor and to quantify the spatial and temporal volume changes in a single cell in primary culture. The technique is based on two- and three-dimensional fluorescence imaging. The temporal information is obtained from a sequence of microscope images, which are analyzed in real time. The spatial data is collected in a sequence of images from the microscope, which is automatically focused up and down through the specimen. The analysis of spatial data is performed off-line and consists of photobleaching compensation, focus restoration, filtering, segmentation and spatial volume estimation.

  8. Measurement of chromosomal breakage in cultured cells by the micronucleus technique

    International Nuclear Information System (INIS)

    The results of a series of experiments in which micronuclei were used to measure the extent of chromosomal damage by ionizing radiation are summarized. The data show that in most situations micronuclei accurately reflect chromosomal breakage and that they may be used for rapid and simple estimates of aberration frequency. The results of some studies on trisomy-21, Fanconi's anaemia, Bloom's syndrome and ataxia telangiectasia are included; the advantages and disadvantages of the technique are discussed. (author)

  9. Fish Stem Cell Cultures

    OpenAIRE

    Ni Hong, Zhendong Li, Yunhan Hong

    2011-01-01

    Stem cells have the potential for self-renewal and differentiation. First stem cell cultures were derived 30 years ago from early developing mouse embryos. These are pluripotent embryonic stem (ES) cells. Efforts towards ES cell derivation have been attempted in other mammalian and non-mammalian species. Work with stem cell culture in fish started 20 years ago. Laboratory fish species, in particular zebrafish and medaka, have been the focus of research towards stem cell cultures. Medaka is th...

  10. Expression of human immunodeficiency virus (HIV) in naturally infected peripheral blood mononuclear cells: comparison of a standard co-culture technique with a newly developed microculture method.

    Science.gov (United States)

    Eberlein, B; Baur, A; Neundorfer, M; Jahn, G

    1991-05-01

    Peripheral blood mononuclear cells (PBMCs) from 29 patients infected with human immunodeficiency virus (HIV) were cultured by two different methods. One was the standard co-culture technique, the other a newly developed microculture method. In this assay 10(6) PBMCs were cultivated in 250 microliters medium, no activating agents or allogeneic cells were present. P24 antigen production measured by this method was found in 7 out of 11 PBMC cultures of patients in the Walter Reed (WR) stage 1 or 2, whereas only 4 samples were positive by the co-culture procedure. Cultures from patients in the later stages of the disease (WR 5/6) showed a higher p24 production by the co-culture method than by the microculture assay. It is assumed that rapidly growing HIV strains can be better assessed by the co-culture method which may select for these strains. P24 expression can be more easily obtained by the microculture technique even in cases where slowly replicating strains may be present. In conclusion, results from the microculture procedure described may be a useful supplementation to findings observed by the co-culture method. PMID:1909827

  11. Optimizing stem cell culture.

    Science.gov (United States)

    van der Sanden, Boudewijn; Dhobb, Mehdi; Berger, François; Wion, Didier

    2010-11-01

    Stem cells always balance between self-renewal and differentiation. Hence, stem cell culture parameters are critical and need to be continuously refined according to progress in our stem cell biology understanding and the latest technological developments. In the past few years, major efforts have been made to define more precisely the medium composition in which stem cells grow or differentiate. This led to the progressive replacement of ill-defined additives such as serum or feeder cell layers by recombinant cytokines or growth factors. Another example is the control of the oxygen pressure. For many years cell cultures have been done under atmospheric oxygen pressure which is much higher than the one experienced by stem cells in vivo. A consequence of cell metabolism is that cell culture conditions are constantly changing. Therefore, the development of high sensitive monitoring processes and control algorithms is required for ensuring cell culture medium homeostasis. Stem cells also sense the physical constraints of their microenvironment. Rigidity, stiffness, and geometry of the culture substrate influence stem cell fate. Hence, nanotopography is probably as important as medium formulation in the optimization of stem cell culture conditions. Recent advances include the development of synthetic bioinformative substrates designed at the micro- and nanoscale level. On going research in many different fields including stem cell biology, nanotechnology, and bioengineering suggest that our current way to culture cells in Petri dish or flasks will soon be outdated as flying across the Atlantic Ocean in the Lindbergh's plane. PMID:20803548

  12. Cell Culture Made Easy.

    Science.gov (United States)

    Dye, Frank J.

    1985-01-01

    Outlines steps to generate cell samples for observation and experimentation. The procedures (which use ordinary laboratory equipment) will establish a short-term primary culture of normal mammalian cells. Information on culture vessels and cell division and a list of questions to generate student interest and involvement in the topics are…

  13. Neutron techniques for cultural heritage

    International Nuclear Information System (INIS)

    Neutron methods are emerging as an innovative and attractive investigative approach to characterising Cultural Heritage artefacts without the need for sampling or invasive procedures. Among Cultural Heritage collections conserved in national and private museums worldwide, metal artefacts of archaeological and historical interest, when properly studied, can reveal secrets of past human history. We are only allowed to treat them using the most delicate care and must avoid, as far as possible, accelerating any natural ageing process. Moreover, as analytical science evolves, we should not investigate these objects using invasive tools that might prevent further analytical techniques being applied to the same objects by future generations of scientists. In particular white beam and energy selective neutron radiography, tomography, and laminography can be successfully used to characterise the structure, morphology and composition of metal artworks by providing detailed three-dimensional information through the reconstruction of macroscopic cross sections of the object under investigation. From this wealth of data it is possible to obtain information on the conservation status of the artefact and reveal its manufacturing procedures, which in many cases may still be a mystery. In fact, the study of forging techniques, and their evolution over time, represents one of the most interesting topics in the investigation of metallic artefacts, which are considerably different within various cultures. In the present work, a comprehensive study of the metallurgy of metal artefacts of historical and archaeological interest in museums was carried out with the aim of investigating the composition, assembly methods, and structural variations in such artefacts from different cultures and different periods of history. The study has been developed in cooperation with several Museum Institutions and Neutron Facilities.

  14. Frequency domain analysis of membrane capacitance of cultured cells (HeLa and myeloma) using the micropipette technique.

    OpenAIRE

    Asami, K; Takahashi, Y.; Takashima, S.

    1990-01-01

    The membrane capacitance and conductance of cultured cells (HeLa and mouse myeloma) are investigated using the micropipette method. Mean values of the membrane capacities were found to be 1.9 microF/cm2 for HeLa cells and 1.0 microF/cm2 for myeloma cells. These values are in agreement with those obtained using the suspension method. Whereas the suspension method is unable to provide the information on membrane conductance, the micropipette method is able to measure even an extremely small mem...

  15. Techniques of the Translation of Culture

    OpenAIRE

    Xiao Geng

    2013-01-01

    This article discusses the basic principle of the translation of the culture and explores some techniques of the translation of culture. It also studies the impact of cultural differences on translation in several aspects. Language is a social behavior that reflects culture. Being ignorant of the target language culture, we would be a failure in translating, thus translating a langue into another foreign language necessarily involves culture integration. It is not enough to be aware of the im...

  16. Bacterial cell culture

    OpenAIRE

    sprotocols

    2014-01-01

    ### Materials 1. Glass culture tubes with metal caps and labels - Growth medium, from media room or customized - Glass pipette tubes - Parafilm ### Equipment 1. Vortexer - Fireboy or Bunsen burner - Motorized pipette - Micropipettes and sterile tips ### Procedure For a typical liquid culture, use 5 ml of appropriate medium. The amount in each tube does not have to be exact if you are just trying to culture cells for their precious DNA. 1. Streak an a...

  17. Fish Stem Cell Cultures

    Directory of Open Access Journals (Sweden)

    Ni Hong, Zhendong Li, Yunhan Hong

    2011-01-01

    Full Text Available Stem cells have the potential for self-renewal and differentiation. First stem cell cultures were derived 30 years ago from early developing mouse embryos. These are pluripotent embryonic stem (ES cells. Efforts towards ES cell derivation have been attempted in other mammalian and non-mammalian species. Work with stem cell culture in fish started 20 years ago. Laboratory fish species, in particular zebrafish and medaka, have been the focus of research towards stem cell cultures. Medaka is the second organism that generated ES cells and the first that gave rise to a spermatogonial stem cell line capable of test-tube sperm production. Most recently, the first haploid stem cells capable of producing whole animals have also been generated from medaka. ES-like cells have been reported also in zebrafish and several marine species. Attempts for germline transmission of ES cell cultures and gene targeting have been reported in zebrafish. Recent years have witnessed the progress in markers and procedures for ES cell characterization. These include the identification of fish homologs/paralogs of mammalian pluripotency genes and parameters for optimal chimera formation. In addition, fish germ cell cultures and transplantation have attracted considerable interest for germline transmission and surrogate production. Haploid ES cell nuclear transfer has proven in medaka the feasibility of semi-cloning as a novel assisted reproductive technology. In this special issue on “Fish Stem Cells and Nuclear Transfer”, we will focus our review on medaka to illustrate the current status and perspective of fish stem cells in research and application. We will also mention semi-cloning as a new development to conventional nuclear transfer.

  18. Optimizing stem cell culture.

    OpenAIRE

    van der Sanden, Boudewijn; Dhobb, Mehdi; Berger, François; Wion, Didier

    2010-01-01

    International audience Stem cells always balance between self-renewal and differentiation. Hence, stem cell culture parameters are critical and need to be continuously refined according to progress in our stem cell biology understanding and the latest technological developments. In the past few years, major efforts have been made to define more precisely the medium composition in which stem cells grow or differentiate. This led to the progressive replacement of ill-defined additives such a...

  19. Mycoplasmas detection in cells cultures

    OpenAIRE

    Rivera-Tapia José Antonio; Castillo-Viveros Linda Valeria; Sánchez-Hernández José Antonio

    2010-01-01

    INTRODUCTION. Cells cultures are widely used in both biomedical and biotechnological research centers and industry, as well as for diagnostic test in hospitals. Contaminations of cells cultures with microbial organisms as well as with virus or other eukaryotic cell lines are a major problem in cell culture related research.OBJECTIVE. Mycoplasmas detection in cells cultures came from biomedical laboratories.MATERIAL AND METHODS. The cells cultures screened for mycoplasmas by using of microbiol...

  20. Perfusion based cell culture chips

    DEFF Research Database (Denmark)

    Heiskanen, Arto; Emnéus, Jenny; Dufva, Martin

    2010-01-01

    Performing cell culture in miniaturized perfusion chambers gives possibilities to experiment with cells under near in vivo like conditions. In contrast to traditional batch cultures, miniaturized perfusion systems provide precise control of medium composition, long term unattended cultures and ti...

  1. Culture of Cells from Amphibian Embryos.

    Science.gov (United States)

    Stanisstreet, Martin

    1983-01-01

    Describes a method for in vitro culturing of cells from amphibian early embryos. Such cells can be used to demonstrate such properties of eukaryote cells as cell motility, adhesion, differentiation, and cell sorting into tissues. The technique may be extended to investigate other factors. (Author/JN)

  2. Oscillating Cell Culture Bioreactor

    Science.gov (United States)

    Freed, Lisa E.; Cheng, Mingyu; Moretti, Matteo G.

    2010-01-01

    To better exploit the principles of gas transport and mass transport during the processes of cell seeding of 3D scaffolds and in vitro culture of 3D tissue engineered constructs, the oscillatory cell culture bioreactor provides a flow of cell suspensions and culture media directly through a porous 3D scaffold (during cell seeding) and a 3D construct (during subsequent cultivation) within a highly gas-permeable closed-loop tube. This design is simple, modular, and flexible, and its component parts are easy to assemble and operate, and are inexpensive. Chamber volume can be very low, but can be easily scaled up. This innovation is well suited to work with different biological specimens, particularly with cells having high oxygen requirements and/or shear sensitivity, and different scaffold structures and dimensions. The closed-loop changer is highly gas permeable to allow efficient gas exchange during the cell seeding/culturing process. A porous scaffold, which may be seeded with cells, is fixed by means of a scaffold holder to the chamber wall with scaffold/construct orientation with respect to the chamber determined by the geometry of the scaffold holder. A fluid, with/without biological specimens, is added to the chamber such that all, or most, of the air is displaced (i.e., with or without an enclosed air bubble). Motion is applied to the chamber within a controlled environment (e.g., oscillatory motion within a humidified 37 C incubator). Movement of the chamber induces relative motion of the scaffold/construct with respect to the fluid. In case the fluid is a cell suspension, cells will come into contact with the scaffold and eventually adhere to it. Alternatively, cells can be seeded on scaffolds by gel entrapment prior to bioreactor cultivation. Subsequently, the oscillatory cell culture bioreactor will provide efficient gas exchange (i.e., of oxygen and carbon dioxide, as required for viability of metabolically active cells) and controlled levels of fluid

  3. The potential of metabolomic analysis techniques for the characterisation of α1-adrenergic receptors in cultured N1E-115 mouse neuroblastoma cells.

    Science.gov (United States)

    Wenner, Maria I; Maker, Garth L; Dawson, Linda F; Drummond, Peter D; Mullaney, Ian

    2016-08-01

    Several studies of neuropathic pain have linked abnormal adrenergic signalling to the development and maintenance of pain, although the mechanisms underlying this are not yet fully understood. Metabolomic analysis is a technique that can be used to give a snapshot of biochemical status, and can aid in the identification of the mechanisms behind pathological changes identified in cells, tissues and biological fluids. This study aimed to use gas chromatography-mass spectrometry-based metabolomic profiling in combination with reverse transcriptase-polymerase chain reaction and immunocytochemistry to identify functional α1-adrenergic receptors on cultured N1E-115 mouse neuroblastoma cells. The study was able to confirm the presence of mRNA for the α1D subtype, as well as protein expression of the α1-adrenergic receptor. Furthermore, metabolomic data revealed changes to the metabolite profile of cells when exposed to adrenergic pharmacological intervention. Agonist treatment with phenylephrine hydrochloride (10 µM) resulted in altered levels of several metabolites including myo-inositol, glucose, fructose, alanine, leucine, phenylalanine, valine, and n-acetylglutamic acid. Many of the changes observed in N1E-115 cells by agonist treatment were modulated by additional antagonist treatment (prazosin hydrochloride, 100 µM). A number of these changes reflected what is known about the biochemistry of α1-adrenergic receptor activation. This preliminary study therefore demonstrates the potential of metabolomic profiling to confirm the presence of functional receptors on cultured cells. PMID:26408527

  4. Microfluidic Cell Culture Device

    Science.gov (United States)

    Takayama, Shuichi (Inventor); Cabrera, Lourdes Marcella (Inventor); Heo, Yun Seok (Inventor); Smith, Gary Daniel (Inventor)

    2014-01-01

    Microfluidic devices for cell culturing and methods for using the same are disclosed. One device includes a substrate and membrane. The substrate includes a reservoir in fluid communication with a passage. A bio-compatible fluid may be added to the reservoir and passage. The reservoir is configured to receive and retain at least a portion of a cell mass. The membrane acts as a barrier to evaporation of the bio-compatible fluid from the passage. A cover fluid may be added to cover the bio-compatible fluid to prevent evaporation of the bio-compatible fluid.

  5. In vitro culture of human thyroid cells

    International Nuclear Information System (INIS)

    Procedures for establishing primary cultures of human thyroid tissue are described. Tissues removed surgically from patients with papillary carcinoma (PC), follicular adenoma (FA), or hyperthyroidism were grown in culture. In addition, normal cells were separated from the margins of excised tumors and were also cultured. For each gram of thyroid tissue cultured, more than 1 x 105 cells attached to culture dishes. A mixture of 2.5 % fetal bovine serum supplemented with insulin, hydrocortisone, transferrin, glycl-1-histidyl-L-lysine acetate, somatostatin and epidermal growth factor was added to nutrient media containing equal parts of Ham's F-12 and minimum essential medium (αMEM). Complete medium selectively supported epithelial cell growth while restricting fibroblast cell growth, especially during the first two weeks of the primary culture. Cells were stimulated with thyroid stimulating hormone (TSH) and produced raised levels of cAMP and thyroid hormone (T3). Culture conditions that affected the response of cells to X-rays were identified. During the culture period, first and second passage cells were compared for differences in their radiosensitivities. In all cases, cells showed differences in their responses to radiation depending on the cell passage number. However, results of replicate experiments of first passage cells that were exposed to X-rays showed good agreement between experiments. This technique makes it possible to quantitate the effects of chemical and physical cytotoxic agents on proliferating human thyroid epithelial cells. (author)

  6. Methods for Maintaining Insect Cell Cultures

    OpenAIRE

    Dwight E. Lynn

    2002-01-01

    Insect cell cultures are now commonly used in insect physiology, developmental biology, pathology, and molecular biology. As the field has advanced from methods development to a standard procedure, so has the diversity of scientists using the technique. This paper describes methods that are effective for maintaining various insect cell lines. The procedures are differentiated between loosely or non-attached cell strains, attached cell strains, and strongly adherent cell strains.

  7. Techniques for quantifying effects of dietary antioxidants on transcription factor translocation and nitric oxide production in cultured cells

    OpenAIRE

    Ewins, B. A.; Vassiliadou, M.; Minihane, A. M.; Rimbach, G. H.; Weinberg, P.D.

    2006-01-01

    Dietary antioxidants can affect cellular processes relevant to chronic inflammatory diseases such as atherosclerosis. We have used non-standard techniques to quantify effects of the antioxidant soy isoflavones genistein and daidzein on translocation of Nuclear Factor-KB (NF-KB) and nitric oxide (NO) production, which are important in these diseases. Translocation was quantified using confocal immunofluoresecence microscopy and ratiometric image analysis. NO was quantified by an electrochemica...

  8. Using Tissue Culture To Investigate Plant Cell Differentiation and Dedifferentiation.

    Science.gov (United States)

    Bozzone, Donna M.

    1997-01-01

    Describes an experimental project that uses plant tissue culture techniques to examine cell differentiation in the carrot. Allows students to gain experience in some important techniques and to explore fundamental questions about cell differentiation. (DDR)

  9. THE METHODS FOR MAINTAINING INSECT CELL CULTURES

    Science.gov (United States)

    Insect cell cultures are now commonly used in insect physiology, developmental biology, pathology, and molecular biology. As the field has advanced from a methods development to a standard procedure, so has the diversity of scientists using the technique. This paper describes techniques that are e...

  10. Glucocorticoid regulation in rat brain cell cultures. Hydrocortisone increases the rate of synthesis of glycerol phosphate dehydrogenase in C6 glioma cells. [Tritium tracer technique

    Energy Technology Data Exchange (ETDEWEB)

    McGinnis, J.F.; de Vellis, J.

    1978-12-10

    Cytoplasmic glycerol phosphate dehydrogenase (sn-glycerol-3-phosphate: NAD/sup +/ 2-oxidoreductase, EC 1.1.1.8) was rapidly purified from rat skeletal muscle in high yield using a combination of classical and affinity techniques. A single band of protein having a molecular weight of 30,000 was found using dodecyl sulfate-polyacrylamide gel electrophoresis. Antisera were generated in rabbits against the purified enzyme and demonstrated to be monospecific by Ouchterlony immunodiffusion against crude homogenates from hydrocortisone-induced and uninduced C6 cells. All of the radioactivity in immunoprecipitates from (/sup 3/H)leucine-labeled cells co-migrated with purified glycerol phosphate dehydrogenase. The amount of radioactivity precipitated was directly proportional to the amount of labeled glycerol phosphate dehydrogenase present, indicating that the assay could be used to quantitate newly synthesized glycerol phosphate dehydrogenase molecules. Using these techniques, the induction of glycerol phosphate dehydrogenase activity by hydrocortisone in the C6 glioma cell line was shown to be due to an increase in the rate of synthesis of the enzyme. Analysis of the kinetics of induction and deinduction supports the above conclusion and suggests that there is essentially no change in the rate of degradation of glycerol phosphate dehydrogenase in the presence and absence of hormone.

  11. Nuclear Techniques for Cultural Heritage Research

    International Nuclear Information System (INIS)

    Cultural heritage ('national heritage' or just 'heritage') is the legacy of physical artefacts and intangible attributes of a group or society that are inherited from past generations, maintained in the present and restored for the benefit of future generations. Physical or 'tangible cultural heritage' includes buildings and historical places, monuments, artefacts, etc., that are considered worthy of preservation for the future. These include preservation and conservation of objects significant to the archaeology, architecture, science or technology of a specific culture. Scientific studies of art and archaeology present a necessary complement for cultural heritage conservation, preservation and investigation. As cultural heritage objects are frequently unique and non-replaceable, non-destructive techniques are mandatory and, hence, nuclear techniques have a high potential to be applied to study these valuable objects. Nuclear techniques, such as neutron activation analysis (NAA), X ray fluorescence (XRF) analysis or ion beam analysis (IBA), have a potential for non-destructive and reliable investigation of precious materials, such as ceramics, stone, metal or pigments from paintings. Such information can help to repair damaged objects adequately, distinguish fraudulent artefacts from real artefacts and assist archaeologists in the appropriate categorization of historical artefacts. Although the application of scientific methods to art and archaeological materials has a long tradition, it is due to the stimulation of institutions such as the United Nations Educational, Scientific and Cultural Organization (UNESCO), the United Nations Environment Programme (UNEP) and the IAEA that applications of natural science techniques are increasingly being accepted by museum curators and cultural heritage researchers. The IAEA as a leading supporter of the peaceful use of nuclear technology assists laboratories in its Member States to apply and develop nuclear methods for

  12. 3D Cell Culture in Alginate Hydrogels

    Directory of Open Access Journals (Sweden)

    Therese Andersen

    2015-03-01

    Full Text Available This review compiles information regarding the use of alginate, and in particular alginate hydrogels, in culturing cells in 3D. Knowledge of alginate chemical structure and functionality are shown to be important parameters in design of alginate-based matrices for cell culture. Gel elasticity as well as hydrogel stability can be impacted by the type of alginate used, its concentration, the choice of gelation technique (ionic or covalent, and divalent cation chosen as the gel inducing ion. The use of peptide-coupled alginate can control cell–matrix interactions. Gelation of alginate with concomitant immobilization of cells can take various forms. Droplets or beads have been utilized since the 1980s for immobilizing cells. Newer matrices such as macroporous scaffolds are now entering the 3D cell culture product market. Finally, delayed gelling, injectable, alginate systems show utility in the translation of in vitro cell culture to in vivo tissue engineering applications. Alginate has a history and a future in 3D cell culture. Historically, cells were encapsulated in alginate droplets cross-linked with calcium for the development of artificial organs. Now, several commercial products based on alginate are being used as 3D cell culture systems that also demonstrate the possibility of replacing or regenerating tissue.

  13. Huanglongbing and psyllid cell cultures

    Science.gov (United States)

    We successfully established cell cultures of the Asian citrus psyllid, Diaphorina citri (Psyllidae: Hemiptera), DcHH-1. The cell culture also supported growth of Candidatus Liberibacter asiaticus. This bacterial pathogen is associated with Huanglongbing, known as citrus greening disease. Research on...

  14. Prevention and Detection of Mycoplasma Contamination in Cell Culture

    Directory of Open Access Journals (Sweden)

    Parvaneh Farzaneh

    2012-01-01

    Full Text Available One of the main problems in cell culture is mycoplasma infection. It can extensively affectcell physiology and metabolism. As the applications of cell culture increase in research,industrial production and cell therapy, more concerns about mycoplasma contaminationand detection will arise. This review will provide valuable information about: 1. the waysin which cells are contaminated and the frequency and source of mycoplasma species incell culture; 2. the ways to prevent mycoplasma contamination in cell culture; 3. the importanceof mycoplasma tests in cell culture; 4. different methods to identify mycoplasmacontamination; 5. the consequences of mycoplasma contamination in cell culture and 6.available methods to eliminate mycoplasma contamination. Awareness about the sourcesof mycoplasma and pursuing aseptic techniques in cell culture along with reliable detectionmethods of mycoplasma contamination can provide an appropriate situation to preventmycoplasma contamination in cell culture.

  15. High density cell culture system

    Science.gov (United States)

    Spaulding, Glenn F. (Inventor)

    1994-01-01

    An annular culture vessel for growing mammalian cells is constructed in a one piece integral and annular configuration with an open end which is closed by an endcap. The culture vessel is rotatable about a horizontal axis by use of conventional roller systems commonly used in culture laboratories. The end wall of the endcap has tapered access ports to frictionally and sealingly receive the ends of hypodermic syringes. The syringes permit the introduction of fresh nutrient and withdrawal of spent nutrients. The walls are made of conventional polymeric cell culture material and are subjected to neutron bombardment to form minute gas permeable perforations in the walls.

  16. Value of two mortality assessment techniques for organ cultured corneal endothelium: trypan blue versus TUNEL technique

    OpenAIRE

    Gain, P; Thuret, G; Chiquet, C.; Dumollard, J.M.; Mosnier, J F; BURILLON, C; Delbosc, B; Hervé, P; Campos, L.

    2002-01-01

    Background/aim: It is known that trypan blue staining is not a good predictor of loss of corneal endothelial cells (ECs) during organ culture. As it is primarily an indicator of membrane integrity, it would also not be expected to identify ECs undergoing apoptosis. The aim of this study was to determine the ability of the in situ TdT dUTP mediated nick end labelling (TUNEL) technique to detect cell death in the corneal endothelium caused by apoptosis during organ culture, compared with conven...

  17. Cell culture purity issues and DFAT cells

    International Nuclear Information System (INIS)

    Highlights: •DFAT cells are progeny cells derived from dedifferentiated mature adipocytes. •Common problems in this research is potential cell contamination of initial cultures. •The initial cell culture purity is crucial in DFAT cell research field. -- Abstract: Dedifferentiation of mature adipocytes, in vitro, has been pursued/documented for over forty years. The subsequent progeny cells are named dedifferentiated adipocyte-derived progeny cells (DFAT cells). DFAT cells are proliferative and likely to possess mutilineage potential. As a consequence, DFAT cells and their progeny/daughter cells may be useful as a potential tool for various aspects of tissue engineering and as potential vectors for the alleviation of several disease states. Publications in this area have been increasing annually, but the purity of the initial culture of mature adipocytes has seldom been documented. Consequently, it is not always clear whether DFAT cells are derived from dedifferentiated mature (lipid filled) adipocytes or from contaminating cells that reside in an impure culture

  18. [CO-CULTURE OF BOAR SPERMATOGONIAL CELLS WITH SERTOLI CELLS].

    Science.gov (United States)

    Savchenkova, I P; Vasil'eva, S A

    2016-01-01

    In the present study, we developed in vitro culture conditions using co-culture of boar spermatogonial cells with Sertoli cells. Testes from 60-day-old crossbred boar were used. A spermatogonia-enriched culture was achieved by enzymatic digestion method and purification by density gradient centrifugation using a discontinuous Percoll gradient and differentiated adherence technique. Lipid drops were detected in isolated Sertoli cells by Oil Red O staining. We have found that the cultivation of boar spermatogonia in the presence of Sertoli cells (up to 35 days) leads to their differentiation as well as in vivo in testis. Association of cells in groups, formation of chains and suspension clusters of the spermatogenic cells were observed on the 10th day. Spermatogonial cellular colonies were noted at the same time. These cellular colonies were analyzed for the expression of genes: Nanog and Plzf in RT PCR. The expression of the Nanog gene in the experimental cellular clones obtained by short-term culture of spermatogonial cells in the presence of Sertoli cells was 200 times higher than the expression of this gene in the freshly isolated spermatogonial cells expression was found in freshly isolated germ cells and in cellular clones derived in vitro. We have found that, in the case of longer cultivation of these cells on Sertoli cells, in vitro process of differentiation of germ cells and formation of single mobile boar spermatozoa occurs at 30-33 days. Cellular population is heterogeneous at this stage. Spermatogenic differentiation in vitro without Sertoli cells stays on the 7th day of cultivation. The results show that co-culture of boar spermatogonia-enriched cells with Sertoli cells can induce their differentiation into spermatozoa in vitro and facilitate obtaining of porcine germ cell culture. PMID:27228660

  19. Nuclear Techniques for Cultural Heritage Research

    International Nuclear Information System (INIS)

    Historical artefacts and art objects are traded world wide and represent a potential source for forgery and false labelling. The market is huge and a large part of the trade is going from developing countries to the developed world where strict regulations can only be applied if convenient methods for checking the authenticity would be available. Portable XRF instruments for rapid screening analysis are already available and have been demonstrated to be useful in analysis of paintings and small artefacts. Laboratory based techniques such as PIXE, INAA and prompt gamma NAA have been applied non-destructively to investigate the provenience of archaeological objects as well as neutron radiography was used to determine different layers of precious paintings. Scientific studies of art and archaeology presents a necessary compliment for cultural heritage conservation, preservation and investigation. Museum curators, however, strongly oppose to sacrifice even small portions of a valuable art object for analysis. Nuclear techniques, such as neutron activation analysis (NAA), x-ray fluorescence analysis (XRF) or ion beam analysis (IBA) have a potential for non-destructive and reliable investigation of precious materials, such as ceramics, stone, metal or pigments from paintings. Such information can help to repair damaged objects adequately, identify fraud from real and assist archaeologists in the appropriate categorisation of historic artefacts. During the Coordinated Research Project 'Applications of Nuclear Analytical Techniques to Investigate the Authenticity of Art Objects' (2004 - 2008) the Agency was supporting Member States (MS) in their effort to apply nuclear techniques for their cultural heritage investigations, e.g. pottery shards from an ancient slave camp in Ghana were analyzed using Compton suppressed NAA. From the results it became clear that the pots were produced locally and were not imported from other places where the slaves were deported from. In China

  20. Cell Phone Detection Techniques

    Energy Technology Data Exchange (ETDEWEB)

    Pratt, Richard M.; Bunch, Kyle J.; Puzycki, David J.; Slaugh, Ryan W.; Good, Morris S.; McMakin, Douglas L.

    2007-10-01

    A team composed of Rick Pratt, Dave Puczyki, Kyle Bunch, Ryan Slaugh, Morris Good, and Doug McMakin teamed together to attempt to exploit cellular telephone features and detect if a person was carrying a cellular telephone into a Limited Area. The cell phone’s electromagnetic properties were measured, analyzed, and tested in over 10 different ways to determine if an exploitable signature exists. The method that appears to have the most potential for success without adding an external tag is to measure the RF spectrum, not in the cell phone band, but between 240 and 400MHz. Figures 1- 7 show the detected signal levels from cell phones from three different manufacturers.

  1. Cell culture compositions

    Science.gov (United States)

    Dunn-Coleman, Nigel; Goedegebuur, Frits; Ward, Michael; Yiao, Jian

    2014-03-18

    The present invention provides a novel endoglucanase nucleic acid sequence, designated egl6 (SEQ ID NO:1 encodes the full length endoglucanase; SEQ ID NO:4 encodes the mature form), and the corresponding endoglucanase VI amino acid sequence ("EGVI"; SEQ ID NO:3 is the signal sequence; SEQ ID NO:2 is the mature sequence). The invention also provides expression vectors and host cells comprising a nucleic acid sequence encoding EGVI, recombinant EGVI proteins and methods for producing the same.

  2. HAIR CELL-LIKE CELL GENERATION INDUCED BY NATURE CULTURE OF ADULT RAT AUDITORY EPITHELIUM

    Institute of Scientific and Technical Information of China (English)

    Liu Hui; Zhu Hongliang; Li Shengli; Yao Xiaobao; Wang Xiaoxia

    2006-01-01

    Objective To establish adult rat auditory epithelial cell culture and try to find precursor cells of auditory hair cells in vitro. Methods With refinement of culture media and techniques, cochlear sensory epithelial cells of adult rat were cultured. Immunocytochemistry and Bromodeoxyuridine (BrdU)labeling were used to detect properties and mitotic status of cultured cells. Results The cultured auditory epithelial cells showed a large, flat epithelial morphotype and expressed F-actin and cytokeratin, a subset of cells generated from auditory epithelium were labeled by calretinin, a specific marker of early hair cell. Conclusion Adult rat auditory epithelium can be induced to generate hair cell-like cells by nature culture, this phenomenon suggests that progenitor cells may exist in rat cochlea and they may give birth to new hair cells. Whether these progenitor cells are tissue specific stem cells is still need more study.

  3. Dynamized Preparations in Cell Culture

    OpenAIRE

    Girija Kuttan; Korengath Chandran Preethi; Ramadasan Kuttan; Ellanzhiyil Surendran Sunila

    2009-01-01

    Although reports on the efficacy of homeopathic medicines in animal models are limited, there are even fewer reports on the in vitro action of these dynamized preparations. We have evaluated the cytotoxic activity of 30C and 200C potencies of ten dynamized medicines against Dalton's Lymphoma Ascites, Ehrlich's Ascites Carcinoma, lung fibroblast (L929) and Chinese Hamster Ovary (CHO) cell lines and compared activity with their mother tinctures during short-term and long-term cell culture. The ...

  4. Insect Cell Culture and Biotechnology

    Institute of Scientific and Technical Information of China (English)

    Robert R.Granados; Guoxun Li; G.W.Blissard

    2007-01-01

    The continued development of new cell culture technology is essential for the future growth and application of insect cell and baculovirus biotechnology. The use of cell lines for academic research and for commercial applications is currently dominated by two cell lines; the Spodoptera frugiperda line, SF21 (and its clonal isolate, SF9), and the Trichoplusia ni line, BTI 5B1-4, commercially known as High Five cells. The long perceived prediction that the immense potential application of the baculovirus-insect cell system, as a tool in cell and molecular biology, agriculture, and animal health, has been achieved. The versatility and recent applications of this popular expression system has been demonstrated by both academia and industry and it is clear that this cell-based system has been widely accepted for biotechnological applications. Numerous small to midsize startup biotechnology companies in North America and the Europe are currently using the baculovirus-insect cell technology to produce custom recombinant proteins for research and commercial applications. The recent breakthroughs using the baculovirus-insect cell-based system for the development of several commercial products that will impact animal and human health will further enhance interest in this technology by pharma. Clearly, future progress in novel cell and engineering advances will lead to fundamental scientific discoveries and serve to enhance the utility and applications of this baculovirus-insect cell system.

  5. Learning about Cells as Dynamic Entities: An Inquiry-Driven Cell Culture Project

    Science.gov (United States)

    Palombi, Peggy Shadduck; Jagger, Kathleen Snell

    2008-01-01

    Using cultured fibroblast cells, undergraduate students explore cell division and the responses of cultured cells to a variety of environmental changes. The students learn new research techniques and carry out a self-designed experiment. Through this project, students enhance their creative approach to scientific inquiry, learn time-management and…

  6. Development, Characterization and Cell Cultural Response of 3D Biocompatible Micro-Patterned Poly-ε-Caprolactone Scaffolds Designed and Fabricated Integrating Lithography and Micromolding Fabrication Techniques

    KAUST Repository

    Limongi, Tania

    2014-12-12

    Scaffold design and fabrication are very important subjects for biomaterial, tissue engineering and regenerative medicine research playing a unique role in tissue regeneration and repair. Among synthetic biomaterials Poly-ε- Caprolactone (PCL) is very attractive bioresorbable polyester due to its high permeability, biodegradability and capacity to be blended with other biopolymers. Thanks to its ability to naturally degrade in tissues, PCL has a great potential as a new material for implantable biomedical micro devices. This work focuses on the establishment of a micro fabrication process, by integrating lithography and micromolding fabrication techniques, for the realization of 3D microstructure PCL devices. Scaffold surface exhibits a combination in the patterned length scale; cylindrical pillars of 10 μm height and 10 μm diameter are arranged in a hexagonal lattice with periodicity of 30 μm and their sidewalls are nano-sculptured, with a regular pattern of grooves leading to a spatial modulation in the z direction. In order to demonstrate that these biocompatible pillared PCL substrates are suitable for a proper cell growth, NIH/3T3 mouse embryonic fibroblasts were seeded on them and cells key adhesion parameters were evaluated. Scanning Electron Microscopy and immunofluorescence analysis were carried out to check cell survival, proliferation and adhesion; cells growing on the PCL substrates appeared healthy and formed a well-developed network in close contact with the micro and nano features of the pillared surface. Those 3D scaffolds could be a promising solution for a wide range of applications within tissue engineering and regenerative medicine applications.

  7. Dynamized Preparations in Cell Culture

    Directory of Open Access Journals (Sweden)

    Ellanzhiyil Surendran Sunila

    2009-01-01

    Full Text Available Although reports on the efficacy of homeopathic medicines in animal models are limited, there are even fewer reports on the in vitro action of these dynamized preparations. We have evaluated the cytotoxic activity of 30C and 200C potencies of ten dynamized medicines against Dalton's Lymphoma Ascites, Ehrlich's Ascites Carcinoma, lung fibroblast (L929 and Chinese Hamster Ovary (CHO cell lines and compared activity with their mother tinctures during short-term and long-term cell culture. The effect of dynamized medicines to induce apoptosis was also evaluated and we studied how dynamized medicines affected genes expressed during apoptosis. Mother tinctures as well as some dynamized medicines showed significant cytotoxicity to cells during short and long-term incubation. Potentiated alcohol control did not produce any cytotoxicity at concentrations studied. The dynamized medicines were found to inhibit CHO cell colony formation and thymidine uptake in L929 cells and those of Thuja, Hydrastis and Carcinosinum were found to induce apoptosis in DLA cells. Moreover, dynamized Carcinosinum was found to induce the expression of p53 while dynamized Thuja produced characteristic laddering pattern in agarose gel electrophoresis of DNA. These results indicate that dynamized medicines possess cytotoxic as well as apoptosis-inducing properties.

  8. Expanding intestinal stem cells in culture

    NARCIS (Netherlands)

    Heo, Inha; Clevers, Hans

    2015-01-01

    Culturing intestinal stem cells into 3D organoids results in heterogeneous cell populations, reflecting the in vivo cell type diversity. In a recent paper published in Nature, Wang et al. established a culture condition for a highly homogeneous population of intestinal stem cells.

  9. Propagation of gnetum by tissue culture technique

    Directory of Open Access Journals (Sweden)

    Te-chato, S.

    2002-07-01

    Full Text Available Plantlet regeneration from tissue culture of gnetum could be induced using two pathways depending upon explant types. Leaf lamina yielded somatic embryos through somatic embryogenesis in basal MS supplemented with the same concentration of BA and TDZ at 0.5 mg/l. Using cluster of flowers or ovules in the same medium formula, however, callus formation took place and developed into meristemoid structures. These calli are called meristematic nodular calli and the pathway was organogenesis. Both pathways of plantlet regeneration could be applied for mass propagation of gnetum on a commercial scale in the future.

  10. The study of cultural objects by nuclear and conventional techniques

    International Nuclear Information System (INIS)

    A survey is given of the techniques that are used at the National Atomic Energy Commission of Argentina for the characterization and study of cultural and archaeological specimens. A short history of these activities is also given. (author)

  11. Simulation of limiting dilution technique in determination of immunocompetent cells frequency in irradiated cell cultures; Simulacao da tecnica de analise por limite de diluicao na determinacao da frequencia de celulas imunocompetentes em culturas contendo celulas irradiadas

    Energy Technology Data Exchange (ETDEWEB)

    Martini Filho, R.J.; Barlette, V.E.; Goes, E.G. [Centro Universitario Franciscano, Santa Maria, RS (Brazil); Covas, D.T.; Orellana, M. [Fundacao Hemocentro de Ribeirao Preto, SP (Brazil)

    2001-07-01

    Limiting dilution techniques (LDA) dose-response data have been used to detect immunocompetent T-Cells in microcultures. In this work, LDA frequencies estimates was obtained using {chi}2 minimization for irradiated cells in a range of 500 to 1,500 cGy. (author)

  12. Detecting mycoplasma contamination in cell cultures by polymerase chain reaction.

    Science.gov (United States)

    Uphoff, Cord C; Drexler, Hans G

    2011-01-01

    The detection of mycoplasmas in human and animal cell cultures is mandatory for every cell culture laboratory, because these bacteria are common contaminants, persist unrecognized in cell cultures for many years, and affect research results as well as the purity of cell culture products. The reliability of the mycoplasma detection depends on the sensitivity and specificity of the method and should also be convenient to be included in the basic routine of cell culture quality assessment. Polymerase chain reaction (PCR) detection is one of the acknowledged methodologies to detect mycoplasmas in cell cultures and cell culture products. Although the PCR offers a fast and simple technique to detect mycoplasmas, the method is also susceptible to errors and can produce false positive as well as false-negative results. Thus, the establishment and the routine application of the PCR assay require optimization and the inclusion of the appropriate control reactions. The presented protocol describes sample preparation, DNA extraction, PCR run, the analysis of the PCR products, and speciation of the contaminant. It also provides detailed information on how to avoid artifacts produced by the method. Established properly, PCR is a reliable, fast, and sensitive method and should be applied regularly to monitor the contamination status of cell cultures. PMID:21516400

  13. Cell Culture as an Alternative in Education.

    Science.gov (United States)

    Nardone, Roland M.

    1990-01-01

    Programs that are intended to inform and provide "hands-on" experience for students and to facilitate the introduction of cell culture-based laboratory exercises into the high school and college laboratory are examined. The components of the CellServ Program and the Cell Culture Toxicology Training Programs are described. (KR)

  14. Comparison of quantitative and semiquantitative culture techniques for burn biopsy.

    OpenAIRE

    Buchanan, K.; Heimbach, D. M.; Minshew, B H; Coyle, M B

    1986-01-01

    Accurate evaluation of bacterial colonization as a predictive index for wound sepsis has relied on a quantitative culture technique that provides exact colony counts per gram of tissue by culture of five serial dilutions of biopsy tissue homogenate. The method, while useful to the physician, is both labor intensive and expensive. In this study 78 eschar biopsies were cultured by a semiquantitative technique that involved the use of 0.1- and 0.01-ml samples of inocula and by the serial dilutio...

  15. Nuclear techniques and in vitro culture for plant improvement

    International Nuclear Information System (INIS)

    The continuous series of food shortages in many parts of the world have led scientists to consider the possibilities of using the new techniques to develop better varieties of plants. The basis for plant breeding is suitable genetic variability and mutation induction as the means to create additional variation. In vitro techniques are a relatively new tool in practical plant breeding. These Proceedings contain 62 papers and posters presented at the symposium, as well as excerpts from the discussions. The Symposium presentations are divided into the following sessions: Genetic variation from in vitro culture; Genetic stability of in vitro cultures; In vitro culture with application of mutagens; Haploids; In vitro mutant selection; Use of genetic variation derived by in vitro culture; In vitro techniques as aids in mutation breeding and Genetic engineering. A separate abstract is prepared for each of these papers and posters

  16. Endothelial cell cultures as a tool in biomaterial research

    NARCIS (Netherlands)

    Kirkpatrick, CJ; Otto, M; Kooten, TV; Krump, [No Value; Kriegsmann, J; Bittinger, F

    1999-01-01

    Progress in biocompatibility and tissue engineering would today be inconceivable without the aid of in vitro techniques. Endothelial cell cultures represent a valuable tool not just in haemocompatibility testing, but also in the concept of designing hybrid organs. In the past endothelial cells (EC)

  17. Metabolic flux rewiring in mammalian cell cultures.

    Science.gov (United States)

    Young, Jamey D

    2013-12-01

    Continuous cell lines (CCLs) engage in 'wasteful' glucose and glutamine metabolism that leads to accumulation of inhibitory byproducts, primarily lactate and ammonium. Advances in techniques for mapping intracellular carbon fluxes and profiling global changes in enzyme expression have led to a deeper understanding of the molecular drivers underlying these metabolic alterations. However, recent studies have revealed that CCLs are not necessarily entrenched in a glycolytic or glutaminolytic phenotype, but instead can shift their metabolism toward increased oxidative metabolism as nutrients become depleted and/or growth rate slows. Progress to understand dynamic flux regulation in CCLs has enabled the development of novel strategies to force cultures into desirable metabolic phenotypes, by combining fed-batch feeding strategies with direct metabolic engineering of host cells. PMID:23726154

  18. Advantages of embryogenic cell cultures of Gramineae

    International Nuclear Information System (INIS)

    Immature embryos and/or explants from very young leaves and inflorescences of 13 species and over 75 cultivars of Gramineae - including wheat, maize, rye, pearl millet, sugar-cane, Napier grass, Guinea grass, etc. - were used to initiate callus cultures. The cultures are white to yellowish white in colour, compact and contain small and thin-walled meristematic cells which are richly cytoplasmic, non-vacuolated and contain prominent starch grains. These embryogenic tissue cultures provide a long-term, highly reliable and efficient means of rapid mass clonal propagation by the formation of somatic embryos that arise from single cells. The cultures consist largely of cytologically normal diploid cells. During the process of plant regeneration via somatic embryogenesis, there is strong selection in favour of normal cells, so that plants recovered from such cultures neither exhibit any morphological abnormalities nor show any evidence of cytological changes in the number or structure of chromosomes. Embryogenic callus cultures have been used successfully to establish highly dispersed and friable cell-suspension cultures. These fast-growing cultures comprise groups of 2-6 embryogenic cells, which adhere together to form larger unorganized aggregates of up to about 75 cells, but do not contain any organized meristems or callus tissues. Plants were regenerated by somatic embryogenesis from embryogenic cell-suspension cultures of pearl millet, Guinea grass, sugar-cane and maize. Finally, embryogenic cell-suspension cultures are the only current source of totipotent protoplasts in Gramineae. Protoplasts isolated from such cultures have been successfully cultured to produce somatic embryos and plants in pearl millet, Guinea grass, Napier grass and sugar-cane. (author)

  19. Cell Suspension Culture of Neem Tree

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The establishment of suspension culture system for neem (Azadirachta indica A. Juss) cells and the suspension culture condition was studied. It shows that the neem cell suspension culture system was best in B5 liquid medium, 2.0~4.0mg/L NAA with direct spill method. Based on the integrated analysis of cell biomass, Azadirachtin content and productivity, the optimum culture conditions were B5 liquid medium, 2.0-4.0 mg/L NAA, 3% sucrose at 25 ℃. The optimum rotating speed of the shaker and broth content d...

  20. 9 CFR 101.6 - Cell cultures.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Cell cultures. 101.6 Section 101.6 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS DEFINITIONS § 101.6 Cell cultures. When used in conjunction with or in reference...

  1. Callus and cell suspension cultures of carnation

    DEFF Research Database (Denmark)

    Engvild, Kjeld Christensen

    1972-01-01

    . Cell suspension cultures worked best in media containing 2,4-D in which they had a doubling time of about 2 days. Filtered suspensions were successfully plated on agar in petri dishes, but division was never observed in single cells. The cultures initiated roots at higher concentrations of IAA or NAA...

  2. Multizone paper platform for 3D cell cultures.

    Directory of Open Access Journals (Sweden)

    Ratmir Derda

    Full Text Available In vitro 3D culture is an important model for tissues in vivo. Cells in different locations of 3D tissues are physiologically different, because they are exposed to different concentrations of oxygen, nutrients, and signaling molecules, and to other environmental factors (temperature, mechanical stress, etc. The majority of high-throughput assays based on 3D cultures, however, can only detect the average behavior of cells in the whole 3D construct. Isolation of cells from specific regions of 3D cultures is possible, but relies on low-throughput techniques such as tissue sectioning and micromanipulation. Based on a procedure reported previously ("cells-in-gels-in-paper" or CiGiP, this paper describes a simple method for culture of arrays of thin planar sections of tissues, either alone or stacked to create more complex 3D tissue structures. This procedure starts with sheets of paper patterned with hydrophobic regions that form 96 hydrophilic zones. Serial spotting of cells suspended in extracellular matrix (ECM gel onto the patterned paper creates an array of 200 micron-thick slabs of ECM gel (supported mechanically by cellulose fibers containing cells. Stacking the sheets with zones aligned on top of one another assembles 96 3D multilayer constructs. De-stacking the layers of the 3D culture, by peeling apart the sheets of paper, "sections" all 96 cultures at once. It is, thus, simple to isolate 200-micron-thick cell-containing slabs from each 3D culture in the 96-zone array. Because the 3D cultures are assembled from multiple layers, the number of cells plated initially in each layer determines the spatial distribution of cells in the stacked 3D cultures. This capability made it possible to compare the growth of 3D tumor models of different spatial composition, and to examine the migration of cells in these structures.

  3. Plant cell cultures and their biotechnological potential

    Energy Technology Data Exchange (ETDEWEB)

    Barz, W.; Ellis, B.E.

    1981-01-01

    The potential of plant cell suspension cultures for the biotechnological production of high-cost, plant-specific compounds is critically evaluated. The basic roles of nutrient media and phytohormones are described followed by a description of the recent progress in mass cultivation of plant cell cultures as measured by biomass and doubling time. The accumulation of secondary constituents in cell cultures is reviewed and methods for the selection of high-producing strains are described. The essential features of the selection strategy are the establishment of cell cultures from high-producing plants and a sensitive assay (e.g. radio-immunoassay) for the screening of microcolonies grown on petri dishes. The accumulation of biosynthetic intermediates of secondary constituents in cell culture strains will possibly lead to the isolation of novel compounds.

  4. Studying cell-cell communication in co-culture

    OpenAIRE

    Bogdanowicz, Danielle R.; Lu, Helen H.

    2013-01-01

    Heterotypic and homotypic cellular interactions are essential for biological function, and co-culture models are versatile tools for investigating these cellular interactions in vitro. Physiologically relevant co-culture models have been used to elucidate the effects of cell-cell physical contact and/or secreted factors, as well as the influence of substrate geometry and interaction scale on cell response. Identifying the relative contribution of each cell population to co-culture is often ex...

  5. Laser-based direct-write techniques for cell printing

    Science.gov (United States)

    Schiele, Nathan R; Corr, David T; Huang, Yong; Raof, Nurazhani Abdul; Xie, Yubing; Chrisey, Douglas B

    2016-01-01

    Fabrication of cellular constructs with spatial control of cell location (±5 μm) is essential to the advancement of a wide range of applications including tissue engineering, stem cell and cancer research. Precise cell placement, especially of multiple cell types in co- or multi-cultures and in three dimensions, can enable research possibilities otherwise impossible, such as the cell-by-cell assembly of complex cellular constructs. Laser-based direct writing, a printing technique first utilized in electronics applications, has been adapted to transfer living cells and other biological materials (e.g., enzymes, proteins and bioceramics). Many different cell types have been printed using laser-based direct writing, and this technique offers significant improvements when compared to conventional cell patterning techniques. The predominance of work to date has not been in application of the technique, but rather focused on demonstrating the ability of direct writing to pattern living cells, in a spatially precise manner, while maintaining cellular viability. This paper reviews laser-based additive direct-write techniques for cell printing, and the various cell types successfully laser direct-written that have applications in tissue engineering, stem cell and cancer research are highlighted. A particular focus is paid to process dynamics modeling and process-induced cell injury during laser-based cell direct writing. PMID:20814088

  6. Surface modified alginate microcapsules for 3D cell culture

    Science.gov (United States)

    Chen, Yi-Wen; Kuo, Chiung Wen; Chueh, Di-Yen; Chen, Peilin

    2016-06-01

    Culture as three dimensional cell aggregates or spheroids can offer an ideal platform for tissue engineering applications and for pharmaceutical screening. Such 3D culture models, however, may suffer from the problems such as immune response and ineffective and cumbersome culture. This paper describes a simple method for producing microcapsules with alginate cores and a thin shell of poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) to encapsulate mouse induced pluripotent stem (miPS) cells, generating a non-fouling surface as an effective immunoisolation barrier. We demonstrated the trapping of the alginate microcapsules in a microwell array for the continuous observation and culture of a large number of encapsulated miPS cells in parallel. miPS cells cultured in the microcapsules survived well and proliferated to form a single cell aggregate. Droplet formation of monodisperse microcapsules with controlled size combined with flow cytometry provided an efficient way to quantitatively analyze the growth of encapsulated cells in a high-throughput manner. The simple and cost-effective coating technique employed to produce the core-shell microcapsules could be used in the emerging field of cell therapy. The microwell array would provide a convenient, user friendly and high-throughput platform for long-term cell culture and monitoring.

  7. A Microwell Cell Culture Platform for the Aggregation of Pancreatic β-Cells

    OpenAIRE

    Bernard, Abigail B.; Lin, Chien-Chi; Anseth, Kristi S.

    2012-01-01

    Cell–cell contact between pancreatic β-cells is important for maintaining survival and normal insulin secretion. Various techniques have been developed to promote cell–cell contact between β-cells, but a simple yet robust method that affords precise control over three-dimensional (3D) β-cell cluster size has not been demonstrated. To address this need, we developed a poly(ethylene glycol) (PEG) hydrogel microwell platform using photolithography. This microwell cell-culture platform promotes t...

  8. Cell Culture for Production of Insecticidal Viruses.

    Science.gov (United States)

    Reid, Steven; Chan, Leslie C L; Matindoost, Leila; Pushparajan, Charlotte; Visnovsky, Gabriel

    2016-01-01

    While large-scale culture of insect cells will need to be conducted using bioreactors up to 10,000 l scale, many of the main challenges for cell culture-based production of insecticidal viruses can be studied using small-scale (20-500 ml) shaker/spinner flasks, either in free suspension or using microcarrier-based systems. These challenges still relate to the development of appropriate cell lines, stability of virus strains in culture, enhancing virus yields per cell, and the development of serum-free media and feeds for the desired production systems. Hence this chapter presents mainly the methods required to work with and analyze effectively insect cell systems using small-scale cultures. Outlined are procedures for quantifying cells and virus and for establishing frozen cells and virus stocks. The approach for maintaining cell cultures and the multiplicity of infection (MOI) and time of infection (TOI) parameters that should be considered for conducting infections are discussed.The methods described relate, in particular, to the suspension culture of Helicoverpa zea and Spodoptera frugiperda cell lines to produce the baculoviruses Helicoverpa armigera nucleopolyhedrovirus, HearNPV, and Anticarsia gemmatalis multicapsid nucleopolyhedrovirus, AgMNPV, respectively, and the production of the nonoccluded Oryctes nudivirus, OrNV, using an adherent coleopteran cell line. PMID:27565495

  9. Towards dynamic metabolic flux analysis in CHO cell cultures.

    Science.gov (United States)

    Ahn, Woo Suk; Antoniewicz, Maciek R

    2012-01-01

    Chinese hamster ovary (CHO) cells are the most widely used mammalian cell line for biopharmaceutical production, with a total global market approaching $100 billion per year. In the pharmaceutical industry CHO cells are grown in fed-batch culture, where cellular metabolism is characterized by high glucose and glutamine uptake rates combined with high rates of ammonium and lactate secretion. The metabolism of CHO cells changes dramatically during a fed-batch culture as the cells adapt to a changing environment and transition from exponential growth phase to stationary phase. Thus far, it has been challenging to study metabolic flux dynamics in CHO cell cultures using conventional metabolic flux analysis techniques that were developed for systems at metabolic steady state. In this paper we review progress on flux analysis in CHO cells and techniques for dynamic metabolic flux analysis. Application of these new tools may allow identification of intracellular metabolic bottlenecks at specific stages in CHO cell cultures and eventually lead to novel strategies for improving CHO cell metabolism and optimizing biopharmaceutical process performance. PMID:22102428

  10. Emulsions Containing Perfluorocarbon Support Cell Cultures

    Science.gov (United States)

    Ju, Lu-Kwang; Lee, Jaw Fang; Armiger, William B.

    1990-01-01

    Addition of emulsion containing perfluorocarbon liquid to aqueous cell-culture medium increases capacity of medium to support mammalian cells. FC-40 Fluorinert (or equivalent) - increases average density of medium so approximately equal to that of cells. Cells stay suspended in medium without mechanical stirring, which damages them. Increases density enough to prevent cells from setting, and increases viscosity of medium so oxygen bubbled through it and nutrients stirred in with less damage to delicate cells.

  11. Preparation of 14C-catechins by tea cell culture

    International Nuclear Information System (INIS)

    The preparation of 14C labelled catechins was studied by feeding 14C labelled precursor to tea cultured cells. Two labelled precursors were tested and their effects were compared. The dynamics of absorption and transformation of fed precursors were analyzed and the effects of pre-culture as well as UV light pretreatment on product labelling rate were evaluated. Product analysis was also made by HSCCC and HPLC techniques

  12. Constructing a High Density Cell Culture System

    Science.gov (United States)

    Spaulding, Glenn F. (Inventor)

    1996-01-01

    An annular culture vessel for growing mammalian cells is constructed in a one piece integral and annular configuration with an open end which is closed by an endcap. The culture vessel is rotatable about a horizontal axis by use of conventional roller systems commonly used in culture laboratories. The end wall of the endcap has tapered access ports to frictionally and sealingly receive the ends of hypodermic syringes. The syringes permit the introduction of fresh nutrient and withdrawal of spent nutrients. The walls are made of conventional polymeric cell culture material and are subjected to neutron bombardment to form minute gas permeable perforations in the walls.

  13. Propagation of Aquilaria malaccensis seedlings through tissue culture techniques

    International Nuclear Information System (INIS)

    Aquilaria malaccensis or karas is the principal source of gaharu resin, which is used in many cultures for incense, perfumes and traditional medicines. The species is mainly propagated conventionally through seeds, cuttings and graftings. Propagation by seeds is usually a reliable method for other forest species, but for karas, this technique is inadequate to meet the current demand of seedling supplies. This is principally due to its low seed viability, low germination rate, delayed rooting of seedlings, long life-cycle and rare seed production. Tissue culture has several advantages over conventional propagation, especially for obtaining large number of uniform and high-yielding plantlets or clones. This paper presents the current progress on mass-propagation of Aquilaria malaccensis seedlings through tissue culture technique at Nuclear Malaysia. (author)

  14. A Modified Technique for Culturing Primary Fetal Rat Cortical Neurons

    Directory of Open Access Journals (Sweden)

    Sui-Yi Xu

    2012-01-01

    Full Text Available The study explored a modified primary culture system for fetal rat cortical neurons. Day E18 embryos from pregnant Sprague Dawley rats were microdissected under a stereoscope. To minimize enzymatic damage to the cultured neurons, we applied a sequential digestion protocol using papain and Dnase I. The resulting sifted cell suspension was seeded at a density of 50,000 cells per cm2 onto 0.1 mg/mL L-PLL-covered vessels. After a four-hour incubation in high-glucose Dulbecco’s Modified Eagle’s Medium (HG-DMEM to allow the neurons to adhere, the media was changed to neurobasal medium that was refreshed by changing half of the volume after three days followed by a complete medium change every week. The cells displayed progressively robust neurite extension, and nonneuronal-like cells could barely be detected by five days in vitro (DIV; cell growth was still substantial at 14 DIV. Neurons were identified by β-tubulin III immunofluorescence, and neuronal purity within the cultures was assessed at over 95% by both flow cytometry and by dark-field counting of β-tubulin III-positive cells. These results suggest that the protocol was successful and that the high purity of neurons in this system could be used as the basis for generating various cell models of neurological disease.

  15. Human hematopoietic cell culture, transduction, and analyses

    DEFF Research Database (Denmark)

    Bonde, Jesper; Wirthlin, Louisa; Kohn, Donald B;

    2008-01-01

    This unit provides methods for introducing genes into human hematopoietic progenitor cells. The Basic Protocol describes isolation of CD34(+) cells, transduction of these cells with a retroviral vector on fibronectin-coated plates, assaying the efficiency of transduction, and establishing long......-term cultures. Support protocols describe methods for maintenance of vector-producing fibroblasts (VPF) and supernatant collection from these cells, screening medium components for the ability to support hematopoietic cell growth, and establishing colonies from long-term cultures. Other protocols provide PCR...

  16. A microwell cell culture platform for the aggregation of pancreatic β-cells.

    Science.gov (United States)

    Bernard, Abigail B; Lin, Chien-Chi; Anseth, Kristi S

    2012-08-01

    Cell-cell contact between pancreatic β-cells is important for maintaining survival and normal insulin secretion. Various techniques have been developed to promote cell-cell contact between β-cells, but a simple yet robust method that affords precise control over three-dimensional (3D) β-cell cluster size has not been demonstrated. To address this need, we developed a poly(ethylene glycol) (PEG) hydrogel microwell platform using photolithography. This microwell cell-culture platform promotes the formation of 3D β-cell aggregates of defined sizes from 25 to 210 μm in diameter. Using this platform, mouse insulinoma 6 (MIN6) β-cells formed aggregates with cell-cell adherin junctions. These naturally formed cell aggregates with controllable sizes can be removed from the microwells for macroencapsulation, implantation, or other biological assays. When removed and subsequently encapsulated in PEG hydrogels, the aggregated cell clusters demonstrated improved cellular viability (>90%) over 7 days in culture, while the β-cells encapsulated as single cells maintained only 20% viability. Aggregated MIN6 cells also exhibited more than fourfold higher insulin secretion in response to a glucose challenge compared with encapsulated single β-cells. Further, the cell aggregates stained positively for E-cadherin, indicative of the formation of cell junctions. Using this hydrogel microwell cell-culture method, viable and functional β-cell aggregates of specific sizes were created, providing a platform from which other biologically relevant questions may be answered. PMID:22320435

  17. Callus and cell suspension cultures of carnation

    DEFF Research Database (Denmark)

    Engvild, Kjeld Christensen

    1972-01-01

    Callus cultures of carnation, Dianthus caryophyllus L. ev. G. J. Sim, were grown on a synthetic medium of half strength Murashige and Skoog salts, 3 % sucrose, 100 mg/l of myo-inositol, 0.5 mg/l each of thiamin, HCl, pyridoxin, HCl and nicotinic acid and 10 g/l agar. Optimal concentrations of....... Cell suspension cultures worked best in media containing 2,4-D in which they had a doubling time of about 2 days. Filtered suspensions were successfully plated on agar in petri dishes, but division was never observed in single cells. The cultures initiated roots at higher concentrations of IAA or NAA...

  18. Cell culture models for study of differentiated adipose cells

    OpenAIRE

    Clynes, Martin

    2014-01-01

    Adipose cells are an important source of mesenchymal stem cells and are important for direct use in research on lipid metabolism and obesity. In addition to use of primary cultures, there is increasing interest in other sources of larger numbers of cells, using approaches including induced pluripotent stem cell differentiation and viral immortalisation.

  19. Human cell culture in a space bioreactor

    Science.gov (United States)

    Morrison, Dennis R.

    1988-01-01

    Microgravity offers new ways of handling fluids, gases, and growing mammalian cells in efficient suspension cultures. In 1976 bioreactor engineers designed a system using a cylindrical reactor vessel in which the cells and medium are slowly mixed. The reaction chamber is interchangeable and can be used for several types of cell cultures. NASA has methodically developed unique suspension type cell and recovery apparatus culture systems for bioprocess technology experiments and production of biological products in microgravity. The first Space Bioreactor was designed for microprocessor control, no gaseous headspace, circulation and resupply of culture medium, and slow mixing in very low shear regimes. Various ground based bioreactors are being used to test reactor vessel design, on-line sensors, effects of shear, nutrient supply, and waste removal from continuous culture of human cells attached to microcarriers. The small Bioreactor is being constructed for flight experiments in the Shuttle Middeck to verify systems operation under microgravity conditions and to measure the efficiencies of mass transport, gas transfer, oxygen consumption and control of low shear stress on cells.

  20. Melphalan metabolism in cultured cells

    International Nuclear Information System (INIS)

    Procedures are presented for the adaptation of reversed-phase-HPLC methods to accomplish separation and isolation of the cancer therapeutic drug melphalan (L-phenylalanine mustard) and its metabolic products from whole cells. Five major degradation products of melphalan were observed following its hydrolysis in phosphate buffer in vitro. The two most polar of these products (or modifications of them) were also found in the cytosol of Chinese hamster CHO cells. The amounts of these two polar products (shown not to be mono- or dihydroxymelphalan) were significantly changed by the pretreatment of cells with ZnC12, one being increased in amount while the other was reduced to an insignificant level. In ZnC12-treated cells, there was also an increased binding of melphalan (or its derivatives) to one protein fraction resolved by gel filtration-HPLC. These observations suggest that changes in polar melphalan products, and perhaps their interaction with a protein, may by involved in the reduction of melphalan cytotoxicity observed in ZnC12-treated cells. While ZnC12 is also known to increase the level of glutathione in cells, no significant amounts of glutathione-melphalan derivatives of the type formed non-enzymatically in vitro could be detected in ZnC12-treated or untreated cells. Formation of derivatives of melphalan with glutathione catabolic products in ZnC12-treated cells has not yet been eliminated, however. 17 refs., 5 figs., 1 tab

  1. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Science.gov (United States)

    Stampfer, Martha R; Garbe, James C

    2015-02-24

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  2. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R.; Garbe, James C.

    2016-06-28

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  3. Establishing a stem cell culture laboratory for clinical trials

    Directory of Open Access Journals (Sweden)

    Elíseo Joji Sekiya

    2012-01-01

    Full Text Available Adult stem/progenitor cells are found in different human tissues. An in vitro cell culture is needed for their isolation or for their expansion when they are not available in a sufficient quantity to regenerate damaged organs and tissues. The level of complexity of these new technologies requires adequate facilities, qualified personnel with experience in cell culture techniques, assessment of quality and clear protocols for cell production. The rules for the implementation of cell therapy centers involve national and international standards of good manufacturing practices. However, such standards are not uniform, reflecting the diversity of technical and scientific development. Here standards from the United States, the European Union and Brazil are analyzed. Moreover, practical solutions encountered for the implementation of a cell therapy center appropriate for the preparation and supply of cultured cells for clinical studies are described. Development stages involved the planning and preparation of the project, the construction of the facility, standardization of laboratory procedures and development of systems to prevent cross contamination. Combining the theoretical knowledge of research centers involved in the study of cells with the practical experience of blood therapy services that manage structures for cell transplantation is presented as the best potential for synergy to meet the demands to implement cell therapy centers.

  4. Impedimetric quantification of cells encapsulated in hydrogel cultured in a paper-based microchamber.

    Science.gov (United States)

    Lei, Kin Fong; Huang, Chia-Hao; Tsang, Ngan-Ming

    2016-01-15

    Recently, 3D cell culture technique was proposed to provide a more physiologically-meaningful environment for cell-based assays. With the development of microfluidics technology, cellular response can be quantified by impedance measurement technique in a real-time and non-invasive manner. However, handling of these microfluidic systems requires a trained engineering personnel and the operation is not compatible to traditional biological research laboratories. In this work, we incorporated the impedance measurement technique to paper-based 3D cell culture model and demonstrated non-invasive quantification of cells encapsulated in hydrogel during the culture course. A cellulose filter paper was patterned with an array of circular microchambers. Cells were encapsulated in hydrogel and loaded to the microchambers for culturing cells in 3D environment. At the preset schedule during the culture course, the paper was placed on a glass substrate with measurement electrodes for the impedance measurement. Cells in each microchamber was represented by impedance magnitude and cell proliferation could be studied over time. Also, conventional bio-assay was performed to further confirm the feasibility of the impedimetric quantification of cells encapsulated in hydrogel cultured in the paper-based microchamber. This technique provides a convenient, fast, and non-invasive approach to monitor cells cultured in 3D environment. It has potential to be developed for routine 3D cell culture protocol in biological research laboratories. PMID:26592655

  5. Cell culture experiments planned for the space bioreactor

    Science.gov (United States)

    Morrison, Dennis R.; Cross, John H.

    1987-01-01

    Culturing of cells in a pilot-scale bioreactor remains to be done in microgravity. An approach is presented based on several studies of cell culture systems. Previous and current cell culture research in microgravity which is specifically directed towards development of a space bioprocess is described. Cell culture experiments planned for a microgravity sciences mission are described in abstract form.

  6. Virus Elimination from Ornamental Plants Using in vitro Culture Techniques

    Directory of Open Access Journals (Sweden)

    Snežana Milošević

    2012-01-01

    Full Text Available Viruses are responsible for numerous epidemics in different crops in all parts of the world.As a consequence of their presence great economic losses are being incurred. In addition tothe development of sensitive techniques for detection, identification and characterization ofviruses, substantial attention has also been paid to biotechnological methods for their eliminationfrom plants. In this review article, the following biotechnological in vitro culture techniquesfor virus elimination from ornamental plants are presented: meristem culture, thermotherapy,chemotherapy, cryotherapy or a combination of these methods. The plant species,as well as the type of virus determine the choice of a most suitable method. The state ofthe art in investigation of virus elimination from Impatiens sp. in Serbia is summarized.

  7. The use of tissue culture techniques to detect irradiated vegetables

    International Nuclear Information System (INIS)

    the ability of two tissue culture methods, callus and vegetable growth induction, to detect irradiated vegetables was evaluated. Potato tubers, carrot roots, garlic cloves and onion bulbs were subjected to various gamma radiation doses (0, 25, 100, 150, 250, 500, 750, and 1000 Gy). Irradiated vegetables were cultured in vitro and in vivo (pots). Gamma irradiation significantly reduced callus-forming ability especially in carrot and potato where no callus was observed in doses higher than 50 Gy. Length of shoots and roots growing from irradiated garlic and onion explants was considerably reduced starting from the 25 Gy dose. No roots were formed on garlic explants at any irradiation dose. Garlic leaves growing from irradiated explants were spotted with purple to brown spots. The intensity of these spots increased as gamma ray dosage increased. In the pot experiment, potato plant appeared in the control only. On the contrary, a complete sprouting of garlic and onion was seen in all irradiation treatments. It was not possible to distinguish between the various irradiation treatments and the control 3 days after planting in pots. The two in vitro techniques, tested in our study, may effectively be used to detect irradiated vegetables and estimate the range of doses used. The callus formation method is more useful for potato and carrot, since regeneration of shoots in vitro from these two plants takes along time, making this method unpractical. The other technique is very useful in the case of onion and garlic since it is rapid. The two techniques can be used with most of the vegetables that can be cultured in vitro. (Author)

  8. Detection of irradiated vegetables using tissue culture techniques

    International Nuclear Information System (INIS)

    The ability of two tissue culture methods, callus and vegetative growth induction, to detect irradiated vegetables was evaluated. Potato tubers, carrot roots, garlic cloves, and onion bulbs were subjected to various gamma irradiation doses (0, 25, 50, 100, 150, 250, 500, 750, and 1000 Gy). Irradiated vegetables were cultured in vitro and in vivo (pots). Gamma irradiation significantly reduced callus-forming ability especially in carrot and carrot and potato where no callus was observed in doses higher than 50 Gy. Length of shoots and roots growing from irradiated garlic and onion explants was considerably reduced starting from the 25 Gy dose. No roots were formed on garlic explants at any irradiation dose. Garlic leaves growing from irradiated explants were spotted with purple to brown spots. The intensity of these spots increased as gamma ray dosage increased. In the pot experiment, potato plants appeared in the control only. On the contrary, a complete sprouting of garlic and onion was seen in all irradiation treatments. It was not possible to distinguish between the various irradiation treatments and the control 3 days after planting in pots. The two in vitro techniques, tested in our study, may effectively be used to detect irradiated vegetables and estimate the range of doses used. The callus formation method is more useful for potato and carrot, since regeneration of shoots in vitro from these two plants takes a long time, making this method unpractical. The other technique is very useful in the case of onion and garlic since it is rapid. The two techniques can be used with most of the vegetables that can be cultured in vitro. (Author)

  9. Analytical techniques applied to study cultural heritage objects

    Energy Technology Data Exchange (ETDEWEB)

    Rizzutto, M.A.; Curado, J.F.; Bernardes, S.; Campos, P.H.O.V.; Kajiya, E.A.M.; Silva, T.F.; Rodrigues, C.L.; Moro, M.; Tabacniks, M.; Added, N., E-mail: rizzutto@if.usp.br [Universidade de Sao Paulo (USP), SP (Brazil). Instituto de Fisica

    2015-07-01

    The scientific study of artistic and cultural heritage objects have been routinely performed in Europe and the United States for decades. In Brazil this research area is growing, mainly through the use of physical and chemical characterization methods. Since 2003 the Group of Applied Physics with Particle Accelerators of the Physics Institute of the University of Sao Paulo (GFAA-IF) has been working with various methodologies for material characterization and analysis of cultural objects. Initially using ion beam analysis performed with Particle Induced X-Ray Emission (PIXE), Rutherford Backscattering (RBS) and recently Ion Beam Induced Luminescence (IBIL), for the determination of the elements and chemical compounds in the surface layers. These techniques are widely used in the Laboratory of Materials Analysis with Ion Beams (LAMFI-USP). Recently, the GFAA expanded the studies to other possibilities of analysis enabled by imaging techniques that coupled with elemental and compositional characterization provide a better understanding on the materials and techniques used in the creative process in the manufacture of objects. The imaging analysis, mainly used to examine and document artistic and cultural heritage objects, are performed through images with visible light, infrared reflectography (IR), fluorescence with ultraviolet radiation (UV), tangential light and digital radiography. Expanding more the possibilities of analysis, new capabilities were added using portable equipment such as Energy Dispersive X-Ray Fluorescence (ED-XRF) and Raman Spectroscopy that can be used for analysis 'in situ' at the museums. The results of these analyzes are providing valuable information on the manufacturing process and have provided new information on objects of different University of Sao Paulo museums. Improving the arsenal of cultural heritage analysis it was recently constructed an 3D robotic stage for the precise positioning of samples in the external beam setup

  10. Analytical techniques applied to study cultural heritage objects

    International Nuclear Information System (INIS)

    The scientific study of artistic and cultural heritage objects have been routinely performed in Europe and the United States for decades. In Brazil this research area is growing, mainly through the use of physical and chemical characterization methods. Since 2003 the Group of Applied Physics with Particle Accelerators of the Physics Institute of the University of Sao Paulo (GFAA-IF) has been working with various methodologies for material characterization and analysis of cultural objects. Initially using ion beam analysis performed with Particle Induced X-Ray Emission (PIXE), Rutherford Backscattering (RBS) and recently Ion Beam Induced Luminescence (IBIL), for the determination of the elements and chemical compounds in the surface layers. These techniques are widely used in the Laboratory of Materials Analysis with Ion Beams (LAMFI-USP). Recently, the GFAA expanded the studies to other possibilities of analysis enabled by imaging techniques that coupled with elemental and compositional characterization provide a better understanding on the materials and techniques used in the creative process in the manufacture of objects. The imaging analysis, mainly used to examine and document artistic and cultural heritage objects, are performed through images with visible light, infrared reflectography (IR), fluorescence with ultraviolet radiation (UV), tangential light and digital radiography. Expanding more the possibilities of analysis, new capabilities were added using portable equipment such as Energy Dispersive X-Ray Fluorescence (ED-XRF) and Raman Spectroscopy that can be used for analysis 'in situ' at the museums. The results of these analyzes are providing valuable information on the manufacturing process and have provided new information on objects of different University of Sao Paulo museums. Improving the arsenal of cultural heritage analysis it was recently constructed an 3D robotic stage for the precise positioning of samples in the external beam setup

  11. A biocompatible micro cell culture chamber (mu CCC) for the culturing and on-line monitoring of eukaryote cells

    DEFF Research Database (Denmark)

    Stangegaard, Michael; Petronis, Sarunas; Jørgensen, Anders Michael;

    2006-01-01

    We have previously shown that a polymeric (PMMA) chip with medium perfusion and integrated heat regulation provides sufficiently precise heat regulation, pH-control and medium exchange to support cell growth for weeks. However, it was unclear how closely the cells cultured in the chip resembled...... cells cultured in the culture flask. In the current study, gene expression profiles of cells cultured in the chip were compared with gene expression profiles of cells cultured in culture flasks. The results showed that there were only two genes that were differently expressed in cells grown in the cell...... culture chip compared to cell culture flasks. The cell culture chip could without further modification support cell growth of two other cell lines. Light coming from the microscope lamp during optical recordings of the cells was the only external factor identified, that could have a negative effect...

  12. A biocompatible micro cell culture chamber (microCCC) for the culturing and on-line monitoring of eukaryote cells

    DEFF Research Database (Denmark)

    Stangegaard, Michael; Petronis, Sarunas; Jørgensen, A M;

    2006-01-01

    We have previously shown that a polymeric (PMMA) chip with medium perfusion and integrated heat regulation provides sufficiently precise heat regulation, pH-control and medium exchange to support cell growth for weeks. However, it was unclear how closely the cells cultured in the chip resembled...... cells cultured in the culture flask. In the current study, gene expression profiles of cells cultured in the chip were compared with gene expression profiles of cells cultured in culture flasks. The results showed that there were only two genes that were differently expressed in cells grown in the cell...... culture chip compared to cell culture flasks. The cell culture chip could without further modification support cell growth of two other cell lines. Light coming from the microscope lamp during optical recordings of the cells was the only external factor identified, that could have a negative effect...

  13. Characterization and Classification of Adherent Cells in Monolayer Culture using Automated Tracking and Evolutionary Algorithms

    OpenAIRE

    Zhang, Z.; Bedder, M; Smith, S L; Walker, D; Shabir, S.; Southgate, J

    2016-01-01

    This paper presents a novel method for tracking and characterizing adherent cells in monolayer culture. A system of cell tracking employing computer vision techniques was applied to time-lapse videos of replicate normal human uro-epithelial cell cultures exposed to different concentrations of adenosine triphosphate (ATP) and a selective purinergic P2X antagonist (PPADS), acquired over a 24hour period. Subsequent analysis following feature extraction demonstrated the ability of the technique t...

  14. Objective Assessments of Temperature Maintenance Using In Vitro Culture Techniques

    OpenAIRE

    Cooke, Simon; Tyler, John P. P.; Driscoll, Geoff

    2002-01-01

    Purpose: To assess the ability of various facets of embryo culture (microscope stage warmers, volumes of culture media, culture vessel lids, and type of culture incubator) to maintain a constant temperature in vitro.

  15. An animal cell culture: Advance technology for modern research

    OpenAIRE

    Sarita Khare; Rajeev Nema

    2012-01-01

    At the present time animal cell culture is more significant and multifarious application tool for current research streams. A lot of field assorted from animal cell culture such: stem cell biology, IVF technology, cancer cell biology, monoclonal antibody production, recombinant protein production, gene therapy, vaccine manufacturing, novel drug selection and improvement. In this review conclude animal cell culture as well as its requirements

  16. Radiation transformation in differentiated human cells in culture

    International Nuclear Information System (INIS)

    A tissue culture technique is described for human thyroid tissue as an approach to studying mechanisms of human radiation carcinogenesis. Normal human tissue obtained from surgery is treated in one of two ways, depending upon size of specimen. Large pieces are completely digested in trypsin/ collagenase solution to a single cell suspension. Small pieces of tissue are plated as explants following partial digestion in trypsin/collagenase solution. Following irradiation of the primary differentiated monolayers (normally 10 days after plating), the development of transformed characteristics is monitored in the subsequent subcultures. A very high level of morphological and functional differentiation is apparent in the primary cultures. Over a period of approx. 6 months, the irradiated surviving cells continue to grow in culture, unlike the unirradiated controls which senesce after 2-3 subcultures. (UK)

  17. Wnt-Dependent Control of Cell Polarity in Cultured Cells.

    Science.gov (United States)

    Runkle, Kristin B; Witze, Eric S

    2016-01-01

    The secreted ligand Wnt5a regulates cell polarity and polarized cell movement during development by signaling through the poorly defined noncanonical Wnt pathway. Cell polarity regulates most aspects of cell behavior including the organization of apical/basolateral membrane domains of epithelial cells, polarized cell divisions along a directional plane, and front rear polarity during cell migration. These characteristics of cell polarity allow coordinated cell movements required for tissue formation and organogenesis during embryonic development. Genetic model organisms have been used to identify multiple signaling pathways including Wnt5a that are required to establish cell polarity and regulate polarized cell behavior. However, the downstream signaling events that regulate these complex cellular processes are still poorly understood. The methods below describe assays to study Wnt5a-induced cell polarity in cultured cells, which may facilitate our understanding of these complex signaling pathways. PMID:27590152

  18. Ion beam analysis and spectrometry techniques for Cultural Heritage studies

    International Nuclear Information System (INIS)

    The implementation of experimental techniques for the characterisation of Cultural heritage materials has to take into account some requirements. The complexity of these past materials requires the development of new techniques of examination and analysis, or the transfer of technologies developed for the study of advanced materials. In addition, due to precious aspect of artwork it is also necessary to use the non-destructive methods, respecting the integrity of objects. It is for this reason that the methods using radiations and/or particles play a important role in the scientific study of art history and archaeology since their discovery. X-ray and γ-ray spectrometry as well as ion beam analysis (IBA) are analytical tools at the service of Cultural heritage. This report mainly presents experimental developments for IBA: PIXE, RBS/EBS and NRA. These developments were applied to the study of archaeological composite materials: layered materials or mixtures composed of organic and non-organic phases. Three examples are shown: evolution of silvering techniques for the production of counterfeit coinage during the Roman Empire and in the 16. century, the characterization of composites or mixed mineral/organic compounds such as bone and paint. In these last two cases, the combination of techniques gave original results on the proportion of both phases: apatite/collagen in bone, pigment/binder in paintings. Another part of this report is then dedicated to the non-invasive/non-destructive characterization of prehistoric pigments, in situ, for rock art studies in caves and in the laboratory. Finally, the perspectives of this work are presented. (author)

  19. Nerve cells culture from lumbar spinal cord on surfaces modified by plasma pyrrole polymerization.

    Science.gov (United States)

    Zuñiga-Aguilar, E; Olayo, R; Ramírez-Fernández, O; Morales, J; Godínez, R

    2014-01-01

    Currently, there are several techniques for modified cell culture surfaces under research to improve cell growth and adhesion. Recently, different methods have been used for surface coating, using biomolecules that enhance cell attachment and growth of nerve cells from spinal cord, such as the use of Poly-DL-Ornithine/Laminin. Plasma-polymerized pyrrole (PPy)-treated surfaces have showed improvement on surfaces biocompatibility with the cells in culture since they do not interfere with any of the biological cell functions. In the present work, we present a novel mouse nerve cell culture technique, using PPy-treated cell culture surfaces. A comparative study of cell survival using Poly-DL-Ornithine/Laminin-treated surfaces was performed. Our results of cell survival when compared with data already reported by other investigators, show that cells cultured on the PPy-modified surface increased survival up to 21 days when compared with Poly-DL-Ornithine/Laminin-coated culture, where 8 days cell survival was obtained. There were electrical and morphological differences in the nerve cells grown in the different surfaces. By comparing the peak ion currents of Poly-DL-Ornithine/Laminin-seeded cells for 8 days with cells grown for 21 days on PPy, an increase of 516% in the Na(+) current and 127% in K(+) currents in cells seeded on PPy were observed. Immunofluorescence techniques showed the presence of cell synapses and culture viability after 21 days. Our results then showed that PPy-modified surfaces are an alternative culture method that increases nerve cells survival from lumbar spinal cord cell culture by preserving its electrical and morphological features. PMID:24650203

  20. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor

    Science.gov (United States)

    Parks, Kelsey

    2009-01-01

    Space flight has been shown to alter the astronauts immune systems. Because immune performance is complex and reflects the influence of multiple organ systems within the host, scientists sought to understand the potential impact of microgravity alone on the cellular mechanisms critical to immunity. Lymphocytes and their differentiated immature form, lymphoblasts, play an important and integral role in the body's defense system. T cells, one of the three major types of lymphocytes, play a central role in cell-mediated immunity. They can be distinguished from other lymphocyte types, such as B cells and natural killer cells by the presence of a special receptor on their cell surface called T cell receptors. Reported studies have shown that spaceflight can affect the expression of cell surface markers. Cell surface markers play an important role in the ability of cells to interact and to pass signals between different cells of the same phenotype and cells of different phenotypes. Recent evidence suggests that cell-cycle regulators are essential for T-cell function. To trigger an effective immune response, lymphocytes must proliferate. The objective of this project is to investigate the changes in growth of human cells cultured in rotating bioreactors and to measure the growth rate and the cell cycle distribution for different human cell types. Human lymphocytes and lymphoblasts will be cultured in a bioreactor to simulate aspects of microgravity. The bioreactor is a cylindrical culture vessel that incorporates the aspects of clinostatic rotation of a solid fluid body around a horizontal axis at a constant speed, and compensates gravity by rotation and places cells within the fluid body into a sustained free-fall. Cell cycle progression and cell proliferation of the lymphocytes will be measured for a number of days. In addition, RNA from the cells will be isolated for expression of genes related in cell cycle regulations.

  1. Integrated biosensors for cell culture monitoring

    OpenAIRE

    De Micheli, Giovanni; Boero, Cristina; Olivo, Jacopo; Carrara, Sandro

    2014-01-01

    Biosensors for endogenous compounds, such as glucose and lactate, are applied to monitor cell cultures. Cells can be cultivated for several purposes, such as understanding and modeling some biological mechanisms, the development of new drugs and therapies, and in the field of regenerative medicine. We have realized a self-contained monitoring system with remote readout. Metabolite detection is based on oxidases immobilized onto carbon nanotubes. We calibrate the system for glucose and lactate...

  2. Characterisation and preservation of cultural heritage artefacts using nuclear techniques

    International Nuclear Information System (INIS)

    This report covers the studies performed for the identification and preservation of cultural heritage using nuclear analytical techniques (NAT). Within the context of the project financed by the IAEA, cultural articles from various excavation regions and from the Anatolian Civilizations Museum were analyzed and identified using the instruments at our Center and information was provided regarding their manufacturing techniques, past restoration history and socioeconomic indicators about the period within which these articles were used. The analysis of the articles which could not be removed from the museum were performed in-situ using portable instruments and support was provided to the experts for some articles from excavation regions for the evaluation of their originality. Within the framework the of this Project, five experts attended to the workshops and meetings organised by the IAEA and in the context of scientific visits and bilateral cooperation, one expert from Greece and three experts from Macedonia visited our Center and Anatolian Civilizations Museum and experimental studies were performed together

  3. Cell culture from sponges: pluripotency and immortality

    NARCIS (Netherlands)

    Caralt Bosch, de S.; Uriz, M.J.; Wijffels, R.H.

    2007-01-01

    Sponges are a source of compounds with potential pharmaceutical applications. In this article, methods of sponge cell culture for production of these bioactive compounds are reviewed, and new approaches for overcoming the problem of metabolite supply are examined. The use of embryos is proposed as a

  4. ANTHOCYANIN (ACN) STABILITY IN CELL CULTURE MEDIA

    Science.gov (United States)

    Anthocyanins (ACNs) are potential oxygen radical scavengers that have coronary vasoactive and vasoprotective properties. Cell or tissue culture systems have been used to examine the bioactivity and mechanisms of action of ACNs on the vascular system. However, due to their unique chemical structure, ...

  5. Cell Culture Microfluidic Biochips: Experimental Throughput Maximization

    DEFF Research Database (Denmark)

    Minhass, Wajid Hassan; Pop, Paul; Madsen, Jan;

    2011-01-01

    Microfluidic biochips offer a promising alternative to a conventional biochemical laboratory, integrating all necessary functionalities on-chip in order to perform biochemical applications. Researchers have started to propose computer-aided design tools for the synthesis of such biochips. Our focus...... Annealing metaheuristic for experimental design generation for the cell culture microfluidic biochips, and we have evaluated our approach using multiple experimental setups....

  6. Multiweek Cell Culture Project for Use in Upper-Level Biology Laboratories

    Science.gov (United States)

    Marion, Rebecca E.; Gardner, Grant E.; Parks, Lisa D.

    2012-01-01

    This article describes a laboratory protocol for a multiweek project piloted in a new upper-level biology laboratory (BIO 426) using cell culture techniques. Human embryonic kidney-293 cells were used, and several culture media and supplements were identified for students to design their own experiments. Treatments included amino acids, EGF,…

  7. Nanotechnology, Cell Culture and Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Kazutoshi Haraguchi

    2011-01-01

    Full Text Available We have fabricated new types of polymer hydrogels and polymer nanocomposites, i.e., nanocomposite gels (NC gels and soft, polymer nanocomposites (M-NCs: solid, with novel organic/inorganic network structures. Both NC gels and M-NCs were synthesized by in-situ free-radical polymerization in the presence of exfoliated clay platelets in aqueous systems and were obtained in various forms such as film, sheet, tube, coating, etc. and sizes with a wide range of clay contents. Here, disk-like inorganic clay nanoparticles act as multi-functional crosslinkers to form new types of network systems. Both NC gels and M-NCs have extraordinary optical and mechanical properties including ultra-high reversible extensibility, as well as a number of new characteristics relating to optical anisotropy, polymer/clay morphology, biocompatibility, stimuli-sensitive surfaces, micro-patterning, etc. For examples, the biological testing of medical devices, comprised of a sensitization test, an irritation test, an intracutaneous test and an in vitro cytotoxicity test,was carried out for NC gels and M-NCs. The safety of NC gels and M-NCs was confirmed in all tests. Also, the interaction of living tissue with NC gel was investigated in vivo by implantation in live goats; neither inflammation nor concrescence occurred around the NC gels. Furthermore, it was found that both N-NC gels consisting of poly(N-isopropylacrylamide(PNIPA/clay network and M-NCs consisting of poly(2-methoxyethyacrylate(PMEA/clay network show characteristic cell culture and subsequent cell detachment on their surfaces, although it was almost impossible to culture cells on conventional, chemically-crosslinked PNIPA hydrogels and chemically crossslinked PMEA, regardless of their crosslinker concentration. Various kinds of cells, such ashumanhepatoma cells (HepG2, normal human dermal fibroblast (NHDF, and human umbilical vein endothelial cells (HUVEC, could be cultured to be confluent on the surfaces of N

  8. Cell culture models using rat primary alveolar type I cells.

    Science.gov (United States)

    Downs, Charles A; Montgomery, David W; Merkle, Carrie J

    2011-10-01

    There is a lack of cell culture models using primary alveolar type I (AT I) cells. The purpose of this study was to develop cell culture models using rat AT I cells and microvascular endothelial cells from the lung (MVECL). Two types of model systems were developed: single and co-culture systems; additionally a 3-dimensional model system was developed. Pure AT I cell (96.3 ± 2.7%) and MVECL (97.9 ± 1.1%) preparations were used. AT I cell morphology, mitochondrial number and distribution, actin filament arrangement and number of apoptotic cells at confluence, and telomere attrition were characterized. AT I cells maintained their morphometric characteristics through at least population doubling (PD) 35, while demonstrating telomere attrition through at least PD 100. Furthermore, AT I cells maintained the expression of their specific markers, T1α and AQ-5, through PD 42. For the co-cultures, AT I cells were grown on the top and MVECL were grown on the bottom of fibronectin-coated 24-well Transwell Fluroblok™ filter inserts. Neither cell type transmigrated the 1 μm pores. Additionally, AT I cells were grown in a thick layer of Matrigel(®) to create a 3-dimensional model in which primary AT I cells form ring-like structures that resemble an alveolus. The development of these model systems offers the opportunities to investigate AT I cells and their interactions with MVECL in response to pharmacological interventions and in the processes of disease, repair and regeneration. PMID:21624488

  9. 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

  10. Dynamic cell culture system (7-IML-1)

    Science.gov (United States)

    Cogoli, Augusto

    1992-01-01

    This experiment is one of the Biorack experiments being flown on the International Microgravity Laboratory 1 (MIL-1) mission as part of an investigation studying cell proliferation and performance in space. One of the objectives of this investigation is to assess the potential benefits of bioprocessing in space with the ultimate goal of developing a bioreactor for continuous cell cultures in space. This experiment will test the operation of an automated culture chamber that was designed for use in a Bioreactor in space. The device to be tested is called the Dynamic Cell Culture System (DCCS). It is a simple device in which media are renewed or chemicals are injected automatically, by means of osmotic pumps. This experiment uses four Type I/O experiment containers. One DCCS unit, which contains a culture chamber with renewal of medium and a second chamber without a medium supply fits in each container. Two DCCS units are maintained under zero gravity conditions during the on-orbit period. The other two units are maintained under 1 gh conditions in a 1 g centrifuge. The schedule for incubator transfer is given.

  11. Advantages and challenges of microfluidic cell culture in polydimethylsiloxane devices

    OpenAIRE

    Halldórsson, Skarphédinn; Lucumi Moreno, Edinson; Gómez-Sjöberg, Rafael; Fleming, Ronan MT

    2015-01-01

    Culture of cells using various microfluidic devices is becoming more common within experimental cell biology. At the same time, a technological radiation of microfluidic cell culture device designs is currently in progress. Ultimately, the utility of microfluidic cell culture will be determined by its capacity to permit new insights into cellular function. Especially insights that would otherwise be difficult or impossible to obtain with macroscopic cell culture in traditional polystyrene dis...

  12. Improving tolerance to Fusarium oxysporum f. sp. melonis in melon using tissue culture and mutation techniques

    International Nuclear Information System (INIS)

    Fusarium wilt is a vascular disease of the Cucurbitaceae family, especially in muskmelon (Cucumis melo L.), caused by the soil fungus Fusarium oxysporum f. sp. melonis (FOM). This pathogen persists in the soil for extended periods of time, and the only effective control is the use of resistant cultivars. During the last three decades, tissue culture techniques have been utilised in crop improvement to generate changes in the genetic material of plants via in vitro somaclonal variations (by organogenesis or somatic embryogenesis) and induced mutagenesis. More recently, researchers have been using in vitro techniques to investigate the effects of fungal culture filtrates or toxins on susceptible and resistant genotypes of different plant species or cultivars to assess disease resistance. This method is effectively used for cucumber and melon. There are various in vitro culture techniques that can be used for cucumber (Malepszy, 1988). In this chapter, we show a method for mass-selection of melon mutants resistant to Fusarium wilt. In vitro selection of resistant cells, from both irradiated and non- irradiated explants, is performed using culture filtrates of different FOM races. This research could lead to the development of new melon cultivars resistant to Fusarium wilt. (author)

  13. Hybridoma cell behaviour in continuous culture under hyperosmotic stress.

    Science.gov (United States)

    Cherlet, M; Marc, A

    1999-01-01

    In this paper, we propose an alternative strategy to the ones proposed before (Oh et al., 1993; Øyaas et al., 1994a) to get real increases of global final antibody titer and production at hyperosmotic stress, by reducing the detrimental effect of such a stress on cell growth, and conserving the stimulating effect on antibody production. It consists of cultivating the cells in continuous culture and increasing the osmolality stepwise. In this way, the cells could progressively adapt to the higher osmolality at each step and antibody titers could be nearly doubled at 370 and 400 mOsm kg-1, compared to the standard osmolality of 335 mOsm kg-1. Surprisingly, the stimulation of antibody production was not confirmed for higher osmolalities, 425 and 450 mOsm kg- 1, despite the minor negative effect on cell growth. Intracellular IgG analysis by flow cytometry revealed at these osmolalities a significant population of non-producing cells. However, even when taking into account this non-producing population, a stimulating effect on antibody production could not be shown at these highest osmolalities. It seems to us that osmolality has a significant effect on the appearance of these non-producing cells, since they were not observed in continuous cultures at standard osmolality, of comparable duration and at an even higher dilution rate. The appearance of the non-producing cells coincides furthermore with modifications of the synthesised antibody, as shown by electrophoretic techniques. It is however not really clear if these two observations reflect actually the same phenomenon. Hyperosmolality affects the cell behaviour in continuous culture in multiple ways, independently of the growth rate, counting all at least partially for the observed stimulation of antibody production: acceleration of the amino acid, and in particular the glutamine metabolism, increase of the cell volume, increase of the intracellular pH and accumulation of cells in the G1 cell cycle phase. PMID

  14. Muskmelon embryo rescue techniques using in vitro embryo culture.

    Science.gov (United States)

    Nuñez-Palenius, Hector Gordon; Ramírez-Malagón, Rafael; Ochoa-Alejo, Neftalí

    2011-01-01

    Among the major cucurbit vegetables, melon (Cucumis melo) has one of the greatest polymorphic fruit types and botanical varieties. Some melon fruits have excellent aroma, variety of flesh colors, deeper flavor, and more juice compared to other cucurbits. Despite numerous available melon cultivars, some of them are exceedingly susceptible to several diseases. The genetic background carrying the genes for tolerance and/or resistance for those diseases is found in wild melon landraces. Unfortunately, the commercial melon varieties are not able to produce viable hybrids when crossed with their wild melon counterparts. Plant tissue culture techniques are needed to surpass those genetic barriers. In vitro melon embryo rescue has played a main role to obtain viable hybrids originated from commercial versus wild melon crosses. In this chapter, an efficient and simple embryo rescue melon protocol is thoroughly described. PMID:21207265

  15. Computational Swarming: A Cultural Technique for Generative Architecture

    Directory of Open Access Journals (Sweden)

    Sebastian Vehlken

    2014-11-01

    Full Text Available After a first wave of digital architecture in the 1990s, the last decade saw some approaches where agent-based modelling and simulation (ABM was used for generative strategies in architectural design. By taking advantage of the self-organisational capabilities of computational agent collectives whose global behaviour emerges from the local interaction of a large number of relatively simple individuals (as it does, for instance, in animal swarms, architects are able to understand buildings and urbanscapes in a novel way as complex spaces that are constituted by the movement of multiple material and informational elements. As a major, zoo-technological branch of ABM, Computational Swarm Intelligence (SI coalesces all kinds of architectural elements – materials, people, environmental forces, traffic dynamics, etc. – into a collective population. Thereby, SI and ABM initiate a shift from geometric or parametric planning to time-based and less prescriptive software tools.Agent-based applications of this sort are used to model solution strategies in a number of areas where opaque and complex problems present themselves – from epidemiology to logistics, and from market simulations to crowd control. This article seeks to conceptualise SI and ABM as a fundamental and novel cultural technique for governing dynamic processes, taking their employment in generative architectural design as a concrete example. In order to avoid a rather conventional application of philosophical theories to this field, the paper explores how the procedures of such technologies can be understood in relation to the media-historical concept of Cultural Techniques.

  16. Protection of the cultural environment by nuclear techniques

    International Nuclear Information System (INIS)

    The use of gamma radiation in the preservation of the cultural environment, and in particular art objects, is examined. In each branch, the present status of research on, and its application to, the treatment of art objects is described. Two major applications - disinfection and consolidation - are reviewed: (1) Disinfection: The destruction by gamma irradiation of xylophagous insects and of fungi which contaminate paintings, pictorial documents and mummies, is discussed. The effect of gamma-rays on the constituent materials of the infected objects is examined. (2) Consolidation: this process represents an extrapolation to the preservation of cultural objects of research performed over the last 20 years with the aim of improving the mechanical properties of porous materials (by impregnation of the material in a vacuum by a liquid plastic substance which is polymerized by radiation in situ). This technique is currently being used to treat ''uncoated'' wooden objects and, to a more limited extent, ''coated'' wood (polychromy and gilt wood), depending on the fragility of the covering. With regard to stone materials, the enhancement of the mechanical properties of a limestone impregnated with a styrene-polyester resin is described; this resin has been used to treat several sculptures. The use of acrylics and methacrylics has caused some samples to crack during radiation polymerization; theories put forward to explain this phenomenon are discussed. The consolidation of terracotta and bone has also been studied. The consolidation of waterlogged wood (archaeological wood) requires a different impregnation technique (diffusion in liquid). This preservation method has already been used to treat archaeological artifacts. (author)

  17. Cell culture: Progenitor cells from human brain after death

    Science.gov (United States)

    Palmer, Theo D.; Schwartz, Philip H.; Taupin, Philippe; Kaspar, Brian; Stein, Stuart A.; Gage, Fred H.

    2001-05-01

    Culturing neural progenitor cells from the adult rodent brain has become routine and is also possible from human fetal tissue, but expansion of these cells from postnatal and adult human tissue, although preferred for ethical reasons, has encountered problems. Here we describe the isolation and successful propagation of neural progenitor cells from human postmortem tissues and surgical specimens. Although the relative therapeutic merits of adult and fetal progenitor cells still need to be assessed, our results may extend the application of these progenitor cells in the treatment of neurodegenerative diseases.

  18. [Wound treatment with autogenous epidermal cell expansion culture].

    Science.gov (United States)

    Bonnekoh, B; Müller, R P; Mahrle, G; Steigleder, G K

    1988-11-11

    Sheets of autologous epidermal cells grown by expansion culture were used to cover small skin defects in seven patients with postoperative necroses, necroses due to temporal arteritis, varicose ulcers or after tangential excision of tattoos. Several transplantation techniques were used: backing of the cultured epithelia with vaseline gauze, Surfasoft, Adaptic, Silastic foil, culturing directly from Petriperm-foil. Meshed Silastic-foil proved to give the best support. Optimal take of the in-vitro epithelia (more than 80% of their surface area) was achieved only for fresh dermal wound-beds. The take was only moderate on chronic granulation tissue, but the transplants reduced the formation of fibrinous-necrotic material and favoured the formation of fresh granulation tissue. PMID:3181024

  19. Vegetal cell cultures under the effect of low intensity microwaves

    International Nuclear Information System (INIS)

    In order to study the microwave effect on the chlorophyll biosynthesis in Papaver rhoes in vitro cultures microwave radiation characterized by a power density of 0.9 mW/cm2 in a frequency range of 9. 75 - 10.75 GHz was used. P. rhoes in vitro cultures, were obtained from explants of leaves and flowers provided by adult individuals, grown in the Botanical garden of AL. I. Cuza University from Iasi. Murashige Skoog agarized medium with a suitable hormone balance was used to conduct cell culture development before as well as after exposure to microwaves. Assimilatory pigment levels (chlorophyll a, chlorophyll b and carotene pigments) have been evaluated by standard spectrophotometric technique. Student t-test (two tailed, pair) gave significant p-values for the modification of chlorophyll a and chlorophyll b levels after microwave treatment (p0.05). Microwave treatment seems to be able to stimulate assimilatory pigment biosynthesis in the vegetal cell cultures. An inhibitory effect may be associated to the phenotypic modifications noticed in the callus growth from 2 exposed vials - probably related to a non-thermal effect of microwaves in living tissues. Further study of sub-cultures derived from exposed vials is needed to clarify if microwaves of low power density are adequate for the stimulation of assimilatory pigment from chloroplast membranes. (authors)

  20. Mouse cell culture - Methods and protocols

    Directory of Open Access Journals (Sweden)

    CarloAlberto Redi

    2010-12-01

    Full Text Available The mouse is, out of any doubt, the experimental animal par excellence for many many colleagues within the scientific community, notably for those working in mammalian biology (in a broad sense, from basic genetic to modeling human diseases, starting at least from 1664 Robert Hooke experiments on air’s propertyn. Not surprising then that mouse cell cultures is a well established field of research itself and that there are several handbooks devoted to this discipline. Here, Andrew Ward and David Tosh provide a necessary update of the protocols currently needed. In fact, nearly half of the book is devoted to stem cells culture protocols, mainly embryonic, from a list of several organs (kidney, lung, oesophagus and intestine, pancreas and liver to mention some........

  1. Differentiation of mammalian skeletal muscle cells cultured on microcarrier beads in a rotating cell culture system

    Science.gov (United States)

    Torgan, C. E.; Burge, S. S.; Collinsworth, A. M.; Truskey, G. A.; Kraus, W. E.

    2000-01-01

    The growth and repair of adult skeletal muscle are due in part to activation of muscle precursor cells, commonly known as satellite cells or myoblasts. These cells are responsive to a variety of environmental cues, including mechanical stimuli. The overall goal of the research is to examine the role of mechanical signalling mechanisms in muscle growth and plasticity through utilisation of cell culture systems where other potential signalling pathways (i.e. chemical and electrical stimuli) are controlled. To explore the effects of decreased mechanical loading on muscle differentiation, mammalian myoblasts are cultured in a bioreactor (rotating cell culture system), a model that has been utilised to simulate microgravity. C2C12 murine myoblasts are cultured on microcarrier beads in a bioreactor and followed throughout differentiation as they form a network of multinucleated myotubes. In comparison with three-dimensional control cultures that consist of myoblasts cultured on microcarrier beads in teflon bags, myoblasts cultured in the bioreactor exhibit an attenuation in differentiation. This is demonstrated by reduced immunohistochemical staining for myogenin and alpha-actinin. Western analysis shows a decrease, in bioreactor cultures compared with control cultures, in levels of the contractile proteins myosin (47% decrease, p < 0.01) and tropomyosin (63% decrease, p < 0.01). Hydrodynamic measurements indicate that the decrease in differentiation may be due, at least in part, to fluid stresses acting on the myotubes. In addition, constraints on aggregate size imposed by the action of fluid forces in the bioreactor affect differentiation. These results may have implications for muscle growth and repair during spaceflight.

  2. Analysis of Culture-Dependent versus Culture-Independent Techniques for Identification of Bacteria in Clinically Obtained Bronchoalveolar Lavage Fluid

    OpenAIRE

    Dickson, Robert P; Erb-Downward, John R.; Prescott, Hallie C.; Martinez, Fernando J; Curtis, Jeffrey L.; Lama, Vibha N.; Huffnagle, Gary B.

    2014-01-01

    The diagnosis and management of pneumonia are limited by the use of culture-based techniques of microbial identification, which may fail to identify unculturable, fastidious, and metabolically active viable but unculturable bacteria. Novel high-throughput culture-independent techniques hold promise but have not been systematically compared to conventional culture. We analyzed 46 clinically obtained bronchoalveolar lavage (BAL) fluid specimens from symptomatic and asymptomatic lung transplant ...

  3. Darwinian Evolution of Prions in Cell Culture*

    OpenAIRE

    Li, Jiali; Browning, Shawn; Mahal, Sukhvir P.; Oelschlegel, Anja M.; Weissmann, Charles

    2009-01-01

    Prions are infectious proteins consisting mainly of PrPSc, a β sheet-rich conformer of the normal host protein PrPC, and occur in different strains. Strain identity is thought to be encoded by PrPSc conformation. We found that biologically cloned prion populations gradually became heterogeneous by accumulating “mutants”, and selective pressures resulted in the emergence of different mutants as major constituents of the evolving population. Thus, when transferred from brain to cultured cells, ...

  4. Bioactive sugar surfaces for hepatocyte cell culture

    OpenAIRE

    Ambury, Rachael

    2010-01-01

    The primary objective of this study was to identify, develop and characterise a novel bioactive surface capable of binding hepatocytes and enabling the retention of hepatocyte-specific cell function during in-vitro culture. The materials were designed to exploit a unique characteristic of hepatocyte biology, with β-galactose moieties displayed to allow cellular adhesion via the specific asialoglycoprotein receptors (ASGP-R) found on hepatocytes. Hydrogels were created by modifying a commercia...

  5. Degradation of TNT by plant cell cultures

    Czech Academy of Sciences Publication Activity Database

    Podlipná, Radka; Nepovím, Aleš; Zeman, S.; Vágner, Martin; Vaněk, Tomáš

    Smolenice, 2003, s. 78-79. [Xenobiochemické sympózium /22./. Smolenice (SK), 09.06.2003-11.06.2003] R&D Projects: GA ČR GP206/02/P065; GA MŠk OC 837.10 Institutional research plan: CEZ:AV0Z5038910; CEZ:AV0Z4055905 Keywords : degradation * plant cell cultures Subject RIV: DK - Soil Contamination ; De-contamination incl. Pesticides

  6. Prevention and Detection of Mycoplasma Contamination in Cell Culture

    OpenAIRE

    Parvaneh Farzaneh; Laleh Nikfarjam

    2011-01-01

    One of the main problems in cell culture is mycoplasma infection. It can extensively affect cell physiology and metabolism. As the applications of cell culture increase in research, industrial production and cell therapy, more concerns about mycoplasma contamination and detection will arise. This review will provide valuable information about: 1. the ways in which cells are contaminated and the frequency and source of mycoplasma species in cell culture; 2. the ways to prevent mycoplasma conta...

  7. Production of putative virulence factors by Renibacterium salmoninarum grown in cell culture.

    Science.gov (United States)

    McIntosh, D; Flaño, E; Grayson, T H; Gilpin, M L; Austin, B; Villena, A J

    1997-10-01

    A cell culture system, employing the fish cell line Epithelioma papillosum cyprini (EPC), was developed to study the synthesis of intracellular antigen and the expression of putative virulence factors by Renibacterium salmoninarum. EPC cultures infected with R. salmoninarum could be maintained for 7 weeks, during which the pathogen multiplied intracellularly. Immunohistochemical examination of infected cultures revealed the production of the p57 antigen, haemolysin and cytolysin. The intracellular nature of the infection was confirmed by transmission electron microscopic examination of EPC monolayers. A comparison of the relative virulence of bacterial cells cultured in EPC cells and on agar plates revealed that the former were markedly more virulent in challenge experiments with juvenile rainbow trout (Oncorhynchus mykiss Walbaum). The EPC cell culture model provided a system for the study of R. salmoninarum under more natural conditions than those achieved with plate culture techniques. PMID:9353936

  8. A Versatile Bioreactor for Dynamic Suspension Cell Culture. Application to the Culture of Cancer Cell Spheroids

    Science.gov (United States)

    Madeddu, Denise; Cerino, Giulia; Falco, Angela; Frati, Caterina; Gallo, Diego; Deriu, Marco A.; Falvo D’Urso Labate, Giuseppe; Quaini, Federico; Audenino, Alberto; Morbiducci, Umberto

    2016-01-01

    A versatile bioreactor suitable for dynamic suspension cell culture under tunable shear stress conditions has been developed and preliminarily tested culturing cancer cell spheroids. By adopting simple technological solutions and avoiding rotating components, the bioreactor exploits the laminar hydrodynamics establishing within the culture chamber enabling dynamic cell suspension in an environment favourable to mass transport, under a wide range of tunable shear stress conditions. The design phase of the device has been supported by multiphysics modelling and has provided a comprehensive analysis of the operating principles of the bioreactor. Moreover, an explanatory example is herein presented with multiphysics simulations used to set the proper bioreactor operating conditions for preliminary in vitro biological tests on a human lung carcinoma cell line. The biological results demonstrate that the ultralow shear dynamic suspension provided by the device is beneficial for culturing cancer cell spheroids. In comparison to the static suspension control, dynamic cell suspension preserves morphological features, promotes intercellular connection, increases spheroid size (2.4-fold increase) and number of cycling cells (1.58-fold increase), and reduces double strand DNA damage (1.5-fold reduction). It is envisioned that the versatility of this bioreactor could allow investigation and expansion of different cell types in the future. PMID:27144306

  9. A Versatile Bioreactor for Dynamic Suspension Cell Culture. Application to the Culture of Cancer Cell Spheroids.

    Science.gov (United States)

    Massai, Diana; Isu, Giuseppe; Madeddu, Denise; Cerino, Giulia; Falco, Angela; Frati, Caterina; Gallo, Diego; Deriu, Marco A; Falvo D'Urso Labate, Giuseppe; Quaini, Federico; Audenino, Alberto; Morbiducci, Umberto

    2016-01-01

    A versatile bioreactor suitable for dynamic suspension cell culture under tunable shear stress conditions has been developed and preliminarily tested culturing cancer cell spheroids. By adopting simple technological solutions and avoiding rotating components, the bioreactor exploits the laminar hydrodynamics establishing within the culture chamber enabling dynamic cell suspension in an environment favourable to mass transport, under a wide range of tunable shear stress conditions. The design phase of the device has been supported by multiphysics modelling and has provided a comprehensive analysis of the operating principles of the bioreactor. Moreover, an explanatory example is herein presented with multiphysics simulations used to set the proper bioreactor operating conditions for preliminary in vitro biological tests on a human lung carcinoma cell line. The biological results demonstrate that the ultralow shear dynamic suspension provided by the device is beneficial for culturing cancer cell spheroids. In comparison to the static suspension control, dynamic cell suspension preserves morphological features, promotes intercellular connection, increases spheroid size (2.4-fold increase) and number of cycling cells (1.58-fold increase), and reduces double strand DNA damage (1.5-fold reduction). It is envisioned that the versatility of this bioreactor could allow investigation and expansion of different cell types in the future. PMID:27144306

  10. Reversible gelling culture media for in-vitro cell culture in three-dimensional matrices

    Science.gov (United States)

    An, Yuehuei H.; Mironov, Vladimir A.; Gutowska, Anna

    2000-01-01

    A gelling cell culture medium useful for forming a three dimensional matrix for cell culture in vitro is prepared by copolymerizing an acrylamide derivative with a hydrophilic comonomer to form a reversible (preferably thermally reversible) gelling linear random copolymer in the form of a plurality of linear chains having a plurality of molecular weights greater than or equal to a minimum gelling molecular weight cutoff, mixing the copolymer with an aqueous solvent to form a reversible gelling solution and adding a cell culture medium to the gelling solution to form the gelling cell culture medium. Cells such as chondrocytes or hepatocytes are added to the culture medium to form a seeded culture medium, and temperature of the medium is raised to gel the seeded culture medium and form a three dimensional matrix containing the cells. After propagating the cells in the matrix, the cells may be recovered by lowering the temperature to dissolve the matrix and centrifuging.

  11. Handbook of plant cell culture. Volume 2. Crop species

    Energy Technology Data Exchange (ETDEWEB)

    Sharp, W.R.; Evans, D.A.; Ammirato, P.V.; Yamada, Y. (eds.)

    1984-01-01

    In this volume the state-of-the-art plant cell culture techniques described in the first volume are applied to several agricultural and horticultural crops. In 21 chapters, they include maize, oats, wheat, beans, red clover and other forage legumes, asparagus, celery, cassava, sweet potato, banana, pawpaw, apple, grapes, conifers, date palm, rubber, sugarcane and tobacco. Each chapter contains (1) detailed protocols to serve as the foundation for current research, (2) a critical review of the literature, and (3) in-depth evaluations of the potential shown by plant cell culture for crop improvement. The history and economic importance of each crop are discussed. This volume also includes an essay, ''Oil from plants'', by M. Calvin.

  12. Development of a pneumatically driven active cover lid for multi-well microplates for use in perfusion three-dimensional cell culture

    OpenAIRE

    Song-Bin Huang; Dean Chou; Yu-Han Chang; Ke-Cing Li; Tzu-Keng Chiu; Yiannis Ventikos; Min-Hsien Wu

    2015-01-01

    Before microfluidic-based cell culture models can be practically utilized for bioassays, there is a need for a transitional cell culture technique that can improve conventional cell culture models. To address this, a hybrid cell culture system integrating an active cover lid and a multi-well microplate was proposed to achieve perfusion 3-D cell culture. In this system, a microfluidic-based pneumatically-driven liquid transport mechanism was integrated into the active cover lid to realize 6-un...

  13. Sodium 22+ washout from cultured rat cells

    International Nuclear Information System (INIS)

    The washout of Na+ isotopes from tissues and cells is quite complex and not well defined. To further gain insight into this process, we have studied 22Na+ washout from cultured Wistar rat skin fibroblasts and vascular smooth muscle cells (VSMCs). In these preparations, 22Na+ washout is described by a general three-exponential function. The exponential factor of the fastest component (k1) and the initial exchange rate constant (kie) of cultured fibroblasts decrease in magnitude in response to incubation in K+-deficient medium or in the presence of ouabain and increase in magnitude when the cells are incubated in a Ca++-deficient medium. As the magnitude of the kie declines (in the presence of ouabain) to the level of the exponential factor of the middle component (k2), 22Na+ washout is adequately described by a two-exponential function. When the kie is further diminished (in the presence of both ouabain and phloretin) to the range of the exponential factor of the slowest component (k3), the washout of 22Na+ is apparently monoexponential. Calculations of the cellular Na+ concentrations, based on the 22Na+ activity in the cells at the initiation of the washout experiments, and the medium specific activity agree with atomic absorption spectrometry measurements of the cellular concentration of this ion. Thus, all three components of 22Na+ washout from cultured rat cells are of cellular origin. Using the exponential parameters, compartmental analyses of two models (in parallel and in series) with three cellular Na+ pools were performed. The results indicate that, independent of the model chosen, the relative size of the largest Na+ pool is 92-93% in fibroblasts and approximately 96% in VSMCs. This pool is most likely to represent the cytosol

  14. Cell response of Chlamydomonas actinochloris culture to repeated microwave irradiation

    Directory of Open Access Journals (Sweden)

    OLESIA O. GRYGORIEVA

    2015-05-01

    Full Text Available Abstract. Grygorieva OO, Berezovsjka MA, Dacenko OI. 2015. Cell response of Chlamydomonas actinochloris culture to repeated microwave irradiation. Nusantara Bioscience 7: 38-42. Two cultures of Chlamydomonas actinochloris Deason et Bold in the lag-phase were exposed to the microwave irradiation. One of them (culture 1 was not treated beforehand, whereas the other (culture 2 was irradiated by microwaves 2 years earlier. The measurement of cell quantity as well as measurement of change of intensities and spectra of cultures photoluminescence (PL in the range of chlorophyll a emission was regularly conducted during the cell cultures development. Cell concentration of culture 1 exposed to the microwave irradiation for the first time has quickly restored while cell concentration of culture 2 which was irradiated repeatedly has fallen significantly. The following increasing of cell concentration of culture 2 is negligible. Cell concentration reaches the steady-state level that is about a half of the cell concentration of control culture. Initially the PL efficiency of cells of both cultures decreases noticeable as a result of irradiation. Then there is the monotonic increase to the values which are significantly higher than the corresponding values in the control cultures. The ratio of the intensities at the maxima of the main emission bands of chlorophyll for control samples of both cultures remained approximately at the same level. At the same time effect of irradiation on the cell PL spectrum appears as a temporary reduction of this magnitude.

  15. The cell-surface proteome of cultured adipose stromal cells.

    Science.gov (United States)

    Donnenberg, Albert D; Meyer, E Michael; Rubin, J Peter; Donnenberg, Vera S

    2015-07-01

    In this technical note we describe a method to evaluate the cell surface proteome of human primary cell cultures and cell lines. The method utilizes the BD Biosciences lyoplate, a system covering 242 surface proteins, glycoproteins, and glycosphingolipids plus relevant isotype controls, automated plate-based flow cytometry, conventional file-level analysis and unsupervised K-means clustering of markers on the basis of percent of positive events and mean fluorescence intensity of positive and total clean events. As an example, we determined the cell surface proteome of cultured adipose stromal cells (ASC) derived from 5 independent clinical isolates. Between-sample agreement of very strongly expressed (n = 32) and strongly expressed (n =16) markers was excellent, constituting a reliable profile for ASC identification and determination of functional properties. Known mesenchymal markers (CD29, CD44, CD73, CD90, CD105) were among the identified strongly expressed determinants. Among other strongly expressed markers are several that are potentially immunomodulatory including three proteins that protect from complement mediated effects (CD46, CD55, and CD59), two that regulate apoptosis (CD77 and CD95) and several with ectoenzymatic (CD10, CD26, CD13, CD73, and CD143) or receptor tyrosine kinase (CD140b (PDGFR), CD340 (Her-2), EGFR) activity, suggesting mechanisms for the anti-inflammatory and tissue remodeling properties of ASC. Because variables are standardized for K-means clustering, results generated using this methodology should be comparable between instrumentation platforms. It is widely generalizable to human primary explant cultures and cells lines and will prove useful to determine how cell passage, culture interventions, and gene expression and silencing affect the cell-surface proteome. PMID:25929697

  16. Cell Formation Techniques – A Study

    Directory of Open Access Journals (Sweden)

    Pawan Kumar Arora,

    2011-02-01

    Full Text Available Cellular manufacturing system has been proved a vital approach for batch and job shop production systems. Group technology has been an essential tool for developing a cellular manufacturing system. Thepaper aims to discuss various cell formation techniques and highlights the significant research work done in past over the years and attempts to points out the gap in research of past studies.

  17. Development of Scalable Culture Systems for Human Embryonic Stem Cells

    OpenAIRE

    Azarin, Samira M.; Palecek, Sean P.

    2010-01-01

    The use of human pluripotent stem cells, including embryonic and induced pluripotent stem cells, in therapeutic applications will require the development of robust, scalable culture technologies for undifferentiated cells. Advances made in large-scale cultures of other mammalian cells will facilitate expansion of undifferentiated human embryonic stem cells (hESCs), but challenges specific to hESCs will also have to be addressed, including development of defined, humanized culture media and su...

  18. Rotating bio-reactor cell culture apparatus

    Science.gov (United States)

    Schwarz, Ray P. (Inventor); Wolf, David A. (Inventor)

    1991-01-01

    A bioreactor system is described in which a tubular housing contains an internal circularly disposed set of blade members and a central tubular filter all mounted for rotation about a common horizontal axis and each having independent rotational support and rotational drive mechanisms. The housing, blade members and filter preferably are driven at a constant slow speed for placing a fluid culture medium with discrete microbeads and cell cultures in a discrete spatial suspension in the housing. Replacement fluid medium is symmetrically input and fluid medium is symmetrically output from the housing where the input and the output are part of a loop providing a constant or intermittent flow of fluid medium in a closed loop.

  19. PHYTOCHEMICAL STUDY OF CELL CULTURE JATROPHA CURCAS

    Directory of Open Access Journals (Sweden)

    KOMAR RUSLAN

    2011-01-01

    Full Text Available Jatropha curcas belongs to the Euphorbiaceae family which has potential economically. This plant has been reported to contain toxic compounds such as curcin and phorbol ester and its derivatives. These compounds may become a problem if J. curcas will be explored as a source of biofuel. In order to provide safety plants, the research on the study of phytochemical and initiation of cell and organ culture have been carried out. J curcas which has been collected from different regions in Indonesia showed to contain relatively the same profile of chemical contents. Dominant compounds that were detected by GCMS are hidrocarbon such as 2-heptenal, decadienal, hexsadecane, pentadecane, cyclooctane etc, fatty acid such as oktadecanoate acid, etthyl linoleate, ethyl stearate, heksadecanoate acid and steroid such as stigmasterol, fucosterol, sitosterol. No phorbol ester and its derivatives have been detected yet by the GCMS method. Callus and suspension cultures of J. curcas have been established to be used for further investigation.

  20. The Effect of Spaceflight on Bone Cell Cultures

    Science.gov (United States)

    Landis, William J.

    1999-01-01

    Understanding the response of bone to mechanical loading (unloading) is extremely important in defining the means of adaptation of the body to a variety of environmental conditions such as during heightened physical activity or in extended explorations of space or the sea floor. The mechanisms of the adaptive response of bone are not well defined, but undoubtedly they involve changes occurring at the cellular level of bone structure. This proposal has intended to examine the hypothesis that the loading (unloading) response of bone is mediated by specific cells through modifications of their activity cytoskeletal elements, and/or elaboration of their extracellular matrices. For this purpose, this laboratory has utilized the results of a number of previous studies defining molecular biological, biochemical, morphological, and ultrastructural events of the reproducible mineralization of a primary bone cell (osteoblast) culture system under normal loading (1G gravity level). These data and the culture system then were examined following the use of the cultures in two NASA shuttle flights, STS-59 and STS-63. The cells collected from each of the flights were compared to respective synchronous ground (1G) control cells examined as the flight samples were simultaneously analyzed and to other control cells maintained at 1G until the time of shuttle launch, at which point they were terminated and studied (defined as basal cells). Each of the cell cultures was assayed in terms of metabolic markers- gene expression; synthesis and secretion of collagen and non-collagenous proteins, including certain cytoskeletal components; assembly of collagen into macrostructural arrays- formation of mineral; and interaction of collagen and mineral crystals during calcification of the cultures. The work has utilized a combination of biochemical techniques (radiolabeling, electrophoresis, fluorography, Western and Northern Blotting, and light microscopic immunofluorescence) and structural

  1. IN VITRO CELL CULTURE AND HORMONE RADIOIMMUNOASSAY OF HUAMAN PITUITARY ADENOMAS

    Institute of Scientific and Technical Information of China (English)

    陆汉魁; 林祥通; 等

    1994-01-01

    Tissues from 30 human pituitary adenomas are monolayer-cell-cultured in vitro.Hormone secretion of GH,PRL,TSH,LH and FSH by cells into medium is detected by radioimmunoassay .The pattern and amount of hormone(s0 in the medium are used to determine the nature of the cells and thus to establish functional classification of pituitary adenomas.The results show that cell culture technique provides and easy and suitable mode for investigating the nature of pituitary adenomas.Hormone radioimmunoassay of culture medium is precise and reliable and represents the whole adenoma tissue.Further studies can lead to clearer understandngs of the pathology of pituitary adenomas.

  2. An introduction to plant cell culture: the future ahead.

    Science.gov (United States)

    Loyola-Vargas, Víctor M; Ochoa-Alejo, Neftalí

    2012-01-01

    Plant cell, tissue, and organ culture (PTC) techniques were developed and established as an experimental necessity for solving important fundamental questions in plant biology, but they currently represent very useful biotechnological tools for a series of important applications such as commercial micropropagation of different plant species, generation of disease-free plant materials, production of haploid and doublehaploid plants, induction of epigenetic or genetic variation for the isolation of variant plants, obtention of novel hybrid plants through the rescue of hybrid embryos or somatic cell fusion from intra- or intergeneric sources, conservation of valuable plant germplasm, and is the keystone for genetic engineering of plants to produce disease and pest resistant varieties, to engineer metabolic pathways with the aim of producing specific secondary metabolites or as an alternative for biopharming. Some other miscellaneous applications involve the utilization of in vitro cultures to test toxic compounds and the possibilities of removing them (bioremediation), interaction of root cultures with nematodes or mycorrhiza, or the use of shoot cultures to maintain plant viruses. With the increased worldwide demand for biofuels, it seems that PTC will certainly be fundamental for engineering different plants species in order to increase the diversity of biofuel options, lower the price marketing, and enhance the production efficiency. Several aspects and applications of PTC such as those mentioned above are the focus of this edition. PMID:22610615

  3. Renotropic stimulation in rat kidney cell culture

    International Nuclear Information System (INIS)

    A circulating renotropic factor specific for renal cells has been described in rats. The addition of sera obtained from unilaterally nephrectomized (uni) rats 24h after operation compared to sham-operated (sham) rats augments 3H-thymidine incorporation into the DNA of incubating kidney slices approximately 10% - 30%. Attempting to amplify the sensitivity of the assay for this renotropic agent, the authors replaced slices with primary rat kidney cultures. The assay system was based on one previously used for rabbits. The cultured cells were synchronized in their growth phase by a period of protein-free starvation. Compared to sera from sham rats, sera from uni rats showed significant stimulation of thymidine incorporation into DNA, 35.5% +/- 9.3 (SEM), p < .0001, at 16 h; 63.3% +/- 10.0 (SEM), p < .001, at 24 h; and 19.5% +/- 6.5 (SEM), p < .01, at 48 h post operation. Accordingly, the maximal stimulation at 24 h was greater than that previously found using the kidney slice assay. Measurable renotropic activity occurred earlier and over a shorter duration than in rabbits. Stimulation was similar when a D-valine medium, relatively specific for renal epithelial cells, replaced DME medium

  4. Optical Oxygen Sensors for Applications in Microfluidic Cell Culture

    OpenAIRE

    Grist, Samantha M.; Lukas Chrostowski; Cheung, Karen C.

    2010-01-01

    The presence and concentration of oxygen in biological systems has a large impact on the behavior and viability of many types of cells, including the differentiation of stem cells or the growth of tumor cells. As a result, the integration of oxygen sensors within cell culture environments presents a powerful tool for quantifying the effects of oxygen concentrations on cell behavior, cell viability, and drug effectiveness. Because microfluidic cell culture environments are a promising alternat...

  5. Cardiac Cells Beating in Culture: A Laboratory Exercise

    Science.gov (United States)

    Weaver, Debora

    2007-01-01

    This article describes how to establish a primary tissue culture, where cells are taken directly from an organ of a living animal. Cardiac cells are taken from chick embryos and transferred to culture dishes. These cells are not transformed and therefore have a limited life span. However, the unique characteristics of cardiac cells are maintained…

  6. Growth of cultured porcine retinal pigment epithelial cells

    DEFF Research Database (Denmark)

    Wiencke, A.K.; Kiilgaard, Jens Folke; Nicolini, Jair;

    2003-01-01

    To establish and characterize cultures of porcine retinal pigment epithelial (pRPE) cells in order to produce confluent monolayers of cells for transplantation.......To establish and characterize cultures of porcine retinal pigment epithelial (pRPE) cells in order to produce confluent monolayers of cells for transplantation....

  7. Equipment for large-scale mammalian cell culture.

    Science.gov (United States)

    Ozturk, Sadettin S

    2014-01-01

    This chapter provides information on commonly used equipment in industrial mammalian cell culture, with an emphasis on bioreactors. The actual equipment used in the cell culture process can vary from one company to another, but the main steps remain the same. The process involves expansion of cells in seed train and inoculation train processes followed by cultivation of cells in a production bioreactor. Process and equipment options for each stage of the cell culture process are introduced and examples are provided. Finally, the use of disposables during seed train and cell culture production is discussed. PMID:24429549

  8. An Important Method in the Investigation of Vascular Pathologies: Endothelial Cell Culture

    Directory of Open Access Journals (Sweden)

    Yusufhan Yazır

    2012-12-01

    Full Text Available Endothelial cells line the interior surface of blood vessels and form an interface between circulating blood in the lumen and the rest of the vessel wall. Endothelial cells are involved in many aspects of vascular biology, including barrier function, vasoconstriction, coagulation and inflamation. The endothelial cells in different organs have different functions and surface phenotype. These cells express prostoglandin-I2, platelet activating factor, collagen, endothelin-1, laminin, fibronectin and growth factors including platelet derived growth factor, fibroblast growth factor. İn the cell culture, cells can be isolated, maintened and proliferate in the laboratory conditions. The techniques of the cell culture have allowed scientists to use the cells in vitro for experimental studies, such as the production of vaccine, antibody and enzime, drug research, cell-cell interactions. Human umbilical vein endothelial cell is a good source for endothelial cell, because it is cheaper, easy to find and has the basic features of the normal endothelial cells.

  9. Effects of ultraviolet irradiation on cultured B-16 melanoma cells

    International Nuclear Information System (INIS)

    Changes of the plasma membrane during exposure to ultraviolet light were studied in cultured B-16 melanoma cells by the method of electron spin resonance with the spin labelling technique. The lipid peroxide content increased immediately after exposure to UV light and returned to the normal level, 3 hrs post-exposure. Plasma membrane fluidity increased significantly, 3 - 6 hrs after exposure, and remained at the increased level in the hydrophobic region of the membrane until 18 hrs. There were no significant differences in phospholipid content before and after exposure, except for cardiolipin. (author)

  10. Development of primary cell culture from Scylla serrata: Primary cell cultures from Scylla serrata

    OpenAIRE

    Sashikumar, Anu; Desai, P. V.

    2008-01-01

    This paper reports for the first time, the Primary cell culture of hepatopancreas from edible crab Scylla serrata using crab saline, L-15 (Leibovitz), 1 × L-15 + crab saline, 2 × L-15 + crab saline, 3 × L-15 and citrate buffer without any serum. We could isolate and maintain E (Embryonalzellen), F (Fibrenzellen), B (Blasenzellen), R (Restzellen) and G (Granular cells). Upon seeding the hepatopancreatic E, F, B, and R cells showed different survival pattern over time than granular cells. A mod...

  11. Design of 3D printed insert for hanging culture of Caco-2 cells

    International Nuclear Information System (INIS)

    A Caco-2 cell culture on Transwell, an alternative testing to animal or human testing used in evaluating drug intestinal permeability, incorrectly estimated the absorption of actively transported drugs due to the low expression of membrane transporters. Similarly, three-dimensional (3D) cultures of Caco-2 cells, which have been recommended to be more physiological relevant, were not superior to the Transwell culture in either accuracy or convenience in drug permeability testing. Using rapid 3D printing prototyping techniques, this study proposed a hanging culture of Caco-2 cells that performed with high accuracy in predicting drug permeability in humans. As found, hanging cultured Caco-2 cells formed a confluent monolayer and maintained high cell viability on the 3D printed insert. Compared with the normal culture on Transwell, the Caco-2 cells on the 3D printed insert presented ∼30–100% higher brush border enzyme activity and ∼2–7 folds higher activity of P-glycoprotein/multidrug resistance-associated protein 2 during 21 days of incubation. For the eight membrane transporter substrates, the predictive curve of the 3D printing culture exhibited better linearity (R2 = 0.92) to the human oral adsorption than that of the Transwell culture (R2 = 0.84), indicating better prediction by the 3D printing culture. In this regard, the 3D printed insert for hanging culture could be potentially developed as a convenient and low-cost tool for testing drug oral absorption. (paper)

  12. Phosphatidylinositol species of suspension cultured plant cells

    Energy Technology Data Exchange (ETDEWEB)

    Heim, S.; Wagner, K.G.

    Suspension cultured Nicotiana tabacum and Catharanthus roseus cells were labeled with (/sup 3/H)inositol, the phospholipid fraction extracted and separated by thin layer chromatography. Three different solvent systems and reference compounds were used to assign the different /sup 3/H-labeled species by autoradiography. The ratio of (/sup 3/H)inositol incorporation into PI, PIP and PIP/sub 2/ was found to be 95:4:1; with some preparations a lyso-PI band was obtained which incorporated about a tenth of the label of the PIP band. With Catharanthus roseus cells a very faint band between PI and lyso-PI was detected which could not be assigned to a reference compound.

  13. Evaluation of osteogenic cell culture and osteogenic/peripheral blood mononuclear human cell co-culture on modified titanium surfaces

    International Nuclear Information System (INIS)

    This study aimed to determine the effect of a bioactive ceramic coating on titanium in the nanothickness range on human osteogenic cells, peripheral blood mononuclear cells (PBMC) and on osteogenic cells co-cultured with PBMC without exogenous stimuli. Cell viability, proliferation, adhesion, cytokine release (IL1β, TGFβ1, IL10 and IL17) and intracellular stain for osteopontin and alkaline phosphatase were assessed. Morphologic evaluation showed smaller and less spread cell aspects in co-culture relative to osteogenic cell culture. Cell viability, proliferation and adhesion kinetics were differently influenced by surface texture/chemistry in culture versus co-culture. Cytokine release was also influenced by the interaction between mononuclear and osteogenic cells (mediators released by mononuclear cells acted on osteogenic cells and vice versa). In general, ‘multi-cell type’ interactions played a more remarkable role than the surface roughness or chemistry utilized on the in vitro cellular events related to initial stages of bone formation. (paper)

  14. Recombinant Protein Production and Insect Cell Culture and Process

    Science.gov (United States)

    Spaulding, Glenn F. (Inventor); Goodwin, Thomas J. (Inventor); OConnor, Kim C. (Inventor); Francis, Karen M. (Inventor); Andrews, Angela D. (Inventor); Prewett, Tracey L. (Inventor)

    1997-01-01

    A process has been developed for recombinant production of selected polypeptides using transformed insect cells cultured in a horizontally rotating culture vessel modulated to create low shear conditions. A metabolically transformed insect cell line is produced using the culture procedure regardless of genetic transformation. The recombinant polypeptide can be produced by an alternative process using virtually infected or stably transformed insect cells containing a gene encoding the described polypeptide. The insect cells can also be a host for viral production.

  15. Cell culture device and microchamber which can be monitored using nuclear magnetic resonance

    OpenAIRE

    Celda-Muñoz, Bernardo; Esteve-Moya, Vicent; Sancho-Bielsa, Francisco; Villa Sanz, Rosa; Fernández Ledesma, Luis José; Berganzo Ruiz, Javier

    2010-01-01

    [EN] The invention relates to a cell culture device and microchamber which can be monitored using nuclear magnetic resonance and other imaging techniques, in which the culture microchamber is encapsulated and housed inside a chip. The microchamber and the device are easy for the user to handle, allowing same to be handled or repositioned without requiring complex mounting operations. In addition, the invention allows cultures to be studied for long periods, great...

  16. Metabolic flux rewiring in mammalian cell cultures

    OpenAIRE

    Young, Jamey D.

    2013-01-01

    Continuous cell lines (CCLs) engage in “wasteful” glucose and glutamine metabolism that leads to accumulation of inhibitory byproducts, primarily lactate and ammonium. Advances in techniques for mapping intracellular carbon fluxes and profiling global changes in enzyme expression have led to a deeper understanding of the molecular drivers underlying these metabolic alterations. However, recent studies have revealed that CCLs are not necessarily entrenched in a glycolytic or glutaminolytic phe...

  17. Optical Management Techniques for Organic Solar Cells

    CERN Document Server

    Rajagopal, Adharsh

    2016-01-01

    In this thesis, two different optical management techniques for organics based solar cells are explored. The first part is focused on the development of a textured rear reflector for OPVs. The use of textured reflector (TR) facilitates an increase in the optical path length along with light trapping within the active layer. TR was fabricated through a relatively simpler technique by depositing metal films over a microlens array (MLA). Zinc oxide nanoparticles were used to minimize the shadowing effect. Using TR, enhancements in short-circuit current density and power conversion efficiencies up to 10-25% were demonstrated for a polymer based organic solar cell. The second part is focused on improving the effectiveness of MLA incorporation in OPVs. The increase in path length achieved using MLA can be improved by increasing the refractive index of MLA and incorporating MLA directly on the transparent electrode instead of glass substrate. This approach could avoid the optical losses occurring at the interface be...

  18. High vacuum cells for classical surface techniques

    International Nuclear Information System (INIS)

    Novel glass cells were designed and built to be able to perform surface potential and surface tension measurements in a contained environment. The cells can withstand pressures of approximately 1x10-6 Torr, providing a reasonable level of control in terms of the amounts of volatile contaminants during experimentation. The measurements can take several hours; thus the cells help maintain the integrity of the sample in the course of the experiment. To test for the feasibility of the cell design, calibration measurements were performed. For the surface potential cell, the modified TREK 6000B-7C probe exhibited performance comparable to its unmodified counterpart. The correlation measurements between applied potential on the test surface and the measured potential showed R-values very close to 1 as well as standard deviation values of less than 1. Results also demonstrate improved measurement values for experiments performed in vacuum. The surface tension cell, on the other hand, which was used to perform the pendant drop method, was tested on common liquids and showed percentage errors of 0.5% when compared to literature values. The fabricated cells redefine measurements using classical surface techniques, providing unique and novel methods of sample preparation, premeasurement preparation, and sample analysis at highly beneficial expenditure cost.

  19. Discarded human fetal tissue and cell cultures for transplantation research

    International Nuclear Information System (INIS)

    A feasibility study has been performed to explore the utility of various tissues from discarded human abortuses for transplantation and related research. Specifically, aborted fetuses plus parental blood samples and all relevant clinical data were obtained through a local hospital complex. Whenever possible, pancreas, skin and skeletal muscle, heart, liver, kidney, cartilage and lung tissues were removed, dissociated and subfractionated for cryopreservation, characterization and cultivation trials in vitro. Existing protocols for these manipulations were compared and improved upon as required. Clonal culture, cell aggregate maintenance techniques and use of feeder cell populations have been utilized where appropriate to develop quantitative comparative data. Histological and biochemical assays were applied both to evaluate separation/cultivation methods and to identify optimal culture conditions for maintaining functional cells. Immunochemical and molecular biological procedures were applied to study expression of Major Histocompatibility Vomplex (MHC) class 1 and 11 molecules on cell lines derived. Tissue and cell culture populations were examined for infections with bacteria, ftingi, mycoplasma, HIV, CMV, hepatitis B and other viruses. Only 1% of the abortuses tested were virally infected. Cytogenetic analyses confin-ned the normal diploid status in the vast majority (>98%) of lines tested. A total of over 250 abortuses have been obtained and processed. Only 25 were found to be contaminated with bacteria or fungi and unsuitable for further cultivation trials. A total of over 200 cell populations were isolated, characterized and cryopreserved for further study. Included were kidney, lung, liver and epidermal epithelia: cartilage-derived cells from the spine and epiphyses plus myogenic myoblasts. Selected lines have been immortalized using HPV I 6E6/E7 sequences. Epithelia from the liver and pancreas and cardiac myocytes were the most problematic in that initial

  20. Teaching Organizational Culture Using a Projective Technique: Collage Construction

    Science.gov (United States)

    Colakoglu, Saba; Littlefield, Jon

    2011-01-01

    Although the topic of "organizational culture" is an integral part of syllabi across a wide range of core business classes such as Principles of Management, Organizational Behavior, and Human Resource Management, few experiential exercises exist that can enhance student understanding and learning of different layers of organizational culture. In…

  1. System-level modeling and simulation of the cell culture microfluidic biochip ProCell

    DEFF Research Database (Denmark)

    Minhass, Wajid Hassan; Pop, Paul; Madsen, Jan;

    2010-01-01

    -defined micro-channels using valves and pumps. We present an approach to the system-level modeling and simulation of a cell culture microfluidic biochip called ProCell, Programmable Cell Culture Chip. ProCell contains a cell culture chamber, which is envisioned to run 256 simultaneous experiments (viewed as a...

  2. Protection of cultured mammalian cells by rebamipide

    Energy Technology Data Exchange (ETDEWEB)

    Antoku, Shigetoshi; Aramaki, Ryoji [Kyushu Univ., Fukuoka (Japan). Faculty of Medicine; Tanaka, Hisashi; Kusumoto, Naotoshi

    1997-06-01

    Rebamipide which is used as a drug for gastritis and stomach ulcer has large capability for OH radical scavenging. It is expected that rebamipide has protective effect against ionizing radiations. The present paper deals with protective effect of rebamipide for cultured mammalian cells exposed to ionizing radiations. As rebamipide is insoluble in water, three solvents were used to dissolve. Rebamipide dissolved in dimethyl sulfoxide (DMSO), dimethyl formamide (DMFA) and 0.02 N NaOH was added to the cells in Eagle`s minimum essential medium (MEM) supplemented with 10% fetal calf serum and the cells were irradiated with X-rays. After irradiation, the cells were trypsinized, plated in MEM with 10% fetal calf serum and incubated for 7 days in a CO{sub 2} incubator to form colonies. Rebamipide dissolved in 0.02 N NaOH exhibited the protective effect expected its OH radical scavenging capability. However, the protective effect of rebamipide dissolved in DMSO was about half of that expected by its radical scavenging capability and that of rebamipide dissolved in DMFA was not observed. Uptake of rebamipide labeled with {sup 14}C increased with increasing contact time with rebamipide. These rebamipide mainly distributed in nucleus rather than cytoplasm. (author)

  3. Determining Cell Number During Cell Culture using the Scepter Cell Counter

    OpenAIRE

    Ongena, Kathleen; Das, Chandreyee; Smith, Janet L.; Gil, Sónia; Johnston, Grace

    2010-01-01

    Counting cells is often a necessary but tedious step for in vitro cell culture. Consistent cell concentrations ensure experimental reproducibility and accuracy. Cell counts are important for monitoring cell health and proliferation rate, assessing immortalization or transformation, seeding cells for subsequent experiments, transfection or infection, and preparing for cell-based assays. It is important that cell counts be accurate, consistent, and fast, particularly for quantitative measuremen...

  4. Analysis of culture-dependent versus culture-independent techniques for identification of bacteria in clinically obtained bronchoalveolar lavage fluid.

    Science.gov (United States)

    Dickson, Robert P; Erb-Downward, John R; Prescott, Hallie C; Martinez, Fernando J; Curtis, Jeffrey L; Lama, Vibha N; Huffnagle, Gary B

    2014-10-01

    The diagnosis and management of pneumonia are limited by the use of culture-based techniques of microbial identification, which may fail to identify unculturable, fastidious, and metabolically active viable but unculturable bacteria. Novel high-throughput culture-independent techniques hold promise but have not been systematically compared to conventional culture. We analyzed 46 clinically obtained bronchoalveolar lavage (BAL) fluid specimens from symptomatic and asymptomatic lung transplant recipients both by culture (using a clinical microbiology laboratory protocol) and by bacterial 16S rRNA gene pyrosequencing. Bacteria were identified in 44 of 46 (95.7%) BAL fluid specimens by culture-independent sequencing, significantly more than the number of specimens in which bacteria were detected (37 of 46, 80.4%, P ≤ 0.05) or "pathogen" species reported (18 of 46, 39.1%, P ≤ 0.0001) via culture. Identification of bacteria by culture was positively associated with culture-independent indices of infection (total bacterial DNA burden and low bacterial community diversity) (P ≤ 0.01). In BAL fluid specimens with no culture growth, the amount of bacterial DNA was greater than that in reagent and rinse controls, and communities were markedly dominated by select Gammaproteobacteria, notably Escherichia species and Pseudomonas fluorescens. Culture growth above the threshold of 10(4) CFU/ml was correlated with increased bacterial DNA burden (P < 0.01), decreased community diversity (P < 0.05), and increased relative abundance of Pseudomonas aeruginosa (P < 0.001). We present two case studies in which culture-independent techniques identified a respiratory pathogen missed by culture and clarified whether a cultured "oral flora" species represented a state of acute infection. In summary, we found that bacterial culture of BAL fluid is largely effective in discriminating acute infection from its absence and identified some specific limitations of BAL fluid culture in the

  5. The influence of organizational culture on the use of quality techniques and its impact on performance

    DEFF Research Database (Denmark)

    Gambi, Lillian; Jørgensen, Frances; Boer, Harry;

    2013-01-01

    to enhanced performance more than if those culture profiles are combined with the use of measurement techniques. This research has important implications for managers in the sense that they need to be actively aware of the need to adopt quality techniques that fit with the culture of their organization....

  6. Documentation of cultural heritage; techniques, potentials, and constraints

    Science.gov (United States)

    Hassani, F.; Moser, M.; Rampold, R.; Wu, C.

    2015-08-01

    Cultural Heritage is known as an invaluable asset of human being, which portrays his achievements over centuries. The need for identification and preservation of cultural heritage is well understood and experts' attempt is to exploit any possible method to fulfill this aim. There are several published literatures and documents, which emphasize on the importance of the documentation of the cultural heritage such as Burra Charter. However, with the development of human and invention of new tools and technologies, the concept of the conservation of cultural heritage has changed considerably. The new technologies such as computers and digital tools have opened new windows and bestowed new opportunities in the process of conservation of cultural heritage. In this regard, it is important to review different technologies in order to make the best advantage of these tools in the cultural heritage field. The focus of this paper would be on the non-technical users who need to gain an overall comprehension of these new emerging tools. The foundation of this paper will be on the existing literatures published by various experts in addition to the author's experience and research in the conservation field.

  7. Three-Dimensional Cell Culture: A Breakthrough in Vivo

    Directory of Open Access Journals (Sweden)

    Delphine Antoni

    2015-03-01

    Full Text Available Cell culture is an important tool for biological research. Two-dimensional cell culture has been used for some time now, but growing cells in flat layers on plastic surfaces does not accurately model the in vivo state. As compared to the two-dimensional case, the three-dimensional (3D cell culture allows biological cells to grow or interact with their surroundings in all three dimensions thanks to an artificial environment. Cells grown in a 3D model have proven to be more physiologically relevant and showed improvements in several studies of biological mechanisms like: cell number monitoring, viability, morphology, proliferation, differentiation, response to stimuli, migration and invasion of tumor cells into surrounding tissues, angiogenesis stimulation and immune system evasion, drug metabolism, gene expression and protein synthesis, general cell function and in vivo relevance. 3D culture models succeed thanks to technological advances, including materials science, cell biology and bioreactor design.

  8. Flow field measurements in the cell culture unit

    Science.gov (United States)

    Walker, Stephen; Wilder, Mike; Dimanlig, Arsenio; Jagger, Justin; Searby, Nancy

    2002-01-01

    The cell culture unit (CCU) is being designed to support cell growth for long-duration life science experiments on the International Space Station (ISS). The CCU is a perfused loop system that provides a fluid environment for controlled cell growth experiments within cell specimen chambers (CSCs), and is intended to accommodate diverse cell specimen types. Many of the functional requirements depend on the fluid flow field within the CSC (e.g., feeding and gas management). A design goal of the CCU is to match, within experimental limits, all environmental conditions, other than the effects of gravity on the cells, whether the hardware is in microgravity ( micro g), normal Earth gravity, or up to 2g on the ISS centrifuge. In order to achieve this goal, two steps are being taken. The first step is to characterize the environmental conditions of current 1g cell biology experiments being performed in laboratories using ground-based hardware. The second step is to ensure that the design of the CCU allows the fluid flow conditions found in 1g to be replicated from microgravity up to 2g. The techniques that are being used to take these steps include flow visualization, particle image velocimetry (PIV), and computational fluid dynamics (CFD). Flow visualization using the injection of dye has been used to gain a global perspective of the characteristics of the CSC flow field. To characterize laboratory cell culture conditions, PIV is being used to determine the flow field parameters of cell suspension cultures grown in Erlenmeyer flasks on orbital shakers. These measured parameters will be compared to PIV measurements in the CSCs to ensure that the flow field that cells encounter in CSCs is within the bounds determined for typical laboratory experiments. Using CFD, a detailed simulation is being developed to predict the flow field within the CSC for a wide variety of flow conditions, including microgravity environments. Results from all these measurements and analyses of the

  9. Primary cell cultures from sea urchin ovaries: a new experimental tool.

    Science.gov (United States)

    Mercurio, Silvia; Di Benedetto, Cristiano; Sugni, Michela; Candia Carnevali, M Daniela

    2014-02-01

    In the present work, primary cell cultures from ovaries of the edible sea urchin Paracentrotus lividus were developed in order to provide a simple and versatile experimental tool for researches in echinoderm reproductive biology. Ovary cell phenotypes were identified and characterized by different microscopic techniques. Although cell cultures could be produced from ovaries at all stages of maturation, the cells appeared healthier and viable, displaying a higher survival rate, when ovaries at early stages of gametogenesis were used. In terms of culture medium, ovarian cells were successfully cultured in modified Leibovitz-15 medium, whereas poor results were obtained in minimum essential medium Eagle and medium 199. Different substrates were tested, but ovarian cells completely adhered only on poly-L-lysine. To improve in vitro conditions and stimulate cell proliferation, different serum-supplements were tested. Fetal calf serum and an originally developed pluteus extract were detrimental to cell survival, apparently accelerating processes of cell death. In contrast, cells cultured with sea urchin egg extract appeared larger and healthier, displaying an increased longevity that allowed maintaining them for up to 1 month. Overall, our study provides new experimental bases and procedures for producing successfully long-term primary cell cultures from sea urchin ovaries offering a good potential to study echinoid oogenesis in a controlled system and to investigate different aspects of echinoderm endocrinology and reproductive biology. PMID:24002666

  10. Organ culture-cell culture system for studying multistage carcinogenesis in respiratory epithelium. [Mice

    Energy Technology Data Exchange (ETDEWEB)

    Steele, Vernon E.; Marchok, Ann C.; Nettesheim, Paul

    1977-01-01

    An organ culture-cell culture system was used to demonstrate carcinogen dose-dependent transformation of tracheal epithelial cells in vitro. Tracheal explants were exposed to MNNG (N-methyl-N/sup 1/-nitro-N-nitrosoguanidine) in organ culture. Outgrowths from these explants provided epithelial cell cultures. The numbers of long term epithelial cell cultures and cell lines that were established per explant increased as MNNG exposure concentration increased. At the present time, more cell lines derived from explants exposed to the highest MNNG concentration have produced palpable tumors than cell lines derived from explants exposed to lower MNNG concentrations. No cell lines were established from primaries derived from control explants. TPA (12-0-tetradecanoyl-phorbol-13-acetate), stimulates DNA synthesis in tracheal epithelium in organ culture in a manner simular to that described for mouse skin. Short exposures to TPA not only stimulated DNA synthesis earlier, but the stimulation was greater than that obtained with continuous exposure. At the present time, exposure of tracheal organ cultures to MNNG followed by TPA has resulted in an enhanced production of morphologically altered cells in primary epithelial cell cultures, than exposure to either agent alone.

  11. Experimental Methodology used by Cell Cultures Laboratory from INRMFB to assess the therapeutic effect of natural factors

    OpenAIRE

    Munteanu Constantin; Munteanu Diana

    2010-01-01

    The experimental study design on cell cultures allows the direct biological evaluation at the cellular level, of the therapeutic effect that natural factors can play over the organism.Techniques for obtaining cell cultures requires a complex and laborious task that starts from live tissue sampling, continuous with isolation of cells and their preparation for sowing a culture plate. This preparation involves mechanical and enzymatic action from the researcher on biological material. Derived ce...

  12. Electrospinning of microbial polyester for cell culture

    International Nuclear Information System (INIS)

    Biodegradable and biocompatible poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), a copolymer of microbial polyester, was fabricated as a nanofibrous mat by electrospinning. The specific surface area and the porosity of electrospun PHBV nanofibrous mat were determined. When the mechanical properties of flat film and electrospun PHBV nanofibrous mats were investigated, both the tensile modulus and strength of electrospun PHBV were less than those of cast PHBV film. However, the elongation ratio of nanofiber mat was higher than that of the cast film. The structure of electrospun nanofibers using PHBV-trifluoroethanol solutions depended on the solution concentrations. When x-ray diffraction patterns of bulk PHBV before and after electrospinning were compared, the crystallinity of PHBV was not significantly affected by the electrospinning process. Chondrocytes adhered and grew on the electrospun PHBV nanofibrous mat better than on the cast PHBV film. Therefore, the electrospun PHBV was considered to be suitable for cell culture

  13. Methods for the isolation and identification of polycyclic aromatic hydrocarbons found in complex mixtures and the determination of their possible toxicity by means of a host mediated bioassay technique. Progress report, July 1, 1976--February 1, 1977. [Cultured mouse leumemia cell bioassay system

    Energy Technology Data Exchange (ETDEWEB)

    Lipsky, S.R.; Alexander, G.; McMurray, W.; Capizzi, R.

    1977-02-01

    Techniques were developed to produce excellent high performance glass capillary columns for gas chromatographic analyses of a wide range of complex mixtures of organic compounds, including those containing a wide array of polycyclic aromatic hydrocarbons (PAH) derived from a coal liquefaction process. Work was begun to assess the potential mutogenicity and/or carcinogenicity of the various isolated PAH fractions utilizing a unique host mediated bioassay system. Preliminary results indicate that further efforts will be required to determine dose response parameters of cultured mouse leukemia cells, as well as suitable vehicles for the satisfactory introduction of certain PAH fractions into this particular bioassay system.

  14. Cells and Culture Systems Used to Model the Small Airway Epithelium.

    Science.gov (United States)

    Bhowmick, Rudra; Gappa-Fahlenkamp, Heather

    2016-06-01

    The pulmonary epithelium is divided into upper, lower, and alveolar (or small) airway epithelia and acts as the mechanical and immunological barrier between the external environment and the underlying submucosa. Of these, the small airway epithelium is the principal area of gas exchange and has high immunological activity, making it a major area of cell biology, immunology, and pharmaceutical research. As animal models do not faithfully represent the human pulmonary system and ex vivo human lung samples have reliability and availability issues, cell lines, and primary cells are widely used as small airway epithelial models. In vitro, these cells are mostly cultured as monolayers (2-dimensional cultures), either media submerged or at air-liquid interface. However, these 2-dimensional cultures lack a three dimension-a scaffolding extracellular matrix, which establishes the intercellular network in the in vivo airway epithelium. Therefore, 3-dimensional cell culture is currently a major area of development, where cells are cultured in a matrix or are cultured in a manner that they develop ECM-like scaffolds between them, thus mimicking the in vivo phenotype more faithfully. This review focuses on the commonly used small airway epithelial cells, their 2-dimensional and 3-dimensional culture techniques, and their comparative phenotype when cultured under these systems. PMID:27071933

  15. Acetic acid bacteria from biofilm of strawberry vinegar visualized by microscopy and detected by complementing culture-dependent and culture-independent techniques.

    Science.gov (United States)

    Valera, Maria José; Torija, Maria Jesús; Mas, Albert; Mateo, Estibaliz

    2015-04-01

    Acetic acid bacteria (AAB) usually develop biofilm on the air-liquid interface of the vinegar elaborated by traditional method. This is the first study in which the AAB microbiota present in a biofilm of vinegar obtained by traditional method was detected by pyrosequencing. Direct genomic DNA extraction from biofilm was set up to obtain suitable quality of DNA to apply in culture-independent molecular techniques. The set of primers and TaqMan--MGB probe designed in this study to enumerate the total AAB population by Real Time--PCR detected between 8 × 10(5) and 1.2 × 10(6) cells/g in the biofilm. Pyrosequencing approach reached up to 10 AAB genera identification. The combination of culture-dependent and culture-independent molecular techniques provided a broader view of AAB microbiota from the strawberry biofilm, which was dominated by Ameyamaea, Gluconacetobacter, and Komagataeibacter genera. Culture-dependent techniques allowed isolating only one genotype, which was assigned into the Ameyamaea genus and which required more analysis for a correct species identification. Furthermore, biofilm visualization by laser confocal microscope and scanning electronic microscope showed different dispositions and cell morphologies in the strawberry vinegar biofilm compared with a grape vinegar biofilm. PMID:25475315

  16. CATEGORICAL IMAGE COMPONENTS IN THE FORMING SYSTEM OF A MARKETING TECHNIQUES MANAGER’S IMAGE CULTURE

    OpenAIRE

    Anna Borisovna Cherednyakova

    2015-01-01

    Based on the understanding of the image culture formation of managers of marketing techniques, as a representative of the social and communication interaction of public structures, categorical apparatus of image culture with an emphasis on the etymology of the image, as an integral component of image culture was analyzed. Categorical components of the image are presented from the standpoint of image culture, as personal new formation, an integral part of the professional activity of the marke...

  17. Particle Trajectories in Rotating Wall Cell Culture Devices

    Science.gov (United States)

    Ramachandran N.; Downey, J. P.

    1999-01-01

    Cell cultures are extremely important to the medical community since such cultures provide an opportunity to perform research on human tissue without the concerns inherent in experiments on individual humans. Development of cells in cultures has been found to be greatly influenced by the conditions of the culture. Much work has focused on the effect of the motions of cells in the culture relative to the solution. Recently rotating wall vessels have been used with success in achieving improved cellular cultures. Speculation and limited research have focused on the low shear environment and the ability of rotating vessels to keep cells suspended in solution rather than floating or sedimenting as the primary reasons for the improved cellular cultures using these devices. It is widely believed that the cultures obtained using a rotating wall vessel simulates to some degree the effect of microgravity on cultures. It has also been speculated that the microgravity environment may provide the ideal acceleration environment for culturing of cellular tissues due to the nearly negligible levels of sedimentation and shear possible. This work predicts particle trajectories of cells in rotating wall vessels of cylindrical and annular design consistent with the estimated properties of typical cellular cultures. Estimates of the shear encountered by cells in solution and the interactions with walls are studied. Comparisons of potential experiments in ground and microgravity environments are performed.

  18. Efficient Electroporation of DNA and Protein into Confluent and Differentiated Epithelial Cells in Culture

    OpenAIRE

    Deora, Ami A.; Diaz, Fernando; Schreiner, Ryan; Rodriguez-Boulan, Enrique

    2007-01-01

    Electroporation-mediated delivery of molecules is a procedure widely used for transfecting complementary DNA in bacteria, mammalian and plant cells. This technique has proven very efficient for the introduction of macromolecules into cells in suspension culture and even into cells in their native tissue environment, e.g. retina and embryonic tissues. However, in spite of several attempts to date, there are no well-established procedures to electroporate polarized epithelial cells adhering to ...

  19. Usability and Applicability of Microfluidic Cell Culture Systems

    DEFF Research Database (Denmark)

    Hemmingsen, Mette

    devices still lack general implementation into biological research laboratories. In this project, the usability and applicability of microfluidic cell culture systems have been investigated. The tested systems display good properties regarding optics and compatibility with standard laboratory equipment...... possibilities for, for example, precise control of the chemical environment, 3D cultures, controlled co-culture of different cell types or automated, individual control of up to 96 cell culture chambers in one integrated system. Despite the great new opportunities to perform novel experimental designs, these...... shown to be needed. This is possibly one of the reasons for the lack of implementation of microfluidic cell culture systems into biological research laboratories. Procedures to perform long-term microfluidic perfusion cell culture experiments have been established. Furthermore, successful application of...

  20. Preparation of labelled lipids by the use of plant cell cultures

    International Nuclear Information System (INIS)

    The preparation of some radioacitvely labelled lipids by the use of plant cell cultures is discussed and further applications of the new method are suggested. Cell suspension cultures of rape (Brassica napus) and soya (Glycine max) have been used for the preparation of lipids labelled with radioisotopes. Radioactive acetic acid as well as various long-chain fatty acids are readily incorporated into the neutral and ionic lipids of plant cell cultures. In addition, 14C-labelled glycerol, ethanolamine and choline are well utilized by the cells. Randomly labelled lipids have been obtained by incubating cell suspension cultures of rape and soya with [1-14C] acetic acid, and uniformly labelled lipids have been isolated from cultures that had been incubated with a mixture of [1-14C] acetic acid plus [2-14C] acetic acid. The use of techniques of plant cell cultures for the preparation of lipds labelled with stable or radioactive isotopesappears particularly rewarding because the uptake of precursors by the cells and their incorporation into various lipid compounds proceeds rapidly and often quanitatively.This new approach should be useful also for the biosynthesis of lipids whose acyl moieties contain a spn radical, a fluorescent group, or a light-sensitive label. Thus, plant cell cultures constitute valuable new tools for the biosynthetic preparation of a great variety of labelled lipids. (A.G.)

  1. THE ALKALOID CYTISINE IN THE CELL CULTURE

    Directory of Open Access Journals (Sweden)

    Gazaliev A.M.

    2012-08-01

    Full Text Available Alkaloids are vegetative establishments of complex and original structure with nitrous heterocycles in the basis. For a long time they drew researchers’ attention because of their unique and specific physiological effect on alive organisms. Not all the representatives of the globe’s flora contain these unique substances. Alkaloid cytisine is to be found mainly in the plants of the fabaceous family - Fabaceae. For the cytisine production the seeds of Thermopsis lanceolata R.Br (T. lanceolata R.Br and Cytisus laburnum (C. laburnum are used as a raw material. The object of the research is T. lanceolata cell culture. Sterile sprouts are used at the first stage of the experiment. Callus genesis is accompanied with dedifferentiation. It leads to the cellular organization simplification. Based on an important property of a plant cell, such as totipotency, there appears the formation of the “de novo” biosynthetic device. The cultivation algorithm consists of two basic stages: (i the cultivation conditions optimization of callus with a high level of the primary metabolites biosynthesis (Aspartat – lysine; (ii the research of cultivation chemical and physical factors influence on the secondary metabolite (cytisine biosynthesis and accumulation. During the cultivation the Murashige and Skoog classical recipe of nutrient medium will be used. Optimization of the cultivation conditions will concern the phytohormones, macro- and micronutrients content, as the purpose of optimization is the production of the determined high-level competence embriogenical callus. The main problem is genetic heterogeneity of a cellular population and instability of morpho-physiological processes. The correct management of higher plants cells population is possible at the synchronization of a cellular cycle phases. The references analysis has shown that it is almost impossible to synchronize cellular cycles in the culture of plant tissue. The application of chemical

  2. Pigment cell differentiation in sea urchin blastula-derived primary cell cultures.

    Science.gov (United States)

    Ageenko, Natalya V; Kiselev, Konstantin V; Dmitrenok, Pavel S; Odintsova, Nelly A

    2014-07-01

    The quinone pigments of sea urchins, specifically echinochrome and spinochromes, are known for their effective antioxidant, antibacterial, antifungal, and antitumor activities. We developed in vitro technology for inducing pigment differentiation in cell culture. The intensification of the pigment differentiation was accompanied by a simultaneous decrease in cell proliferation. The number of pigment cells was two-fold higher in the cells cultivated in the coelomic fluids of injured sea urchins than in those intact. The possible roles of the specific components of the coelomic fluids in the pigment differentiation process and the quantitative measurement of the production of naphthoquinone pigments during cultivation were examined by MALDI and electrospray ionization mass spectrometry. Echinochrome A and spinochrome E were produced by the cultivated cells of the sand dollar Scaphechinus mirabilis in all tested media, while only spinochromes were found in the cultivated cells of another sea urchin, Strongylocentrotus intermedius. The expression of genes associated with the induction of pigment differentiation was increased in cells cultivated in the presence of shikimic acid, a precursor of naphthoquinone pigments. Our results should contribute to the development of new techniques in marine biotechnology, including the generation of cell cultures producing complex bioactive compounds with therapeutic potential. PMID:24979272

  3. Pigment Cell Differentiation in Sea Urchin Blastula-Derived Primary Cell Cultures

    Directory of Open Access Journals (Sweden)

    Natalya V. Ageenko

    2014-06-01

    Full Text Available The quinone pigments of sea urchins, specifically echinochrome and spinochromes, are known for their effective antioxidant, antibacterial, antifungal, and antitumor activities. We developed in vitro technology for inducing pigment differentiation in cell culture. The intensification of the pigment differentiation was accompanied by a simultaneous decrease in cell proliferation. The number of pigment cells was two-fold higher in the cells cultivated in the coelomic fluids of injured sea urchins than in those intact. The possible roles of the specific components of the coelomic fluids in the pigment differentiation process and the quantitative measurement of the production of naphthoquinone pigments during cultivation were examined by MALDI and electrospray ionization mass spectrometry. Echinochrome A and spinochrome E were produced by the cultivated cells of the sand dollar Scaphechinus mirabilis in all tested media, while only spinochromes were found in the cultivated cells of another sea urchin, Strongylocentrotus intermedius. The expression of genes associated with the induction of pigment differentiation was increased in cells cultivated in the presence of shikimic acid, a precursor of naphthoquinone pigments. Our results should contribute to the development of new techniques in marine biotechnology, including the generation of cell cultures producing complex bioactive compounds with therapeutic potential.

  4. Systems Biology for Organotypic Cell Cultures

    Energy Technology Data Exchange (ETDEWEB)

    Grego, Sonia [RTI International, Research Triangle Park, NC (United States); Dougherty, Edward R. [Texas A & M Univ., College Station, TX (United States); Alexander, Francis J. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Auerbach, Scott S. [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Berridge, Brian R. [GlaxoSmithKline, Research Triangle Park, NC (United States); Bittner, Michael L. [Translational Genomics Research Inst., Phoenix, AZ (United States); Casey, Warren [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Cooley, Philip C. [RTI International, Research Triangle Park, NC (United States); Dash, Ajit [HemoShear Therapeutics, Charlottesville, VA (United States); Ferguson, Stephen S. [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Fennell, Timothy R. [RTI International, Research Triangle Park, NC (United States); Hawkins, Brian T. [RTI International, Research Triangle Park, NC (United States); Hickey, Anthony J. [RTI International, Research Triangle Park, NC (United States); Kleensang, Andre [Johns Hopkins Univ., Baltimore, MD (United States). Center for Alternatives to Animal Testing; Liebman, Michael N. [IPQ Analytics, Kennett Square, PA (United States); Martin, Florian [Phillip Morris International, Neuchatel (Switzerland); Maull, Elizabeth A. [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Paragas, Jason [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Qiao, Guilin [Defense Threat Reduction Agency, Ft. Belvoir, VA (United States); Ramaiahgari, Sreenivasa [National Inst. of Environmental Health Sciences, Research Triangle Park, NC (United States); Sumner, Susan J. [RTI International, Research Triangle Park, NC (United States); Yoon, Miyoung [The Hamner Inst. for Health Sciences, Research Triangle Park, NC (United States); ScitoVation, Research Triangle Park, NC (United States)

    2016-08-04

    Translating in vitro biological data into actionable information related to human health holds the potential to improve disease treatment and risk assessment of chemical exposures. While genomics has identified regulatory pathways at the cellular level, translation to the organism level requires a multiscale approach accounting for intra-cellular regulation, inter-cellular interaction, and tissue/organ-level effects. Tissue-level effects can now be probed in vitro thanks to recently developed systems of three-dimensional (3D), multicellular, “organotypic” cell cultures, which mimic functional responses of living tissue. However, there remains a knowledge gap regarding interactions across different biological scales, complicating accurate prediction of health outcomes from molecular/genomic data and tissue responses. Systems biology aims at mathematical modeling of complex, non-linear biological systems. We propose to apply a systems biology approach to achieve a computational representation of tissue-level physiological responses by integrating empirical data derived from organotypic culture systems with computational models of intracellular pathways to better predict human responses. Successful implementation of this integrated approach will provide a powerful tool for faster, more accurate and cost-effective screening of potential toxicants and therapeutics. On September 11, 2015, an interdisciplinary group of scientists, engineers, and clinicians gathered for a workshop in Research Triangle Park, North Carolina, to discuss this ambitious goal. Participants represented laboratory-based and computational modeling approaches to pharmacology and toxicology, as well as the pharmaceutical industry, government, non-profits, and academia. Discussions focused on identifying critical system perturbations to model, the computational tools required, and the experimental approaches best suited to generating key data. This consensus report summarizes the discussions held.

  5. Induced engulfment of Neisseria gonorrhoeae by tissue culture cells.

    OpenAIRE

    Richardson, W P; Sadoff, J C

    1988-01-01

    Engulfment of gonococci by mammalian tissue culture cells was examined as a model of the penetration of host cells in gonorrhea. Engulfment required viable organisms; killing the gonococci with heat or refrigeration abolished the process. Engulfment also required tissue culture cell microtubule- and microfilament-dependent movement; treating the cells with cytochalasin B (0.5 micrograms/ml) or demecolcine (Colcemid; Ciba-Geigy AG, Basel, Switzerland) (10 micrograms/ml) also prevented his proc...

  6. Increased exosome production from tumour cell cultures using the Integra CELLine Culture System.

    Science.gov (United States)

    Mitchell, J Paul; Court, Jacqueline; Mason, Malcolm David; Tabi, Zsuzsanna; Clayton, Aled

    2008-06-01

    Exosomes are nanometer-sized vesicles, secreted from most cell types, with documented immune-modulatory functions. Exosomes can be purified from cultured cells but to do so effectively, requires maintenance of cells at high density in order to obtain sufficient accumulation of exosomes in the culture medium, prior to purification. Whilst high density cultures can be achieved with cells in suspension, this remains difficult with adherent cells, resulting in low quantity of exosomes for subsequent study. We have used the Integra CELLine culture system, originally designed for hybridoma cultures, to achieve a significant increase in obtainable exosomes from adherent and non-adherent tumour cells. Traditional cultures of mesothelioma cells (cultured in 75 cm(2) flasks) gave an average yield of 0.78 microg+/-0.14 microg exosome/ml of conditioned medium. The CELLine Adhere 1000 (CLAD1000) flask, housing the same cell line, increased exosome yield approximately 12 fold to 10.06 microg+/-0.97 microg/ml. The morphology, phenotype and immune function of these exosomes were compared, and found to be identical in all respects. Similarly an 8 fold increase in exosome production was obtained from NKL cells (a suspension cell line) using a CELLine 1000 (CL1000) flask. The CELLine system also incurred ~5.5 fold less cost and reduced labour for cell maintenance. This simple culture system is a cost effective, useful method for significantly increasing the quantity of exosomes available from cultured cells, without detrimental effects. This tool should prove advantageous in future studies of exosome-immune modulation in cancer and other settings. PMID:18423480

  7. Lethal graft-versus-host disease: modification with allogeneic cultured donor cells

    International Nuclear Information System (INIS)

    The use of the bone marrow culture technique was studied as a means to prepare donor marrow for bone marrow transplantation to avoid lethal graft-versus-host disease (GVHD). Preliminary experiments demonstrated the rapid loss of theta-positive cells in such cultures, so that theta-positive cells were not detected after 6 days. Initial experiments in C3H/HeJ (H-2k, Hbbd) recipients prepared with 900 rad demonstrated improved survival when 3-day cultured C57BL/6 (H-2b, Hbbs) donor cells were used in place of hind limb marrow for transplantation. However, hemoglobin typing of recipient animals revealed only short-term donor engraftment, with competitive repopulation of recipient marrow occurring. Subsequent experiments were done in 1,200-rad prepared recipients, with long-term donor engraftment demonstrated. The majority of 1,200-rad prepared animals receiving cultured allogeneic cells died of GVHD, but animals receiving 28-day cultured cells had an improved 90-day survival and a delay in GVHD development over animals receiving hind limb marrow or marrow from shorter times in culture. In addition, animals receiving anti-theta-treated, 3-day nonadherent cells had an improved survival (44%) over animals receiving anti-theta-treated hind limb marrow (20%). These experiments demonstrate modest benefit for the use of cultured cells in bone marrow transplantation across major H-2 histocompatibility complex differences

  8. Optimization of porcine urothelial cell cultures: Best practices, recommendations, and threats.

    Science.gov (United States)

    Pokrywczynska, Marta; Czapiewska, Monika; Jundzill, Arkadiusz; Bodnar, Magdalena; Balcerczyk, Daria; Kloskowski, Tomasz; Nowacki, Maciej; Marszalek, Andrzej; Drewa, Tomasz

    2016-07-01

    Many experimental approaches have been conducted in order to isolate urothelial cells from bladder tissue biopsies, but each method described has utilized different protocols and sources of bladder tissue. In this study, we compared the different methods of urothelial cell isolation available in literature together with standardized methods in order to obtain more unified results. Five methods for primary porcine urothelial culture establishment were compared: tissue explants and four enzymatic methods utilizing collagenase II, dispase II, combination of dispase II and trypsin, and trypsin alone. The average number of isolated cells, cell morphology, success of established culture, average number of cells from the first passage, expression of p63 and pancytokeratin and the characterization of urothelial cell growth, and aging were analyzed during the in vitro culture. The method utilizing dispase II was the most efficient and reproducible method for the isolation and culture of porcine urothelial cells when compared to the other tested methods. Urothelial cells obtained by this method grew considerably well and the cultures were established with high efficiency, which enabled us in obtaining a large quantity of cells with normal morphology. Contamination with fibroblasts in this method was the lowest. The utilization of a proper method for urothelial cell isolation is a critical step in the urinary tract regeneration when using tissue engineering techniques. In summary, this study demonstrated that by utilizing the described method with dispase II, a suitable number of cells was achieved, proving the method useful for tissue regeneration. PMID:27079486

  9. Tissue culture technique as new approach to combat Striga hermonthica

    OpenAIRE

    M.M. Hassan; Osman, M. G.; Mohammed Ahmed, M. M.; Abdalaleem, K. G.; Abdelgain, M. E.; Babiker, A. G. T.

    2011-01-01

    This study was conducted to evaluate the effect of media type, sugars, Ammonium nitrate, Potassium phosphate, auxins and cytokinine in different concentrations on in vitro Striga seed germination. Medium B5 was showed to induce Striga germination after short period of time. Striga germination reached 100% after 10 day of culture. This may be due to chemical components of this media that induced Striga germination. Results indicated that Striga seeds germination was affected by sugars. Sucrose...

  10. Experimental Methodology used by Cell Cultures Laboratory from INRMFB to assess the therapeutic effect of natural factors

    Directory of Open Access Journals (Sweden)

    Munteanu Constantin

    2010-11-01

    Full Text Available The experimental study design on cell cultures allows the direct biological evaluation at the cellular level, of the therapeutic effect that natural factors can play over the organism.Techniques for obtaining cell cultures requires a complex and laborious task that starts from live tissue sampling, continuous with isolation of cells and their preparation for sowing a culture plate. This preparation involves mechanical and enzymatic action from the researcher on biological material. Derived cell cultures are monitored morphologically by high-performance inverted biological microscope, with video camera for image acquisition. In the final stage, the cells are scraped, and through biochemical and molecular techniques, the therapeutic efficiency hypothesis of the investigated natural factor is verified experimentally. The cell cultures can be crioconservated in special containers with liquid nitrogen.

  11. High-Aspect-Ratio Rotating Cell-Culture Vessel

    Science.gov (United States)

    Wolf, David A.; Sams, Clarence; Schwarz, Ray P.

    1992-01-01

    Cylindrical rotating cell-culture vessel with thin culture-medium layer of large surface area provides exchange of nutrients and products of metabolism with minimal agitation. Rotation causes averaging of buoyant forces otherwise separating components of different densities. Vessel enables growth of cells in homogeneous distribution with little agitation and little shear stress.

  12. Growth and Plating of Cell Suspension Cultures of Datura Innoxia

    DEFF Research Database (Denmark)

    Engvild, Kjeld Christensen

    1974-01-01

    ammonium malate) or on NO3−-N alone. Dry weight yield was proportional to the amount of nitrate-N added (47 mg/mg N). Filtered suspension cultures containing single cells (plating cultures) could be grown in agar in petri dishes when NAA or 2,4-D were used as growth substances. Cells grew at densities...

  13. Batch variation between branchial cell cultures: An analysis of variance

    DEFF Research Database (Denmark)

    Hansen, Heinz Johs. Max; Grosell, M.; Kristensen, L.

    2003-01-01

    We present in detail how a statistical analysis of variance (ANOVA) is used to sort out the effect of an unexpected batch-to-batch variation between cell cultures. Two separate cultures of rainbow trout branchial cells were grown on permeable filtersupports ("inserts"). They were supposed to be...

  14. 21 CFR 864.2280 - Cultured animal and human cells.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cultured animal and human cells. 864.2280 Section 864.2280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products §...

  15. Cell culture monitoring by impedance mapping using a multielectrode scanning impedance spectroscopy system (CellMap)

    International Nuclear Information System (INIS)

    We report on the impedance mapping of in vitro cellular morphology by electrical impedance spectroscopy, using microelectrodes. A micro multielectrode system was designed, fabricated, assembled, tested and demonstrated for the monitoring of anchorage-dependent cell behavior and morphology. This system allowed continuous, label-free, quantitative monitoring and visualization of cell adhesion, spreading, proliferation and detachment due to cell cycle processes as well as cell–drug interaction, with spatio-temporal resolution. OvCa429 ovarian cancer cells were monitored in vitro over a period of 70 hours by inoculating the cell suspension directly on the multielectrode device. The phase angle of impedance was observed to develop a distinctive shape as a result of cell attachment and proliferation. The shape of the phase angle curve reverted back to the pre-attachment shape upon detachment of cells from the substrate, caused by the addition of trypsin to the cell culture medium. The impedance data of the cell culture were then successfully modeled as a multi-parametric equivalent circuit. The model incorporated both interfacial and cell-layer impedance parameters. Upon addition of trypsin, the cell-layer parameters showed a marked decline and were eventually eliminated from the multi-parametric model, confirming the correlation of the model to the electrode–cell–electrolyte system. These experiments demonstrate the applicability of the impedance mapping technique in visualizing and quantifying physiological changes in the cell layer due to cellular processes as well as the effect of external chemical stimulus on cells (cell–drug interaction)

  16. Comparison of defined culture systems for feeder cell free propagation of human embryonic stem cells

    OpenAIRE

    Akopian, Veronika; Andrews, Peter W.; Beil, Stephen; Benvenisty, Nissim; Brehm, Jennifer; Christie, Megan; Ford, Angela; Fox, Victoria; Gokhale, Paul J; Healy, Lyn; Holm, Frida; Hovatta, Outi; Knowles, Barbara B; Ludwig, Tenneille E; Ronald D G McKay

    2010-01-01

    There are many reports of defined culture systems for the propagation of human embryonic stem cells in the absence of feeder cell support, but no previous study has undertaken a multi-laboratory comparison of these diverse methodologies. In this study, five separate laboratories, each with experience in human embryonic stem cell culture, used a panel of ten embryonic stem cell lines (including WA09 as an index cell line common to all laboratories) to assess eight cell culture methods, with pr...

  17. New fabrication technique of conductive polymer/insulating polymer composite films and evaluation of biocompatibility in neuron cultures

    International Nuclear Information System (INIS)

    Poly(vinyl alcohol), PVA, produces a flexible composite polymer film with electrical, optical and electrochemical properties very similar to those of polypyrrole (PPy). The rate of electrochemical polymerization depends on the diffusion rate of the electrolyte across the PVA film to the indium tin oxide (ITO) electrode. In particular, a solvent with a hydrophilic nature easily penetrates into the PVA film. By applying this new process, we demonstrate a unique method of forming an electrically conductive pattern in PVA film. It will be possible to develop electrodes for electrical stimulation of the nervous system using the conducting polymer, PPy. Then, by applying a similar technique, we fabricated poly(3,4-ethylenedioxythiophene), PEDOT/PVA, composite films and investigated their basic electrochemical properties. Moreover, in this study, in order to develop a novel cell-culture system which makes it possible to communicate with cultured cells, fibroblasts were cultured on PPy- and PEDOT-coated ITO conductive glass plates for 7 days. The result reveals that the PPy and PEDOT films support the secretory functions of the cells cultured on its surface. The PPy- and PEDOT-coated electrodes may be useful to culture the cells on.

  18. New fabrication technique of conductive polymer/insulating polymer composite films and evaluation of biocompatibility in neuron cultures

    Energy Technology Data Exchange (ETDEWEB)

    Onoda, Mitsuyoshi, E-mail: onoda@eng.u-hyogo.ac.j [Department of Electrical Engineering and Computer Sciences, Graduate School of Engineering, University of Hyogo, Himwji Shosha Campus, 2167 Shosha, Himeji, Hyogo 671-2280 (Japan); Abe, Yayoi; Tada, Kazuya [Department of Electrical Engineering and Computer Sciences, Graduate School of Engineering, University of Hyogo, Himwji Shosha Campus, 2167 Shosha, Himeji, Hyogo 671-2280 (Japan)

    2009-11-30

    Poly(vinyl alcohol), PVA, produces a flexible composite polymer film with electrical, optical and electrochemical properties very similar to those of polypyrrole (PPy). The rate of electrochemical polymerization depends on the diffusion rate of the electrolyte across the PVA film to the indium tin oxide (ITO) electrode. In particular, a solvent with a hydrophilic nature easily penetrates into the PVA film. By applying this new process, we demonstrate a unique method of forming an electrically conductive pattern in PVA film. It will be possible to develop electrodes for electrical stimulation of the nervous system using the conducting polymer, PPy. Then, by applying a similar technique, we fabricated poly(3,4-ethylenedioxythiophene), PEDOT/PVA, composite films and investigated their basic electrochemical properties. Moreover, in this study, in order to develop a novel cell-culture system which makes it possible to communicate with cultured cells, fibroblasts were cultured on PPy- and PEDOT-coated ITO conductive glass plates for 7 days. The result reveals that the PPy and PEDOT films support the secretory functions of the cells cultured on its surface. The PPy- and PEDOT-coated electrodes may be useful to culture the cells on.

  19. Autophagic response to cell culture stress in pluripotent stem cells.

    Science.gov (United States)

    Gregory, Siân; Swamy, Sushma; Hewitt, Zoe; Wood, Andrew; Weightman, Richard; Moore, Harry

    2016-05-01

    Autophagy is an important conserved cellular process, both constitutively as a recycling pathway for long lived proteins and as an upregulated stress response. Recent findings suggest a fundamental role for autophagic processes in the maintenance of pluripotent stem cell function. In human embryonic stem cells (hESCS), autophagy was investigated by transfection of LC3-GFP to visualize autophagosomes and with an antibody to LC3B protein. The presence of the primary cilium (PC) in hESCs as the site of recruitment of autophagy-related proteins was also assessed. HESCs (mShef11) in vitro displayed basal autophagy which was upregulated in response to deprivation of culture medium replacement. Significantly higher levels of autophagy were exhibited on spontaneous differentiation of hESCs in vitro. The PC was confirmed to be present in hESCs and therefore may serve to coordinate autophagy function. PMID:26385182

  20. Horizontally rotated cell culture system with a coaxial tubular oxygenator

    Science.gov (United States)

    Wolf, David A. (Inventor); Schwarz, Ray P. (Inventor); Trinh, Tinh T. (Inventor)

    1991-01-01

    The present invention relates to a horizontally rotating bioreactor useful for carrying out cell and tissue culture. For processing of mammalian cells, the system is sterilized and fresh fluid medium, microcarrier beads, and cells are admitted to completely fill the cell culture vessel. An oxygen containing gas is admitted to the interior of the permeable membrane which prevents air bubbles from being introduced into the medium. The cylinder is rotated at a low speed within an incubator so that the circular motion of the fluid medium uniformly suspends the microbeads throughout the cylinder during the cell growth period. The unique design of this cell and tissue culture device was initially driven by two requirements imposed by its intended use for feasibility studies for three dimensional culture of living cells and tissues in space by JSC. They were compatible with microgravity and simulation of microgravity in one G. The vessels are designed to approximate the extremely quiescent low shear environment obtainable in space.

  1. Biona-C Cell Culture pH Monitoring System

    Science.gov (United States)

    Friedericks, C.

    1999-01-01

    Sensors 2000! is developing a system to demonstrate the ability to perform accurate, real-time measurements of pH and CO2 in a cell culture media in Space. The BIONA-C Cell Culture pH Monitoring System consists of S2K! developed ion selective sensors and control electronics integrated with the fluidics of a cell culture system. The integrated system comprises a "rail" in the Cell Culture Module (CCM) of WRAIR (Space Biosciences of Walter Read Army Institute of Research). The CCM is a Space Shuttle mid-deck locker experiment payload. The BIONA-C is displayed along with associated graphics and text explanations. The presentation will stimulate interest in development of sensor technology for real-time cell culture measurements. The transfer of this technology to other applications will also be of interest. Additional information is contained in the original document.

  2. PECULIARITIES OF SECONDARY METABOLITES BIOSYNTHESIS IN PLANT CELL CULTURES

    Directory of Open Access Journals (Sweden)

    A.M. NOSOV

    2014-06-01

    Full Text Available metabolites formation in plant cell cultures of Panax spp., (ginsenosides; Dioscorea deltoidea (steroid glycosides; Ajuga reptans, Serratula coronata, Rhaponticum carthamoides (ecdisteroids; Polyscias spp., (triterpene glycosides, Taxus spp. (taxoids, Stevia rebaudiana (diterpene steviol-glycosides, Stephania glabra (alkaloids. They are some regular trends of secondary metabolites synthesis in the plant cell culture:It can be noted the stable synthesis of the compound promoting cell proliferation. Indeed, cell cultures of Dioscorea deltoidea were demonstrated to accumulate only furostanol glycosides, which promoted cell division. Furostanol glycoside content of Dioscorea strain DM-0.5 was up to 6 - 12% by dry biomass.Panax ginseng and P. japonicus plant cell cultures synthesize as minimum seven triterpene glycosides (ginsenosides, the productivity of these compounds was up to 6.0 - 8.0% on dry biomass.By contrast, the detectable synthesis of diterpene steviol-glycosides in cultivated cells of Stevia rebaudiana initiated in the mixotrophic cultures during chloroplast formation only.Despite these differences, or mainly due to them, plant cell cultures have become an attractive source of phytochemicals in alternative to collecting wild plants. It provides a guideline to bioreactor-based production of isoprenoids using undifferentiated plant cell cultures

  3. A Continuous-Exchange Cell-Free Protein Synthesis System Based on Extracts from Cultured Insect Cells

    OpenAIRE

    Marlitt Stech; Quast, Robert B.; Rita Sachse; Corina Schulze; Wüstenhagen, Doreen A.; Stefan Kubick

    2014-01-01

    In this study, we present a novel technique for the synthesis of complex prokaryotic and eukaryotic proteins by using a continuous-exchange cell-free (CECF) protein synthesis system based on extracts from cultured insect cells. Our approach consists of two basic elements: First, protein synthesis is performed in insect cell lysates which harbor endogenous microsomal vesicles, enabling a translocation of de novo synthesized target proteins into the lumen of the insect vesicles or, in the case ...

  4. Morphological and Immunohistochemical Characterization of Canine Osteosarcoma Spheroid Cell Cultures.

    Science.gov (United States)

    Gebhard, C; Gabriel, C; Walter, I

    2016-06-01

    Spheroid cell culture emerges as powerful in vitro tool for experimental tumour research. In this study, we established a scaffold-free three-dimensional spheroid system built from canine osteosarcoma (OS) cells (D17). Spheroids (7, 14 and 19 days of cultivation) and monolayer cultures (2 and 7 days of cultivation) were evaluated and compared on light and electron microscopy. Monolayer and spheroid cultures were tested for vimentin, cytokeratin, alkaline phosphatase, osteocalcin and collagen I by means of immunohistochemistry. The spheroid cell culture exhibited a distinct network of collagen I in particular after 19-day cultivation, whereas in monolayer cultures, collagen I was arranged as a lamellar basal structure. Necrotic centres of large spheroids, as observed in 14- and 19-day cultures, were characterized by significant amounts of osteocalcin. Proliferative activity as determined by Ki-67 immunoreactivity showed an even distribution in two-dimensional cultures. In spheroids, proliferation was predominating in the peripheral areas. Metastasis-associated markers ezrin and S100A4 were shown to be continuously expressed in monolayer and spheroid cultures. We conclude that the scaffold-free spheroid system from canine OS cells has the ability to mimic the architecture of the in vivo tumour, in particular cell-cell and cell-matrix interactions. PMID:26287450

  5. Obtaining barley haploid embryos and seedlings using anther culture technique

    International Nuclear Information System (INIS)

    The effect of three barley genotypes (Igri, Arabi abiad, and Taqa 76), three irradiation doses (0, 5, and 10 Gy), and two media (FW, modified FW), on the number of formed embryos, and the ratio between regenerated embryos to green seedlings and albinos, were studied using anther culture. Also the study involved the compatibility between seedling morphology and chromosome number. results indicated significant differences among the genotypes, and media in callus and embryos formation and also in the ratio and albino seedlings. However, the effect of gamma rays dose was significant only on embryos regeneration. A high percentage of compatibility (90%) was obtained between the seedling morphology and chromosome number. (author)

  6. Culturing of PC12 Cells, Neuronal Cells, Astrocytes Cultures and Brain Slices in an Open Microfluidic System

    DEFF Research Database (Denmark)

    Al Atraktchi, Fatima Al-Zahraa; Bakmand, Tanya; Rømer Sørensen, Ane;

    culturing PC12 cells, neuronal cells, astrocytes cultures and brain slices. The microfluidic system provides efficient nutrient delivery, waste removal, access to oxygen, fine control over the neurochemical environment and access to modern microscopy. Additionally, the setup consists of an in vitro...

  7. Human skeletal muscle-derived stem cells retain stem cell properties after expansion in myosphere culture

    International Nuclear Information System (INIS)

    Human skeletal muscle contains an accessible adult stem-cell compartment in which differentiated myofibers are maintained and replaced by a self-renewing stem cell pool. Previously, studies using mouse models have established a critical role for resident stem cells in skeletal muscle, but little is known about this paradigm in human muscle. Here, we report the reproducible isolation of a population of cells from human skeletal muscle that is able to proliferate for extended periods of time as floating clusters of rounded cells, termed 'myospheres' or myosphere-derived progenitor cells (MDPCs). The phenotypic characteristics and functional properties of these cells were determined using reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry and immunocytochemistry. Our results showed that these cells are clonogenic, express skeletal progenitor cell markers Pax7, ALDH1, Myod, and Desmin and the stem cell markers Nanog, Sox2, and Oct3/4 significantly elevated over controls. They could be maintained proliferatively active in vitro for more than 20 weeks and passaged at least 18 times, despite an average donor-age of 63 years. Individual clones (4.2%) derived from single cells were successfully expanded showing clonogenic potential and sustained proliferation of a subpopulation in the myospheres. Myosphere-derived cells were capable of spontaneous differentiation into myotubes in differentiation media and into other mesodermal cell lineages in induction media. We demonstrate here that direct culture and expansion of stem cells from human skeletal muscle is straightforward and reproducible with the appropriate technique. These cells may provide a viable resource of adult stem cells for future therapies of disease affecting skeletal muscle or mesenchymal lineage derived cell types.

  8. Human skeletal muscle-derived stem cells retain stem cell properties after expansion in myosphere culture

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Yan [Department of Otolaryngology, Head and Neck Surgery Charite-Universitaetsmedizin Berlin, Berlin (Germany); Department of Otolaryngology, Head and Neck Surgery, The Third Affiliated Hospital of Sun Yat-sen University, Guang Zhou (China); Li, Yuan [Department of Otolaryngology, Head and Neck Surgery, The Third Affiliated Hospital of Sun Yat-sen University, Guang Zhou (China); Chen, Chao; Stoelzel, Katharina [Department of Otolaryngology, Head and Neck Surgery Charite-Universitaetsmedizin Berlin, Berlin (Germany); Kaufmann, Andreas M. [Clinic for Gynecology CCM/CBF, Charite-Universitaetsmedizin Berlin, Berlin (Germany); Albers, Andreas E., E-mail: andreas.albers@charite.de [Department of Otolaryngology, Head and Neck Surgery Charite-Universitaetsmedizin Berlin, Berlin (Germany)

    2011-04-15

    Human skeletal muscle contains an accessible adult stem-cell compartment in which differentiated myofibers are maintained and replaced by a self-renewing stem cell pool. Previously, studies using mouse models have established a critical role for resident stem cells in skeletal muscle, but little is known about this paradigm in human muscle. Here, we report the reproducible isolation of a population of cells from human skeletal muscle that is able to proliferate for extended periods of time as floating clusters of rounded cells, termed 'myospheres' or myosphere-derived progenitor cells (MDPCs). The phenotypic characteristics and functional properties of these cells were determined using reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry and immunocytochemistry. Our results showed that these cells are clonogenic, express skeletal progenitor cell markers Pax7, ALDH1, Myod, and Desmin and the stem cell markers Nanog, Sox2, and Oct3/4 significantly elevated over controls. They could be maintained proliferatively active in vitro for more than 20 weeks and passaged at least 18 times, despite an average donor-age of 63 years. Individual clones (4.2%) derived from single cells were successfully expanded showing clonogenic potential and sustained proliferation of a subpopulation in the myospheres. Myosphere-derived cells were capable of spontaneous differentiation into myotubes in differentiation media and into other mesodermal cell lineages in induction media. We demonstrate here that direct culture and expansion of stem cells from human skeletal muscle is straightforward and reproducible with the appropriate technique. These cells may provide a viable resource of adult stem cells for future therapies of disease affecting skeletal muscle or mesenchymal lineage derived cell types.

  9. Degradation of high density lipoprotein in cultured rat luteal cells

    International Nuclear Information System (INIS)

    In rat ovary luteal cells, degradation of high density lipoprotein (HDL) to tricholoracetic acid (TCA)-soluble products accounts for only a fraction of the HDL-derived cholesterol used for steroidogenesis. In this study the authors have investigated the fate of 125I]HDL bound to cultured luteal cells using pulse-chase technique. Luteal cell cultures were pulse labeled with [125I]HDL3 and reincubated in the absence of HDL. By 24 h about 50% of the initallay bound radioactivity was released into the medium, of which 60-65% could be precipitated with 10% TCA. Gel filtration of the chase incubation medium on 10% agarose showed that the amount of TCA-soluble radioactivity was nearly completely accounted for by a sharp peak in the low molecular weight region which was identified as 96% monoiodotyrosine by paper chromatography. The TCA-precipitable radioactivity was nearly completely accounted for by a sharp peak in the low molecular weight region which was identified as 96% monoiodotyrosine by paper chromatography. The TCA-precipitable radioactivity eluted over a wide range of molecular weights (15,000-80,000), and there was very little intact HDL present. Electrophoresis of the chase medium showed that component of the TCA-precipitable portion had mobility similar to apo AI. Lysosomal inhibitors of receptor-mediated endocytosis had no effect on the composition or quantity of radioactivity released during chase incubation. The results show that HDL3 binding to luteal cells is followed by complete degradation of the lipoprotein, although the TCA-soluble part does not reflect the extent of degradation

  10. Project 'Use of nuclear techniques in investigation, conservation and management of the cultural historical patrimony

    International Nuclear Information System (INIS)

    This project is aimed at solving problems of conservation of the cultural historical patrimony through the active participation of the member countries of ARCAL by the application of Analytic Nuclear Techniques

  11. Microglial cells in astroglial cultures: a cautionary note

    Directory of Open Access Journals (Sweden)

    Saura Josep

    2007-10-01

    Full Text Available Abstract Primary rodent astroglial-enriched cultures are the most popular model to study astroglial biology in vitro. From the original methods described in the 1970's a great number of minor modifications have been incorporated into these protocols by different laboratories. These protocols result in cultures in which the astrocyte is the predominant cell type, but astrocytes are never 100% of cells in these preparations. The aim of this review is to bring attention to the presence of microglia in astroglial cultures because, in my opinion, the proportion of and the role that microglial cells play in astroglial cultures are often underestimated. The main problem with ignoring microglia in these cultures is that relatively minor amounts of microglia can be responsible for effects observed on cultures in which the astrocyte is the most abundant cell type. If the relative contributions of astrocytes and microglia are not properly assessed an observed effect can be erroneously attributed to the astrocytes. In order to illustrate this point the case of NO production in activated astroglial-enriched cultures is examined. Lipopolysaccharide (LPS induces nitric oxide (NO production in astroglial-enriched cultures and this effect is very often attributed to astrocytes. However, a careful review of the published data suggests that LPS-induced NO production in rodent astroglial-enriched cultures is likely to be mainly microglial in origin. This review considers cell culture protocol factors that can affect the proportion of microglial cells in astroglial cultures, strategies to minimize the proportion of microglia in these cultures, and specific markers that allow the determination of such microglial proportions.

  12. Cultural Variations in Mothers' Acceptance of and Intent to Use Behavioral Child Management Techniques

    Science.gov (United States)

    Mah, Janet W. T.; Johnston, Charlotte

    2012-01-01

    We examined cultural differences in mothers' acceptance of and intent to use behavioral parenting techniques for managing disruptive child behavior, and the possible roles of parenting styles and implicit theories in explaining these cultural differences. A community sample of 117 Euro-Canadian and Chinese-immigrant mothers of boys aged 4- to…

  13. EFFECTS OF PDGF-BB ON INTRACELLULAR CALCIUM CONCENTRATION AND PROLIFERATION IN CULTURED GLOMERULAR MESANGIAL CELLS

    Institute of Scientific and Technical Information of China (English)

    WEN Li-ping; ZHANG Chong; BIAN Fan; ZOU Jun; JIANG Geng-ru; ZHU Han-wei

    2006-01-01

    Objective To investigate the relationship between the alteration of intracellular calcium concentration and proliferation in cultured glomerular mesangial cells. Methods Rat mesangial cells were cultured.Intracellular calcium concentrations were measured by confocal Laser Scanning Microscopy and Fura-3 fluorescence dyeing techniques. Cell growth was measured by MTT assay. Results PDGF-BB increased intracellular calcium concentrations in a dose-dependent manner, and at the same time promote the proliferation of mesangial cells. After preincubation with calcium channel blocker nifedipine or angiotensin converting enzyme inhibitor captopril, both the increase of intracellular calcium concentrations and cell proliferations induced by PDGF-BB were inhibited. Tripterigium Wilfordii Glycosides (TMG) significantly inhibited the mesangial cell proliferations, but it had no significant effect on intracellular calcium concentrations. Conclusion There was a positive relationship between the elevation of intracellular calcium concentration and cell proliferation in glomerular mesangial cells, but the increase of in- tracellular calcium concentrations wasn't the only way for proliferation.

  14. Isolation, Culture, and Maintenance of Mouse Intestinal Stem Cells

    Science.gov (United States)

    O’Rourke, Kevin P.; Ackerman, Sarah; Dow, Lukas E; Lowe, Scott W

    2016-01-01

    In this protocol we describe our modifications to a method to isolate, culture and maintain mouse intestinal stem cells as crypt-villus forming organoids. These cells, isolated either from the small or large intestine, maintain self-renewal and multilineage differentiation potential over time. This provides investigators a tool to culture wild type or transformed intestinal epithelium, and a robust assay for stem cell tissue homeostasis in vitro.

  15. SPONTANEOUS TRANSFORMATION OF CULTURED PORCINE BONE MARROW STROMAL CELLS

    DEFF Research Database (Denmark)

    Zou, Lijin; Zou, Xuenong; Li, Haisheng;

    -term culture are transformed into malignant cells. MATERIAL AND METHODS BMSC from 6 pigs were isolated and propagated continuously. Cell morphology was observed. Transformation properties were evaluated by means of serum dependence assay, Ki- 67 immunostaining, soft agar colony assay, karyotyping, telomerase...... was increased and TGF‚ signaling pathway was upregulated. However, telomerase activity maintained negative during culture. CONCLUSION Porcine BMSC can undergo spontaneous transformation, which provides a useful model to study the mechanisms associated with the tumorigenic potential of adult stem cells....

  16. Effects of methyl isocyanate on rat brain cells in culture.

    Science.gov (United States)

    Anderson, D; Goyle, S; Phillips, B J; Tee, A; Beech, L; Butler, W H

    1990-09-01

    Since the disaster in Bhopal, India, people exposed to methyl isocyanate (MIC) have complained of various disorders including neuromuscular dysfunction. In an attempt to get information about such dysfunction we have previously shown that MIC can affect muscle cells in culture. The present communication reports investigations into the effect of MIC on brain cells in culture. MIC was toxic to brain cells and the response was dose related. The observations were supported by light and electron microscopy. PMID:2207030

  17. Cultural Techniques of Cognitive Capitalism: Metaprogramming and the Labour of Code

    OpenAIRE

    Jussi Parikka

    2014-01-01

    This article addresses cultural techniques of cognitive capitalism. The author argues that to understand the full implications of the notion of cognitive capitalism we need to address the media and cultural techniques which conditions its range and applications. The article offers an expanded understanding of the labour of code and programming through a case study of 'metaprogramming', a software related organisation practice that offered a way to think of software creativity and programming ...

  18. Micropropagation of Dalbergia sissoo Roxb. through tissue culture technique.

    Science.gov (United States)

    Sahu, Jyoti; Khan, Shagufta; Sahu, Ram Kumar; Roy, Amit

    2014-04-01

    Multiple shoots of Dalbergia sissoo Roxb. (Sissoo) were incited from seeds through indirect somatic embryogenesis method. Seeds were inoculated in Murashige and Skoog's medium without any growth hormone. Than cotyledonary leaves were struck and used for callus induction on MS medium amplified with 2, 4-dichlorophenoxyacetic acid (0.5 to 4 mg mL(-1)). After 3 to 4 weeks the embryogenic callus clumps was transferred to medium supplemented with cytokinin (BAP 1 to 5 mg L(-1), kinetin 1-5.0 mg L(-1)) for embryo maturation and germination. The high-frequency shoot proliferation (82%) and maximum number of shoots per explants were recorded in MS medium containing NAA (0.5)+BAP (0.5). The findings of recent investigations have shown that, it is possible to induce indirect somatic embryogenesis in Dalbergia sissoo and plant regeneration from callus cultures derived from cotyledonary leaves as explants. PMID:25911856

  19. Chemotherapy in heterogeneous cultures of cancer cells with interconversion

    International Nuclear Information System (INIS)

    Recently, the interconversion between differentiated and stem-like cancer cells has been observed. Here, we model the in vitro growth of heterogeneous cell cultures in the presence of interconversion from differentiated cancer cells to cancer stem cells (CSCs), showing that, by targeting only CSC with cytotoxic agents, it is not always possible to eradicate cancer. We have determined the kinetic conditions under which cytotoxic agents in in vitro heterogeneous cultures of cancer cells eradicate cancer. In particular, we have shown that the chemotherapeutic elimination of in vitro cultures of heterogeneous cancer cells is effective only if it targets all cancer cell types, and if the induced death rates for the different subpopulations of cancer cell types are large enough. The quantitative results of the model are compared and validated with experimental data. (paper)

  20. A sliding cell technique for diffusion measurements in liquid metals

    OpenAIRE

    Yongliang Geng; Chunao Zhu; Bo Zhang

    2014-01-01

    The long capillary and shear cell techniques are the usual methods for diffusion measurements in liquid metals. Here we present a new “sliding cell technique” to measure interdiffusion in liquid alloys, which combines the merits of these two methods. Instead of a number of shear cells, as used in the shear cell method, only one sliding cell is designed to separate and join the liquid diffusion samples. Using the sliding cell technique, the influence of the heating process (which affects liqui...

  1. Feeding Frequency Affects Cultured Rat Pituitary Cells in Low Gravity

    Science.gov (United States)

    Hymer, W. C.; Grindeland, R. E.; Salada, T.; Cenci, R.; Krishnan, K.; Mukai, C.; Nagaoka, S.

    1996-01-01

    In this report, we describe the results of a rat pituitary cell culture experiment done on STS-65 in which the effect of cell feeding on the release of the six anterior pituitary hormones was studied. We found complex microgravity related interactions between the frequency of cell feeding and the quantity and quality (i.e. biological activity) of some of the six hormones released in flight. Analyses of growth hormone (GH) released from cells into culture media on different mission days using gel filtration and ion exchange chromatography yielded qualitatively similar results between ground and flight samples. Lack of cell feeding resulted in extensive cell clumping in flight (but not ground) cultures. Vigorous fibroblast growth occurred in both ground and flight cultures fed 4 times. These results are interpreted within the context of autocrine and or paracrine feedback interactions. Finally the payload specialist successfully prepared a fresh trypsin solution in microgravity, detached the cells from their surface and reinserted them back into the culture chamber. These cells reattached and continued to release hormone in microgravity. In summary, this experiment shows that pituitary cells are microgravity sensitive and that coupled operations routinely associated with laboratory cel1 culture can also be accomplished in low gravity.

  2. Mammosphere culture of cancer stem cells in a microfluidic device

    Science.gov (United States)

    Saadin, Katayoon; White, Ian M.

    2012-03-01

    It is known that tumor-initiating cells with stem-like properties will form spherical colonies - termed mammospheres - when cultured in serum-free media on low-attachment substrates. Currently this assay is performed in commercially available 96-well trays with low-attachment surfaces. Here we report a novel microsystem that features on-chip mammosphere culture on low attachment surfaces. We have cultured mammospheres in this microsystem from well-studied human breast cancer cell lines. To enable the long-term culture of these unattached cells, we have integrated diffusion-based delivery columns that provide zero-convection delivery of reagents, such as fresh media, staining agents, or drugs. The multi-layer system consists of parallel cell-culture chambers on top of a low-attachment surface, connected vertically with a microfluidic reagent delivery layer. This design incorporates a reagent reservoir, which is necessary to reduce evaporation from the cell culture micro-chambers. The development of this microsystem will lead to the integration of mammosphere culture with other microfluidic functions, including circulating tumor cell recovery and high throughput drug screening. This will enable the cancer research community to achieve a much greater understanding of these tumor initiating cancer stem cells.

  3. Epithelial cell detachment by Porphyromonas gingivalis biofilm and planktonic cultures.

    Science.gov (United States)

    Huang, Lijia; van Loveren, Cor; Ling, Junqi; Wei, Xi; Crielaard, Wim; Deng, Dong Mei

    2016-04-01

    Porphyromonas gingivalis is present as a biofilm at the sites of periodontal infections. The detachment of gingival epithelial cells induced by P. gingivalis biofilms was examined using planktonic cultures as a comparison. Exponentially grown planktonic cultures or 40-h biofilms were co-incubated with epithelial cells in a 24-well plate for 4 h. Epithelial cell detachment was assessed using imaging. The activity of arginine-gingipain (Rgp) and gene expression profiles of P. gingivalis cultures were examined using a gingipain assay and quantitative PCR, respectively. P. gingivalis biofilms induced significantly higher cell detachment and displayed higher Rgp activity compared to the planktonic cultures. The genes involved in gingipain post-translational modification, but not rgp genes, were significantly up-regulated in P. gingivalis biofilms. The results underline the importance of including biofilms in the study of bacterial and host cell interactions. PMID:26963862

  4. [Continuous perfusion culture hybridoma cells for production of monoclonal antibody].

    Science.gov (United States)

    Mi, Li; Li, Ling; Feng, Qiang; Yu, Xiao-Ling; Chen, Zhi-Nan

    2002-05-01

    Hybridoma cells were cultured by continuous perfusion in Fibra-Cel of 5L packed-bed bioreactor for 22 days in low serum or serum-free media. The corresponded amino acids were fed and serum concentration was decreased by analyzing glucose concentration, oxygen uptake rate, secretary antibody amount and amino acids concentration in culture supernatant. Comparing with continuous perfusion culture that amino acids were not fed, antibody amount of production was increased about 2-3 times. The inoculated cell density was 2.5 x 10(5) cells/mL, while the final cell density was 8.79 x 10(8) cells/mL. Antibody production was reached 295 mg/L/d at average level, and the highest level was reached 532 mg/L/d. These results provided a primary mode of enlarge culture for monoclonal antibody industralization. PMID:12192875

  5. A practical guide to hydrogels for cell culture.

    Science.gov (United States)

    Caliari, Steven R; Burdick, Jason A

    2016-04-28

    There is growing appreciation of the role that the extracellular environment plays in regulating cell behavior. Mechanical, structural, and compositional cues, either alone or in concert, can drastically alter cell function. Biomaterials, and particularly hydrogels, have been developed and implemented to present defined subsets of these cues for investigating countless cellular processes as a means of understanding morphogenesis, aging, and disease. Although most scientists concede that standard cell culture materials (tissue culture plastic and glass) do a poor job of recapitulating native cellular milieus, there is currently a knowledge barrier for many researchers in regard to the application of hydrogels for cell culture. Here, we introduce hydrogels to those who may be unfamiliar with procedures to culture and study cells with these systems, with a particular focus on commercially available hydrogels. PMID:27123816

  6. THE ULTRASTRUCTURE OF SEPARATED AND CULTURED CELL OF PORPHYRA YEZOENSIS

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    There are many reports that cells (protoplasts) separated from the thallus of Porphyra by enzyme can develop to normal leafy thalli in the same way as monospores. But there are few investigations on the subcellular structure of the isolated vegetative cell for comparison with the subcellular structure of monospores. To clarify whether the separated and cultured cells undergo the same or similar ultrastructure changes during culture and germination as monospores undergo in their formation and germination, we observed their ultrastructure, compared them with those of the monospore and found that the ultrastructure of separated and cultured cells did not have the characteristic feature as that of monospore formation, such as production of small and large fibrous vesicles, but was accompanied by vacuolation and starch mobilization like that in monospore germination. The paper also discusses the relations between monospores and separated and cultured cells.

  7. Crystal Violet Assay for Determining Viability of Cultured Cells.

    Science.gov (United States)

    Feoktistova, Maria; Geserick, Peter; Leverkus, Martin

    2016-01-01

    Adherent cells detach from cell culture plates during cell death. This characteristic can be used for the indirect quantification of cell death and to determine differences in proliferation upon stimulation with death-inducing agents. One simple method to detect maintained adherence of cells is the staining of attached cells with crystal violet dye, which binds to proteins and DNA. Cells that undergo cell death lose their adherence and are subsequently lost from the population of cells, reducing the amount of crystal violet staining in a culture. This protocol describes a quick and reliable screening method that is suitable for the examination of the impact of chemotherapeutics or other compounds on cell survival and growth inhibition. However, characterization of the cause of reduced crystal violet staining requires additional methods detailed elsewhere. PMID:27037069

  8. Use of MALDI-TOF MS technique for rapid identification of bacteria from positive blood cultures

    Directory of Open Access Journals (Sweden)

    Sung Kuk Hong

    2014-01-01

    Full Text Available We evaluated the feasibility of same-day routine aerobic bacterial identification using the following procedures: Picking colonies from 4 and 6 h incubated subculture from positive blood culture bottle and analyzing them by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS. The matched identification rate of this procedure at the species level was 80.6% (141/175 for the 4-h cultures compared with overnight cultures and 90.9% (159/175 for the 6-h cultures. Thus, our technique provides an easy and rapid method for identification of aerobic bacteria in routine clinical microbiology laboratories.

  9. Controlling the diversity of cell populations in a stem cell culture

    NARCIS (Netherlands)

    Heo, Inha; Clevers, Hans

    2015-01-01

    Culturing intestinal stem cells into 3D organoids results in heterogeneous cell populations, reflecting the in vivo cell type diversity. In a recent paper published in Nature, Wang et al. established a culture condition for a highly homogeneous population of intestinal stem cells.

  10. A Place to Call Home: Bioengineering Pluripotential Stem Cell Cultures

    OpenAIRE

    Mark Weingarten; Nathan Akhavan; Joshua Hanau; Yakov Peter

    2015-01-01

    Pluripotent stem cells (PSCs) have the power to revolutionize the future of cell-based therapies and regenerative medicine. However, stem/progenitor cell use in the clinical arsenal has been hampered by discrepancies resulting from stem cell engineering and expansion, as well as in their (mass) differentiation in culture. Moreover, the manner in which external conditions affect PSC and induced-pluripotent stem cell lineage establishment as well as maturation remains controversial. In this rev...

  11. New technique to register proliferation of clonogenic cells from brain tumors

    Energy Technology Data Exchange (ETDEWEB)

    Unsgaard, G.; Larsen, B.; Dalen, A.; Vik, R.; Ringkjob, R.

    1985-01-01

    The soft agar technique for culturing human clonogenic tumor cells has been usefully applied for predicting individual clinical responses to chemotherapy, for screening of new antineoplastic drugs, and in basic biological research. The counting of colonies formed by clonogenic cells is, however, a rather time consuming and inaccurate procedure. The authors report a method to combine the easy and precise registration of DNA-synthesis by TH-thymidine incorporation with the ability of soft agar to permit proliferation of clonogenic cells and inhibit proliferation of non-neoplastic cells. The glioma cell lines U 251 MG and T-MG 1, the benignant glia cells T-BG 1, T-BG 2, T-BG 3 and fibroblasts were cultured in Furcellaran gel. Twenty hours before harvesting TH-thymidine was added. TH-thymidine incorporation in malignant cells increased exponentially with time, while TH-thymidine incorporation in the benignant glia cells and fibroblasts was inhibited. The correlation between number of colonies counted after 16 days and TH-thymidine incorporation registered after different culture times was very good. The correlation was best when the cultures were harvested after 8 days. The most intense proliferation seemed to take place during the first week in culture. The good correlation between TH-thymidine incorporation on day 7 and colony number on day 14, indicate that reduction of assay time is possible also for the glioma biopsies.

  12. Air pollutant production by algal cell cultures

    Science.gov (United States)

    Fong, F.; Funkhouser, E. A.

    1982-01-01

    The production of phytotoxic air pollutants by cultures of Chlorella vulgaris and Euglena gracilis is considered. Algal and plant culture systems, a fumigation system, and ethylene, ethane, cyanide, and nitrogen oxides assays are discussed. Bean, tobacco, mustard green, cantaloupe and wheat plants all showed injury when fumigated with algal gases for 4 hours. Only coleus plants showed any resistance to the gases. It is found that a closed or recycled air effluent system does not produce plant injury from algal air pollutants.

  13. Insect cell culture and applications to research and pest management

    Science.gov (United States)

    Building on earlier research, insect cell culture began with the successful establishment of one cell line from pupal ovarian tissue. The field has grown to the extent that now over 500 insect cell lines have been established from many insect species representing numerous insect Orders and from seve...

  14. Primary cell culture from human oral tissue: gingival keratinocytes,gingival fibroblasts and periodontal ligament fibroblasts

    Directory of Open Access Journals (Sweden)

    Supreya Wanichpakorn

    2010-08-01

    Full Text Available Primary cell culture of human oral tissue has many applications for oral biology research. There are two techniques in primary culture, which includes the enzymatic and direct explant technique. The objectives of this study were (1 to isolate and investigate the difference in percentage the success in culturing three cell types from human oral tissue: gingival keratinocytes, gingival fibroblasts and periodontal ligament fibroblasts by using the direct explant technique; (2 to compare the effect of sex and age on the success of tissue culturing. Twenty seven tissue samples were obtained from healthy human gingival tissue, 19 female and 8 male patients aged 14-67 years (37.7±17.5. The tissue was cut into 1x1 mm pieces and placed on plastic culture plates containing Dulbecco’s Modified Eagle’s Medium supplemented with 10% fetal calf serum, 100 U/ml penicillin, 100 µg/ml streptomycin and 1% amphotericin B. For the keratinocytes culture, after the epithelial cells started to multiply around the gingival origin and the diameter was 2-5 mm., the fibroblasts were liminated by mechanical removal under inverted microscope to prevent fibroblast overgrowth and the medium was changed to keratinocyte-SFM (Gibco, BRL supplemented with 5 µg/ml gentamycin. The results revealed that gingival fibroblast gave the highest success rate in culture (96.3%, followed by gingival keratinocytes (88.9% and periodontal ligament fibroblasts (81.5%. There was no significant difference in the success rate of cultivation between younger and older individuals, as between sex of the subjects (p>0.05. The risk of failure in culture techniques is mainly caused by microbiological contamination from the tissue samples.

  15. Detection and Treatment of Mycoplasma Contamination in Cultured Cells

    Directory of Open Access Journals (Sweden)

    Hsuan Jung

    2003-04-01

    Full Text Available Background: Mycoplasmas, the smallest and simplest prokaryotes that reside in endosomesof mammalian cells, are widespread contaminants found in cell cultures.About 30% of all cell cultures, varying from 15 to 80%, are reportedlycontaminated with mycoplasmas. Here, we present our experience in successfullydetecting and treating mycoplasmal infection in various cell lines.Methods: The nested polymerase chain reaction (PCR detection and microscopicexamination, including phase-contrast, fluorescent, as well as differentialinterference contrast, were used for detecting potential mycoplasma contaminationof cell lines used in our laboratory. As soon as mycoplasma was identified,antibiotic treatment was initiated.Results: Mycoplasmal contamination was detected in six of 15 cell lines using thenested PCR amplification of mycoplasma DNA, which was further demonstratedusing 4, 6-Diamidino-2-phenylindole (DAPI staining and fluorescentmicroscopy. Alternate treatment with two antibiotics, macrolide (tiamulinand tetracycline (minocycline, effectively eliminated mycoplasma, whichwas validated by both PCR and microscopic studies.Conclusions: The nested PCR using genomic DNA extracted from cultured cells as templatesis a rapid and sensitive method for detecting mycoplasma contamination.Treatment with combined antibiotics can completely eradicatemycoplasmal infection from cultured cells. For the ease of use, PCR and/orDAPI staining appear suitable for detecting potential mycoplasmal contaminationin laboratories that rely heavily on the cell culture system.

  16. The effects of glucocorticoids on cultured human endothelial cells.

    Science.gov (United States)

    Maca, R D; Fry, G L; Hoak, J C

    1978-04-01

    The effects of hydrocortisone, dexamethasone and prednisone on the morphology, replication, DNA synthesis, cell protein content and protein synthesis of cultured, human endothelial cells were evaluated. After culturing the cells with these glucocorticoids for 24-48 h, the cells covered a greater portion of the culture surface area. The mean surface area of the individual endothelial cell treated with glucocorticoids was 1.53 times greater than that of the untreated control endothelial cell. When compared with controls, the endothelial cover provided by the cells treated with glucocorticoids was more extensive and in many instances covered the entire culture surface. The change in morphology was associated with an increase in protein synthesis and protein content of the cells without an increase in DNA synthesis or cellular replication. Dexamethasone was approximately 10-fold more effective than hydrocortisone, while prednisone was the least effective. Aldosterone, DOCA, testosterone, progesterone, oestradiol and oestriol were ineffective. These studies indicate that glucocorticoids can alter the morphology and biochemistry of cultured endothelial cells and may have implications for the effects of steroids in the treatment of thrombocytopenic states and vascular disorders in man. PMID:646949

  17. Preliminary analysis of cellular sociology of co-cultured glioma initiating cells and macrophages in vitro

    Institute of Scientific and Technical Information of China (English)

    Mingxia Zhang; Xingliang Dai; Xiaonan Li; Qiang Huang; Jun Dong; Junjie Chen; Lin Wang; Xiaoyan Ji; Lin Yang; Yujing Sheng; Hairui Liu; Haiyang Wang; Aidong Wang

    2016-01-01

    Objective:Real-time monitoring of cytokine secretion at the single immunocyte level, based on the concept of immune cells, sociology has been recently reported. However, the relationships between glioma-initiating cells (GICs) and host immune cells and their mutual interactions in the tumor microenvironment have not been directly observed and remain unclear. Methods:The dual fluorescence tracing technique was applied to label the co-cultured GICs and host macrophages (Mø), and the interactions between the two types of cells were observed using a live cell imaging system. Fusion cells in the co-culture system were monocloned and proliferated in vitro and their social interactions were observed and recorded. Results:Using real-time dynamic observation of target cells, 6 types of intercellular conjunction microtubes were found to function in the transfer of intercellular information between GICs and Mø;GICs and host Mø can fuse into hybrid cells after several rounds of mutual interactions, and then these fusion cells fused with each other;Fusion cells generated offspring cells through symmetrical and asymmetrical division or underwent apoptosis. A“cell in cell” phenomenon was observed in the fusion cells, which was often followed by cell release, namely entosis. Conclusions:Preliminary studies revealed the patterns of cell conjunction via microtubes between GICs and host Mø and the processes of cell fusion, division, and entosis. The results revealed malignant transformation of host Mø, induced by GICs, suggesting complex social relationships among tumor-immune cells in gliomas.

  18. Novel culturing platform for brain slices and neuronal cells

    DEFF Research Database (Denmark)

    Svendsen, Winnie Edith; Al Atraktchi, Fatima Al-Zahraa; Bakmand, Tanya;

    2015-01-01

    In this paper we demonstrate a novel culturing system for brain slices and neuronal cells, which can control the concentration of nutrients and the waste removal from the culture by adjusting the fluid flow within the device. The entire system can be placed in an incubator. The system has been te...

  19. Cell/Tissue Culture Radiation Exposure Facility Project

    Data.gov (United States)

    National Aeronautics and Space Administration — We propose to develop a Cell/Tissue Culture Radiation Exposure Facility (CTC-REF) to enable radiobiologists to investigate the real-time radiation effects on...

  20. Optimization of Human Corneal Endothelial Cells for Culture: The Removal of Corneal Stromal Fibroblast Contamination Using Magnetic Cell Separation

    Directory of Open Access Journals (Sweden)

    Gary S. L. Peh

    2012-01-01

    Full Text Available The culture of human corneal endothelial cells (CECs is critical for the development of suitable graft alternative on biodegradable material, specifically for endothelial keratoplasty, which can potentially alleviate the global shortage of transplant-grade donor corneas available. However, the propagation of slow proliferative CECs in vitro can be hindered by rapid growing stromal corneal fibroblasts (CSFs that may be coisolated in some cases. The purpose of this study was to evaluate a strategy using magnetic cell separation (MACS technique to deplete the contaminating CSFs from CEC cultures using antifibroblast magnetic microbeads. Separated “labeled” and “flow-through” cell fractions were collected separately, cultured, and morphologically assessed. Cells from the “flow-through” fraction displayed compact polygonal morphology and expressed Na+/K+ATPase indicative of corneal endothelial cells, whilst cells from the “labeled” fraction were mostly elongated and fibroblastic. A separation efficacy of 96.88% was observed. Hence, MACS technique can be useful in the depletion of contaminating CSFs from within a culture of CECs.

  1. Effect of Co-Culturing of Mice Liver Cells and Embryonic Carcinomatous Stem Cells on the Rate of Differentiation to Hematopoietic Cells

    Directory of Open Access Journals (Sweden)

    AA Pourfatollah

    2005-10-01

    Full Text Available Introduction: Considering the importance of co-culture in differentiation of embryonic stem cells, the aim of this study was evaluation of the effect of co-culturing fetal liver stroma cells with P19 cells on the line of differentiation. Materials and Methods: For this purpose, P19 cells were cultured directly in semisolid medium. These cells proliferated and primarily differentiated to colonies know as embryoid bodies (EBs after 8-12 days. The Ebs cells were trypsinized and dissociated to single or double cells. Then these cells were co-cultured on the mouse fetal liver feeder layer in the absence of exogenous factors. After 14-18 days, the colonies were studied morphologically by benzidine and giemsa staining and also counted under invert microscope. Results: The percentages of benzidine positive (or erythroid and negative colonies were 94% and 6% respectively and also the cells of colonies were studied by Giemsa staining. Results showed that they were myeloid or lymphoid type cells. Thus, the results show that in the presence of mouse fetal liver feeder layer, the number of erythroid colonies was increased. Conclusions: Therefore, this technique may be effective for differentiation of stem cells from different sources into hematopoietic cells and can be used in future for human cell therapy.

  2. Convoluted cells as a marker for maternal cell contamination in CVS cultures

    DEFF Research Database (Denmark)

    Hertz, Jens Michael; Jensen, P K; Therkelsen, A J

    1987-01-01

    In order to identify cells of maternal origin in CVS cultures, tissue from 1st trimester abortions were cultivated and the cultures stained in situ for X-chromatin. Convoluted cells and maternal fibroblasts were found to be positive. By chromosome analysis of cultures from 105 diagnostic placenta...... biopsies, obtained by the transabdominal route, metaphases of maternal origin were found in nine cases. In eight of these cases colonies of convoluted cells were observed. We conclude that convoluted cells are of maternal origin and are a reliable marker for maternal cell contamination in CVS cultures....

  3. Effects of UVC-irradiation on cultured mouse embryonic cells

    International Nuclear Information System (INIS)

    Effects of UVC-irradiation on the cultured differentiating mouse embryonic cells were investigated. Embryonic mesenchymal cells, isolated from fore-and hind-limbs or mid brain of Day 11 mouse embryos, and 3T3 cells, a reference mouse fibroblast cell line, were irradiated with UVC at a dose range of 0∼30 J/m2. Dose-dependent inhibition was found for both cellular proliferation and differentiation, dose-dependent induction of DNA cyclobutane pyrimidine dimers and (6-4) photoproducts were found in the embryonic cells. Mesenchymal chondrogenesis was more sensitive to the UVC than proliferation, and the UVC-induced DNA damage and their repair kinetics in the cultured embryonic cells were similar to those in mouse 3T3 cells. No effects of treatments by the fluorescent light pre or post UVC-irradiation were found on the repair kinetics of DNA damage in all of the cells

  4. Three-dimensional hydrogel cell culture systems for modeling neural tissue

    Science.gov (United States)

    Frampton, John

    designed for use as a tool to predict the transport and processing that occurs prior to drug uptake in the central nervous system (CNS), and to predict BBB permeability. Electrochemical techniques and immunohistochemistry were used to validate this model and provide detailed information about cellular organization and function. Electrochemical impedance spectroscopy (EIS) provided evidence that endothelial cells cultured in the presence of astrocytes formed tight junctions capable of occluding the flow of electrical current. In a second series of experiments, a microglia-astrocyte co-culture system was developed to assess the effects of glial cells on electrode impedance recorded from neural prosthetic devices in vitro. Impedance measurements were compared with confocal images to determine the effects of glial cell density and cell type on electrode performance. The results indicate that EIS data can be used to model components of the reactive cell responses in brain tissue, and that impedance measurements recorded in vitro can be compared to measurements recorded in vivo. Taken together, these results demonstrate that alginate hydrogels can be used for the creation of 3-D neural cell scaffolds, and that such cell scaffolds can be used to model a variety of three-dimensional neural tissues in vitro, that cannot be studied in 2-D cultures.

  5. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture

    Science.gov (United States)

    Lestard, Nathalia R.

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  6. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture.

    Science.gov (United States)

    Lestard, Nathalia R; Capella, Marcia A M

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  7. Time-lapse imaging assay using the BioStation CT: A sensitive drug-screening method for three-dimensional cell culture

    OpenAIRE

    Sakamoto, Ruriko; Rahman, M. Mamunur; SHIMOMURA, MANAMI; Itoh, Manabu; Nakatsura, Tetsuya

    2015-01-01

    Three-dimensional (3D) cell culture is beneficial for physiological studies of tumor cells, due to its potential to deliver a high quantity of cell culture information that is representative of the cancer microenvironment and predictive of drug responses in vivo. Currently, gel-associated or matrix-associated 3D cell culture is comprised of intricate procedures that often result in experimental complexity. Therefore, we developed an innovative anti-cancer drug sensitivity screening technique ...

  8. Qualitative study of three cell culture methods.

    Science.gov (United States)

    Wang, Aiguo; Xia, Tao; Ran, Peng; Chen, Xuemin; Nuessler, Andreas K

    2002-01-01

    Primary rat hepatocytes were cultured using different in vitro models and the enzyme leakage, albumin secretion, and cytochrome P450 1A (CYP 1A) activity were observed. The results showed that the level of LDH was decreased over time in culture. However, on day 5, LDH showed a significant increase in monolayer culture (MC) while after day 8 no LDH was detectable in sandwich culture (SC). The levels of AST and ALT did not change significantly over the investigated time. The CYP 1A activity was gradually decreased in a time-dependent manner in MC and SC. The decline of CYP 1A was faster in MC than in SC. This effect was partially reversed by using cytochrome P450 (CYP450) inducer such as Omeprazol and 3-methylcholanthrene (3-MC) and the CYP 1A induction was always higher in MC than in SC. In bioreactor basic CYP 1A activity was preserved over 2 weeks and the highest albumin production was observed in bioreactor followed by SC and MC. Taken together, it was indicated each investigated model had its advantages and disadvantages. It was also underlined that various in vitro models may address different questions. PMID:12674760

  9. [Effect evaluation of three cell culture models].

    Science.gov (United States)

    Wang, Aiguo; Xia, Tao; Yuan, Jing; Chen, Xuemin

    2003-11-01

    Primary rat hepatocytes were cultured using three kinds of models in vitro and the enzyme leakage, albumin secretion, and cytochrome P450 1A (CYP 1A) activity were observed. The results showed that the level of LDH in the medium decreased over time in the period of culture. However, on 5 days, LDH showed a significant increase in monolayer culture (MC) while after 8 days LDH was not detected in sandwich culture (SC). The levels of AST and ALT in the medium did not change significantly over the investigated time. The basic CYP 1A activity gradually decreased with time in MC and SC. The decline of CYP 1A in rat hepatocytes was faster in MC than that in SC. This effect was partially reversed by using cytochrome P450 (CYP450) inducers such as omeprazol and 3-methylcholanthrene (3-MC) and the CYP 1A induction was always higher in MC than that in SC. Basic CYP 1A activity in bioreactor was keeped over 2 weeks and the highest albumin production was observed in bioreactor, and next were SC and MC. In conclusion, our results clearly indicated that there have some advantages and disadvantages in each of models in which can address different questions in metabolism of toxicants and drugs. PMID:14963896

  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. Relationship between sensitivity to ultraviolet light and budding in yeast cells of different culture ages

    International Nuclear Information System (INIS)

    Subpopulations of yeast cells, consisting of cells of different sizes and different percentages of budding cells, were prepared by centrifugation through sucrose solutions with linear density gradients of cultures at different phases of the growth cycle. Ultraviolet survival of these cells was determined by colony counting, and the survival rate was compared with the cells' respiratory rates. Individual budding cells and interdivisional cells, and also mother cells and daughter cells derived from irradiated budding cells, were isolated by the micromanipulation technique. The number of divisions in each cell was measured during a 21-hr incubation period immediately after irradiation. In the population in the logarithmic phase consisting of homogeneous cells of middle size, no difference in uv sensitivity was observed between mother cells and daughter cells, irrespective of mutual adhesion. Budding cell resistance was observed in the population in the transitional phase; this was due to the lesser uv sensitivity of daughter cells in the fresh medium. In the stationary phase, daughter cells were rather more sensitive than mother cells or interdivisional cells, so there was little difference in uv sensitivity between budding cells and interdivisional cells

  12. Tumor targeting of humanized fragment antibody secreted from transgenic rice cell suspension culture

    DEFF Research Database (Denmark)

    Hong, Shin-Young; Lee, Tae-Sup; Kim, Ju; Jung, Jae-Ho; Choi, Chang-Woon; Kim, Tae-Geum; Kwon, Tae-Ho; Jang, Yong-Suk; Yang, Moon-Sik

    The tumor-associated glycoprotein 72 (TAG 72) has been shown to be expressed in the majority of human adenocarcinomas. In an effort to develop a technique for the safe and inexpensive production of large quantities of anti-TAG 72 humanized antibody fragments (hzAb) as a future source of clinical......-grade proteins, we developed a transgenic rice cell suspension culture system. The in vivo assembly and secretion of hzAb were achieved in a transgenic rice cell culture under the control of the rice alpha amylase 3D (RAmy 3D) expression system, and the biological activities of plant-derived hzAb were determined...

  13. Examination of rat embryo cell cultures for mycoplasma pulmonis with an autoradiographic method

    International Nuclear Information System (INIS)

    As a contribution to the possibility of a safe method for detection of Mycoplasma pulmonis contamination of rat embryos, embryos of experimentally infected, naturally infected and germfree rats were examined with a culture method, in cell culture for 3-thymidine incorporation and with the DNA(H-)stain, in tissue sections with immunofluorescence (FITC) and the peroxidase technique, and fetal membranes with scanning electron microscopy (SEM). Embryos of experimentally infected animals were always, of naturally infected animals mostly, and such of germfree animals often Mycoplasma pulmonis infected. 3-thymidine incorporation and H-stain in cell culture but also a two-phase culture system showed high sensitivity. For demonstration of the antigen in situ immunofluorescence, immunoperoxidase, and SEM were useful. (author)

  14. Immunoselection techniques in hematopoietic stem cell transplantation.

    Science.gov (United States)

    Li Pira, Giuseppina; Biagini, Simone; Cicchetti, Elisabetta; Merli, Pietro; Brescia, Letizia Pomponia; Milano, Giuseppe Maria; Montanari, Mauro

    2016-06-01

    Hematopoietic Stem Cells Transplantation (HSCT) is an effective treatment for hematological and non-hematological diseases. The main challenge in autologous HSCT is purging of malignant cells to prevent relapse. In allogeneic HSCT graft-versus-host disease (GvHD) and opportunistic infections are frequent complications. Two types of graft manipulation have been introduced: the first one in the autologous context aimed at separating malignant cells from hematopoietic stem cells (HSC), and the second one in allogeneic HSCT aimed at reducing the incidence of GvHD and at accelerating immune reconstitution. Here we describe the manipulations used for cell purging in autologous HSCT or for T Cell Depletion (TCD) and T cell selection in allogeneic HSCT. More complex manipulations, requiring a Good Manufacturing Practice (GMP) facility, are briefly mentioned. PMID:27209628

  15. Stability of resazurin in buffers and mammalian cell culture media

    DEFF Research Database (Denmark)

    Rasmussen, Eva; Nicolaisen, G.M.

    1999-01-01

    The utility of a ferricyanide/ferrocyanide system used in the AlamarBlue(TM) (Serotec, Oxford, UK) vital. dye to inhibit the reduction of resazurin by mammalian cell culture media is questioned. Resazurin was found to be relatively stable when dissolved in phosphate-buffered saline (PBS). The use...... of HEPES resulted in a huge immediate dye reduction, which was significantly enhanced by exposure to diffuse light from fluorescent tubes in the laboratory 8 h per day. The reduction of resazurin by various cell culture media was time and temperature dependent, and it was significantly enhanced by......'s nutrient mixture F-10 and F-12. Fetal calf serum (5-20%) slightly decreased resazurin reduction during the first 2 days of incubation. The reduction of resazurin by mammalian cell culture media do not appear to be problematic under normal culture conditions, and it is primarily dependent upon the presence...

  16. Detection of multiple mycoplasma infection in cell cultures by PCR

    Directory of Open Access Journals (Sweden)

    J. Timenetsky

    2006-07-01

    Full Text Available A total of 301 cell cultures from 15 laboratories were monitored for mycoplasma (Mollicutes using PCR and culture methodology. The infection was detected in the cell culture collection of 12 laboratories. PCR for Mollicutes detected these bacteria in 93 (30.9% samples. Although the infection was confirmed by culture for 69 (22.9% samples, PCR with generic primers did not detect the infection in five (5.4%. Mycoplasma species were identified with specific primers in 91 (30.2% of the 98 samples (32.6% considered to be infected. Mycoplasma hyorhinis was detected in 63.3% of the infected samples, M. arginini in 59.2%, Acholeplasma laidlawii in 20.4%, M. fermentans in 14.3%, M. orale in 11.2%, and M. salivarium in 8.2%. Sixty (61.2% samples were co-infected with more than one mycoplasma species. M. hyorhinis and M. arginini were the microorganisms most frequently found in combination, having been detected in 30 (30.6% samples and other associations including up to four species were detected in 30 other samples. Failure of the treatments used to eliminate mycoplasmas from cell cultures might be explained by the occurrence of these multiple infections. The present results indicate that the sharing of non-certified cells among laboratories may disseminate mycoplasma in cell cultures.

  17. Pedagigocal technique of building the culture of interpersonal relations in preschool children at art classes

    OpenAIRE

    Svetlana Vyacheslavovna Kahnovich

    2013-01-01

    The article looks at the pedagogical technique of building the culture of interpersonal relations in preschool children at the local and modular level. Interpersonal relations are viewed as the module and art classes as the local level. The research is timely as it can assist in studying the problem of moral development of preschool children by building the culture of interpersonal relations by artistic education means. The study presents novelty concluding from the survey of scientific liter...

  18. Voltage effects on cells cultured on metallic biomedical implants

    Science.gov (United States)

    Haerihosseini, Seyed Morteza

    Electrochemical voltage shifts in metallic biomedical implants occur in-vivo due to a number of processes including mechanically assisted corrosion. Surface potential of biomedical implants and excursions from resting open circuit potential (OCP), which is the voltage they attain while in contact with an electrolyte, can significantly change the interfacial properties of the metallic surfaces and alter the behavior of the surrounding cells, compromising the biocompatibility of metallic implants. Voltages can also be controlled to modulate cell function and fate. To date, the details of the physico-chemical phenomena and the role of different biomaterial parameters involved in the interaction between cells and metallic surfaces under cathodic bias have not been fully elucidated. In this work, changes in the interfacial properties of a CoCrMo biomedical alloy (ASTM F-1537) in phosphate-buffered saline (PBS) (pH 7.4) at different voltages was studied. Step polarization impedance spectroscopy technique was used to apply 50 mV voltage steps to samples, and the time-based current transients were recorded. A new equation was derived based on capacitive discharge through a Tafel element and generalized to deal with non-ideal impedance behavior. The new function compared to the KWW-Randles function, better matched the time-transient response. The results also showed a voltage dependent oxide resistance and capacitance behavior. Additionally, the in-vitro effect of static voltages on the behavior of MC3T3-E1 pre-osteoblasts cultured on CoCrMo alloy (ASTM-1537) was studied to determine the range of cell viability and mode of cell death beyond the viable range. Cell viability and morphology, changes in actin cytoskeleton, adhesion complexes and nucleus, and mode of cell death (necrosis, or intrinsic or extrinsic apoptosis) were characterized at different voltages ranging from -1000 to +500 mV (Ag/AgCl). Moreover, electrochemical currents and metal ion concentrations at each

  19. Integrin VLA-3: ultrastructural localization at cell-cell contact sites of human cell cultures

    OpenAIRE

    1989-01-01

    The integrin VLA-3 is a cell surface receptor, which binds to fibronectin, laminin, collagen type I and VI (Takada, Y., E. A. Wayner, W. G. Carter, and M. E. Hemler. 1988. J. Cell. Biochem. 37:385-393) and is highly expressed in substrate adherent cultures of almost all human cell types. The ligand specificity of VLA-3 and the inhibition of cell adhesion by anti-VLA-3 monoclonal antibodies suggest its involvement in cell-substrate interaction. In normal tissues, VLA-3 is restricted to few cel...

  20. High cell density cultures produced by internal retention: application in continuous ethanol fermentation

    Directory of Open Access Journals (Sweden)

    Berta Carola Pérez

    2007-04-01

    Full Text Available Ethanol has provoked great interest due to its potential as an alternative fuel. Nevertheless, fermentation processes must be developed by increasing the low volumetric productivity achieved in conventional cultures (batch or continuous to make this product become economically competitive. This can be achieved by using techniques leading to high cell concentration and reducing inhibition by the end-product. One of the frequently employed methods involves cell recycling. This work thus developed a membrane reactor incorporating a filtration module with 5 u,m stainless steel tubular units inside a 3L stirred jar fermenter for investigating its application in continuous ethanol production. The effects of cell concentration and transmembrane pressure difference on permeate flux were evaluated for testing the filtration module's performance. The internal cell retention system was operated in Saccharomyces cerevisiae continuous culture derived from sucrose, once fermentation conditions had been selected (30 °C, 1.25 -1.75 vvm, pH 4.5. Filter unit permeability was maintained by applying pulses of air. More than 97% of the grown cells were retained in the fermenter, reaching 51 g/L cell concentration and 8.51 g/L.h average ethanol productivity in culture with internal cell retention; this was twice that obtained in a conventional continuous culture. Key words: Membrane reactor, Saccharomyces cerevisiae, alcoholic fermentation, cell recycling.

  1. 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.

  2. Metabolism Kinetics of Glucose in Anchorage-dependent Cell Cultures

    Institute of Scientific and Technical Information of China (English)

    孙祥明; 张元兴

    2001-01-01

    The kinetic model of glucose metabolism was established and successfully applied to batchcultures of rCHO and rBHK cells. It was found that a large amount of glucose was utilized for cellmaintenance, and the overwhelming majority of maintenance energy from glucose was by its anaerobicmetabolism in both rBHK and rCHO cell cultures. The overall maintenance coefficients from aerobicmetabolism were 1.9×10-13 mmol/(cell.h) for rCHO cells and 7×10-13 mmol/(cell.h) for rBHK cells. Inaddition, all Go/T and Eo/T gradually increased with the same trend as the cell growth in the culture ofboth rCHO and rBHK cells. The overall molecule yield coefficients of lactate to glucose were 1.61 for rCHO cells and 1.38 for rBHK cells. The yield coefficients of cell to glucose were 4.5×108 cells/mmol for rCHO cells and 1.9 × 108 cells/mmol for rBHK cells, respectively.

  3. Improvement of rice anther culture and application of the technique in mutation breeding

    International Nuclear Information System (INIS)

    The ability of callus formation and green plant regeneration was very different for different rice type and varieties in anther culture. The differentiation and regeneration of green plants were obviously improved when the rice anthers at about 30 d after culture on induction medium were irradiated with 20 Gy of γ-rays and calli were cultured on the differentiation medium containing 30 mg/L colchicines. The stimulation effect of γ-irradiation combined with colchicines was much better than that of their single use. Mutation frequency and selective efficiency in M2 were obviously increased by application of the technique

  4. PEDAGIGOCAL TECHNIQUE OF BUILDING THE CULTURE OF INTERPERSONAL RELATIONS IN PRESCHOOL CHILDREN AT ART CLASSES

    Directory of Open Access Journals (Sweden)

    Svetlana Vyacheslavovna Kahnovich

    2013-10-01

    Full Text Available The article looks at the pedagogical technique of building the culture of interpersonal relations in preschool children at the local and modular level. Interpersonal relations are viewed as the module and art classes as the local level. The research is timely as it can assist in studying the problem of moral development of preschool children by building the culture of interpersonal relations by artistic education means. The study presents novelty concluding from the survey of scientific literature. The process of building the culture of interpersonal relations in children has not been properly studied by preschool pedagogy. The task of the present study is to elaborate a pedagogical technique to build the culture of interpersonal relations between children at art classes. The article discusses ‘technological’ criteria (term by G.K. Selevko and presents interactive principles of the pedagogical technique. Group activities alongside with individual ones were viewed as organizational forms of art classes. Building the culture of interpersonal relations in preschool children at art classes is closely connected with the development of their personality, a child’s  consciousness, their motivational and conceptual spheres during their gradual moral development at various levels - emotional (attitude, axiological level, psychic (intentional cognitive processes, activity (artistic and interpersonal literacy. Graphic (projective methods were used to analyze age dynamics of ethical and moral development. The conclusion describes a set of pedagogical conditions for efficient building of the culture of interpersonal relations in children at art classes.  Goal. To elaborate a pedagogical technique for building the culture of interpersonal relations in preschool children at art classes. The technique can be applied at local and modular level.Methods and Methodology. The pedagogical technique is aimed at building the culture of interpersonal relations

  5. A Simple Hydrophilic Treatment of SU-8 Surfaces for Cell Culturing and Cell Patterning

    DEFF Research Database (Denmark)

    Wang, Zhenyu; Stangegaard, Michael; Dufva, Hans Martin;

    2005-01-01

    SU-8, an epoxy-based photoresist, widely used in constitution different mTAS systems, is incompatible with mammalian cell adhesion and culture in its native form. Here, we demonstrate a simple, cheap and robust two-step method to render a SU-8 surface hydrophilic and compatible with cell culture....... The contact angle of SU-8 surface was significantly reduced from 90° to 25° after the surface modification. The treated SU-8 surfaces provided a cell culture environment that was comparable with cell culture flask surface in terms of generation time and morphology....

  6. Replica-moulded polydimethylsiloxane culture vessel lids attenuate osmotic drift in long-term cell cultures

    Indian Academy of Sciences (India)

    Axel Blau; Tanja Neumann; Christiane Ziegler; Fabio Benfenati

    2009-03-01

    An imbalance in medium osmolarity is a determinant that affects cell culture longevity. Even in humidified incubators, evaporation of water leads to a gradual increase in osmolarity overtime. We present a simple replica-moulding strategy for producing self-sealing lids adaptable to standard, small-size cell-culture vessels. They are made of polydimethylsiloxane (PDMS), a flexible, transparent and biocompatible material, which is gas-permeable but largely impermeable to water. Keeping cell cultures in a humidified 5% CO2 incubator at 37°C, medium osmolarity increased by +6.86 mosmol/kg/day in standard 35 mm Petri dishes, while PDMS lids attenuated its rise by a factor of four to changes of +1.72 mosmol/kg/ day. Depending on the lid membrane thickness, pH drifts at ambient CO2 levels were attenuated by a factor of 4 to 9. Comparative evaporation studies at temperatures below 60°C yielded a 10-fold reduced water vapour flux of 1.75 g/day/dm2 through PDMS lids as compared with 18.69 g/day/dm2 with conventional Petri dishes. Using such PDMS lids, about 2/3 of the cell cultures grew longer than 30 days in vitro. Among these, the average survival time was 69 days with the longest survival being 284 days under otherwise conventional cell culture conditions.

  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...

  8. Cell cultures from the symbiotic soft coral Sinularia flexibilis

    NARCIS (Netherlands)

    Khalesi, M.K.; Vera-Jimenez, N.I.; Aanen, D.K.; Beeftink, H.H.; Wijffels, R.H.

    2008-01-01

    The symbiotic octocoral Sinularia flexibilis is a producer of potential pharmaceuticals. Sustainable mass production of these corals as a source of such compounds demands innovative approaches, including coral cell culture. We studied various cell dissociation methodologies and the feasibility of cu

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

    DEFF Research Database (Denmark)

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

    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-...

  10. Duchenne muscular dystrophy: normal ATP turnover in cultured cells

    International Nuclear Information System (INIS)

    This paper examines ATP metabolism in cultured muscle cells and fibroblasts from patients with Duchenne dystrophy. ATP and ADP levels were the same in cultured cells from normal subjects and patients and there was no difference in ATP synthesis or degradation. The ATP synthesis was measured by the incorporation of C 14-U-adenine into aTP and ADP. although there was a significant decrease in radioactively labelled ATP after incubation with deoxyglucose in Duchenne muscle cells, there was no difference in ATP concentration of ADP metabolism

  11. Schwann cell cultures from human fetal dorsal root ganglia

    Institute of Scientific and Technical Information of China (English)

    Yaping Feng; Hui Zhu; Jiang Hao; Xinmin Wang; Shengping Wu; Li Bai; Xiangming Li; Yun Zha

    2009-01-01

    BACKGROUND:Previous studies have used many methods for in vitro Schwann cells (SCs) cul-tures and purification,such as single cell suspension and cytosine arabinoside.However,it has been difficult to obtain sufficient cellular density,and the procedures have been quite tedious.OBJECTIVE:To investigate the feasibility of culturing high-density SCs using fetal human dorsal root ganglion tissue explants.DESIGN,TIME AND SETTING:Cell culture and immunohistochemistry were performed at the Cen-tral Laboratory of Kunming General Hospital of Chinese PLA between March 2001 and October 2008.MATERIALS:Culture media containing 10% fetal bovine serum,as well as 0.2% collagenase and 0.25% trypsin were purchased from Gibco,USA;mouse anti-human S-100 monoclonal antibody and goat anti-mouse IgG labeled with horseradish peroxidase were provided by Beijing Institute of Bi-ological Products,China.METHODS:Primarily cultured SCs were dissociated from dorsal root ganglia of human aborted fe-tuses at 4-6 months pregnancy.Following removal of the dorsal root ganglion perineurium,the gan-glia were dissected into tiny pieces and digested with 0.2% collagenase and 0.25% trypsin (volume ratio 1:1),then explanted and cultured.SC purification was performed with 5 mL 10% fetal bovine serum added to the culture media,followed by differential adhesion.MAIN OUTCOME MEASURES:SCs morphology was observed under inverted phase contrast light microscopy.SC purity was evaluated according to percentage of S-100 immunostained cells.RESULTS:SCs were primarily cultured for 5-6 days and then subcultured for 4-5 passages.The highly enriched SC population reached > 95% purity and presented with normal morphology.CONCLUSION:A high purity of SCs was obtained with culture methods using human fetal dorsal root ganglion tissue explants.

  12. Miniature Bioreactor System for Long-Term Cell Culture

    Science.gov (United States)

    Gonda, Steve R.; Kleis, Stanley J.; Geffert, Sandara K.

    2010-01-01

    A prototype miniature bioreactor system is designed to serve as a laboratory benchtop cell-culturing system that minimizes the need for relatively expensive equipment and reagents and can be operated under computer control, thereby reducing the time and effort required of human investigators and reducing uncertainty in results. The system includes a bioreactor, a fluid-handling subsystem, a chamber wherein the bioreactor is maintained in a controlled atmosphere at a controlled temperature, and associated control subsystems. The system can be used to culture both anchorage-dependent and suspension cells, which can be either prokaryotic or eukaryotic. Cells can be cultured for extended periods of time in this system, and samples of cells can be extracted and analyzed at specified intervals. By integrating this system with one or more microanalytical instrument(s), one can construct a complete automated analytical system that can be tailored to perform one or more of a large variety of assays.

  13. Hydrodynamic effects on cells in agitated tissue culture reactors

    Science.gov (United States)

    Cherry, R. S.; Papoutsakis, E. T.

    1986-01-01

    The mechanisms by which hydrodynamic forces can affect cells grown on microcarrier beads in agitated cell culture reactors were investigated by analyzing the motion of microcarriers relative to the surrounding fluid, to each other, and to moving or stationary solid surfaces. It was found that harmful effects on cell cultures that have been previously attributed to shear can be better explained as the effects of turbulence (of a size scale comparable to the microcarriers or the spacing between them) or collisions. The primary mechanisms of cell damage involve direct interaction between microcarriers and turbulent eddies, collisions between microcarriers in turbulent flow, and collisions against the impeller or other solid surfaces. The implications of these analytical results for the design of tissue culture reactors are discussed.

  14. Culture and immortalization of pancreatic ductal epithelial cells.

    Science.gov (United States)

    Lawson, Terence; Ouellette, Michel; Kolar, Carol; Hollingsworth, Michael

    2005-01-01

    Some populations of the epithelial cells from the duct and ductular network of the mammalian pancreas have been isolated and maintained in vitro for up to 3 mo. These cells express many of the surface factors that are unique to them in vivo. They also retain significant drug- and carcinogen-metabolizing capacity in vitro. In this chapter we review the progression of the methods for the isolation, culture and maintenance in vitro for these cells from the earliest when only duct/ductular fragments were obtainable to the current ones which provide epithelial cells. The critical steps in the isolation process are identified and strategies are provided to facilitate these steps. These include the selection of tissue digestive enzymes, the importance of extensive mincing before culture and the importance of roles of some co-factors used in the culture medium. PMID:15542901

  15. Microfluidic bioreactors for culture of non-adherent cells

    DEFF Research Database (Denmark)

    Shah, Pranjul Jaykumar; Vedarethinam, Indumathi; Kwasny, Dorota;

    2011-01-01

    Microfluidic bioreactors (μBR) are becoming increasingly popular for cell culture, sample preparation and analysis in case of routine genetic and clinical diagnostics. We present a novel μBR for non-adherent cells designed to mimic in vivo perfusion of cells based on diffusion of media through a...... sandwiched membrane. The culture chamber and perfusion chamber are separated by a sandwiched membrane and each chamber has separate inlet/outlets for easy loading/unloading of cells and perfusion of the media. The perfusion of media and exchange of nutrients occur through the sandwiched membrane, which was...... of CFSE staining and subsequent counting in a flow cytometer. To conclude on the applicability of μBR for genetic diagnostics, we prepare chromosome spreads on glass slides from the cultured samples, which is the primary step for metaphase FISH analysis....

  16. Designing Neural Networks in Culture: Experiments are described for controlled growth, of nerve cells taken from rats, in predesigned geometrical patterns on laboratory culture dishes

    OpenAIRE

    Wheeler, Bruce C; Brewer, Gregory J.

    2010-01-01

    Technology has advanced to where it is possible to design and grow—with predefined geometry and surprisingly good fidelity—living networks of neurons in culture dishes. Here we overview the elements of design, emphasizing the lithographic techniques that alter the cell culture surface which in turn influences the attachment and growth of the neural networks. Advanced capability in this area makes it possible to design networks of desired complexity. Other issues addressed include the influenc...

  17. Cultured focusidosis fibroblasts: A simple technique demonstrating storage of tritiated-fucose labeled material

    International Nuclear Information System (INIS)

    Incorporation of tritiated fucose into the acid-soluble and acid-insoluble fractions of cultured fucosidosis and control fibroblast monolayers was determined over a 10-day period. Fucosidosis cells incorporated significantly more label into the acid-soluble, but not the acid-insoluble fraction, than control cells. The ratio of acid-soluble to acid-insoluble radioactivity increased linearly from 1 to 10 days for the fucosidosis cells, indicating the storage of low molecular weight partial degradation products. (author)

  18. Dose verification by OSLDs in the irradiation of cell cultures

    International Nuclear Information System (INIS)

    The determination of value of irradiation dose presents difficulties when targets are irradiated located in regions where electronic equilibrium of charged particle is not reached, as in the case of irradiation -in vitro- of cell lines monolayer-cultured, in culture dishes or flasks covered with culture medium. The present study aimed to implement a methodology for dose verification in irradiation of cells in culture media by optically stimulated luminescence dosimetry (OSLD). For the determination of the absorbed dose in terms of cell proliferation OSL dosimeters of aluminum oxide doped with carbon (Al2O3:C) were used, which were calibrated to the irradiation conditions of culture medium and at doses that ranged from 0.1 to 15 Gy obtained with a linear accelerator of 6 MV photons. Intercomparison measurements were performed with an ionization chamber of 6 cm3. Different geometries were evaluated by varying the thicknesses of solid water, air and cell culture medium. The results showed deviations below 2.2% when compared with the obtained doses of OSLDs and planning system used. Also deviations were observed below 3.4% by eccentric points of the irradiation plane, finding homogeneous dose distribution. Uncertainty in the readings was less than 2%. The proposed methodology contributes a contribution in the dose verification in this type of irradiations, eliminating from the calculation uncertainties, potential errors in settling irradiation or possible equipment failure with which is radiating. It also provides certainty about the survival curves to be plotted with the experimental data. (Author)

  19. Application of dynamic speckle interferometry for diagnosis of metabolism change in cultured cells

    Science.gov (United States)

    Vladimirov, A. P.; Mikhailova, J. A.; Baharev, A. A.; Malygin, A. S.; Novosyolova, I. A.; Yakin, D. I.

    2015-08-01

    Previously, a speckle interferometry technique and a device that allows quantitative evaluation of the metabolic activity of cultured cells were theoretically grounded and successfully tested. A speckle time-averaging technique was proposed to separate and study the processes occurring in cells at various velocities. The objective of the present research was comparing the parameters of speckle dynamics used to evaluate cell metabolism and averaged in areas of various size. Areas inside the speckle image of a cell as well as areas of the image plane with various numbers of cells were averaging areas. Defrosted L-41 cells precipitated on a glass substrate were the target of the research. Time-average value T of digital radiation intensity in the TV camera pixels I~ (1) and the correlation coefficient of two digital images η (2) were used as speckle field change parameters. The digital images corresponded to a single frame area at two time moments.

  20. Radiosensitivity of cultured insect cells: I. Lepidoptera

    International Nuclear Information System (INIS)

    The radiosensitivity of five lepidopteran insect cell lines representing five different genera has been investigated. These lines are: (1) TN-368, Trichoplusia ni; (2) IPLB-SF-1254, Spodoptera frugiperda; (3) IPLB-1075, Heliothis zea; (4) MRRL-CHl, clone GVl, Manduca sexta; and (5) IAL-PID2, Plodia interpunctella. The cell lines grew at different rates and had population doubling times that ranged from 19 to 52 hr. All of the lines are highly heteroploid and have approximate chromosome numbers near or above 100. The chromosomes are very small. All of the lines are extremely radioresistant; cell populations are able to recover from 260 kVp X-ray exposures up to and including 400 Gy, the highest dose examined. Cell survival curves were obtainable for only the TN-368 and IPLB-SF-1254 lines. The TN-368 cells displayed a biphasic survival response with D0, d/sub q/, and n values of 65.7 and 130.2 Gy, 9.0 and -36.1 Gy, and 1.2 and 0.8, respectively, for the steep and shallow portions of the curve. The IPLB-SF-1254 cells had a D0 of 63.9 Gy. D/sub q/ of 19.0 Gy, and n value of 1.4. These studies provide definitive evidence of the radioresistance of lepidopteran cells, and suggest that this radioresistance is a characteristic of lepidopteran insects

  1. Determination of the synthesis of uptake of α2-macroglobulin by cultured human glioma cells

    International Nuclear Information System (INIS)

    Using immunological techniques, the synthesis of α2-macroglobulin was studied in established cell lines derived from human glioblastomas multiform. α2-Macroglobulin was detected in cytoplasm and in the culture medium of the analyzed cell lines. Radioimmunoprecipitation, revealed a protein with Mr corresponding to α2-macroglobulin in the medium conditioned by U-118MG and U-343MG cells. On the other hand, using immunoblot analysis, α2-macroglobulin was detected in all of the analyzed lines. In immunofluorescence test, α2-macroglobulin was determined also in all four cell lines, but with different staining pattern. Conditioned culture medium of U-536MG cells with the lowest level of α2-macroglobulin exerted the lowest mitogenic activity for human fibroblasts. (author)

  2. A novel voltage clamp technique for mapping ionic currents from cultured skeletal myotubes.

    OpenAIRE

    Anson, B D; Roberts, W M

    1998-01-01

    The biophysical properties and cellular distribution of ion channels largely determine the input/output relationships of electrically excitable cells. A variety of patch pipette voltage clamp techniques are available to characterize ionic currents. However, when used by themselves, such techniques are not well suited to the task of mapping low-density channel distributions. We describe here a new voltage clamp method (the whole cell loose patch (WCLP) method) that combines whole-cell recordin...

  3. The replacement of serum by hormones in cell culture media.

    Science.gov (United States)

    Sato, G; Hayashi, I

    1976-12-01

    The replacement of serum by hormones in cell culture media. (Reemplazo del suero por hormonas en el medio de cultivo de células). Arch. Biol. Med. Exper. 10: 120-121, 1976. The serum used in cell culture media can be replaced by a mixture of hormones and some accesory blood factors. The pituitary cell line GH3 can be grown in a medium in which serum is replaced by triiodothyronine, transferrin, parathormone, tyrotrophin releasing hormone and somatomedins. Hela and BHK cell strains can also be grown in serum free medium supplemented with hormones. Each cell type appears to have different hormonal requirements yet it may found that some hormones are required for most cell types. PMID:1026199

  4. Continuous culture of immobilized streptomyces cells for kasugamycin production.

    Science.gov (United States)

    Kim, C J; Chang, Y K; Chun, G T; Jeong, Y H; Lee, S J

    2001-01-01

    Continuous cultures of immobilized Streptomyces kasugaensis, a kasugamycin producer, were carried out on Celite beads. When using a prototype separator for immobilized-cell separation and recycling, the continuous operation could not be sustained for an extended period as a result of an excessive loss of immobilized cells caused by the poor performance of the separator. Accordingly, the immobilized-cell separator was revised to provide better immobilized-cell settling and thus recycling into the reactor. In a subsequent culture using the revised separator, a stable operation was maintained for over 820 h with a high kasugamycin productivity. The kasugamycin productivity ranged from 9.8 to 16.1 mg/L/h, which was about 14- to 23-fold higher than that in a batch suspended-cell culture. When the original feeding medium concentration was doubled at the end of the continuous culture, the productivity became severely impaired for several reasons, which will be discussed. An excessive formation of free cells and loss of immobilized cells through the separator were also observed. PMID:11386865

  5. Isolation, culture and characterization of primary mouse RPE cells.

    Science.gov (United States)

    Fernandez-Godino, Rosario; Garland, Donita L; Pierce, Eric A

    2016-07-01

    Mouse models are powerful tools for the study of ocular diseases. Alterations in the morphology and function of the retinal pigment epithelium (RPE) are common features shared by many ocular disorders. We report a detailed protocol to collect, seed, culture and characterize RPE cells from mice. We describe a reproducible method that we previously developed to collect and culture murine RPE cells on Transwells as functional polarized monolayers. The collection of RPE cells takes ∼3 h, and the cultures mimic in vivo RPE cell features within 1 week. This protocol also describes methods to characterize the cells on Transwells within 1-2 weeks by transmission and scanning electron microscopy (TEM and SEM, respectively), immunostaining of vibratome sections and flat mounts, and measurement of transepithelial electrical resistance. The RPE cell cultures are suitable to study the biology of the RPE from wild-type and genetically modified strains of mice between the ages of 10 d and 12 months. The RPE cells can also be manipulated to investigate molecular mechanisms underlying the RPE pathology in the numerous mouse models of ocular disorders. Furthermore, modeling the RPE pathology in vitro represents a new approach to testing drugs that will help accelerate the development of therapies for vision-threatening disorders such as macular degeneration (MD). PMID:27281648

  6. Evaluating the role of low-speed centrifugation towards transfecting human peripheral blood mononuclear cell culture

    Directory of Open Access Journals (Sweden)

    M Majumdar

    2014-01-01

    Full Text Available The conventional method of transfection of suspension cells by chemical has proven to be very difficult. We present a new transfection protocol, wherein, low-speed centrifugation of cell culture plates immediately after adding the lipid: DNA complex significantly enhances the transfection efficiency. Peripheral blood mononuclear cells (PBMCs were transfected with BLOCK-iT™ Fluorescent Oligo (scrambled siRNA and lipofectamine complex using conventional and low-speed centrifugation modified transfection protocols. The efficiency of transfection was determined using flowcytometer and cell viability was checked using MTT assay. Incorporation of low-speed centrifugation significantly enhances the transfection efficiency of BLOCK-iT™ in the suspension culture of PBMCs as compared to conventional transfection method (99.8% vs 28.3%; P < 0.0001, even at a low concentration of 40 picomoles without affecting the cell viability. Centrifugation enhanced transfection (CET technique is simple, time-saving and novel application without compromising the cell viability in the context of recently popular RNA interference in suspension cultures of PBMCs. This undemanding modification might be applicable to a wide variety of cell lines and solve crucial problem of researchers working with RNA interference in suspension cultures.

  7. Differential heat shock response of primary human cell cultures and established cell lines

    DEFF Research Database (Denmark)

    Richter, W W; Issinger, O G

    1986-01-01

    degrees C treatment, whereas in immortalized cell lines usually 90% of the cells were found in suspension. Enhanced expression of the major heat shock protein (hsp 70) was found in all heat-treated cells. In contrast to the primary cell cultures, established and transformed cell lines synthesized a...

  8. An improved protocol for isolation and culture of mesenchymal stem cells from mouse bone marrow

    Directory of Open Access Journals (Sweden)

    Shuo Huang

    2015-01-01

    Full Text Available Mesenchymal stem cells (MSCs from bone marrow are main cell source for tissue repair and engineering, and vehicles of cell-based gene therapy. Unlike other species, mouse bone marrow derived MSCs (BM-MSCs are difficult to harvest and grow due to the low MSCs yield. We report here a standardised, reliable, and easy-to-perform protocol for isolation and culture of mouse BM-MSCs. There are five main features of this protocol. (1 After flushing bone marrow out of the marrow cavity, we cultured the cells with fat mass without filtering and washing them. Our method is simply keeping the MSCs in their initial niche with minimal disturbance. (2 Our culture medium is not supplemented with any additional growth factor. (3 Our method does not need to separate cells using flow cytometry or immunomagnetic sorting techniques. (4 Our method has been carefully tested in several mouse strains and the results are reproducible. (5 We have optimised this protocol, and list detailed potential problems and trouble-shooting tricks. Using our protocol, the isolated mouse BM-MSCs were strongly positive for CD44 and CD90, negative CD45 and CD31, and exhibited tri-lineage differentiation potentials. Compared with the commonly used protocol, our protocol had higher success rate of establishing the mouse BM-MSCs in culture. Our protocol may be a simple, reliable, and alternative method for culturing MSCs from mouse bone marrow tissues.

  9. Formation and action of oxygen activated species in cell cultures

    International Nuclear Information System (INIS)

    The differences of hydrogen peroxide sensibility of mammal cell lineages (man, mouse, chinese hamster) in culture are studied. The cellular survival and the frequency of DNA induced breaks by hydrogen peroxide are analysed. The efficiency of elimination of DNA breaks by cells is determined. The possible relation between the cell capacity of repair and its survival to hydrogen peroxide action is also discussed. (M.A.)

  10. Polyphosphoinositides are present in plant tissue culture cells

    International Nuclear Information System (INIS)

    Polyphosphoinositides have been isolated from wild carrot cells grown in suspension culture. This is the first report of polyphosphoinositides in plant cells. The phospholipids were identified by comigration with known standards on thin-layer plates. After overnight labeling of the cells with myo-[2-3H] inositol, the phosphoinositides as percent recovered inositol were 93% phosphatidylinositol., 3.7% lysophosphatidylinositol, 1.7% phosphatidylinositol monophosphate, 0.8% phosphatidylinositol bisphosphate

  11. Culture of Neural Stem Cells in Calcium-alginate Microbeads

    Institute of Scientific and Technical Information of China (English)

    Li-Song YAO; Tian-Qing LIU; Dan GE; Xue-Hu MA; Zhan-Feng CUI

    2005-01-01

    @@ 1 Introduction Recent research shows that neural stem cells may play an important role in the nerve injury reparation and nerve disease treatment. The shortage of the source and the number of NSCs, however, is the main challenge for its clinic application. In this situation, expansion of NSCs in large scale and culture in three dimensional environment are very worth of exploration. Notablely, the shear stress existed in bioreactors can cause serious cell injury especially for the shear sensitive cells like NSCs.

  12. Culture of Neural Stem Cells in Calcium-alginate Microbeads

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    1 Introduction Recent research shows that neural stem cells may play an important role in the nerve injury reparation and nerve disease treatment. The shortage of the source and the number of NSCs, however, is the main challenge for its clinic application. In this situation, expansion of NSCs in large scale and culture in three dimensional environment are very worth of exploration. Notablely, the shear stress existed in bioreactors can cause serious cell injury especially for the shear sensitive cells like ...

  13. Epithelial morphogenesis in three-dimensional cell culture system

    OpenAIRE

    Liu, Mengfei; 刘梦菲

    2014-01-01

    In human body, the most common structures formed by epithelial cells are hollow cysts or tubules. The key feature of the cysts and tubules is the central lumen, which is lined by epithelial cell sheets. The central lumen allows material exchange, thus it is indispensable for the proper function of the epithelial tissue. In order to understand the way that the epithelial cells form highly specialized structure, an in vitro three-dimensional (3D) culture system was established. The Caco-2 c...

  14. Carbon Nanotubes-Based Electrochemical Sensing for Cell Culture Monitoring

    OpenAIRE

    Boero, Cristina; Carrara, Sandro; Del Vecchio, Giovanna; Albini, Giuseppe D.; Calzà, Laura; De Micheli, Giovanni

    2010-01-01

    Monitoring of metabolic compounds, such as glucose and lactate, is extensively reported in literature, especially for clinical purposes. Instead, the application of such technologies for monitoring metabolites in cell cultures has not been explored. From one side, such devices can provide information to the current state-of-the-art of cell lines, particularly those which are not fully known, as stem and embryonic cells. On the other hand, those systems can pave the way to fully automation for...

  15. Optimization of Seeding Density in Microencapsulated Recombinant CHO Cell Culture

    OpenAIRE

    Zhang, Ying; Zhou, Jing; Zhang, Xulang; Yu, Weiting; Guo, Xin; Wang, Wei; Ma, Xiaojun

    2008-01-01

    Microencapsulation technology is an alternative large-scale mammalian cell culture method. The semi-permeable membrane of the microcapsule allows free diffusion of nutrients, oxygen and toxic metabolites to support cell growth, and the microcapsule membrane can protect the cells from the mechanical damage of shear forces associated with agitation and aeration. Many polymers have been used to make microcapsules, such as chitosan, polyacrylates, alginate, polyamino acids, and polyamides. One of...

  16. Culturing Human Pluripotent and Neural Stem Cells in an Enclosed Cell Culture System for Basic and Preclinical Research

    Science.gov (United States)

    Stover, Alexander E.; Herculian, Siranush; Banuelos, Maria G.; Navarro, Samantha L.; Jenkins, Michael P.; Schwartz, Philip H.

    2016-01-01

    This paper describes how to use a custom manufactured, commercially available enclosed cell culture system for basic and preclinical research. Biosafety cabinets (BSCs) and incubators have long been the standard for culturing and expanding cell lines for basic and preclinical research. However, as the focus of many stem cell laboratories shifts from basic research to clinical translation, additional requirements are needed of the cell culturing system. All processes must be well documented and have exceptional requirements for sterility and reproducibility. In traditional incubators, gas concentrations and temperatures widely fluctuate anytime the cells are removed for feeding, passaging, or other manipulations. Such interruptions contribute to an environment that is not the standard for cGMP and GLP guidelines. These interruptions must be minimized especially when cells are utilized for therapeutic purposes. The motivation to move from the standard BSC and incubator system to a closed system is that such interruptions can be made negligible. Closed systems provide a work space to feed and manipulate cell cultures and maintain them in a controlled environment where temperature and gas concentrations are consistent. This way, pluripotent and multipotent stem cells can be maintained at optimum health from the moment of their derivation all the way to their eventual use in therapy. PMID:27341536

  17. Investigation progress of imaging techniques monitoring stem cell therapy

    International Nuclear Information System (INIS)

    Recently stem cell therapy has showed potential clinical application in diabetes mellitus, cardiovascular diseases, malignant tumor and trauma. Efficient techniques of non-invasively monitoring stem cell transplants will accelerate the development of stem cell therapies. This paper briefly reviews the clinical practice of stem cell, in addition, makes a review of monitoring methods including magnetic resonance and radionuclide imaging which have been used in stem cell therapy. (authors)

  18. Human cell culture models for investigating molecular and cytogenetic changes in radiation carcinogenesis

    International Nuclear Information System (INIS)

    Primary cultures of human epithelial cells have proved difficult to transform because of the inherent short duration that these cells can be cultured. However, primary cultures of human cells can be immortalised using the catalytic sub-unit of telomerase (hTERT). Radiation carcinogenesis has been investigated using a human retinal pigment epithelial cell line (340RPE-T53 hTERT). Transformants can be selected using anchorage independent growth and cell lines derived from these are tumourigenic in immunosupressed mice. Molecular cytogenetic changes using CGH, SKY and FISH with breakpoint-specific YAC- and BAC- probes revealed a high level amplification on 10p11.2 in several clones which has been identified as an atypical protein kinase C binding protein using FISH gene-specific PCR products. Patterns of gene expression were studied using HuGen Human cDNA arrays using indirect labelling. The control parent RPE cell line could then be compared with cloned radiation-induced tumour cell lines derived from it following fractionated doses of gamma irradiation. Osteonectin was down regulated in 4 different tumour lines. This gene maps to a region of chromosome 5q that is commonly deleted in leukaemia. Nexin and p105 were down regulated in 3 lines and tumour suppressing subtransferable candidate 1 in I line. Further hTERT immortalised cell lines have been derived from primary cultures of human mammary epithelial cells. The breast epithelium contains a number of different cell types and the lines have been characterised using immunocytochemical techniques. The cells are cytokeratin 19 negative but CD10, cytokeratin 5 and p63 positive indicating a basal cell phenotype. Following exposure to fractionated doses of gamma irradiation anchorage independent colonies are formed. Thus human cell lines immortalised with hTERT are providing a useful model system for investigating radiation carcinogenesis and the molecular and cytogenetic changes induced. Supported by EC Nuclear Fission

  19. Specimen Sample Preservation for Cell and Tissue Cultures

    Science.gov (United States)

    Meeker, Gabrielle; Ronzana, Karolyn; Schibner, Karen; Evans, Robert

    1996-01-01

    The era of the International Space Station with its longer duration missions will pose unique challenges to microgravity life sciences research. The Space Station Biological Research Project (SSBRP) is responsible for addressing these challenges and defining the science requirements necessary to conduct life science research on-board the International Space Station. Space Station will support a wide range of cell and tissue culture experiments for durations of 1 to 30 days. Space Shuttle flights to bring experimental samples back to Earth for analyses will only occur every 90 days. Therefore, samples may have to be retained for periods up to 60 days. This presents a new challenge in fresh specimen sample storage for cell biology. Fresh specimen samples are defined as samples that are preserved by means other than fixation and cryopreservation. The challenge of long-term storage of fresh specimen samples includes the need to suspend or inhibit proliferation and metabolism pending return to Earth-based laboratories. With this challenge being unique to space research, there have not been any ground based studies performed to address this issue. It was decided hy SSBRP that experiment support studies to address the following issues were needed: Fixative Solution Management; Media Storage Conditions; Fresh Specimen Sample Storage of Mammalian Cell/Tissue Cultures; Fresh Specimen Sample Storage of Plant Cell/Tissue Cultures; Fresh Specimen Sample Storage of Aquatic Cell/Tissue Cultures; and Fresh Specimen Sample Storage of Microbial Cell/Tissue Cultures. The objective of these studies was to derive a set of conditions and recommendations that can be used in a long duration microgravity environment such as Space Station that will permit extended storage of cell and tissue culture specimens in a state consistent with zero or minimal growth, while at the same time maintaining their stability and viability.

  20. Cryopreservation in situ of cell monolayers on collagen vitrigel membrane culture substrata: ready-to-use preparation of primary hepatocytes and ES cells.

    Science.gov (United States)

    Miyamoto, Yoshitaka; Enosawa, Shin; Takeuchi, Tomoyo; Takezawa, Toshiaki

    2009-01-01

    Cryopreservation is generally performed on cells in suspension. In the case of adherent cells such as hepatocytes, a loss of their ability to attach is a more serious problem than a decreased viability after cryopreservation. We herein report a novel technology of direct in situ cryopreservation of cells cultured on collagen vitrigel membranes, which have excellent mechanical strength and can be easily handled by tweezers even when coated with cultured cells. Rat primary hepatocytes, mitomycin C-treated mouse fibroblasts (feeder cells for ES cells), and mouse ES cells on the feeder cells were cultured on collagen vitrigel membranes for 1 day. The membranes with cells attached were then plucked up from the dish, soaked in cryopreservation medium containing 10% dimethyl sulfoxide, frozen using a controlled-rate freezer, and transferred to liquid nitrogen. The cells cultured on plastic cell culture dishes were also frozen as controls. After storage in liquid nitrogen for periods from 1 week to 3 months, the cryopreserved membranes with the cells still attached were thawed by adding warmed culture medium. Cell viability estimated by morphology and functional staining with calcein showed significant improvement in comparison to cells cryopreserved without the collagen vitrigel membrane. The recoveries of living cells after cryopreservation were 26.7%, 76.2%, and 58.6% for rat hepatocytes, mitomycin C-treated mouse fibroblasts, and mouse ES cells on collagen vitrigel membranes, respectively. In contrast, essentially no cells at all remained on the plastic cell culture dishes after thawing. Because adherent cell storage under these conditions is very convenient, the use of this technique employing collagen vitrigel membranes should be generally applicable to the cryopreservation of adherent cells that are otherwise problematic to store as frozen stocks. PMID:19775524

  1. Expression of CD44 in Cultured Human Trabecular Meshwork Cells

    Institute of Scientific and Technical Information of China (English)

    Zhongguo Li; Hong Zhang

    2004-01-01

    Purpose:To determine whether cultured human trabecular meshwork cells express CD44 and to discuss their possible relationship with primary open angle glaucoma.Methods:Human trabecular meshwork cells were cultured in DMEM/F12 media. Total RNAs from the cells were extracted with Trizol reagent. Messenger RNA expression of CD44 in human trabecular meshwork cells was examined by using reverse transcriptasepolymerase chain reaction ( RT-PCR ) analysis. Expression of CD44 was confirmed by Western-blotting and immunofiuorescent microscopy. Effect of CD44-specific antisense oligonucleotide on adhesion of trabecular meshwork cells to hyaluronate was determined by MTT assay.Results:A single RT-PCR product whose size was 471bp was obtained.A band about 80kD was stained by Western-blot. Immunofiuorescent examination of expression of CD44 on the cell surface was positive and reactions were mainly localized in cell membranes.Adhesion of trabecular meshwork cells to hyaluronate was inhibited by CD44-specific antisense oligonucleotide.Conclusions: Cultured human trabecular meshwork cells express CD44. CD44 may play a role in pathogenesis of primary open angle glaucoma. Eye Science 2004;20:52-56.

  2. Enhanced casein kinase II activity in human tumour cell cultures

    DEFF Research Database (Denmark)

    Prowald, K; Fischer, H; Issinger, O G

    1984-01-01

    Casein kinase II (CKII) activity is enhanced as much as 2-3 fold in established and 4-5-fold in transformed human cell lines when compared to that of fibroblasts and primary human tumour cell cultures where CKII activity never exceeded a basic level. The high activity of CKII in transformed cells...... and in established cell lines was reduced to about the same basic level after treatment with heparin, a highly specific inhibitor of CKII activity. The activity of the cAMP-dependent protein kinase was virtually the same in fibroblasts and various human tumour cell lines investigated....

  3. A co-cultured skin model based on cell support membranes

    International Nuclear Information System (INIS)

    Tissue engineering of skin based on collagen: PCL biocomposites using a designed co-culture system is reported. The collagen: PCL biocomposites having collagen: PCL (w/w) ratios of 1:4, 1:8, and 1:20 have been proven to be biocompatible materials to support both adult normal human epidermal Keratinocyte (NHEK) and mouse 3T3 fibroblast growth in cell culture, respectively, by Dai, Coombes, et al. in 2004. Films of collagen: PCL biocomposites were prepared using non-crosslinking method by impregnation of lyophilized collagen mats with PCL/dichloromethane solutions followed by solvent evaporation. To mimic the dermal/epidermal structure of skin, the 1:20 collagen: PCL biocomposites were selected for a feasibility study of a designed co-culture technique that would subsequently be used for preparing fibroblast/biocomposite/keratinocyte skin models. A 55.3% increase in cell number was measured in the designed co-culture system when fibroblasts were seeded on both sides of a biocomposite film compared with cell culture on one surface of the biocomposite in the feasibility study. The co-culture of human keratinocytes and 3T3 fibroblasts on each side of the membrane was therefore studied using the same co-culture system by growing keratinocytes on the top surface of membrane for 3 days and 3T3 fibroblasts underneath the membrane for 6 days. Scanning electron microscopy (SEM) and immunohistochemistry assay revealed good cell attachment and proliferation of both human keratinocytes and 3T3 fibroblasts with these two types of cells isolated well on each side of the membrane. Using a modified co-culture technique, a co-cultured skin model presenting a confluent epidermal sheet on one side of the biocomposite film and fibroblasts populated on the other side of the film was developed successfully in co-culture system for 28 days under investigations by SEM and immunohistochemistry assay. Thus, the design of a co-culture system based on 1:20 (w/w) collagen: PCL biocomposite

  4. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell...

  5. Hollow fiber clinostat for simulating microgravity in cell culture

    Science.gov (United States)

    Rhodes, Percy H. (Inventor); Miller, Teresa Y. (Inventor); Snyder, Robert S. (Inventor)

    1992-01-01

    A clinostat for simulating microgravity on cell systems carried in a fiber fixedly mounted in a rotatable culture vessel is disclosed. The clinostat is rotated horizontally along its longitudinal axis to simulate microgravity or vertically as a control response. Cells are injected into the fiber and the ends of the fiber are sealed and secured to spaced end pieces of a fiber holder assembly which consists of the end pieces, a hollow fiber, a culture vessel, and a tension spring with three alignment pins. The tension spring is positioned around the culture vessel with its ends abutting the end pieces for alignment of the spring. After the fiber is secured, the spring is decompressed to maintain tension on the fiber while it is being rotated. This assures that the fiber remains aligned along the axis of rotation. The fiber assembly is placed in the culture vessel and culture medium is added. The culture vessel is then inserted into the rotatable portion of the clinostat and subjected to rotate at selected rpms. The internal diameter of the hollow fiber determines the distance the cells are from the axis of rotation.

  6. Agricultural production - Phase 2. Indonesia. Rice - azolla - fish culture - use of nuclear technique

    International Nuclear Information System (INIS)

    The primary aim of the expert mission was to provide advice on the use of nuclear techniques to study rice-azolla-fish culture. Results of the work performed so far show that basal application of azolla gives similar or better yields of rice than basal application of urea. Fish productivity was also found to be significantly higher when azolla is present. 2 tabs

  7. Technique of the 'in vitro' fertilization and the culture of mouse embryos at preimplantation

    International Nuclear Information System (INIS)

    The mammal embryo is an intensive cellular proliferating system, very radiosensitive and therefore adequate to the study of the biological effects of ionizing radiation. The technique of the in vitro fertilization and the culture of mouse embryos at preimplantation period, modified by Yamada et al (1982) to improve the efficiency of more than 95% of blastocyst formation is described. (author)

  8. [The potential of three-dimensional tumor models and cell culturing in cancer research and diagnostics].

    Science.gov (United States)

    Alföldi, Róbert; Szebeni, János Gábor; Puskás, G László

    2015-12-01

    In vitro testing of antitumor agents on human cancer cell lines has become essential in pharmaceutical research and in clinical practice. Although the most widely used technique is the two-dimensional cell growing protocol (in tissue culture plates), the new three-dimensional methods are becoming more and more popular as their structure and complexity is more similar to the microenvironment of the real tumor. The aim of the present study is to describe the most widely used in vitro three-dimensional tumor models and to compare a RAFT(TM) three dimensional in vitro tumor model with the traditional two-dimensional tumor cell cultures. In the study, the viability and the enzyme activity of cultured A549 non-small cell lung cancer (NSCLC) cells under different conditions were compared. The results show that while the number of necrotic cells increased significantly (20-fold; 2D/A549 T75 conventional tissue culture flask 1.6%; 2D/A549-collagen coated T75 tissue culture flask 1.45%, RAFT(TM) 22.11%) during long culturing period in the RAFT(TM) three-dimensional in vitro tumor model, there was no significant difference during the conventional antitumor screening period (3-5 day) compared to the traditional two-dimensional cell cultures. The structure of the tumor cell islets grown with RAFT(TM) is much more complex than that of the traditional two-dimensional cultures. Thus, similarly to the in vivo tumor microenvironment, there is also a collagen matrix in the extracellular space which can have significant effect on the diffusion of the antitumor agents to cells. In conclusion, it can be stated that testing of antitumor agents on tumor cells cultured in three-dimensional systems can be an important complementary method to the traditional two-dimensional in vitro analyses. The results of the new three-dimensional method can be more easily applied in the in vivo analysis and translated into clinical practice. PMID:26665190

  9. Surface-engineered substrates for improved human pluripotent stem cell culture under fully defined conditions

    OpenAIRE

    Saha, Krishanu; Mei, Ying; Reisterer, Colin M.; Pyzocha, Neena Kenton; Yang, Jing; Muffat, Julien; Davies, Martyn C.; Alexander, Morgan R.; Langer, Robert; Anderson, Daniel G.; Jaenisch, Rudolf

    2011-01-01

    The current gold standard for the culture of human pluripotent stem cells requires the use of a feeder layer of cells. Here, we develop a spatially defined culture system based on UV/ozone radiation modification of typical cell culture plastics to define a favorable surface environment for human pluripotent stem cell culture. Chemical and geometrical optimization of the surfaces enables control of early cell aggregation from fully dissociated cells, as predicted from a numerical model of cell...

  10. Lingual Epithelial Stem Cells and Organoid Culture of Them

    Directory of Open Access Journals (Sweden)

    Hiroko Hisha

    2016-01-01

    Full Text Available As tongue cancer is one of the major malignant cancers in the world, understanding the mechanism of maintenance of lingual epithelial tissue, which is known to be the origin of tongue cancer, is unquestionably important. However, the actual stem cells that are responsible for the long-term maintenance of the lingual epithelium have not been identified. Moreover, a simple and convenient culture method for lingual epithelial stem cells has not yet been established. Recently, we have shown that Bmi1-positive cells, residing at the second or third layer of the epithelial cell layer at the base of the interpapillary pit (IPP, were slow-cycling and could supply keratinized epithelial cells for over one year, indicating that Bmi1-positive cells are long-term lingual epithelial stem cells. In addition, we have developed a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Here, we discuss current progress in the identification of lingual stem cells and future applications of the lingual culture system for studying the regulatory mechanisms of the lingual epithelium and for regenerative medicine.

  11. Cancer Stem Cells: Cell Culture, Markers and Targets for New Therapies

    OpenAIRE

    Gilbert, Candace A.; Ross, Alonzo H.

    2009-01-01

    A cancer stem cell is defined as an undifferentiated cell with the ability to self-renew, differentiate to multiple lineages and initiate tumors that mimic the parent tumor. In this review, we focus on glioblastomas, describing recent progress and problems in characterizing these cells. There have been advances in cancer stem cell culture, but tumor cell heterogeneity has made purification of cancer stem cells difficult. Indeed, it may be that cancer stem cells significantly vary from tumor t...

  12. Regulatory Effect of Dexamethasone on Aquaporin-1 Expression in Cultured Bovine Trabecular Meshwork Cells

    Institute of Scientific and Technical Information of China (English)

    XIONG Xinchun; MIAO Juan; XI Zulian; ZHANG Haijiang; HAN Bo; HU Yizhen

    2005-01-01

    To evaluate the effect of dexamethasone on the expression of aquaporin-1 (AQP-1) in cultured bovine trabecular meshwork cells, bovine trabecular meshwork cells were cultured in vitro and reproduced to the third and the fourth generation, then treated with dexamethasone at the concentrations of 5, 25, 50, 250 μg/L respectively for 7 days. Immunohistochemical technique-supervision method was employed to measure, and image analysis system to analyze the expression of AQP-1 in normal cultured bovine trabecular meshwork cells and those treated with dexamethasone.In normal bovine trabecular meshwork cells, the grayscale of AQP-1 positive staining was 167.94±1.18, while it was 168.92±0.91, 176.72±1.80, 180.64±1.31, 185.64±1.58 in cells treated with 5, 25, 50, 250 tg/L concentrations of dexamethasone. When the concentration of dexamethasone was higher than 25 μg/L, the expression of AQP-1 was significantly inhibited (P<0.05).The regulation of AQP-1 expression by dexamethasone in cultured bovine trabecular meshwork cells in vitro may be one of causes that retard the aqueous outflow in glucocorticoid induced glaucoma.

  13. Isolation and culture of cells from the nephrogenic zone of the embryonic mouse kidney.

    Science.gov (United States)

    Brown, Aaron C; Blank, Ulrika; Adams, Derek C; Karolak, Michele J; Fetting, Jennifer L; Hill, Beth L; Oxburgh, Leif

    2011-01-01

    Embryonic development of the kidney has been extensively studied both as a model for epithelial-mesenchymal interaction in organogenesis and to gain understanding of the origins of congenital kidney disease. More recently, the possibility of steering naïve embryonic stem cells toward nephrogenic fates has been explored in the emerging field of regenerative medicine. Genetic studies in the mouse have identified several pathways required for kidney development, and a global catalog of gene transcription in the organ has recently been generated http://www.gudmap.org/, providing numerous candidate regulators of essential developmental functions. Organogenesis of the rodent kidney can be studied in organ culture, and many reports have used this approach to analyze outcomes of either applying candidate proteins or knocking down the expression of candidate genes using siRNA or morpholinos. However, the applicability of organ culture to the study of signaling that regulates stem/progenitor cell differentiation versus renewal in the developing kidney is limited as cultured organs contain a compact extracellular matrix limiting diffusion of macromolecules and virus particles. To study the cell signaling events that influence the stem/progenitor cell niche in the kidney we have developed a primary cell system that establishes the nephrogenic zone or progenitor cell niche of the developing kidney ex vivo in isolation from the epithelial inducer of differentiation. Using limited enzymatic digestion, nephrogenic zone cells can be selectively liberated from developing kidneys at E17.5. Following filtration, these cells can be cultured as an irregular monolayer using optimized conditions. Marker gene analysis demonstrates that these cultures contain a distribution of cell types characteristic of the nephrogenic zone in vivo, and that they maintain appropriate marker gene expression during the culture period. These cells are highly accessible to small molecule and recombinant

  14. Metabolism of quercetin in cell suspension culture of Nicotiana tabacum

    International Nuclear Information System (INIS)

    Quercetin was oxidized on a rotating glassy-carbon electrode in phosphate-methanol buffer. The half-wave potentials for several pH were determined. Oxidation of quercetin by one of Nicotiana tabacum cell suspension cultures was carried out in vitro and final products were characterized by means of UV spectra and mass-spectrophotometry. Each product of oxidation was assayed for oxygen consumption inhibiting activity, using a 3 days old cell suspension culture of Nicotiana tabacum. Dimers and polymers showed strong inhibiting activity in O-2 consumption

  15. Preparation of cells cultured on silicon wafers for mass spectrometry analysis.

    Science.gov (United States)

    Wittig, Andrea; Wiemann, Martin; Fartmann, Michael; Kriegeskotte, Christian; Arlinghaus, Heinrich F; Zierold, Karl; Sauerwein, Wolfgang

    2005-04-01

    The distribution of specific atoms and molecules within living cells is of high interest in bio-medical research. Laser secondary neutral mass spectrometry (laser-SNMS) and time-of-flight secondary ion mass spectrometry (TOF-SIMS) detect atoms with high sensitivity and spatial resolution. The application of these methods to cultured cells requires special preparation techniques preserving morphological and chemical integrity of the living cells. The cells should, therefore, be grown on a conducting material preventing charging of the sample during ion bombardment. Silicon is currently used as the preferred support material for non-biological samples in mass spectrometry. This study investigates (1) the influence of silicon surfaces on cell growth and (2) the suitability of a sandwiched, rapid freezing method to analyse transmembrane ion gradients. Human melanoma cells were grown on silicon with polished or etched surfaces. Growth kinetics were studied using the Sulforhodamine-B assay. Number, shape, and morphology of the cells were assessed by epifluorescence microscopy of calcein AM- and DAPI-stained cells. Cells were subjected to rapid freezing, freeze-fracturing, and freeze-drying prior to analysis by TOF-SIMS and laser-SNMS. While cell numbers and morphology on the rough silicon wafers were impaired, morphology and growth kinetics of cells on polished silicon were identical to control cells on cell culture tested polystyrene. TOF-SIMS and laser-SNMS resulted in high-resolution elemental images and mass spectra. Measurement of the intracellular Na+ and K+ concentrations revealed a ratio as observed in living cells. In conclusion, culturing cells on polished silicon wafers followed by sandwiched, rapid freezing is an adequate preparation method to study intracellular ion distribution with mass spectrometry. PMID:15940684

  16. Dynamic cell culture system: a new cell cultivation instrument for biological experiments in space

    Science.gov (United States)

    Gmunder, F. K.; Nordau, C. G.; Tschopp, A.; Huber, B.; Cogoli, A.

    1988-01-01

    The prototype of a miniaturized cell cultivation instrument for animal cell culture experiments aboard Spacelab is presented (Dynamic cell culture system: DCCS). The cell chamber is completely filled and has a working volume of 200 microliters. Medium exchange is achieved with a self-powered osmotic pump (flowrate 1 microliter h-1). The reservoir volume of culture medium is 230 microliters. The system is neither mechanically stirred nor equipped with sensors. Hamster kidney (Hak) cells growing on Cytodex 3 microcarriers were used to test the biological performance of the DCCS. Growth characteristics in the DCCS, as judged by maximal cell density, glucose consumption, lactic acid secretion and pH, were similar to those in cell culture tubes.

  17. Cell sources for in vitro human liver cell culture models.

    Science.gov (United States)

    Zeilinger, Katrin; Freyer, Nora; Damm, Georg; Seehofer, Daniel; Knöspel, Fanny

    2016-09-01

    In vitro liver cell culture models are gaining increasing importance in pharmacological and toxicological research. The source of cells used is critical for the relevance and the predictive value of such models. Primary human hepatocytes (PHH) are currently considered to be the gold standard for hepatic in vitro culture models, since they directly reflect the specific metabolism and functionality of the human liver; however, the scarcity and difficult logistics of PHH have driven researchers to explore alternative cell sources, including liver cell lines and pluripotent stem cells. Liver cell lines generated from hepatomas or by genetic manipulation are widely used due to their good availability, but they are generally altered in certain metabolic functions. For the past few years, adult and pluripotent stem cells have been attracting increasing attention, due their ability to proliferate and to differentiate into hepatocyte-like cells in vitro However, controlling the differentiation of these cells is still a challenge. This review gives an overview of the major human cell sources under investigation for in vitro liver cell culture models, including primary human liver cells, liver cell lines, and stem cells. The promises and challenges of different cell types are discussed with a focus on the complex 2D and 3D culture approaches under investigation for improving liver cell functionality in vitro Finally, the specific application options of individual cell sources in pharmacological research or disease modeling are described. PMID:27385595

  18. Pedigree analysis of proliferation kinetics in cultured mammalian cells

    International Nuclear Information System (INIS)

    Quantitative analysis of proliferation kinetics in cultured mammalian cells was given an outline by extremely low speed photography (pedigree method). Photographing method of this analysis, camera used in this analysis, cultivation method and apparatus for cultivation, and film analysis were explained. As to changes of generation time by this analysis, relationship of each stage of cell cycle to colony formation or to proliferation capacity were explained in non-irradiated cells and irradiated cells. On abnormal cell division in time of large dose irradiation, a condition from cell fusion to cell death via multipolar division was explained. Mechanisms of proliferation death and interphase death were explained by analysis of pedigree data on radiation injuries in time of division and by mentioning division probability. Some information about inhibition of cell proliferation by radiation and lethal effect of radiation was described. (Kanao, N.)

  19. Differential Effect of Culture Temperature and Specific Growth Rate on CHO Cell Behavior in Chemostat Culture

    OpenAIRE

    Vergara, Mauricio; Becerra, Silvana; Berrios, Julio; Osses, Nelson; Reyes, Juan; Rodríguez-Moyá, María; Gonzalez, Ramon; Altamirano, Claudia

    2014-01-01

    Mild hypothermia condition in mammalian cell culture technology has been one of the main focuses of research for the development of breeding strategies to maximize productivity of these production systems. Despite the large number of studies that show positive effects of mild hypothermia on specific productivity of r-proteins, no experimental approach has addressed the indirect effect of lower temperatures on specific cell growth rate, nor how this condition possibly affects less specific pro...

  20. Mass spectrometric characterization of elements and molecules in cell cultures and tissues

    Science.gov (United States)

    Arlinghaus, H. F.; Kriegeskotte, C.; Fartmann, M.; Wittig, A.; Sauerwein, W.; Lipinsky, D.

    2006-07-01

    Time-of-flight secondary ion mass spectrometry (ToF-SIMS) and laser post-ionization secondary neutral mass spectrometry (laser-SNMS) have been used to image and quantify targeted compounds, intrinsic elements and molecules with subcellular resolution in single cells of both cell cultures and tissues. Special preparation procedures for analyzing cell cultures and tissue materials were developed. Cancer cells type MeWo, incubated with boronated compounds, were sandwiched between two substrates, cryofixed, freeze-fractured and freeze-dried. Also, after injection with boronated compounds, different types of mouse tissues were extracted, prepared on a special specimen carrier and plunged with high velocity into LN 2-cooled propane for cryofixation. After trimming, these tissue blocks were freeze-dried. The measurements of the K/Na ratio demonstrated that for both cell cultures and tissue materials the special preparation techniques used were appropriate for preserving the chemical and structural integrity of the living cell. The boron images show inter- and intracellular boron signals with different intensities. Molecular images show distinct features partly correlated with the cell structure. A comparison between laser-SNMS and ToF-SIMS showed that especially laser-SNMS is particularly well-suited for identifying specific cell structures and imaging ultratrace element concentrations in tissues.

  1. Mass spectrometric characterization of elements and molecules in cell cultures and tissues

    Energy Technology Data Exchange (ETDEWEB)

    Arlinghaus, H.F. [Physikalisches Institut, Universitaet Muenster, Wilhelm-Klemm-Str. 10, D-48149 Muenster (Germany)]. E-mail: arlinghaus@uni-muenster.de; Kriegeskotte, C. [Physikalisches Institut, Universitaet Muenster, Wilhelm-Klemm-Str. 10, D-48149 Muenster (Germany); Fartmann, M. [Physikalisches Institut, Universitaet Muenster, Wilhelm-Klemm-Str. 10, D-48149 Muenster (Germany); Wittig, A. [Strahlenklinik, Universitaetsklinikum Essen, D-45122 Essen (Germany); Sauerwein, W. [Strahlenklinik, Universitaetsklinikum Essen, D-45122 Essen (Germany); Lipinsky, D. [Physikalisches Institut, Universitaet Muenster, Wilhelm-Klemm-Str. 10, D-48149 Muenster (Germany)

    2006-07-30

    Time-of-flight secondary ion mass spectrometry (ToF-SIMS) and laser post-ionization secondary neutral mass spectrometry (laser-SNMS) have been used to image and quantify targeted compounds, intrinsic elements and molecules with subcellular resolution in single cells of both cell cultures and tissues. Special preparation procedures for analyzing cell cultures and tissue materials were developed. Cancer cells type MeWo, incubated with boronated compounds, were sandwiched between two substrates, cryofixed, freeze-fractured and freeze-dried. Also, after injection with boronated compounds, different types of mouse tissues were extracted, prepared on a special specimen carrier and plunged with high velocity into LN{sub 2}-cooled propane for cryofixation. After trimming, these tissue blocks were freeze-dried. The measurements of the K/Na ratio demonstrated that for both cell cultures and tissue materials the special preparation techniques used were appropriate for preserving the chemical and structural integrity of the living cell. The boron images show inter- and intracellular boron signals with different intensities. Molecular images show distinct features partly correlated with the cell structure. A comparison between laser-SNMS and ToF-SIMS showed that especially laser-SNMS is particularly well-suited for identifying specific cell structures and imaging ultratrace element concentrations in tissues.

  2. Culture and characterization of rat hair follicle stem cells.

    Science.gov (United States)

    Quan, Renfu; Zheng, Xuan; Ni, Yueming; Xie, Shangju; Li, Changming

    2016-08-01

    The purpose of this study was to establish methods for isolation, culture, expansion, and characterization of rat hair follicle stem cells (rHFSCs). Hair follicles were harvested from 1-week-old Sprague-Dawley rats and digested with dispase and collagenase IV. The bulge of the hair follicle was dissected under a microscope and cultured in Dulbecco's modified Eagle's medium/F12 supplemented with KnockOut™ Serum Replacement serum substitute, penicillin-streptomycin, L-glutamine, non-essential amino acids, epidermal growth factor, basic fibroblast growth factor, polyhydric alcohol, and hydrocortisone. The rHFSCs were purified using adhesion to collagen IV. Cells were characterized by detecting marker genes with immunofluorescent staining and real-time polymerase chain reaction (PCR). The proliferation and vitality of rHFSCs at different passages were evaluated. The cultured rHFSCs showed typical cobblestone morphology with good adhesion and colony-forming ability. Expression of keratin 15, integrin α6, and integrin β1 were shown by immunocytochemistry staining. On day 1-2, the cells were in the latent phase. On day 5-6, the cells were in the logarithmic phase. Cell vitality gradually decreased from the 7th passage. Real-time PCR showed that the purified rHFSCs had good vitality and proliferative capacity and contained no keratinocytes. Highly purified rHFSCs can be obtained using tissue culture and adhesion to collagen IV. The cultured cells had good proliferative capacity and could therefore be a useful cell source for tissue-engineered hair follicles, vessels, and skin. PMID:25407732

  3. Cell culture systems for the hepatitis C virus

    Institute of Scientific and Technical Information of China (English)

    Gilles Duverlie; Czeslaw Wychowski

    2007-01-01

    Since the discovery of HCV in 1989, the lack of a cell culture system has hampered research progress on this important human pathogen. No robust system has been obtained by empiric approaches, and HCV cell culture remained hypothetical until 2005. The construction of functional molecular clones has served as a starting point to reconstitute a consensus infectious cDNA that was able to transcribe infectious HCV RNAs as shown by intrahepatic inoculation in a chimpanzee. Other consensus clones have been selected and established in a human hepatoma cell line as replicons, i.e. self-replicating subgenomic or genomic viral RNAs. However, these replicons did not support production of infectious virus. Interestingly, some full-length replicons could be established without adaptive mutations and one of them was able to replicate at very high levels and to release virus particles that are infectious in cell culture and in vivo. This new cell culture system represents a major breakthrough in the HCV field and should enable a broad range of basic and applied studies to be achieved.

  4. Testicular Sertoli cells influence the proliferation and immunogenicity of co-cultured endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Ping, E-mail: fanpinggoodluck@163.com [Department of Rheumatism and Immunity, The First Affiliated Hospital Xi' an Jiaotong University School of Medicine, Xi' an, Shaanxi 710061 (China); He, Lan; Pu, Dan; Lv, Xiaohong; Zhou, Wenxu; Sun, Yining; Hu, Nan [Department of Rheumatism and Immunity, The First Affiliated Hospital Xi' an Jiaotong University School of Medicine, Xi' an, Shaanxi 710061 (China)

    2011-01-21

    Research highlights: {yields} The proliferation of dramatic increased by co-cultured with Sertoli cells. {yields} VEGF receptor-2 expression of ECs was up-regulated by co-cultured with Sertoli cells. {yields} The MHC expression of ECs induced by INF-{gamma} and IL-6, IL-8 and sICAM induced by TNF-{alpha} decreased respectively after co-cultured with Sertoli cells. {yields} ECs co-cultured with Sertoli cells also didn't increase the stimulation index of spleen lymphocytes. -- Abstract: The major problem of the application of endothelial cells (ECs) in transplantation is the lack of proliferation and their immunogenicity. In this study, we co-cultured ECs with Sertoli cells to monitor whether Sertoli cells can influence the proliferation and immunogenicity of co-cultured ECs. Sertoli cells were isolated from adult testicular tissue. ECs were divided into the control group and the experimental group, which included three sub-groups co-cultured with 1 x 10{sup 3}, 1 x 10{sup 4} or 1 x 10{sup 5} cell/ml of Sertoli cells. The growth and proliferation of ECs were observed microscopically, and the expression of vascular endothelial growth factor (VEGF) receptor-2 (KDR) was examined by Western blotting. In another experiment, ECs were divided into the control group, the single culture group and the co-culture group with the optimal concentration of Sertoli cells. After INF-{gamma} and TNF-{alpha} were added to the culture medium, MHC II antigen expression was detected by immunofluorescence staining and western blotting; interleukin (IL)-6, IL-8 and soluble intercellular adhesion molecule (sICAM) were measured in the culture medium by ELISA. We demonstrated that 1 x 10{sup 4} cell/ml Sertoli cells promoted the proliferation of co-cultured ECs more dramatically than that in other groups (P < 0.05). Western blotting showed that 1 x 10{sup 4} cell/ml of the Sertoli cells was most effective in the up-regulation of KDR expression in the co-cultured ECs (P < 0.05). Sertoli

  5. Testicular Sertoli cells influence the proliferation and immunogenicity of co-cultured endothelial cells

    International Nuclear Information System (INIS)

    Research highlights: → The proliferation of dramatic increased by co-cultured with Sertoli cells. → VEGF receptor-2 expression of ECs was up-regulated by co-cultured with Sertoli cells. → The MHC expression of ECs induced by INF-γ and IL-6, IL-8 and sICAM induced by TNF-α decreased respectively after co-cultured with Sertoli cells. → ECs co-cultured with Sertoli cells also didn't increase the stimulation index of spleen lymphocytes. -- Abstract: The major problem of the application of endothelial cells (ECs) in transplantation is the lack of proliferation and their immunogenicity. In this study, we co-cultured ECs with Sertoli cells to monitor whether Sertoli cells can influence the proliferation and immunogenicity of co-cultured ECs. Sertoli cells were isolated from adult testicular tissue. ECs were divided into the control group and the experimental group, which included three sub-groups co-cultured with 1 x 103, 1 x 104 or 1 x 105 cell/ml of Sertoli cells. The growth and proliferation of ECs were observed microscopically, and the expression of vascular endothelial growth factor (VEGF) receptor-2 (KDR) was examined by Western blotting. In another experiment, ECs were divided into the control group, the single culture group and the co-culture group with the optimal concentration of Sertoli cells. After INF-γ and TNF-α were added to the culture medium, MHC II antigen expression was detected by immunofluorescence staining and western blotting; interleukin (IL)-6, IL-8 and soluble intercellular adhesion molecule (sICAM) were measured in the culture medium by ELISA. We demonstrated that 1 x 104 cell/ml Sertoli cells promoted the proliferation of co-cultured ECs more dramatically than that in other groups (P 4 cell/ml of the Sertoli cells was most effective in the up-regulation of KDR expression in the co-cultured ECs (P < 0.05). Sertoli cells can effectively suppress INF-γ-induced MHC II antigen expression in co-cultured ECs compared with single culture

  6. Enhancement effect of shikonin in cell suspension culture and transfermanant culture by radiation application

    International Nuclear Information System (INIS)

    The cell lines 679, 679-29 and 622-46 of L. erythrorhizon could be selected on LS agar medium for the production shikonin in cell suspension culture. The shikonin was increased moderately in suspension culture of cell line 622-46 in LS liquid medium containing BA 2 mg·L-1 and IAA 0.2 mg·L-1 in the dark, and was increased by adding 1 μM Cu2+ and 100 μM methyl jasmonate The accumulation of shikonin in the liquid medium was increased significantly by 2 Gy irradiation to callus of cell line 622-46 and culture in LS liquid medium containing BA 2 mg·L-1 and IAA 0.2 mg·L-1 in the dark and shikonin in cell debris was higher by 16 Gy irradiation. The activity of p-hydroxybenzoate geranyltransferase was increased by irradiation of 2 Gy and 16 Gy of γ radiation. Seedling hypocotyles of L. erythrorhizon were infected with Agrogacterium rhizogenes strain 15834 harboring a binary vector with an intron bearing the GUS (β-glucuronidase) gene driven by cauliflower mosaic virus (CaMV) 35S promotor as well as the HPT (hygromycin phosphotransferase) gene as the selection marker. Hairy roots isolated were hygromycin resistant and had integrated GUS gene in DNA. The root tip grown on M-9 medium showed normal pigment production pattern in border cells and root hairs

  7. Detection of Infectious Bovine Rhinotracheitis and Bovine Viral Diarrhea Viruses in the Nasal Epithelial Cells by the Direct Immunofluorescence Technique

    OpenAIRE

    Silim, A.; Elazhary, M. A. S. Y.

    1983-01-01

    Nasal epithelial cells were collected by cotton swabs for the diagnosis in experimental and field cases of infectious bovine rhinotracheitis and field cases of bovine viral diarrhea in calves. A portion of the cells was washed twice in phosphate buffered saline and a 25 µL drop was placed on microscope slides. The cells were dried, fixed and stained according to the direct fluorescent antibody technique. Another portion of the same specimen was inoculated onto primary bovine skin cell culture...

  8. Expansion and properties of human bone marrow stromal cells cultured in different culture media

    Czech Academy of Sciences Publication Activity Database

    Syrová, Zdeňka; Glogarová, Kateřina; Jendelová, Pavla; Syková, Eva

    2006-01-01

    Roč. 8, Supplement 1 (2006), s. 238-238. ISSN 1465-3249. [ ISCT 2006. 04.05.2006-07.05.2006, Berlin] R&D Projects: GA MZd(CZ) NR8339; GA MŠk 1M0538 Institutional research plan: CEZ:AV0Z50390512 Keywords : Cells cultured Subject RIV: FH - Neurology

  9. Co-culture of vascular endothelial cells and smooth muscle cells by hyaluronic acid micro-pattern on titanium surface

    Science.gov (United States)

    Li, Jingan; Li, Guicai; Zhang, Kun; Liao, Yuzhen; Yang, Ping; Maitz, Manfred F.; Huang, Nan

    2013-05-01

    Micro-patterning as an effective bio-modification technique is increasingly used in the development of biomaterials with superior mechanical and biological properties. However, as of now, little is known about the simultaneous regulation of endothelial cells (EC) and smooth muscle cells (SMC) by cardiovascular implants. In this study, a co-culture system of EC and SMC was built on titanium surface by the high molecular weight hyaluronic acid (HMW-HA) micro-pattern. Firstly, the micro-pattern sample with a geometry of 25 μm wide HMW-HA ridges, and 25 μm alkali-activated Ti grooves was prepared by microtransfer molding (μTM) for regulating SMC morphology. Secondly, hyaluronidase was used to decompose high molecular weight hyaluronic acid into low molecular weight hyaluronic acid which could promote EC adhesion. Finally, the morphology of the adherent EC was elongated by the SMC micro-pattern. The surface morphology of the patterned Ti was imaged by SEM. The existence of high molecular weight hyaluronic acid on the modified Ti surface was demonstrated by FTIR. The SMC micro-pattern and EC/SMC co-culture system were characterized by immunofluorescence microscopy. The nitric oxide release test and cell retention calculation were used to evaluate EC function on inhibiting hyperplasia and cell shedding, respectively. The results indicate that EC in EC/SMC co-culture system displayed a higher NO release and cell retention compared with EC cultured alone. It can be suggested that the EC/SMC co-culture system possessed superiority to EC cultured alone in inhibiting hyperplasia and cell shedding at least in a short time of 24 h.

  10. Arsenic exposure induces the Warburg effect in cultured human cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Fei; Severson, Paul; Pacheco, Samantha; Futscher, Bernard W.; Klimecki, Walter T., E-mail: klimecki@pharmacy.arizona.edu

    2013-08-15

    Understanding how arsenic exacts its diverse, global disease burden is hampered by a limited understanding of the particular biological pathways that are disrupted by arsenic and underlie pathogenesis. A reductionist view would predict that a small number of basic pathways are generally perturbed by arsenic, and manifest as diverse diseases. Following an initial observation that arsenite-exposed cells in culture acidify their media more rapidly than control cells, the report here shows that low level exposure to arsenite (75 ppb) is sufficient to induce aerobic glycolysis (the Warburg effect) as a generalized phenomenon in cultured human primary cells and cell lines. Expanded studies in one such cell line, the non-malignant pulmonary epithelial line, BEAS-2B, established that the arsenite-induced Warburg effect was associated with increased accumulation of intracellular and extracellular lactate, an increased rate of extracellular acidification, and inhibition by the non-metabolized glucose analog, 2-deoxy-D-glucose. Associated with the induction of aerobic glycolysis was a pathway-wide induction of glycolysis gene expression, as well as protein accumulation of an established glycolysis master-regulator, hypoxia-inducible factor 1A. Arsenite-induced alteration of energy production in human cells represents the type of fundamental perturbation that could extend to many tissue targets and diseases. - Highlights: • Chronic arsenite exposure induces aerobic glycolysis, dubbed the “Warburg effect”. • Arsenite-induced Warburg effect is a general phenomenon in cultured human cells. • HIF-1A may mediate arsenite induced Warburg effect.

  11. Testing of serum atherogenicity in cell cultures: questionable data published

    Directory of Open Access Journals (Sweden)

    Sergei V. Jargin

    2012-01-01

    Full Text Available In a large series of studies was reported that culturing of smooth muscle cells with serum from atherosclerosis patients caused intracellular lipid accumulation, while serum from healthy controls had no such effect. Cultures were used for evaluation of antiatherogenic drugs. Numerous substances were reported to lower serum atherogenicity: statins, trapidil, calcium antagonists, garlic derivatives etc. On the contrary, beta-blockers, phenothiazines and oral hypoglycemics were reported to be pro-atherogenic. Known antiatherogenic agents can influence lipid metabolism and cholesterol synthesis, intestinal absorption or endothelium-related mechanisms. All these targets are absent in cell monocultures. Inflammatory factors, addressed by some antiatherogenic drugs, are also not reproduced. In vivo, relationship between cholesterol uptake by cells and atherogenesis must be inverse rather than direct: in familial hypercholesterolemia, inefficient clearance of LDL-cholesterol by cells predisposes to atherosclerosis. Accordingly, if a pharmacological agent reduces cholesterol uptake by cells in vitro, it should be expected to elevate cholesterol in vivo. Validity of clinical recommendations, based on serum atherogenicity testing in cell monocultures, is therefore questionable. These considerations pertain also to the drugs developed on the basis of the cell culture experiments.

  12. Arsenic exposure induces the Warburg effect in cultured human cells

    International Nuclear Information System (INIS)

    Understanding how arsenic exacts its diverse, global disease burden is hampered by a limited understanding of the particular biological pathways that are disrupted by arsenic and underlie pathogenesis. A reductionist view would predict that a small number of basic pathways are generally perturbed by arsenic, and manifest as diverse diseases. Following an initial observation that arsenite-exposed cells in culture acidify their media more rapidly than control cells, the report here shows that low level exposure to arsenite (75 ppb) is sufficient to induce aerobic glycolysis (the Warburg effect) as a generalized phenomenon in cultured human primary cells and cell lines. Expanded studies in one such cell line, the non-malignant pulmonary epithelial line, BEAS-2B, established that the arsenite-induced Warburg effect was associated with increased accumulation of intracellular and extracellular lactate, an increased rate of extracellular acidification, and inhibition by the non-metabolized glucose analog, 2-deoxy-D-glucose. Associated with the induction of aerobic glycolysis was a pathway-wide induction of glycolysis gene expression, as well as protein accumulation of an established glycolysis master-regulator, hypoxia-inducible factor 1A. Arsenite-induced alteration of energy production in human cells represents the type of fundamental perturbation that could extend to many tissue targets and diseases. - Highlights: • Chronic arsenite exposure induces aerobic glycolysis, dubbed the “Warburg effect”. • Arsenite-induced Warburg effect is a general phenomenon in cultured human cells. • HIF-1A may mediate arsenite induced Warburg effect

  13. Adenosine formation in contracting primary rat skeletal muscle cells and endothelial cells in culture

    DEFF Research Database (Denmark)

    Hellsten, Ylva; Frandsen, Ulrik

    1997-01-01

    1. The present study examined the capacity for adenosine formation, uptake and metabolism in contracting primary rat muscle cells and in microvascular endothelial cells in culture. 2. Strong and moderate electrical simulation of skeletal muscle cells led to a significantly greater increase in the...... extracellular adenosine concentration (421 +/- 91 and 235 +/- 30 nmol (g protein)-1, respectively; P < 0.05) compared with non-stimulated muscle cells (161 +/- 20 nmol (g protein)-1). The ATP concentration was lower (18%; P < 0.05) in the intensely contracted, but not in the moderately contracted muscle cells....... 3. Addition of microvascular endothelial cells to the cultured skeletal muscle cells enhanced the contraction-induced accumulation of extracellular adenosine (P < 0.05), whereas endothelial cells in culture alone did not cause extracellular accumulation of adenosine. 4. Skeletal muscle cells were...

  14. PDMS/glass microfluidic cell culture system for cytotoxicity tests and cells passage

    DEFF Research Database (Denmark)

    Ziolkowska, K.; Jedrych, E.; Kwapiszewski, R.; Lopacinska, Joanna M.; Skolimowski, Maciej; Chudy, M.

    2010-01-01

    In this paper, hybrid (PDMS/glass) microfluidic cell culture system (MCCS) integrated with the concentration gradient generator (CGG) is presented. PDMS gas permeability enabled cells' respiration in the fabricated microdevices and excellent glass hydrophilicity allowed successful cells' seeding....... The human lung carcinoma cells (A549) were cultured in the microdevice for several days. The growth and proliferation of cells was monitored using an inverted fluorescence microscope. After the cells' confluence was achieved in the microchambers, the novel method of cells' passaging in the designed...... microdevice was developed and successfully tested. The MCCS microdevice is fully reusable, i.e. it can be used several times for various cell culture and cytotoxic experiments. The suitability of designed MCCS for cell-based cytotoxicity assay application was verified using 1,4-dioxane as a model toxic agent...

  15. Studies on the Developmental Potentiality of Cultured Cell Nuclei of Fish

    Directory of Open Access Journals (Sweden)

    Hongxi Chen, Yonglan Yi, Minrong Chen, Xingqi Yang

    2010-01-01

    Full Text Available By means of the serial nuclear transplantation technique, the authors obtained a nuclear transplant fish from subcultured cell originated from the blastula cells of the crucian carp (Carassius auratus Linnaeus. This nuclear transplant fish survived for three years, but its sexual glands were undifferentiated. The authors have also obtained a sexually mature adult fish from short-term cultured kidney cell nucleus of an adult crucian carp. Results of the experiment implied that the subcultured cell nuclei of fish blastula cells and the specialized somatic cell nuclei of adult fish still retained their developmental totipotency, and thus, it indicated that there is a possibility of fish somatic cell breeding through the use of nuclear transplantation.

  16. Adenosine-induced hyperpolarization of the membrane voltage in rat mesangial cells in primary culture.

    OpenAIRE

    Pavenstädt, H. (Hermann); Ruh, J; Greger, R; Schollmeyer, P.

    1994-01-01

    1. The effect of adenosine on membrane voltage and ion currents was studied in rat mesangial cells in primary culture. Membrane voltage was measured with the patch clamp technique in the slow- or fast whole cell configuration. The resting membrane voltage of mesangial cells was -48 +/- 0.5 mV. Adenosine (10(-8)-10(-3) M) induced a sustained and concentration-dependent hyperpolarization of membrane voltage (ED50 approximately 6 x 10(-7) M). Adenosine (10(-5) M) hyperpolarized the membrane volt...

  17. The effects of hypoxia on in vitro culture of dental-derived stem cells.

    Science.gov (United States)

    Werle, Stefanie Bressan; Chagastelles, Pedro; Pranke, Patricia; Casagrande, Luciano

    2016-08-01

    The culture of cells under hypoxia is considered one of the hot topics of tissue engineering, especially when exploring the proliferation capacity, a critical step for cellular-based therapies. The use of in vitro hypoxic environment aims to simulate the oxygen concentrations found in stem cell niches. Dental tissues are attractive sources of stem cells, as they are obtained from discarded tissue, after third molar extraction and exfoliation deciduous teeth, respectively. However, small amounts of cells are obtained from these sources. Thus, optimizing the in vitro conditions for proliferation and differentiation of these cells is essential for future regenerative strategies. This review presents a summary of the results regarding the effect of hypoxia on dental-derived stem cells after an electronic search on PubMed databases. The studies show increased differentiation potential and paracrine action of dental-derived stem cells under hypoxic environment. There are controversies related to proliferation of dental-derived stem cells under induced hypoxia. The lack of standardization in cell culture techniques contributes to these biases and future studies should describe in more detail the protocols used. The knowledge regarding the effect of hypoxia on dental-derived stem cells needs further clarification for assisting the clinical application of these cells. PMID:27045351

  18. Effects of Visible Light on Cultured Bovine Trabecular Cells

    Institute of Scientific and Technical Information of China (English)

    姜发纲; 郝风芹; 魏厚仁; 许德胜

    2004-01-01

    To explore the biological effects of light on trabecular cells, cultured bovine trabecular cells were exposed to visible light of different wavelength with different energy. Cellular morphology, structure, proliferation, and phagocytosis were observed. The cells showed no remarkable changes when the energy was low. When the exposure energy reached 1. 12 mW/cm2 , the cytoplasm showed a rough appearance, and cell proliferation and phagocytosis decreased. This phototoxicity was strong with white light (compound chromatic light), moderate with violet light or yellow light, and mild with red light.

  19. Kinetics of meta-iodo-benzylguanidine in neuroblastoma cell cultures

    International Nuclear Information System (INIS)

    The continuously cultured human neuroblastoma cell line SK-N-SH possesses an active transport mechanism for uptake of meta-iodobenzylguanidine (MIBG). The uptake rate at an MIBG-concentration of 6,4x10-8 M was 0.35x10-12 mol/minx106 cells, corresponding to values as measured in human pheochromocytoma cell lines. MIBG is released from the cells with a biological half-life of 81,3 h in correspondence to half-life values as measured in vivo in neuroblastoma patients. (orig.)

  20. Cultural Techniques of Cognitive Capitalism: Metaprogramming and the Labour of Code

    Directory of Open Access Journals (Sweden)

    Jussi Parikka

    2014-03-01

    Full Text Available This article addresses cultural techniques of cognitive capitalism. The author argues that to understand the full implications of the notion of cognitive capitalism we need to address the media and cultural techniques which conditions its range and applications. The article offers an expanded understanding of the labour of code and programming through a case study of 'metaprogramming', a software related organisation practice that offered a way to think of software creativity and programming in organisations. The ideas from the 1970s that are discussed offer a different way to approach creativity and collaborative and post-Fordist capitalism. The author brings together different theoretical perspectives, including German media theory and Yann Moulier Boutang’s thesis about cognitive capitalism. The wider argument is that we should pay more attention to the media archaeological conditions of practices of labour and value appropriation of contemporary technological capitalism as well as the cultural techniques which include 'ontological and aesthetic operations' that produce cultural, material situations.

  1. Load Cell Response Correction Using Analog Adaptive Techniques

    OpenAIRE

    Jafaripanah, Mehdi; Al-Hashimi, Bashir; White, Neil M.

    2003-01-01

    Load cell response correction can be used to speed up the process of measurement. This paper investigates the application of analog adaptive techniques in load cell response correction. The load cell is a sensor with an oscillatory output in which the measurand contributes to response parameters. Thus, a compensation filter needs to track variation in measurand whereas a simple, fixed filter is only valid at one load value. To facilitate this investigation, computer models for the load cell a...

  2. Rapport d'activité 2008-2010 : Culture Scientifique et Technique

    OpenAIRE

    Le Gall, Alain-Hervé; Jézéquel, Isabelle; Bernard, Dominique; Canard, Alain

    2011-01-01

    C'est à cet objectif que tente de répondre l'Université de Rennes 1 en organisant des activités regroupées sous le terme générique de " culture scientifique et technique ". Notre université compte de nombreux laboratoires mixtes réunissant à la fois universitaires (enseignants-chercheurs et personnels BIATOSS), chercheurs et ITA des grands organismes de recherche qui ont tous cette mission ou cette passion de diffusion des connaissances et d'éducation à la culture scientifique. Ceux-ci n'hési...

  3. Human adipose-derived stromal/stem cell isolation, culture, and osteogenic differentiation.

    Science.gov (United States)

    Qureshi, Ammar T; Chen, Cong; Shah, Forum; Thomas-Porch, Caasy; Gimble, Jeffrey M; Hayes, Daniel J

    2014-01-01

    Annually, more than 200,000 elective liposuction procedures are performed in the United States and over a million worldwide. The ease of harvest and abundance make human adipose-derived stromal/stem cells (hASCs) isolated from lipoaspirates an attractive, readily available source of adult stem cells that have become increasingly popular for use in many studies. Here, we describe common methods for hASC culture, preservation, and osteogenic differentiation. We introduce methods of ceramic, polymer, and composite scaffold synthesis with a description of morphological, chemical, and mechanical characterization techniques. Techniques for scaffold loading are compared, and methods for determining cell loading efficiency and proliferation are described. Finally, we provide both qualitative and quantitative techniques for in vitro assessment of hASC osteogenic differentiation. PMID:24529434

  4. Isolation and culture of umbilical vein mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    D.T. Covas

    2003-09-01

    Full Text Available Bone marrow contains a population of stem cells that can support hematopoiesis and can differentiate into different cell lines including adipocytes, osteocytes, chondrocytes, myocytes, astrocytes, and tenocytes. These cells have been denoted mesenchymal stem cells. In the present study we isolated a cell population derived from the endothelium and subendothelium of the umbilical cord vein which possesses morphological, immunophenotypical and cell differentiation characteristics similar to those of mesenchymal stem cells isolated from bone marrow. The cells were isolated from three umbilical cords after treatment of the umbilical vein lumen with collagenase. The cell population isolated consisted of adherent cells with fibroblastoid morphology which, when properly stimulated, gave origin to adipocytes and osteocytes in culture. Immunophenotypically, this cell population was found to be positive for the CD29, CD13, CD44, CD49e, CD54, CD90 and HLA-class 1 markers and negative for CD45, CD14, glycophorin A, HLA-DR, CD51/61, CD106, and CD49d. The characteristics described are the same as those presented by bone marrow mesenchymal stem cells. Taken together, these findings indicate that the umbilical cord obtained from term deliveries is an important source of mesenchymal stem cells that could be used in cell therapy protocols.

  5. Cannabinoids induce incomplete maturation of cultured human leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Murison, G.; Chubb, C.B.H.; Maeda, S.; Gemmell, M.A.; Huberman, E.

    1987-08-01

    Monocyte maturation markers were induced in cultured human myeloblastic ML-2 leukemia cells after treatment for 1-6 days with 0.03-30 ..mu..M ..delta../sup 9/-tetrahydrocannabinol (THC), the major psychoactive component of marijuana. After a 2-day or longer treatment, 2- to 5-fold increases were found in the percentages of cells exhibiting reactivity with either the murine OKM1 monoclonal antibody of the Leu-M5 monoclonal antibody, staining positively for nonspecific esterase activity, and displaying a promonocyte morphology. The increases in these differentiation markers after treatment with 0.03-1 ..mu..M THC were dose dependent. At this dose range, THC did not cause an inhibition of cell growth. The THC-induced cell maturation was also characterized by specific changes in the patterns of newly synthesized proteins. The THC-induced differentiation did not, however, result in cells with a highly developed mature monocyte phenotype. However, treatment of these incompletely matured cells with either phorbol 12-myristate 13-acetate of 1..cap alpha..,25-dihydroxycholecalciferol, which are inducers of differentiation in myeloid leukemia cells (including ML-2 cells), produced cells with a mature monocyte morphology. The ML-2 cell system described here may be a useful tool for deciphering critical biochemical events that lead to the cannabinoid-induced incomplete cell differentiation of ML-2 cells and other related cell types. Findings obtained from this system may have important implications for studies of cannabinoid effects on normal human bone-marrow progenitor cells.

  6. Culturing intestinal stem cells: applications for colorectal cancer research

    OpenAIRE

    Fujii, Masayuki; Sato, Toshiro

    2014-01-01

    Recent advance of sequencing technology has revealed genetic alterations in colorectal cancer (CRC). The biological function of recurrently mutated genes has been intensively investigated through mouse genetic models and CRC cell lines. Although these experimental models may not fully reflect biological traits of human intestinal epithelium, they provided insights into the understanding of intestinal stem cell self-renewal, leading to the development of novel human intestinal organoid culture...

  7. New Generation of Cell Culture Assay for Smallpox Vaccine Potency

    OpenAIRE

    Leparc-Goffart, Isabelle; Poirier, Bertrand; El Zaouk, Annie; Tissier, Marie-Hélène; Fuchs, Florence

    2003-01-01

    The potency of smallpox vaccines produced in the 1970s was tested by titration onto chorioallantoic membranes of fertilized hen eggs (CAM assay). The potency specification commonly approved for these vaccines was a titer above 108 pock-forming units per milliliter. We developed and validated a cell culture titration assay to have a more reliable potency test. The cell titration assay and the CAM assay were tested in parallel on 34 first-generation smallpox vaccine lots. These allowed us to de...

  8. Endocytic activity of Sertoli cells grown in bicameral culture chambers

    International Nuclear Information System (INIS)

    Immature rat Sertoli cells were cultured for 7 to 14 days on Millipore filters impregnated with a reconstituted basement membrane extract in dual-environment (bicameral) culture chambers. Electron microscopy of the cultured cells revealed the presence of rod-shaped mitochondria, Golgi apparatus, rough endoplasmic reticulum, and Sertoli-Sertoli tight junctions, typical of these cells in vivo. The endocytic activity of both the apical and basal surfaces of the Sertoli cells was examined by either adding alpha 2-macroglobulin (alpha 2-M) conjugated to 20 nm gold particles to the apical chamber or by adding 125I labeled alpha 2-M to the basal chamber. During endocytosis from the apical surface of Sertoli cells, the alpha 2-M-gold particles were bound initially to coated pits and then internalized into coated vesicles within 5 minutes. After 10 minutes, the alpha 2-M-gold was found in multi-vesicular bodies (MVBs) and by 30 minutes it was present in the lysosomes. The proportion of alpha 2-M-gold found within endocytic cell organelles after 1 hour of uptake was used to estimate the approximate time that this ligand spent in each type of organelle. The alpha 2-M-gold was present in coated pits, coated vesicles, multivesicular bodies, and lysosomes for approximately 3, 11, 22, and 24 minutes, respectively. This indicates that the initial stages of endocytosis are rapid, whereas MVBs and lysosomes are relatively long-lived

  9. Staurosporine induces different cell death forms in cultured rat astrocytes

    International Nuclear Information System (INIS)

    Astroglial cells are frequently involved in malignant transformation. Besides apoptosis, necroptosis, a different form of regulated cell death, seems to be related with glioblastoma genesis, proliferation, angiogenesis and invasion. In the present work we elucidated mechanisms of necroptosis in cultured astrocytes, and compared them with apoptosis, caused by staurosporine. Cultured rat cortical astrocytes were used for a cell death studies. Cell death was induced by different concentrations of staurosporine, and modified by inhibitors of apoptosis (z-vad-fmk) and necroptosis (nec-1). Different forms of a cell death were detected using flow cytometry. We showed that staurosporine, depending on concentration, induces both, apoptosis as well as necroptosis. Treatment with 10−7 M staurosporine increased apoptosis of astrocytes after the regeneration in a staurosporine free medium. When caspases were inhibited, apoptosis was attenuated, while necroptosis was slightly increased. Treatment with 10−6 M staurosporine induced necroptosis that occurred after the regeneration of astrocytes in a staurosporine free medium, as well as without regeneration period. Necroptosis was significantly attenuated by nec-1 which inhibits RIP1 kinase. On the other hand, the inhibition of caspases had no effect on necroptosis. Furthermore, staurosporine activated RIP1 kinase increased the production of reactive oxygen species, while an antioxidant BHA significantly attenuated necroptosis. Staurosporine can induce apoptosis and/or necroptosis in cultured astrocytes via different signalling pathways. Distinction between different forms of cell death is crucial in the studies of therapy-induced necroptosis

  10. Rapid method for culturing embryonic neuron-glial cell cocultures

    DEFF Research Database (Denmark)

    Svenningsen, Åsa Fex; Shan, Wei-Song; Colman, David R;

    2003-01-01

    enteric neurons is seen after 3 weeks (2 weeks in ascorbic acid), suggesting that basal lamina production is important even for glial ensheathment in the enteric nervous system. No overgrowth of fibroblasts or other nonneuronal cells was noted in any cultures, and myelination of the peripheral nervous...

  11. Plant Cell Cultures as Source of Cosmetic Active Ingredients

    Directory of Open Access Journals (Sweden)

    Ani Barbulova

    2014-04-01

    Full Text Available The last decades witnessed a great demand of natural remedies. As a result, medicinal plants have been increasingly cultivated on a commercial scale, but the yield, the productive quality and the safety have not always been satisfactory. Plant cell cultures provide useful alternatives for the production of active ingredients for biomedical and cosmetic uses, since they represent standardized, contaminant-free and biosustainable systems, which allow the production of desired compounds on an industrial scale. Moreover, thanks to their totipotency, plant cells grown as liquid suspension cultures can be used as “biofactories” for the production of commercially interesting secondary metabolites, which are in many cases synthesized in low amounts in plant tissues and differentially distributed in the plant organs, such as roots, leaves, flowers or fruits. Although it is very widespread in the pharmaceutical industry, plant cell culture technology is not yet very common in the cosmetic field. The aim of the present review is to focus on the successful research accomplishments in the development of plant cell cultures for the production of active ingredients for cosmetic applications.

  12. Biodegradable Mg corrosion and osteoblast cell culture studies

    International Nuclear Information System (INIS)

    Magnesium (Mg) is a biodegradable metal that has significant potential advantages as an implant material. In this paper, corrosion and cell culture experiments were performed to evaluate the biocompatibility of Mg. The corrosion current and potential of a Mg disk were measured in different physiological solutions including deionized (DI) water, phosphate-buffered saline (PBS), and McCoy's 5A culture medium. The corrosion currents in the PBS and in the McCoy's 5A-5% FBS media were found to be higher than in DI water, which is expected because corrosion of Mg occurs faster in a chloride solution. Weight loss, open-circuit potential, and electrochemical impedance spectroscopy measurements were also performed. The Mg specimens were also characterized using an environmental scanning electron microscope and energy-dispersive X-ray analysis (EDAX). The X-ray analysis showed that in the cell culture media a passive interfacial layer containing oxygen, chloride, phosphate, and potassium formed on the samples. U2OS cells were then co-cultured with a Mg specimen for up to one week. Cytotoxicity results of magnesium using MTT assay and visual observation through cell staining were not significantly altered by the presence of the corroding Mg sample. Further, bone tissue formation study using von Kossa and alkaline phosphatase staining indicates that Mg may be suitable as a biodegradable implant material.

  13. Test chambers for cell culture in static magnetic field

    International Nuclear Information System (INIS)

    Article presents a test chamber intended to be used for in vitro cell culture in homogenous constant magnetic field with parametrically variable magnitude. We constructed test chambers with constant parameters of control homeostasis of cell culture for the different parameters of static magnetic field. The next step was the computer calculation of 2D and 3D simulation of the static magnetic field distribution in the chamber. The analysis of 2D and 3D calculations of magnetic induction in the cells' exposition plane reveals, in comparison to the detection results, the greater accuracy of 2D calculations (Figs. 9 and 10). The divergence in 2D method was 2–4% and 8 to 10% in 3D method (reaching 10% only out of the cells′ cultures margins). -- Highlights: ► We present test chamber to be used for in vitro cell culture in static magnetic field. ► The technical data of the chamber construction was presented. ► 2D versus 3D simulation of static magnetic field distribution in chamber was reported. ► We report the accuracy of 2D calculation than 3D

  14. System-level modeling and simulation of the cell culture microfluidic biochip ProCell

    OpenAIRE

    Minhass, Wajid Hassan; Pop, Paul; Madsen, Jan; Hemmingsen, Mette; Dufva, Martin

    2010-01-01

    Microfluidic biochips offer a promising alternative to a conventional biochemical laboratory. There are two technologies for the microfluidic biochips: droplet-based and flow-based. In this paper we are interested in flow-based microfluidic biochips, where the liquid flows continuously through pre-defined micro-channels using valves and pumps. We present an approach to the system-level modeling and simulation of a cell culture microfluidic biochip called ProCell, Programmable Cell Culture Chi...

  15. Fabrication of a thermoresponsive cell culture dish: a key technology for cell sheet tissue engineering

    OpenAIRE

    Jun Kobayashi and Teruo Okano

    2010-01-01

    This article reviews the properties and characterization of an intelligent thermoresponsive surface, which is a key technology for cell sheet-based tissue engineering. Intelligent thermoresponsive surfaces grafted with poly(N-isopropylacrylamide) exhibit hydrophilic/hydrophobic alteration in response to temperature change. Cultured cells are harvested on thermoresponsive cell culture dishes by decreasing the temperature without the use of digestive enzymes or chelating agents. Our group has d...

  16. INCREASED OSMOLARITY AND CELL CLUSTERING PRESERVES CANINE NOTOCHORDAL CELL PHENOTYPE IN CULTURE

    OpenAIRE

    Spillekom, S; Smolders, L.A.; Grinwis, G.C.M.; Arkesteijn, I.; Ito, K.; Meij, B. P.; Tryfonidou, M.A.

    2013-01-01

    Abstract Degeneration of the intervertebral disc (IVD) is associated with a loss of notochordal cells (NCs) from the nucleus pulposus (NP) and their replacement by chondrocyte-like cells. NCs are known to maintain extracellular matrix quality and stimulate the chondrocyte-like NP cells, making NCs attractive for designing new tissue engineering approaches for IVD regeneration. However, optimal conditions, such as osmolarity and other characteristics of the culture media, for long-term culture...

  17. Tailor-made three-dimensional hybrid scaffolds for cell cultures

    International Nuclear Information System (INIS)

    The construction of the ideal three-dimensional scaffold for cell culture is one of the most intriguing topics in tissue engineering. It has been shown that cells can be cultured on most organic biomimetic materials, which now are losing popularity in favour of novel, hybrid systems. In this study, a series of photosensitive sol-gel hybrid materials, based on silicon-zirconium and silicon-titanium oxides, have been investigated for their suitability in three-dimensional scaffold fabrication. These materials can be structured by two-photon polymerization, a laser-based technique allowing the fabrication of micrometre-size structures with submicron resolution. The work presented here examined the effect of the organic/inorganic composition of the materials on cell behaviour and the establishment of a 'cell-culture friendly' environment. This is vital for cell adhesion, growth and differentiation, as the organic part of the material provides the soft matrix for cell growth, whereas the inorganic component gives the mechanical stability and rigidity of the three-dimensional structures. In addition, the use of femtosecond laser structuring permits the fabrication of a wide range of mechanically stable scaffolds of different sizes and shapes to be tested in terms of cell viability, proliferation and orientation.

  18. Neural differentiation of human placenta-derived mesenchymal stem cells following neural cell co-culture

    Institute of Scientific and Technical Information of China (English)

    Nailong Yang; Hongyan Zhang; Xiaojuan Sun; Lili Xu

    2011-01-01

    We induced human placenta-derived mesenchymal stem cells (hPMSCs) to differentiate into neural cells by adding chemical reagents,despite the fact that toxic chemicals induce cell shrinkage or cytoskeletal formation,which does not represent a proper cell differentiation process.The present study established a co-culture system with hPMSCs and neural cells and analyzed the influence of neural cells on hPMSC differentiation in a co-culture system.hPMSCs were isolated and purified from human full-term placenta using collagenase digestion.Fetal neural cells were co-cultured with hPMSCs for 48 hours using the Transwell co-culture system.hPMSCs co-cultured with neural cells exhibited a slender morphology with a filament.After 96 hours,hPMSCs expressed neuron-specific enolase,which suggested that co-culture of hPMSCs and neural cells induced neural differentiation of hPMSCs.

  19. Selective cell destruction and precise neurite transection in neuroblastoma cultures with pulsed ultraviolet laser microbeam irradiation

    International Nuclear Information System (INIS)

    Ultraviolet (UV) laser pulses at 337 nm and 12 nsec were used as an effective surgical tool for the elimination of cells and the transection of cellular processes in culture. Analyses of lased neuroblastoma cells with light and scanning electron microscopy show that precise, immediate lesions of 0.5 μm to 2 μm can be produced using either of two techniques: (1) direct laser transections at energy densities above 12 μJ/μm2; and (2) indirect transections by involving the substrate in the surgery process at energy densities ranging from 1 to 12 μJ/μm2. The first technique uses cytoplasmic cavitations in the laser focus; the second depends on shock waves created by vaporizing substrate. Damage thresholds have been established for commonly used tissue culture materials and for unpigmented cytoplasm. Thresholds for all plastics tested lie below that of unpigmented cytoplasm making the direct transection technique inapplicable because of the formation of large, destructive shock waves. A reduction of energy densities to the damage thresholds of specific substrate materials allows the production of minute shock waves that create the highly localized damage required for reliable, precise and non-lethal cell surgery in culture. (Auth.)

  20. Mutation breeding through in vitro culture techniques for resistance to cocoa swollen shoot virus disease

    International Nuclear Information System (INIS)

    The Cocoa Swollen Shoot Virus (CSSV) disease is epidemic in cocoa (Theobroma cacao) in Ghana, but there is no resistant material in the genetic resources available. In order to widen the genetic variability in this crop, efforts were made to apply conventional mutation breeding and in combination with in vitro culture technique. Gamma irradiation with doses less than 100 Gy were applied to pollen, beans and vegetative buds, and more than 1,000 M2/F1 or M1V3 plants for each experiments were screened for CSSV resistance by patch grafting method. The results indicated that it is feasible to use vegetative buds and in vitro culture to generate the desired variation. Some promising CSSV resistant clones are under further testing. In vitro culture conditions are also reported. (author). 7 refs, 7 tabs

  1. Osteoinductive activity of insulin-functionalized cell culture surfaces obtained using diazonium chemistry

    Science.gov (United States)

    Mikulska, Anna; Filipowska, Joanna; Osyczka, Anna; Nowakowska, Maria; Szczubiałka, Krzysztof

    2014-12-01

    Polymeric surfaces suitable for cell culture (DR/Pec) were constructed from diazoresin (DR) and pectin (Pec) in a form of ultrathin films using the layer-by-layer (LbL) technique. The surfaces were functionalized with insulin using diazonium chemistry. Such functionalized surfaces were used to culture human mesenchymal stem cells (hMSCs) to assess their suitability for bone tissue engineering and regeneration. The activity of insulin immobilized on the surfaces (DR/Pec/Ins) was compared to that of insulin dissolved in the culture medium. Human MSC grown on insulin-immobilized DR/Pec surfaces displayed increased proliferation and higher osteogenic activity. The latter was determined by means of alkaline phosphatase (ALP) activity, which increases at early stages of osteoblasts differentiation. Insulin dissolved in the culture medium did not stimulate cell proliferation and its osteogenic activity was significantly lower. Addition of recombinant human bone morphogenetic protein 2 (rhBMP-2) to the culture medium further increased ALP activity in hMSCs indicating additive osteogenic action of immobilized insulin and rhBMP-2

  2. The molecularly crowded cytoplasm of bacterialcCells : Dividing cells contrasted with viable but non-culturable (VBNC) bacterial cells

    NARCIS (Netherlands)

    Trevors, J. T.; van Elsas, J. D.; Bej, A. K.

    2013-01-01

    In this perspective, we discuss the cytoplasm in actively growing bacterial cells contrasted with viable but non-culturable (VBNC) cells. Actively growing bacterial cells contain a more molecularly crowded and organized cytoplasm, and are capable of completing their cell cycle resulting in cell divi

  3. Human disc cells in monolayer vs 3D culture: cell shape, division and matrix formation

    Directory of Open Access Journals (Sweden)

    Hanley Edward N

    2000-10-01

    Full Text Available Abstract Background The relationship between cell shape, proliferation, and extracellular matrix (ECM production, important aspects of cell behavior, is examined in a little-studied cell type, the human annulus cell from the intervertebral disc, during monolayer vs three-dimensional (3D culture. Results Three experimental studies showed that cells respond specifically to culture microenvironments by changes in cell shape, mitosis and ECM production: 1 Cell passages showed extensive immunohistochemical evidence of Type I and II collagens only in 3D culture. Chondroitin sulfate and keratan sulfate were abundant in both monolayer and 3D cultures. 2 Cells showed significantly greater proliferation in monolayer in the presence of platelet-derived growth factor compared to cells in 3D. 3 Cells on Matrigel™-coated monolayer substrates became rounded and formed nodular colonies, a finding absent during monolayer growth. Conclusions The cell's in vivo interactions with the ECM can regulate shape, gene expression and other cell functions. The shape of the annulus cell changes markedly during life: the young, healthy disc contains spindle shaped cells and abundant collagen. With aging and degeneration, many cells assume a strikingly different appearance, become rounded and are surrounded by unusual accumulations of ECM products. In vitro manipulation of disc cells provides an experimental window for testing how disc cells from given individuals respond when they are grown in environments which direct cells to have either spindle- or rounded-shapes. In vitro assessment of the response of such cells to platelet-derived growth factor and to Matrigel™ showed a continued influence of cell shape even in the presence of a growth factor stimulus. These findings contribute new information to the important issue of the influence of cell shape on cell behavior.

  4. Characterization of the microbial acid mine drainage microbial community using culturing and direct sequencing techniques.

    Science.gov (United States)

    Auld, Ryan R; Myre, Maxine; Mykytczuk, Nadia C S; Leduc, Leo G; Merritt, Thomas J S

    2013-05-01

    We characterized the bacterial community from an AMD tailings pond using both classical culturing and modern direct sequencing techniques and compared the two methods. Acid mine drainage (AMD) is produced by the environmental and microbial oxidation of minerals dissolved from mining waste. Surprisingly, we know little about the microbial communities associated with AMD, despite the fundamental ecological roles of these organisms and large-scale economic impact of these waste sites. AMD microbial communities have classically been characterized by laboratory culturing-based techniques and more recently by direct sequencing of marker gene sequences, primarily the 16S rRNA gene. In our comparison of the techniques, we find that their results are complementary, overall indicating very similar community structure with similar dominant species, but with each method identifying some species that were missed by the other. We were able to culture the majority of species that our direct sequencing results indicated were present, primarily species within the Acidithiobacillus and Acidiphilium genera, although estimates of relative species abundance were only obtained from direct sequencing. Interestingly, our culture-based methods recovered four species that had been overlooked from our sequencing results because of the rarity of the marker gene sequences, likely members of the rare biosphere. Further, direct sequencing indicated that a single genus, completely missed in our culture-based study, Legionella, was a dominant member of the microbial community. Our results suggest that while either method does a reasonable job of identifying the dominant members of the AMD microbial community, together the methods combine to give a more complete picture of the true diversity of this environment. PMID:23485423

  5. Hetero-cellular prototyping by synchronized multi-material bioprinting for rotary cell culture system.

    Science.gov (United States)

    Snyder, Jessica; Son, Ae Rin; Hamid, Qudus; Wu, Honglu; Sun, Wei

    2016-03-01

    Bottom-up tissue engineering requires methodological progress of biofabrication to capture key design facets of anatomical arrangements across micro, meso and macro-scales. The diffusive mass transfer properties necessary to elicit stability and functionality require hetero-typic contact, cell-to-cell signaling and uniform nutrient diffusion. Bioprinting techniques successfully build mathematically defined porous architecture to diminish resistance to mass transfer. Current limitations of bioprinted cell assemblies include poor micro-scale formability of cell-laden soft gels and asymmetrical macro-scale diffusion through 3D volumes. The objective of this work is to engineer a synchronized multi-material bioprinter (SMMB) system which improves the resolution and expands the capability of existing bioprinting systems by packaging multiple cell types in heterotypic arrays prior to deposition. This unit cell approach to arranging multiple cell-laden solutions is integrated with a motion system to print heterogeneous filaments as tissue engineered scaffolds and nanoliter droplets. The set of SMMB process parameters control the geometric arrangement of the combined flow's internal features and constituent material's volume fractions. SMMB printed hepatocyte-endothelial laden 200 nl droplets are cultured in a rotary cell culture system (RCCS) to study the effect of microgravity on an in vitro model of the human hepatic lobule. RCCS conditioning for 48 h increased hepatocyte cytoplasm diameter 2 μm, increased metabolic rate, and decreased drug half-life. SMMB hetero-cellular models present a 10-fold increase in metabolic rate, compared to SMMB mono-culture models. Improved bioprinting resolution due to process control of cell-laden matrix packaging as well as nanoliter droplet printing capability identify SMMB as a viable technique to improve in vitro model efficacy. PMID:26759993

  6. Unique cell culture systems for ground based research

    Science.gov (United States)

    Lewis, Marian L.

    1990-01-01

    The horizontally rotating fluid-filled, membrane oxygenated bioreactors developed at NASA Johnson for spacecraft applications provide a powerful tool for ground-based research. Three-dimensional aggregates formed by cells cultured on microcarrier beads are useful for study of cell-cell interactions and tissue development. By comparing electron micrographs of plant seedlings germinated during Shuttle flight 61-C and in an earth-based rotating bioreactor it is shown that some effects of microgravity are mimicked. Bioreactors used in the UAH Bioreactor Laboratory will make it possible to determine some of the effects of altered gravity at the cellular level. Bioreactors can be valuable for performing critical, preliminary-to-spaceflight experiments as well as medical investigations such as in vitro tumor cell growth and chemotherapeutic drug response; the enrichment of stem cells from bone marrow; and the effect of altered gravity on bone and muscle cell growth and function and immune response depression.

  7. Milk stimulates growth of prostate cancer cells in culture.

    Science.gov (United States)

    Tate, Patricia L; Bibb, Robert; Larcom, Lyndon L

    2011-11-01

    Concern has been expressed about the fact that cows' milk contains estrogens and could stimulate the growth of hormone-sensitive tumors. In this study, organic cows' milk and two commercial substitutes were digested in vitro and tested for their effects on the growth of cultures of prostate and breast cancer cells. Cows' milk stimulated the growth of LNCaP prostate cancer cells in each of 14 separate experiments, producing an average increase in growth rate of over 30%. In contrast, almond milk suppressed the growth of these cells by over 30%. Neither cows' milk nor almond milk affected the growth of MCF-7 breast cancer cells or AsPC-1 pancreatic cancer cells significantly. Soy milk increased the growth rate of the breast cancer cells. These data indicate that prostate and breast cancer patients should be cautioned about the possible promotional effects of commercial dairy products and their substitutes. PMID:22043817

  8. Microspectroscopic investigation of the membrane clogging during the sterile filtration of the growth media for mammalian cell culture.

    Science.gov (United States)

    Cao, Xiaolin; Loussaert, James A; Wen, Zai-qing

    2016-02-01

    Growth media for mammalian cell culture are very complex mixtures of several dozens of ingredients, and thus the preparation of qualified media is critical to viable cell density and final product titers. For liquid media prepared from powdered ingredients, sterile filtration is required prior to use to safeguard the cell culture process. Recently one batch of our prepared media failed to pass through the sterile filtration due to the membrane clogging. In this study, we report the root cause analysis of the failed sterile filtration based on the investigations of both the fouling media and the clogged membranes with multiple microspectroscopic techniques. Cellular particles or fragments were identified in the fouling media and on the surfaces of the clogged membranes, which were presumably introduced to the media from the bacterial contamination. This study demonstrated that microspectroscopic techniques may be used to rapidly identify both microbial particles and inorganic precipitates in the cell culture media. PMID:26637950

  9. Lethal impacts of cigarette smoke in cultured tobacco cells

    Directory of Open Access Journals (Sweden)

    Kawano Tomonori

    2011-07-01

    Full Text Available Abstract Background In order to understand and generalize the toxic mechanism of cigarette smoke in living cells, comparison of the data between animal systems and other biological system such as microbial and plant systems is highly beneficial. Objective By employing the tobacco cells as model materials for cigarette smoke toxicity assay, the impacts of the combustion by-products such as nitrogen oxides could be highlighted as the toxic impacts of the plant-derived endogenous chemicals could be excluded in the plant cells. Methods Cigarette smoke-induced cell death was assessed in tobacco cell suspension cultures in the presence and absence of pharmacological inhibitors. Results Cigarette smoke was effective in induction of cell death. The smoke-induced cell death could be partially prevented by addition of nitric oxide (NO scavenger, suggesting the role for NO as the cell death mediator. Addition of NO donor to tobacco cells also resulted in development of partial cell death further confirming the role of NO as cell death mediator. Members of reactive oxygen species and calcium ion were shown to be protecting the cells from the toxic action of smoke-derived NO.

  10. Improved Cell Culture Method for Growing Contracting Skeletal Muscle Models

    Science.gov (United States)

    Marquette, Michele L.; Sognier, Marguerite A.

    2013-01-01

    An improved method for culturing immature muscle cells (myoblasts) into a mature skeletal muscle overcomes some of the notable limitations of prior culture methods. The development of the method is a major advance in tissue engineering in that, for the first time, a cell-based model spontaneously fuses and differentiates into masses of highly aligned, contracting myotubes. This method enables (1) the construction of improved two-dimensional (monolayer) skeletal muscle test beds; (2) development of contracting three-dimensional tissue models; and (3) improved transplantable tissues for biomedical and regenerative medicine applications. With adaptation, this method also offers potential application for production of other tissue types (i.e., bone and cardiac) from corresponding precursor cells.

  11. Stimulating the production of homoisoflavonoids in cell suspension cultures of Caesalpinia pulcherrima using cork tissue.

    Science.gov (United States)

    Zhao, Ping; Iwamoto, Yuko; Kouno, Isao; Egami, Yasukuni; Yamamoto, Hirobumi

    2004-09-01

    It has previously been demonstrated that cork tissue increases the efficiency of the production of lipophilic secondary metabolites in diverse plant cell suspension cultures. In the present study, three new homoisoflavonoids--named dihydrobonducellin, 2'-methoxydihydrobonducellin, and 2'-methoxybonducellin--and bonducellin and isobonducellin were isolated from Caesalpinia pulcherrima cultured cells coincubated with cork tissue. Cork tissue increased the production of 2'-methoxybonducellin by about 7-fold relative to control cells, and more than 80% of the product was recoverable from the cork tissue. When cork tissue and methyl jasmonate or yeast extract were added simultaneously to the medium, the amount of 2'-methoxybonducellin produced increased further. The production of the other four homoisoflavonoids was enhanced by variable amounts. Our results indicate that the addition of cork tissue would be an effective technique for investigating formation of secondary metabolites that usually accumulate only in trace amounts. PMID:15381409

  12. Molecular technique identifies the pathogen responsible for culture negative infective endocarditis

    OpenAIRE

    SHIN, G. Y.; Manuel, R J; Ghori, S; Brecker, S; Breathnach, A. S.

    2005-01-01

    A case of culture negative endocarditis complicated by immune complex glomerulonephritis and severe aortic regurgitation necessitated aortic valve replacement. Empirical treatment with penicillin and gentamicin according to UK guidelines was started. The pathogen, Streptococcus sanguis, was later identified by polymerase chain reaction amplification and sequencing of bacterial 16S ribosomal RNA. This molecular technique is likely to be of increasing importance in determining the aetiology of ...

  13. Growing Three-Dimensional Cartilage-Cell Cultures

    Science.gov (United States)

    Spaulding, Glenn F.; Prewett, Tacey L.; Goodwin, Thomas J.

    1995-01-01

    Process for growing three-dimensional cultures of mammalian cartilage from normal mammalian cells devised. Effected using horizontal rotating bioreactor described in companion article, "Simplified Bioreactor for Growing Mammalian Cells" (MSC-22060). Bioreactor provides quiescent environment with generous supplies of nutrient and oxygen. Initiated with noncartilage cells. Artificially grown tissue resembles that in mammalian cartilage. Potential use in developing therapies for damage to cartilage by joint and back injuries and by such inflammatory diseases as arthritis and temporal-mandibular joint disease. Also used to test nonsteroid anti-inflammation medicines.

  14. Suspension Culture Alters Insulin Secretion in Induced Human Umbilical Cord Matrix-Derived Mesenchymal Cells

    Directory of Open Access Journals (Sweden)

    Fatemeh Seyedi

    2016-04-01

    Full Text Available Objective: Worldwide, diabetes mellitus (DM is an ever-increasing metabolic disorder. A promising approach to the treatment of DM is the implantation of insulin producing cells (IPC that have been derived from various stem cells. Culture conditions play a pivotal role in the quality and quantity of the differentiated cells. In this experimental study, we have applied various culture conditions to differentiate human umbilical cord matrix-derived mesenchymal cells (hUCMs into IPCs and measured insulin production. Materials and Methods: In this experimental study, we exposed hUCMs cells to pancreatic medium and differentiated them into IPCs in monolayer and suspension cultures. Pancreatic medium consisted of serum-free Dulbecco’s modified eagle’s medium Nutrient mixture F12 (DMEM/F12 medium with 17.5 mM glucose supplemented by 10 mM nicotinamide, 10 nM exendin-4, 10 nM pentagastrin, 100 pM hepatocyte growth factor, and B-27 serum-free supplement. After differentiation, insulin content was analyzed by gene expression, immunocytochemistry (IHC and the chemiluminesence immunoassay (CLIA. Results: Reverse transcription-polymerase chain reaction (RT-PCR showed efficient expressions of NKX2.2, PDX1 and INSULIN genes in both groups. IHC analysis showed higher expression of insulin protein in the hanging drop group, and CLIA revealed a significant higher insulin production in hanging drops compared with the monolayer group following the glucose challenge test. Conclusion: We showed by this novel, simple technique that the suspension culture played an important role in differentiation of hUCMs into IPC. This culture was more efficient than the conventional culture method commonly used in IPC differentiation and cultivation.

  15. Basis for Cumulative Cultural Evolution in Chimpanzees: Social Learning of a More Efficient Tool-Use Technique

    OpenAIRE

    Yamamoto, Shinya; Humle, Tatyana; TANAKA, MASAYUKI

    2013-01-01

    Background: The evidence for culture in non-human animals has been growing incrementally over the past two decades. However, the ability for cumulative cultural evolution, with successive generations building on earlier achievements, in non-human animals remains debated. Faithful social learning of incremental improvements in technique is considered to be a defining feature of human culture, differentiating human from non-human cultures. This study presents the first experimental evidence for...

  16. Basis for Cumulative Cultural Evolution in Chimpanzees: Social Learning of a More Efficient Tool-Use Technique

    OpenAIRE

    Shinya Yamamoto; Tatyana Humle; Masayuki Tanaka

    2013-01-01

    BACKGROUND: The evidence for culture in non-human animals has been growing incrementally over the past two decades. However, the ability for cumulative cultural evolution, with successive generations building on earlier achievements, in non-human animals remains debated. Faithful social learning of incremental improvements in technique is considered to be a defining feature of human culture, differentiating human from non-human cultures. This study presents the first experimental evidence for...

  17. Effects of Shuanghuangbu on the total protein content and ultrastructure in cultured human periodontal ligament cells

    Institute of Scientific and Technical Information of China (English)

    许彦枝; 邹慧儒; 王小玲; 刘世正; 王永军

    2004-01-01

    Background Successful periodontal regeneration depends on the migration, proliferation and differentiation of periodontal ligament cells in periodontal defects. The total protein content and the ultrastructure demonstrate the metabolizability and activity of periodontal ligament cells. This study was conducted to observe the effects of Shuanghuangbu, a mixture of medicinal herbs, on the total protein content and the ultrastructure of human periodontal ligament cells.Methods Periodontal ligament cells were grown to confluence and then cultured in Dulbecco's modified eagle medium (DMEM) supplemented with Shuanghuangbu over the concentration range of 0 to 1000 μg/ml. The total protein content in cultured cells was determined by using Coommasie brilliant blue technique. Periodontal ligament cells were incubated in 0 and 100 μg/ml Shuanghuangbu decoction for 5 days, then observed through transmission electron microscope.Results The total protein content of human periodontal ligament cells increased in each experiment group added 10-1000 μg/ml Shuanghuangbu respectively, and the effect of 100 μg/ml was excellent. Under the transmission electron microscope, there were more rough endoplasmic reticulums and mitochodrias in the experiment group than those in the control group. Conclusion Shuanghuangbu stimulates the protein synthesis of human periodontal ligament cells and improves human periodontal ligament cells' metabolizability and activity.

  18. Human neuronal cells in culture: from concepts to basic methodology.

    Science.gov (United States)

    Silani, V; Pizzuti, A; Donato, M F; Falini, A; Bassani, R; Strada, O; Causarano, R I; Mariani, D; Villani, R M; Scarlato, G

    1990-01-01

    The paper reviews some conceptual and methodological aspects of the tissue culture models which, during the past three decades, demonstrated a remarkable mimicry of many important structures and functions of the mammalian Central Nervous System (CNS) and related peripheral sensory and motor elements. Emphasis is placed on an original human neuronal tissue culture model obtained from selective CNS areas. The different cell types were identified and the neurotrophic interactions preliminary characterized. Neuropathological findings suggest hypothesis that can be fully tested using in vitro human models of affected cerebral specific areas. PMID:2102114

  19. De novo identification of viral pathogens from cell culture hologenomes

    Directory of Open Access Journals (Sweden)

    Patowary Ashok

    2012-01-01

    Full Text Available Abstract Background Fast, specific identification and surveillance of pathogens is the cornerstone of any outbreak response system, especially in the case of emerging infectious diseases and viral epidemics. This process is generally tedious and time-consuming thus making it ineffective in traditional settings. The added complexity in these situations is the non-availability of pure isolates of pathogens as they are present as mixed genomes or hologenomes. Next-generation sequencing approaches offer an attractive solution in this scenario as it provides adequate depth of sequencing at fast and affordable costs, apart from making it possible to decipher complex interactions between genomes at a scale that was not possible before. The widespread application of next-generation sequencing in this field has been limited by the non-availability of an efficient computational pipeline to systematically analyze data to delineate pathogen genomes from mixed population of genomes or hologenomes. Findings We applied next-generation sequencing on a sample containing mixed population of genomes from an epidemic with appropriate processing and enrichment. The data was analyzed using an extensive computational pipeline involving mapping to reference genome sets and de-novo assembly. In depth analysis of the data generated revealed the presence of sequences corresponding to Japanese encephalitis virus. The genome of the virus was also independently de-novo assembled. The presence of the virus was in addition, verified using standard molecular biology techniques. Conclusions Our approach can accurately identify causative pathogens from cell culture hologenome samples containing mixed population of genomes and in principle can be applied to patient hologenome samples without any background information. This methodology could be widely applied to identify and isolate pathogen genomes and understand their genomic variability during outbreaks.

  20. Long term maintenance of myeloid leukemic stem cells cultured with unrelated human mesenchymal stromal cells

    Directory of Open Access Journals (Sweden)

    Sawa Ito

    2015-01-01

    Full Text Available Mesenchymal stromal cells (MSCs support the growth and differentiation of normal hematopoietic stem cells (HSCs. Here we studied the ability of MSCs to support the growth and survival of leukemic stem cells (LSCs in vitro. Primary leukemic blasts isolated from the peripheral blood of 8 patients with acute myeloid leukemia (AML were co-cultured with equal numbers of irradiated MSCs derived from unrelated donor bone marrow, with or without cytokines for up to 6 weeks. Four samples showed CD34+CD38− predominance, and four were predominantly CD34+CD38+. CD34+ CD38− predominant leukemia cells maintained the CD34+ CD38− phenotype and were viable for 6 weeks when co-cultured with MSCs compared to co-cultures with cytokines or medium only, which showed rapid differentiation and loss of the LSC phenotype. In contrast, CD34+ CD38+ predominant leukemic cells maintained the CD34+CD38+ phenotype when co-cultured with MSCs alone, but no culture conditions supported survival beyond 4 weeks. Cell cycle analysis showed that MSCs maintained a higher proportion of CD34+ blasts in G0 than leukemic cells cultured with cytokines. AML blasts maintained in culture with MSCs for up to 6 weeks engrafted NSG mice with the same efficiency as their non-cultured counterparts, and the original karyotype persisted after co-culture. Chemosensitivity and transwell assays suggest that MSCs provide pro-survival benefits to leukemic blasts through cell–cell contact. We conclude that MSCs support long-term maintenance of LSCs in vitro. This simple and inexpensive approach will facilitate basic investigation of LSCs and enable screening of novel therapeutic agents targeting LSCs.

  1. Raman spectroscopy:an evolving technique for live cell studies

    OpenAIRE

    Smith, Rachael; Wright, Karen Leslie; Ashton, Lorna

    2016-01-01

    One of the most exciting developments in Raman spectroscopy in the last decade has been its application to cells and tissues for diagnostic and pharmaceutical applications, and in particular its use in the analysis of cellular dynamics. Raman spectroscopy is rapidly advancing as a cell imaging method that overcomes many of the limitations of current techniques and is earning its place as a routine tool in cell biology. In this review we focus on important developments in Raman spectroscopy th...

  2. Keratocytes Derived from Spheroid Culture of Corneal Stromal Cells Resemble Tissue Resident Keratocytes

    OpenAIRE

    Byun, Yong-Soo; Tibrewal, Sapna; Kim, Eunjae; Yco, Lisette; Sarkar, Joy; Ivanir, Yair; Liu, Chia-Yang; Sano, Cecile M.; Jain, Sandeep

    2014-01-01

    Purpose Corneal stromal cells transform to precursor cells in spheroid culture. We determined whether keratocytes derived from spheroid culture of murine corneal stromal cells resemble tissue resident keratocytes. Methods Spheroid culture was performed by seeding dissociated stromal cells onto ultra-low attachment plates containing serum-free mesenchymal stem cell culture medium. Spheroids were characterized with phenotype specific markers and stemness transcription factor genes. Spheroids an...

  3. Characterizing parameters of Jatropha curcas cell cultures for microgravity studies

    Science.gov (United States)

    Vendrame, Wagner A.; Pinares, Ania

    2013-06-01

    Jatropha (Jatropha curcas) is a tropical perennial species identified as a potential biofuel crop. The oil is of excellent quality and it has been successfully tested as biodiesel and in jet fuel mixes. However, studies on breeding and genetic improvement of jatropha are limited. Space offers a unique environment for experiments aiming at the assessment of mutations and differential gene expression of crops and in vitro cultures of plants are convenient for studies of genetic variation as affected by microgravity. However, before microgravity studies can be successfully performed, pre-flight experiments are necessary to characterize plant material and validate flight hardware environmental conditions. Such preliminary studies set the ground for subsequent spaceflight experiments. The objectives of this study were to compare the in vitro growth of cultures from three explant sources (cotyledon, leaf, and stem sections) of three jatropha accessions (Brazil, India, and Tanzania) outside and inside the petriGAP, a modified group activation pack (GAP) flight hardware to fit petri dishes. In vitro jatropha cell cultures were established in petri dishes containing a modified MS medium and maintained in a plant growth chamber at 25 ± 2 °C in the dark. Parameters evaluated were surface area of the explant tissue (A), fresh weight (FW), and dry weight (DW) for a period of 12 weeks. Growth was observed for cultures from all accessions at week 12, including subsequent plantlet regeneration. For all accessions differences in A, FW and DW were observed for inside vs. outside the PetriGAPs. Growth parameters were affected by accession (genotype), explant type, and environment. The type of explant influenced the type of cell growth and subsequent plantlet regeneration capacity. However, overall cell growth showed no abnormalities. The present study demonstrated that jatropha in vitro cell cultures are suitable for growth inside PetriGAPs for a period of 12 weeks. The parameters

  4. Isolation and culture of protoplasts of Ma-phut (Garcinia dulcis derived from cell suspension culture

    Directory of Open Access Journals (Sweden)

    Sompong Te-chato

    2008-09-01

    Full Text Available Friable callus induced from young leaves of Ma-phut on Murashige and Skoog (MS medium containing 3% sucrose,1 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D, 0.5 mg/l benzyladenine (BA and 500 mg/l polyvinylpyrrolidone (PVP, was cultured in liquid medium with the same components. Various ages of cell suspension at weekly intervals were then incubated in various kinds and concentrations of cell wall digestion enzymes combined with 1% macerozyme R-10 on a rotary shaker at 100 rpm under 1500 lux illumination at 26±4oC. Purified protoplasts were cultured at various densities in MS medium (adjusted osmoticum to 0.4 M by mannitol supplemented with 3% sucrose and two types of auxin, 2,4-D and NAA at four concentrations (1, 2, 3 and 4 mg/l together with 1 mg/l BA. The results revealed that a four-day old cell suspension culture incubated in 2% cellulase Onozuka R-10 (CR10 in combination with 1% macerozyme R-10 gave an optimum result in both yield and viability of protoplasts at 5.7x106/1 ml PCV and 80%, respectively. Embedding protoplasts at a density of 2.5x105/ml in 0.2% phytagel containing MS medium supplemented with 3 mg/l NAA and 1 mg/l BA promoted the most effective division of the protoplasts (20%. The first division of the protoplasts was obtained after 2 days of culture and further divisions to form micro- and macro-colonies could be observed after 7-10 days of culture. However, callusformation and plantlet regeneration was not obtained.

  5. Examination, characterisation and analysis techniques for the knowledge and the conservation / restoration of cultural heritage - importance of ionising radiation techniques

    International Nuclear Information System (INIS)

    For the examination, characterisation and analysis of cultural heritage artefacts or art objects and their component materials, the conservation scientist needs a palette of non destructive and non invasive techniques, in order to improve our knowledge concerning their elaboration, their evolution and/or degradation during time, and to give rational basis for their restoration and conservation. A general survey and illustrations showing the usefulness of these techniques will be presented. Among these methods, many are based on the use of ionising radiation. 1. Radiography (using X-rays, gamma rays, beta particles, secondary electrons, neutrons), electron emission radiography, tomodensimetry, 2. Scanning electron microscope associated with X-ray spectrometry, 3. X-ray diffraction, 4. Synchrotron radiation characterisation, 5. X-ray fluorescence analysis, 6. Activation analysis, 7. Ion beam analysis (PIXE, PIGE, RBS, secondary X-ray fluorescence), 8. Thermoluminescence dating, 9. Carbon-14 dating. These methods are used alone or in connection with other analytical methods. Any kind of materials can be encountered, for instance: i. stones, gems, ceramics, terracotta, enamels, glasses, i i. wood, paper, textile, bone, ivory, i i i. metals, jewellery, i v. paint layers, canvas and wooden backings, pigments, dyers, oils, binding media, varnishes, glues. Some examples will be taken, among recent work done at the Centre of Research and Restoration of the Museums of France (C2RMF), from various geographical origins, various ages and different art disciplines. This will illustrate the kind of assistance that science and technology can provide to a better knowledge of mankind's cultural heritage and also to the establishment of rational basis for its better conservation for the future generations. (Author)

  6. MEMS-based dynamic cell-to-cell culture platforms using electrochemical surface modifications

    International Nuclear Information System (INIS)

    MEMS-based biological platforms with the capability of both spatial placements and time releases of living cells for cell-to-cell culture experiments have been designed and demonstrated utilizing electrochemical surface modification effects. The spatial placement is accomplished by electrochemical surface modification of substrate surfaces to be either adhesive or non-adhesive for living cells. The time control is achieved by the electrical activation of the selective indium tin oxide co-culture electrode to allow the migration of living cells onto the electrode to start the cell-to-cell culture studies. Prototype devices have a three-electrode design with an electrode size of 50 × 50 µm2 and the separation gaps of 2 µm between them. An electrical voltage of −1.5 V has been used to activate the electrodes independently and sequentially to demonstrate the dynamic cell-to-cell culture experiments of NIH 3T3 fibroblast and Madin Darby canine kidney cells. As such, this MEMS platform could be a basic yet versatile tool to characterize transient cell-to-cell interactions

  7. The major bovine mastitis pathogens have different cell tropisms in cultures of bovine mammary gland cells

    NARCIS (Netherlands)

    Lammers, A.; Vorstenbosch, van C.J.; Erkens, J.H.F.; Smith, H.E.

    2001-01-01

    We previously showed that Staphylococcus aureus cells adhered mainly to an elongated cell type, present in cultures of bovine mammary gland cells. Moreover. we showed that this adhesion was mediated by binding to fibronectin. The same in vitro model was used here, to study adhesion of other importan

  8. Isolation and culture of porcine neural progenitor cells from embryos and pluripotent stem cells

    DEFF Research Database (Denmark)

    Rasmussen, Mikkel Aabech; Hall, Vanessa Jane; Hyttel, Poul

    2013-01-01

    The isolation and culture of neural progenitor cells (NPCs) from pluripotent stem cells has facilitated in vitro mechanistic studies of diseases related to the nervous system, as well as discovery of new medicine. In addition, NPCs are envisioned to play a crucial role in future cell replacement...

  9. Arsenite maintains germinative state in cultured human epidermal cells

    International Nuclear Information System (INIS)

    Arsenic is a well-known carcinogen for human skin, but its mechanism of action and proximal macromolecular targets remain to be elucidated. In the present study, low micromolar concentrations of sodium arsenite maintained the proliferative potential of epidermal keratinocytes, decreasing their exit from the germinative compartment under conditions that promote differentiation of untreated cells. This effect was observed in suspension and in post-confluent surface cultures as measured by colony-forming ability and by proportion of rapidly adhering colony-forming cells. Arsenite-treated cultures exhibited elevated levels of β1-integrin and β-catenin, two proteins enriched in cells with high proliferative potential. Levels of phosphorylated (inactive) glycogen synthase kinase 3β were higher in the treated cultures, likely accounting for the increased levels of transcriptionally available β-catenin. These findings suggest that arsenic could have co-carcinogenic and tumor co-promoting activities in the epidermis as a result of increasing the population and persistence of germinative cells targeted by tumor initiators and promoters. These findings also identify a critical signal transduction pathway meriting further exploration in pursuit of this phenomenon

  10. Culture and purification of human fetal olfactory bulb ensheathing cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective: To obtain high purity of human fetal olfactory bulb ensheathing cells (OB-hOECs) in vitro and to develop a simple and effective method for primary culture of OB-hOECs. Methods: OB-hOECs were cultured based on the differential rates of attachment of the various harvested cell types. Then the method was combined with arabinoside cytosine (Ara-C)inhibition, serum-free starvation or intermittent neurotrophin 3 (NT3) nutrition method to observe cell states in different cultural environments. The purity of OB-hOECs was assessed with immunocytochemical analysis. Results: OB-hOECs appeared bipolar and tripolar shape, with slender processes forming network. The purity of OECs reached 88% with the selective attachment method on day 6, and then fibroblast proliferated quickly and reduced the purity. When combined with the starvation method, the purity of OECs was 91% on day 6 and 86% on day 9, however, OECs were in a poor state. While combined with the NT3 method, the purity reached 95% on day 9 and 83% on day 12, respectively. The cells still remained in a good state. Conclusion: A combination of selective attachment and intermittent NT3 nutrition is an effective method to obtain OECs with higher purity and quality.

  11. Dexamethasone Modulation on Cultured Human Retinal Pigment Epithelial Cell

    Institute of Scientific and Technical Information of China (English)

    Bing Liu; Yannian; Hui Yusheng Wang; Hong Wei

    2001-01-01

    Purpose: Dexamethasone(DEX) was tested for its ability to modulate human retinal pigment epithelium (hRPE) cell proliferation in cell culture. Methods: DEX in different concentrations was added to cultured hRPE cells. The effects were measured with MTT method, 3H-thymidine(3H-TdR) incorporation and flow cytometw. Results: DEX increased survival rate and DNA synthesis from 32 mg/L to 320 mg/L under hRPE culture conditions, but paradoxically reduced them at 1 000 mg/L and 3 200 mg/L in dose and time dependent fashion by both MTT assay and 3 H-TdR incorporation. The cell numbers in S phase and G2/M phase increased 28.32 % at DEX concentration 320 mg/L, in contrast, reduced 41.84 % at 1 000 mg/L. Conclusion: DEX increased proliferation from 32 mg/L to 320 mg/L, and inhibited proliferation at concentrations greater than 320 mg/L. The inhibiting effect of DEX may happen in s phase and G2/M phase. Eye Sciernce 2001; 17:27 ~ 30.

  12. Enhancement effect of shikonin in cell suspension culture and transfermanant culture by radiation application

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jae Sung; Lee, Young Keun; Chung, Byung Yeoup; Lee, Young Bok; Hwang Hye Yeon

    2004-10-01

    The cell lines 679, 679-29 and 622-46 of L. erythrorhizon could be selected on LS agar medium for the production shikonin in cell suspension culture. The shikonin was increased moderately in suspension culture of cell line 622-46 in LS liquid medium containing BA 2 mg{center_dot}L{sup -1} and IAA 0.2 mg{center_dot}L{sup -1} in the dark, and was increased by adding 1 {mu}M Cu{sup 2+} and 100 {mu}M methyl jasmonate The accumulation of shikonin in the liquid medium was increased significantly by 2 Gy irradiation to callus of cell line 622-46 and culture in LS liquid medium containing BA 2 mg{center_dot}L{sup -1} and IAA 0.2 mg{center_dot}L{sup -1} in the dark and shikonin in cell debris was higher by 16 Gy irradiation. The activity of p-hydroxybenzoate geranyltransferase was increased by irradiation of 2 Gy and 16 Gy of {gamma} radiation. Seedling hypocotyles of L. erythrorhizon were infected with Agrogacterium rhizogenes strain 15834 harboring a binary vector with an intron bearing the GUS ({beta}-glucuronidase) gene driven by cauliflower mosaic virus (CaMV) 35S promotor as well as the HPT (hygromycin phosphotransferase) gene as the selection marker. Hairy roots isolated were hygromycin resistant and had integrated GUS gene in DNA. The root tip grown on M-9 medium showed normal pigment production pattern in border cells and root hairs.

  13. Rapid fabricating technique for multi-layered human hepatic cell sheets by forceful contraction of the fibroblast monolayer.

    Directory of Open Access Journals (Sweden)

    Yusuke Sakai

    Full Text Available Cell sheet engineering is attracting attention from investigators in various fields, from basic research scientists to clinicians focused on regenerative medicine. However, hepatocytes have a limited proliferation potential in vitro, and it generally takes a several days to form a sheet morphology and multi-layered sheets. We herein report our rapid and efficient technique for generating multi-layered human hepatic cell (HepaRG® cell sheets using pre-cultured fibroblast monolayers derived from human skin (TIG-118 cells as a feeder layer on a temperature-responsive culture dish. Multi-layered TIG-118/HepaRG cell sheets with a thick morphology were harvested on day 4 of culturing HepaRG cells by forceful contraction of the TIG-118 cells, and the resulting sheet could be easily handled. In addition, the human albumin and alpha 1-antitrypsin synthesis activities of TIG-118/HepaRG cells were approximately 1.2 and 1.3 times higher than those of HepaRG cells, respectively. Therefore, this technique is considered to be a promising modality for rapidly fabricating multi-layered human hepatocyte sheets from cells with limited proliferation potential, and the engineered cell sheet could be used for cell transplantation with highly specific functions.

  14. On-chip clearing of arrays of 3-D cell cultures and micro-tissues.

    Science.gov (United States)

    Grist, S M; Nasseri, S S; Poon, T; Roskelley, C; Cheung, K C

    2016-07-01

    Three-dimensional (3-D) cell cultures are beneficial models for mimicking the complexities of in vivo tissues, especially in tumour studies where transport limitations can complicate response to cancer drugs. 3-D optical microscopy techniques are less involved than traditional embedding and sectioning, but are impeded by optical scattering properties of the tissues. Confocal and even two-photon microscopy limit sample imaging to approximately 100-200 μm depth, which is insufficient to image hypoxic spheroid cores. Optical clearing methods have permitted high-depth imaging of tissues without physical sectioning, but they are difficult to implement for smaller 3-D cultures due to sample loss in solution exchange. In this work, we demonstrate a microfluidic platform for high-throughput on-chip optical clearing of breast cancer spheroids using the SeeDB, Clear(T2), and ScaleSQ clearing methods. Although all three methods are able to effectively clear the spheroids, we find that SeeDB and ScaleSQ more effectively clear the sample than Clear(T2); however, SeeDB induces green autofluorescence while ScaleS causes sample expansion. Our unique on-chip implementation permits clearing arrays of 3-D cultures using perfusion while monitoring the 3-D cultures throughout the process, enabling visualization of the clearing endpoint as well as monitoring of transient changes that could induce image artefacts. Our microfluidic device is compatible with on-chip 3-D cell culture, permitting the use of on-chip clearing at the endpoint after monitoring the same spheroids during their culture. This on-chip method has the potential to improve readout from 3-D cultures, facilitating their use in cell-based assays for high-content drug screening and other applications. PMID:27493703

  15. Radiation Response of Cultured Human Cells Is Unaffected by Johrei

    Directory of Open Access Journals (Sweden)

    Zach Hall

    2007-01-01

    Full Text Available Johrei has been credited with healing thousands from radiation wounds after the Hiroshima and Nagasaki bombs in 1945. This alternative medical therapy is becoming increasingly popular in the United States, as are other Energy Medicine modalities that purport to influence a universal healing energy. Human brain cells were cultured and exposed to increasing doses of ionizing radiation. Experienced Johrei practitioners directed healing intentionality toward the cells for 30 min from a distance of 20 cm and the fate of the cells was observed by computerized time-lapse microscopy. Cell death and cell divisions were tallied every 30 min before, during and after Johrei treatment for a total of 22.5 h. An equal number of control experiments were conducted in which cells were irradiated but did not receive Johrei treatment. Samples were assigned to treatment conditions randomly and data analysis was conducted in a blinded fashion. Radiation exposure decreased the rate of cell division (cell cycle arrest in a dose-dependent manner. Division rates were estimated for each 30 min and averaged over 8 independent experiments (4 control and 4 with Johrei treatment for each of 4 doses of X-rays (0, 2, 4 and 8 Gy. Because few cell deaths were observed, pooled data from the entire observation period were used to estimate death rates. Analysis of variance did not reveal any significant differences on division rate or death rate between treatment groups. Only radiation dose was statistically significant. We found no indication that the radiation response of cultured cells is affected by Johrei treatment.

  16. Radiation response of cultured human cells is unaffected by Johrei.

    Science.gov (United States)

    Hall, Zach; Luu, Tri; Moore, Dan; Yount, Garret

    2007-06-01

    Johrei has been credited with healing thousands from radiation wounds after the Hiroshima and Nagasaki bombs in 1945. This alternative medical therapy is becoming increasingly popular in the United States, as are other Energy Medicine modalities that purport to influence a universal healing energy. Human brain cells were cultured and exposed to increasing doses of ionizing radiation. Experienced Johrei practitioners directed healing intentionality toward the cells for 30 min from a distance of 20 cm and the fate of the cells was observed by computerized time-lapse microscopy. Cell death and cell divisions were tallied every 30 min before, during and after Johrei treatment for a total of 22.5 h. An equal number of control experiments were conducted in which cells were irradiated but did not receive Johrei treatment. Samples were assigned to treatment conditions randomly and data analysis was conducted in a blinded fashion. Radiation exposure decreased the rate of cell division (cell cycle arrest) in a dose-dependent manner. Division rates were estimated for each 30 min and averaged over 8 independent experiments (4 control and 4 with Johrei treatment) for each of 4 doses of X-rays (0, 2, 4 and 8 Gy). Because few cell deaths were observed, pooled data from the entire observation period were used to estimate death rates. Analysis of variance did not reveal any significant differences on division rate or death rate between treatment groups. Only radiation dose was statistically significant. We found no indication that the radiation response of cultured cells is affected by Johrei treatment. PMID:17549235

  17. A Place to Call Home: Bioengineering Pluripotential Stem Cell Cultures

    Directory of Open Access Journals (Sweden)

    Mark Weingarten

    2015-03-01

    Full Text Available Pluripotent stem cells (PSCs have the power to revolutionize the future of cell-based therapies and regenerative medicine. However, stem/progenitor cell use in the clinical arsenal has been hampered by discrepancies resulting from stem cell engineering and expansion, as well as in their (mass differentiation in culture. Moreover, the manner in which external conditions affect PSC and induced-pluripotent stem cell lineage establishment as well as maturation remains controversial. In this review, we examine novel methods of cell engineering and the role of reprogramming transcription factors in PSC development. In addition, we explore the effect of external environmental signals on PSC cultivation and differentiation by elucidating key components of the primordial stem cell microenvironment, the blastocyst. Furthermore, we assess the effects of hypoxic conditions on DNA editing, gene expression, and protein function in PSC self-renewal and growth. Finally, we speculate on the principal use of gap junction subunit expression as relevant biomarkers of PSC fate. Improving bioreactor design and pertinent cell biomarker classification could vastly enhance manufactured stem cell yield and quality, thereby increasing the potency and safety of therapeutic cells to be used in regenerative medicine.

  18. Sunlight-induced killing of nondividing human cells in culture

    International Nuclear Information System (INIS)

    Nondividing populations of human diploid fibroblasts that are DNA excision repair proficient and repair deficient were exposed to mid-day summer sunlight and their survival determined based on their ability to remain attached to a culture vessel surface. Whereas mid- and far-UV wavelengths and sunlamp radiation cause a gradual degeneration and detachment of cells in a dose-dependent manner, sunlight does not promote cell killing in repair proficient cells. Detachment of repair deficient cells is promoted to a limited extent but only at sunlight exposure times that are low with respect to the amount of DNA damage induced. Repair proficient and deficient cells exposed to sunlight for longer times do not detach. Pyrimidine dimer levels in these sunlight irradiated cells were great enough to have promoted detachment had these levels been induced by UV (254 nm) alone. Other photodamage induced by these exposures evidently inhibits the dimer induced cell degeneration that leads to cell detachment. It was concluded that pyrimidine dimers are responsible for cell killing at short sunlight exposure times (80 min) cells are killed by a different mechanism. (author)

  19. Long-term Cultured Human Neural Stem Cells Undergo Spontaneous Transformation to Tumor-Initiating Cells

    Directory of Open Access Journals (Sweden)

    Weihua Wu, Qihua He, Xiaoxia Li, Xiaoyan Zhang, Aili Lu, Ruimin Ge, HongYing Zhen, Alfred E. Chang, Qiao Li, Li Shen

    2011-01-01

    Full Text Available In this report, we describe the spontaneous malignant transformation of long-term cultured human fetal striatum neural stem cells (hsNSCs, passage 17. After subcutaneous transplantation of long-term cultured hsNSCs into immunodeficient nude mice, 2 out of 15 mice formed xenografts which expressed neuroendocrine tumor markers CgA and NSE. T1 cells, a cell line that we derived from one of the two subcutaneous xenografts, have undergone continuous expansion in vitro. These T1 cells showed stem cell-like features and expressed neural stem cell markers nestin and CD133. The T1 cells were involved in abnormal karyotype, genomic instability and fast proliferation. Importantly, after long-term in vitro culture, the T1 cells did not result in subcutaneous xenografts, but induced intracranial tumor formation, indicating that they adjusted themselves to the intracranial microenvironment. We further found that the T1 cells exhibited an overexpressed level of EGFR, and the CD133 positive T1 cells showed a truncation mutation in the exons 2-7 of the EGFR (EGFRvIII gene. These results suggest that continuous expansion of neural stem cells in culture may lead to malignant spontaneous transformation. This phenomenon may be functionally related to EGFR by EGFRvIII gene mutation.

  20. Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

    Directory of Open Access Journals (Sweden)

    Magnusson Magnus K

    2010-07-01

    Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

  1. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    International Nuclear Information System (INIS)

    Highlights: ► Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). ► Presence of SCs dramatically increased proliferation and migration of UCMSCs. ► Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of “nurse” cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  2. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Fenxi, E-mail: fxzhang0824@gmail.com [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Hong, Yan; Liang, Wenmei [Department of Histology and Embryology, Guiyang Medical University, Guizhou 550004, People' s Republic of China (China); Ren, Tongming [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Jing, Suhua [ICU Center, The Third Hospital of Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Lin, Juntang [Stem Cell Center, Xinxiang Medical University, Henan 453003, People' s Republic of China (China)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). Black-Right-Pointing-Pointer Presence of SCs dramatically increased proliferation and migration of UCMSCs. Black-Right-Pointing-Pointer Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of 'nurse' cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

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

    Directory of Open Access Journals (Sweden)

    Cuicui ZHANG

    2015-11-01

    Full Text Available Background and objective Unlike normal tissue-derived microvascular endothelial cells, tumor microvessel endothelial cells are highly reactive to growth factors and exhibit more adhesion molecules. Thus, vascular tumors are highly permeable and grow vigorously; this occurrence results in rapid growth and metastasis cancer cells. Therefore, understanding the characteristics of endothelial cells in the tumor microenvironment guides anti-angiogenic therapy. To this end, we explore the effect of the supernatant obtained from cultured Anip973 cells (high-metastatic human lung adenocarcinoma cells on the biological behavior and on the cell surface markers of the human umbilical vein endothelial cell (HUVEC. Methods The HUVEC that was cultured in a medium (RPMI-1640 + 10% fetal bovine serum containing various concentrations of Anip973 supernatants was categorized into experimental groups. The HUVEC cultured in a medium without Anip973 supernatants served as the control group. Proliferation was determined with CCK-8; blood vessel formation was investigated with three-dimensional culture techniques in vitro; and HUVEC migration was observed via transwell assay. At the same time, the expressions of CD105, CD31, and the apoptotic marker of Annexin V were detected through flow cytometry for analyzing the relationship between the expression of cell surface markers and biological behavior. Results Following incubation with the supernatant obtained from cultured Anip973 cells, HUVEC proliferated more than the control group did, and the proliferation rate was maximized when incubated in a supernatant concentration of 250 μL/mL for 24 h (P=0.002. In addition, the experimental groups exhibited varying degrees of migration and forms of vascular lumen sample structure, especially at supernatant concentrations of 125 µL/mL (P<0.001 and 250 µL/mL (P=0.002, respectively. CD105 expression was optimized at 250 μL/mL (P=0.028, and CD31 expression also increased with

  4. Calcium exchange, structure, and function in cultured adult myocardial cells

    International Nuclear Information System (INIS)

    Cells digested from adult rat heart and cultured for 14 days demonstrate all the structural elements, in mature form, associated with the process of excitation-contraction (EC) coupling. The transverse tubular (TT) system is well developed with an extensive junctional sarcoplasmic reticulum (JSR). In nonphosphate-containing buffer contraction of the cells is lost as rapidly as zero extracellular Ca concentration ([Ca]0) solution is applied and a negative contraction staircase is produced on increase of stimulation frequency. Structurally and functionally the cells have the characteristics of adult cells in situ. 45Ca exchange and total 45Ca measurement in N-2-hydroxyethylpiperazine N'-2-ethanesulfonic acid (HEPES)-buffered perfusate define three components of cellular Ca: 1) a rapidly exchangeable component accounting for 36% of total Ca, 2) a slowly exchangeable component (t/sub 1/2/ 53 min) accounting for 7% total Ca, and 3) the remaining 57% cellular Ca is inexchangeable (demonstrates no significant exchange within 60 min). The slowly exchangeable component can be increased 10-fold within 60 min by addition of phosphate to the perfusate. The Ca distribution and exchange characteristics are little different from those of 3-day cultures of neonatal rat heart previously studied. The results suggest that the cells are representative of adult cells in situ and that both sarcolemmal-bound and sarcoplasmic reticular Ca contribute to the component of Ca that is rapidly exchangeable

  5. Aquaporin-1 Expressed in Cultured Human Trabecular Meshwork Cells

    Institute of Scientific and Technical Information of China (English)

    Mingkai Lin; Jian Ge; Yehong Zhuo; Yuqing Lan; Keming Yu; Jianliang Zheng

    2002-01-01

    Objective:To determine if aquaporin-1 could be detected in cultures of human trabecularshwork cells. Methods: Using primers specific for aquaporin-1, reverse transcription combined withpolymerase chain reaction (RT-PCR) yielded a product and its size with total RNAprepared from the human trabecular meshwork cells. SDS-PAGE and immunoblottingwere also used in this study to detect the specific water channel.Results: The presence of this product and its size (298 base pairs) are consistent withthat of an aquaporin-1 message in these cells. A band of 28 kD in agreement with themolecular size of aquaporin-1 was showed in a film by immunoblotting.Conclusion: The presence of aquaporin-1 in human trabecular meshwork cells, thepredominant cell-type of the primary outflow region of the human eye, suggests that waterchannels may be involved in the movement of aqueous fluid out of the eye. In addition,the existence of aquaporin-1 on cultures of human trabecular meshwork cells provides anin vitro model to study the endogenous expression of aquaporin-1 and its possible role inthe regulation of aqueous outflow.

  6. Methyl Jasmonate Represses Growth and Affects Cell Cycle Progression in Cultured Taxus Cells

    OpenAIRE

    Patil, Rohan A.; Lenka, Sangram K.; Normanly, Jennifer; Walker, Elsbeth L.; Roberts, Susan C.

    2014-01-01

    Methyl jasmonate (MeJA) elicitation is an effective strategy to induce and enhance synthesis of the anticancer agent paclitaxel (Taxol®) in Taxus cell suspension cultures; however, concurrent decreases in growth are often observed, which is problematic for large scale bioprocessing. Here, increased accumulation of paclitaxel in Taxus cuspidata suspension cultures with MeJA elicitation was accompanied by a concomitant decrease in cell growth, evident within the first three days post-elicitatio...

  7. Eight-Shaped Hatching Increases the Risk of Inner Cell Mass Splitting in Extended Mouse Embryo Culture

    OpenAIRE

    Yan, Zheng; Liang, Hongxing; Deng, Li; Long, Hui; Chen, Hong; Chai, Weiran; Suo, Lun; Xu, Chen; Kuang, Yanping; Wu, Lingqian; Lu, Shengsheng; Lyu, Qifeng

    2015-01-01

    Increased risk of monozygotic twinning (MZT) has been shown to be associated with assisted reproduction techniques, particularly blastocyst culture. Interestingly, inner cell mass (ICM) splitting in human ‘8’-shaped hatching blastocysts that resulted in MZT was reported. However, the underlying cause of MZT is not known. In this study, we investigated in a mouse model whether in vitro culture leads to ICM splitting and its association with hatching types. Blastocyst hatching was observed in: ...

  8. Comprehensive analysis of signal transduction in three-dimensional ECM-based tumor cell cultures

    Directory of Open Access Journals (Sweden)

    Iris Eke

    2015-11-01

    Full Text Available Analysis of signal transduction and protein phosphorylation is fundamental to understand physiological and pathological cell behavior as well as identification of novel therapeutic targets. Despite the fact that more physiological three-dimensional cell culture assays are increasingly used, particularly proteomics and phosphoproteomics remain challenging due to easy, robust and reproducible sample preparation. Here, we present an easy-to-perform, reliable and time-efficient method for the production of 3D cell lysates without compromising cell adhesion before cell lysis. The samples can be used for Western blotting as well as phosphoproteome array technology. This technique would be of interest for researchers working in all fields of biology and drug development.

  9. Effect of Aflatoxin B1 on Growth of Bovine Mammary Epithelial Cells in 3D and Monolayer Culture System

    Directory of Open Access Journals (Sweden)

    Babak Qasemi-Panahi

    2013-02-01

    Full Text Available Purpose: Many studies have been showed transfer of aflatoxins, toxins produced by Aspergillus flvaus and Aspergillus parasiticus fungi, into milk. These toxins are transferred into the milk through digestive system by eating contaminated food. Due to the toxicity of these materials, it seems that it has side effects on the growth of mammary cells. Therefore, the present work aimed to investigate possible toxic effects of aflatoxin B1 (AFB1 on bovine mammary epithelial cells in monolayer and three-dimensional cultures. Methods: Specimens of the mammary tissue of bovine were sized out in size 2×2 cm in slaughterhouse. After disinfection and washing in sterile PBS, primary cell culture was performed by enzymatic digestion of tissue with collagenase. When proper numbers of cells were achieved in monolayer culture, cells were seeded in a 24-well culture plate for three-dimensional (3D culture in Matrigel matrix. After 21 days of 3D culture and reaching the required number of cells, the concentrations of 15, 25 and 35 μL of AFB1 were added to the culture in quadruplicate and incubated for 8 hours. Cellular cytotoxicity was examined using standard colorimetric assay and finally, any change in the morphology of the cells was studied by microscopic technique. Results: Microscopic investigations showed necrosis of the AFB1-exposed cells compared to the control cells. Also, bovine mammary epithelial cells were significantly affected by AFB1 in dose and time dependent manner in cell viability assays. Conclusion: According to the results, it seems that AFB1 can induce cytotoxicity and necrosis in bovine mammary epithelial cells.

  10. Effects of simulated microgravity on mouse Sertoli cells in culture

    Science.gov (United States)

    Angela, Masini Maria; Prato, Paola; Linda, Scarabelli; Lanza, Cristina; Palmero, Silvio; Pointis, Georges; Ricci, Franco; Strollo, Felice

    With the advent of space flights questions concerning the effects of microgravity (0xG) on hu-man reproduction physiology have got priority Spermatogenesis is a complex, highly ordered process of cell division and differentiation by which spermatogonial cells give rise to mature spermatozoa. Sertoli cells play a crucial role in the development of germ cells and the regulation of spermatogenesis. In this study the influence of 0xG on Sertoli cells was evaluated. A Sertoli cell line from mouse testis (42GPA9) was analyzed for cytoskeletal (using the 3D reconstruction generated from a stack of confocal images) and SHBG changes by immunohistochemistry, for antioxidant agents by RT-PCR and for culture medium lactate concentrations by wet chemistry. Cells were cultured for 6, 24 and 48 hrs on a three-dimensional Random Positioning Machine (3D-RPM); static controls (1xG) were positioned on the supporting frame. At the end of each experiment, cultured cells were either fixed in paraformaldehyde or RNA-extracted or used for culture medium lactate measurements as needed. At 0xG Sertoli cytoskeleton got disorganized, microtubules fragmented and SHBG undetectable already after 24 hrs, with alterations wors-ening further until 48 hrs; various antioxidant systems (SOD, GST, PARP, MTs) appreciably increased during the first 24 hrs but significantly decreased at 48 hrs. No changes occurred in 1xG samples. At least initially, 0xG seems to perturb antioxidant protection strategies allowing the testes to support sperm production, thus generating an aging-like state of oxidative stress. Lactate production at 0xG slightly decreased only after 24 hrs. Further experiments need to be carried out in space to investigate upon steroidogenesis and germ cell differentiation within the testis, to rule out eventually pending male infertility consequences, which would be a problem nowadays, when life expectancy increases and male fertility might become a social issue often extending into 60 years

  11. A modular suite of hardware enabling spaceflight cell culture research

    Science.gov (United States)

    Hoehn, Alexander; Klaus, David M.; Stodieck, Louis S.

    2004-01-01

    BioServe Space Technologies, a NASA Research Partnership Center (RPC), has developed and operated various middeck payloads launched on 23 shuttle missions since 1991 in support of commercial space biotechnology projects. Modular cell culture systems are contained within the Commercial Generic Bioprocessing Apparatus (CGBA) suite of flight-qualified hardware, compatible with Space Shuttle, SPACEHAB, Spacelab and International Space Station (ISS) EXPRESS Rack interfaces. As part of the CGBA family, the Isothermal Containment Module (ICM) incubator provides thermal control, data acquisition and experiment manipulation capabilities, including accelerometer launch detection for automated activation and thermal profiling for culture incubation and sample preservation. The ICM can accommodate up to 8 individually controlled temperature zones. Command and telemetry capabilities allow real-time downlink of data and video permitting remote payload operation and ground control synchronization. Individual cell culture experiments can be accommodated in a variety of devices ranging from 'microgravity test tubes' or standard 100 mm Petri dishes, to complex, fed-batch bioreactors with automated culture feeding, waste removal and multiple sample draws. Up to 3 levels of containment can be achieved for chemical fixative addition, and passive gas exchange can be provided through hydrophobic membranes. Many additional options exist for designing customized hardware depending on specific science requirements.

  12. Gene probes to detect cross-culture contamination in hormone producing cell lines

    DEFF Research Database (Denmark)

    Matsuba, I; Lernmark, A; Madsen, Ole Dragsbæk;

    1988-01-01

    Cross-culture contamination of cell lines propagated in continuous culture is a frequent event and particularly difficult to resolve in cells expressing similar phenotypes. We demonstrate that DNA-DNA hybridization to blotted endonuclease-digested cell DNA effectively detects cross-culture contam...... effective use of gene probes to control the origin of cell cultures.......Cross-culture contamination of cell lines propagated in continuous culture is a frequent event and particularly difficult to resolve in cells expressing similar phenotypes. We demonstrate that DNA-DNA hybridization to blotted endonuclease-digested cell DNA effectively detects cross......-culture contamination to monitor inter-species as well as intra-species cross contamination. An insulin-producing cell-line, Clone-16, originally cloned from a human fetal endocrine pancreatic cell line did not produce human c-peptide as anticipated. DNA from these cells showed no hybridization to the human ALU...

  13. Gill cell culture systems as models for aquatic environmental monitoring.

    Science.gov (United States)

    Bury, Nic R; Schnell, Sabine; Hogstrand, Christer

    2014-03-01

    A vast number of chemicals require environmental safety assessments for market authorisation. To ensure acceptable water quality, effluents and natural waters are monitored for their potential harmful effects. Tests for market authorisation and environmental monitoring usually involve the use of large numbers of organisms and, for ethical, cost and logistic reasons, there is a drive to develop alternative methods that can predict toxicity to fish without the need to expose any animals. There is therefore a great interest in the potential to use cultured fish cells in chemical toxicity testing. This review summarises the advances made in the area and focuses in particular on a system of cultured fish gill cells grown into an epithelium that permits direct treatment with water samples. PMID:24574380

  14. Long-term Cultured Human Neural Stem Cells Undergo Spontaneous Transformation to Tumor-Initiating Cells

    OpenAIRE

    Weihua Wu, Qihua He, Xiaoxia Li, Xiaoyan Zhang, Aili Lu, Ruimin Ge, HongYing Zhen, Alfred E. Chang, Qiao Li, Li Shen

    2011-01-01

    In this report, we describe the spontaneous malignant transformation of long-term cultured human fetal striatum neural stem cells (hsNSCs, passage 17). After subcutaneous transplantation of long-term cultured hsNSCs into immunodeficient nude mice, 2 out of 15 mice formed xenografts which expressed neuroendocrine tumor markers CgA and NSE. T1 cells, a cell line that we derived from one of the two subcutaneous xenografts, have undergone continuous expansion in vitro. These T1 cells showed stem ...

  15. Intracellular boron localization and uptake in cell cultures using imaging secondary ion mass spectrometry (ion microscopy) for neutron capture therapy for cancer.

    Science.gov (United States)

    Bennett, B D; Zha, X; Gay, I; Morrison, G H

    1992-01-01

    Quantitative ion microscopy of freeze-fractured, freeze-dried cultured cells is a technique for single cell and subcellular elemental analysis. This review describes the technique and its usefulness in determining the uptake and subcellular distribution of the boron from boron neutron capture therapy drugs. PMID:1511239

  16. Microfluidic synthesis of microfibers for magnetic-responsive controlled drug release and cell culture.

    Directory of Open Access Journals (Sweden)

    Yung-Sheng Lin

    Full Text Available This study demonstrated the fabrication of alginate microfibers using a modular microfluidic system for magnetic-responsive controlled drug release and cell culture. A novel two-dimensional fluid-focusing technique with multi-inlets and junctions was used to spatiotemporally control the continuous laminar flow of alginate solutions. The diameter of the manufactured microfibers, which ranged from 211 µm to 364 µm, could be well controlled by changing the flow rate of the continuous phase. While the model drug, diclofenac, was encapsulated into microfibers, the drug release profile exhibited the characteristic of a proper and steady release. Furthermore, the diclofenac release kinetics from the magnetic iron oxide-loaded microfibers could be controlled externally, allowing for a rapid drug release by applying a magnetic force. In addition, the successful culture of glioblastoma multiforme cells in the microfibers demonstrated a good structural integrity and environment to grow cells that could be applied in drug screening for targeting cancer cells. The proposed microfluidic system has the advantages of ease of fabrication, simplicity, and a fast and low-cost process that is capable of generating functional microfibers with the potential for biomedical applications, such as drug controlled release and cell culture.

  17. Ion-selective microelectrode arrays for cell culture monitoring

    OpenAIRE

    Generelli, Silvia; De Rooij, Nicolas-F.

    2008-01-01

    The design, microfabrication and characterization of a platform comprising an array of ion-selective microelectrodes (µISE) aimed at in vitro cellular physiology and toxicology is described. This study focusses on K+ and Ca2+ monitoring in cell culture environments. A potential promising application of such a platform is based on recent findings in molecular biology, revealing connections between certain diseases, as for example some types of cancer or parkinsonism, and a malfunction in cellu...

  18. Transparent, biocompatible nanostructured surfaces for cancer cell capture and culture

    Directory of Open Access Journals (Sweden)

    Cheng BR

    2014-05-01

    Full Text Available Boran Cheng,1,* Zhaobo He,2,* Libo Zhao,2,* Yuan Fang,1 Yuanyuan Chen,1 Rongxiang He,2 Fangfang Chen,1 Haibin Song,1 Yuliang Deng,2 Xingzhong Zhao,2 Bin Xiong1 1Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors, Hubei Cancer Clinical Study Center, Wuhan, Hubei, People’s Republic of China; 2Key Laboratory of Artificial Micro- and Nano-Structures of Ministry of Education, School of Physics and Technology, Wuhan University, Wuhan, Hubei, People’s Republic of China *These authors contributed equally to this work Abstract: Circulating tumor cells (CTCs in the blood which have detached from both the primary tumor and any metastases may be considered as a “liquid biopsy” and are expected to replace tumor biopsies in the monitoring of treatment response and determining patient prognosis. Here, we introduce a facile and efficient CTC detection material made of hydroxyapatite/chitosan (HA/CTS, which is beneficial because of its transparency and excellent biological compatibility. Atomic force microscopy images show that the roughness of the HA/CTS nanofilm (HA/CTSNF substrates can be controlled by changing the HA:CTS ratio. Enhanced local topographic interactions between nano-components on cancer cell membranes, and the antibody coated nanostructured substrate lead to improved CTC capture and separation. This remarkable nanostructured substrate has the potential for CTC culture in situ and merits further analysis. CTCs captured from artificial blood samples were observed in culture on HA/CTSNF substrates over a period of 14 days by using conventional staining methods (hematoxylin eosin and Wright’s stain. We conclude that these substrates are multifunctional materials capable of isolating and culturing CTCs for subsequent studies. Keywords: cell capture, cell culture, nanofilms, hydroxyapatite/chitosan

  19. Mammalian Cell Culture Process for Monoclonal Antibody Production: Nonlinear Modelling and Parameter Estimation

    Directory of Open Access Journals (Sweden)

    Dan Selişteanu

    2015-01-01

    Full Text Available Monoclonal antibodies (mAbs are at present one of the fastest growing products of pharmaceutical industry, with widespread applications in biochemistry, biology, and medicine. The operation of mAbs production processes is predominantly based on empirical knowledge, the improvements being achieved by using trial-and-error experiments and precedent practices. The nonlinearity of these processes and the absence of suitable instrumentation require an enhanced modelling effort and modern kinetic parameter estimation strategies. The present work is dedicated to nonlinear dynamic modelling and parameter estimation for a mammalian cell culture process used for mAb production. By using a dynamical model of such kind of processes, an optimization-based technique for estimation of kinetic parameters in the model of mammalian cell culture process is developed. The estimation is achieved as a result of minimizing an error function by a particle swarm optimization (PSO algorithm. The proposed estimation approach is analyzed in this work by using a particular model of mammalian cell culture, as a case study, but is generic for this class of bioprocesses. The presented case study shows that the proposed parameter estimation technique provides a more accurate simulation of the experimentally observed process behaviour than reported in previous studies.

  20. Mammalian cell culture process for monoclonal antibody production: nonlinear modelling and parameter estimation.

    Science.gov (United States)

    Selişteanu, Dan; Șendrescu, Dorin; Georgeanu, Vlad; Roman, Monica

    2015-01-01

    Monoclonal antibodies (mAbs) are at present one of the fastest growing products of pharmaceutical industry, with widespread applications in biochemistry, biology, and medicine. The operation of mAbs production processes is predominantly based on empirical knowledge, the improvements being achieved by using trial-and-error experiments and precedent practices. The nonlinearity of these processes and the absence of suitable instrumentation require an enhanced modelling effort and modern kinetic parameter estimation strategies. The present work is dedicated to nonlinear dynamic modelling and parameter estimation for a mammalian cell culture process used for mAb production. By using a dynamical model of such kind of processes, an optimization-based technique for estimation of kinetic parameters in the model of mammalian cell culture process is developed. The estimation is achieved as a result of minimizing an error function by a particle swarm optimization (PSO) algorithm. The proposed estimation approach is analyzed in this work by using a particular model of mammalian cell culture, as a case study, but is generic for this class of bioprocesses. The presented case study shows that the proposed parameter estimation technique provides a more accurate simulation of the experimentally observed process behaviour than reported in previous studies. PMID:25685797

  1. A Technique of Culturing the Olive Fly, Dacus Oleae Gmel., on Synthetic Media Under Xenic Conditions

    International Nuclear Information System (INIS)

    Five generations of Dacus oleae have been cultured on an agar-dehydrated carrot base medium that contains an enzymatic protein hydrolysate of soya or casein, brewer's yeast, choline chloride and olive oil. Although the culture technique is xenic, an attempt is made to control microorganisms by chemical means. A bacterial species replaced the normal symbiote within D. oleae and was believed to be essential to maintaining the stock, but two generations of D. oleae have been cultured without any bacteria being present in the sites normally occupied within the larva or adult by its typical symbiote. Streptomycin is now being incorporated into the adult food to control bacterial infection of the eggs. Normal larval development, size and reproduction of D. oleae is obtained. Mass culture is possible using the larval medium developed, but further research is necessary to find a faster method of placing the eggs on the medium. Further screening of mould inhibitors is desirable, as well as seeking cheaper substitutes for the medium. (author)

  2. Optimized exosome isolation protocol for cell culture supernatant and human plasma

    Directory of Open Access Journals (Sweden)

    Richard J. Lobb

    2015-07-01

    Full Text Available Extracellular vesicles represent a rich source of novel biomarkers in the diagnosis and prognosis of disease. However, there is currently limited information elucidating the most efficient methods for obtaining high yields of pure exosomes, a subset of extracellular vesicles, from cell culture supernatant and complex biological fluids such as plasma. To this end, we comprehensively characterize a variety of exosome isolation protocols for their efficiency, yield and purity of isolated exosomes. Repeated ultracentrifugation steps can reduce the quality of exosome preparations leading to lower exosome yield. We show that concentration of cell culture conditioned media using ultrafiltration devices results in increased vesicle isolation when compared to traditional ultracentrifugation protocols. However, our data on using conditioned media isolated from the Non-Small-Cell Lung Cancer (NSCLC SK-MES-1 cell line demonstrates that the choice of concentrating device can greatly impact the yield of isolated exosomes. We find that centrifuge-based concentrating methods are more appropriate than pressure-driven concentrating devices and allow the rapid isolation of exosomes from both NSCLC cell culture conditioned media and complex biological fluids. In fact to date, no protocol detailing exosome isolation utilizing current commercial methods from both cells and patient samples has been described. Utilizing tunable resistive pulse sensing and protein analysis, we provide a comparative analysis of 4 exosome isolation techniques, indicating their efficacy and preparation purity. Our results demonstrate that current precipitation protocols for the isolation of exosomes from cell culture conditioned media and plasma provide the least pure preparations of exosomes, whereas size exclusion isolation is comparable to density gradient purification of exosomes. We have identified current shortcomings in common extracellular vesicle isolation methods and provide a

  3. Autoradiographic investigations on cell shape-mediated growth regulation of lens epithelial cells in culture

    International Nuclear Information System (INIS)

    An autoradiographic method is described which is well suited for the determination of the labelling index in flattened as well as rounded cells. Using this method DNA synthesis of lens epithelial cells in culture was found to be dependent on cell attachment, cell flattening and intact microfilaments. Thus previous results on cell shape-mediated growth regulation could be confirmed. Moreover, considering the labelling index it was possible to conclude that cell rounding or a disintegration of microfilaments did not impair ongoing DNA synthesis but did prevent cells from entering the S-phase of the cycle. (author)

  4. Propagation and isolation of ranaviruses in cell culture

    DEFF Research Database (Denmark)

    Ariel, Ellen; Nicolajsen, Nicole; Christophersen, Maj-Britt;

    2009-01-01

    The optimal in vitro propagation procedure for a panel of ranavirus isolates and the best method for isolation of Epizootic haematopoietic necrosis virus (EHNV) from organ material in cell-culture were investigated. The panel of ranavirus isolates included: Frog virus 3 (FV3), Bohle iridovirus (BIV......), Pike-perch iridovirus (PPIV), European catfish virus (ECV), European sheatfish virus (ESV), EHNV, Doctor fish virus (DFV), Guppy virus 6 (GF6), short-finned eel virus (SERV) and Rana esculenta virus Italy 282/102 (REV 282/102). Each isolate was titrated in five cell lines: bluegill fry (BF-2...

  5. [Inhibition of adenovirus reproduction in cell culture by specific antibodies].

    Science.gov (United States)

    Povnytsia, O Iu; Nosach, L M; Zhovnovata, V L; Zahorodnia, S D; Vantsak, N P; Tokarchuk, L V; Polishchuk, O M; Diachenko, N S

    2009-01-01

    The capacity of specific antibodies to inhibit the reproduction of homo- and heterologous adenoviruses in Hela cell added to culture medium after virus adsorption was studied. The inhibiting effect of polyclonal antivirus and monospecific antihexone antibodies to homo- and heterologous adenoviruses was shown. The effect was more expressed when using antibodies to homologous antibodies. The intensity of inhibition depended on antibodies concentration in the medium and infecting dose of the virus. Essential reduction of the quantity of infected cells and a decrease of the titer of adenovirus synthesized in the presence of homo- and heterologous antibodies was shown but adenovirus reproduction was not inhibited completely. PMID:19663330

  6. In Vitro Cell Culture Infectivity Assay for Human Noroviruses

    Energy Technology Data Exchange (ETDEWEB)

    Straub, Tim M.; Honer Zu Bentrup, Kerstin A.; Orosz Coghlan, Patricia A.; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza; Nickerson, Cheryl A.

    2007-01-30

    Human noroviruses (NoV) cause severe, self-limiting gastroenteritis that typically lasts 24 - 48 hours. The true nature of NoV pathogenesis remains unknown due to the lack of suitable tissue culture or animal models. Here we show, for the first time, that NoV can infect and replicate in an organoid, three-dimensional (3-D) model of human small intestinal epithelium (INT-407). Cellular differentiation for this model was achieved by growing the cells in 3-D on porous collagen I-coated microcarrier beads under conditions of physiological fluid shear in rotating wall vessel bioreactors. Microscopy, PCR, and fluorescent in-situ hybridization were employed to provide evidence of NoV infection. CPE and norovirus RNA was detected at each of the five cell passages for both genogroup I and II viruses. Our results demonstrate that the highly differentiated 3-D cell culture model can support the natural growth of human noroviruses, whereas previous attempts using differentiated monolayer cultures failed.

  7. A Three-Dimensional Cell Culture Model To Study Enterovirus Infection of Polarized Intestinal Epithelial Cells.

    Science.gov (United States)

    Drummond, Coyne G; Nickerson, Cheryl A; Coyne, Carolyn B

    2016-01-01

    Despite serving as the primary entry portal for coxsackievirus B (CVB), little is known about CVB infection of the intestinal epithelium, owing at least in part to the lack of suitable in vivo models and the inability of cultured cells to recapitulate the complexity and structure associated with the gastrointestinal (GI) tract. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of Caco-2 cells to model CVB infection of the gastrointestinal epithelium. We show that Caco-2 cells grown in 3-D using the rotating wall vessel (RWV) bioreactor recapitulate many of the properties of the intestinal epithelium, including the formation of well-developed tight junctions, apical-basolateral polarity, brush borders, and multicellular complexity. In addition, transcriptome analyses using transcriptome sequencing (RNA-Seq) revealed the induction of a number of genes associated with intestinal epithelial differentiation and/or intestinal processes in vivo when Caco-2 cells were cultured in 3-D. Applying this model to CVB infection, we found that although the levels of intracellular virus production were similar in two-dimensional (2-D) and 3-D Caco-2 cell cultures, the release of infectious CVB was enhanced in 3-D cultures at early stages of infection. Unlike CVB, the replication of poliovirus (PV) was significantly reduced in 3-D Caco-2 cell cultures. Collectively, our studies show that Caco-2 cells grown in 3-D using the RWV bioreactor provide a cell culture model that structurally and transcriptionally represents key aspects of cells in the human GI tract and can thus be used to expand our understanding of enterovirus-host interactions in intestinal epithelial cells. IMPORTANCE Coxsackievirus B (CVB), a member of the enterovirus family of RNA viruses, is associated with meningitis, pericarditis, diabetes, dilated cardiomyopathy, and myocarditis, among other pathologies. CVB is transmitted via the fecal-oral route and encounters the

  8. A Three-Dimensional Cell Culture Model To Study Enterovirus Infection of Polarized Intestinal Epithelial Cells

    Science.gov (United States)

    Drummond, Coyne G.

    2015-01-01

    ABSTRACT Despite serving as the primary entry portal for coxsackievirus B (CVB), little is known about CVB infection of the intestinal epithelium, owing at least in part to the lack of suitable in vivo models and the inability of cultured cells to recapitulate the complexity and structure associated with the gastrointestinal (GI) tract. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of Caco-2 cells to model CVB infection of the gastrointestinal epithelium. We show that Caco-2 cells grown in 3-D using the rotating wall vessel (RWV) bioreactor recapitulate many of the properties of the intestinal epithelium, including the formation of well-developed tight junctions, apical-basolateral polarity, brush borders, and multicellular complexity. In addition, transcriptome analyses using transcriptome sequencing (RNA-Seq) revealed the induction of a number of genes associated with intestinal epithelial differentiation and/or intestinal processes in vivo when Caco-2 cells were cultured in 3-D. Applying this model to CVB infection, we found that although the levels of intracellular virus production were similar in two-dimensional (2-D) and 3-D Caco-2 cell cultures, the release of infectious CVB was enhanced in 3-D cultures at early stages of infection. Unlike CVB, the replication of poliovirus (PV) was significantly reduced in 3-D Caco-2 cell cultures. Collectively, our studies show that Caco-2 cells grown in 3-D using the RWV bioreactor provide a cell culture model that structurally and transcriptionally represents key aspects of cells in the human GI tract and can thus be used to expand our understanding of enterovirus-host interactions in intestinal epithelial cells. IMPORTANCE Coxsackievirus B (CVB), a member of the enterovirus family of RNA viruses, is associated with meningitis, pericarditis, diabetes, dilated cardiomyopathy, and myocarditis, among other pathologies. CVB is transmitted via the fecal-oral route and

  9. Ethanolamine metabolism in cultured bovine aortic endothelial cells

    International Nuclear Information System (INIS)

    The role of extracellular ethanolamine in phospholipid synthesis was examined in cultured bovine aortic endothelial cells. Serine and ethanolamine were both readily accumulated by these cells and incorporated into phospholipid. Exposing cells to extracellular ethanolamine for 4-6 weeks had no effect on cell growth, yet increased the phosphatidylethanolamine content of these cells by 31% as compared to control cells. The intracellular content of ethanolamine was measured by high performance liquid chromatography, and results showed that the ethanolamine-treated cells contained a significantly greater amount of free ethanolamine compared to control cells. Ethanolamine-treated cells also had decreased accumulation and incorporation into lipid of [3H]ethanolamine throughout a 48-h incubation and increased K'm and V'max parameters of ethanolamine transport as compared to control cells. Studies were also done to examine the effect of ethanolamine on the generation of free ethanolamine from phosphatidylserine. In pulse-chase experiments with [3H]serine, a physiological concentration of ethanolamine decreased the amount of 3H-labeled phosphatidylethanolamine produced from 3H-labeled phosphatidylserine by 12 h as compared to the amount of 3H-labeled phosphatidyl-ethanolamine produced in the absence of ethanolamine in the chase incubation. Furthermore, ethanolamine-treated cells accumulated 20% less labeled ethanolamine in the aqueous pool from [3H]serine after 24 h of incubation than did control cells. These results can be explained by isotope dilution with the ethanolamine pool that accumulates in these cells with time when exposed to media supplemented with a physiological concentration of ethanolamine and by an effect of ethanolamine on ethanolamine generation from phosphatidylserine

  10. Renal mesangial cell cultures as a model for study of erythropoietin production.

    OpenAIRE

    Kurtz, Armin; Jelkmann, W; Sinowatz, F.; Bauer, Christian

    1983-01-01

    Mesangial cells derived from isolated glomeruli of rat kidney were grown as homogeneous cell lines in culture. They released, into the culture medium, erythropoietin that had free terminal galactosyl residues and was therefore not active in vivo. The production of erythropoietin by these cells was significantly enhanced by either lowering the PO2 in the incubation atmosphere or by adding cobalt chloride to the culture medium. Therefore, mesangial cells in culture may be considered as an in vi...

  11. An Assessment of Cell Culture Plate Surface Chemistry for in Vitro Studies of Tissue Engineering Scaffolds

    OpenAIRE

    Alexander Röder; Elena García-Gareta; Christina Theodoropoulos; Nikola Ristovski; Keith A. Blackwood; Woodruff, Maria A.

    2015-01-01

    The use of biopolymers as a three dimensional (3D) support structure for cell growth is a leading tissue engineering approach in regenerative medicine. Achieving consistent cell seeding and uniform cell distribution throughout 3D scaffold culture in vitro is an ongoing challenge. Traditionally, 3D scaffolds are cultured within tissue culture plates to enable reproducible cell seeding and ease of culture media change. In this study, we compared two different well-plates with different surface ...

  12. THE STUDY OF VIRUSES REPRODUCTION IN CELL CULTURES BY THE METHOD OF MATHEMATICAL MODELING

    OpenAIRE

    N. A. Kontarov; S. A. Grishunina; N. V. Balaev; N. V. Yuminova; V. V. Zverev

    2014-01-01

    Abstract. In the study the mathematical analysis of viruses reproduction in cell culture using the Marchuk’ mathematical model to predict  reproduction of virus in one or another cell cultures has been conducted. The obtained theoretical results are corresponded to the experimental data on reproduction of rubella virus in cell cultures RK-13 and BHK-21. The sum of theoretical and experimental results can be used to select the optimal cell cultures for virus cumulation.

  13. Benchmarking of commercially available CHO cell culture media for antibody production

    OpenAIRE

    Reinhart, David; Damjanovic, Lukas; Kaisermayer, Christian; Kunert, Renate

    2015-01-01

    In this study, eight commercially available, chemically defined Chinese hamster ovary (CHO) cell culture media from different vendors were evaluated in batch culture using an IgG-producing CHO DG44 cell line as a model. Medium adaptation revealed that the occurrence of even small aggregates might be a good indicator of cell growth performance in subsequent high cell density cultures. Batch experiments confirmed that the culture medium has a significant impact on bioprocess performance, but hi...

  14. Pigment Cell Differentiation in Sea Urchin Blastula-Derived Primary Cell Cultures

    OpenAIRE

    Natalya V. Ageenko; Konstantin V. Kiselev; Dmitrenok, Pavel S.; Odintsova, Nelly A.

    2014-01-01

    The quinone pigments of sea urchins, specifically echinochrome and spinochromes, are known for their effective antioxidant, antibacterial, antifungal, and antitumor activities. We developed in vitro technology for inducing pigment differentiation in cell culture. The intensification of the pigment differentiation was accompanied by a simultaneous decrease in cell proliferation. The number of pigment cells was two-fold higher in the cells cultivated in the coelomic fluids of injured sea urchin...

  15. An IMRT/VMAT Technique for Nonsmall Cell Lung Cancer

    OpenAIRE

    Nan Zhao; Ruijie Yang; Junjie Wang; Xile Zhang; Jinna Li

    2015-01-01

    The study is to investigate a Hybrid IMRT/VMAT technique which combines intensity modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT) for the treatment of nonsmall cell lung cancer (NSCLC). Two partial arcs VMAT, 5-field IMRT, and hybrid plans were created for 15 patients with NSCLC. The hybrid plans were combination of 2 partial arcs VMAT and 5-field IMRT. The dose distribution of planning target volume (PTV) and organs at risk (OARs) for hybrid technique was compa...

  16. Cryopreservation of primary cell cultures of marine invertebrates.

    Science.gov (United States)

    Odintsova, N; Kiselev, K; Sanina, N; Kostetsky, E

    2001-01-01

    Primary cell cultures obtained from somatic and larval tissues of bivalve molluscs and from embryos of sea urchins were frozen to -196 degrees C by two-step freezing using 10% dimethyl sulfoxide (DMSO) or/and trehalose (3-30 mg/ml) as cryoprotectants. We estimated both cell viability and the RNA synthetic activity after freeze-thaw. Total lipid extracts from the tissues of echinoderms examined as possible cryoprotective agents demonstrated a weak cryoprotective capacity. Mussel lipid extract was found to possess a considerable cryoprotective activity. Cryoprotective capacity of tested lipids correlated with their thermotropic behaviour. DMSO + trehalose combination was shown to be a favourable cryoprotectant and sea urchin blastula cells the most freezing-tolerant cells. PMID:11788872

  17. Characterisation of solar cells by ion beam analysis techniques

    International Nuclear Information System (INIS)

    Several ion beam analysis techniques were applied for the characterisation of amorphous (a- Si) and polycrystalline silicon solar cells. Thickness and composition of thin layers in thin film a-Si cells were analysed by RBS (Rutherford backscattering) using 5 MeV Li beam and ERDA (Elastic recoil detection analysis) using 12 MeV C beam. Nuclear microprobe technique IBIC (Ion beam induced charge) was used for imaging a charge collection efficiency of EFG (edge-defined film-fed grown) silicon in attempt to correlate charge loss with a spatial distribution of structural defects in the material. (author)

  18. Nanocrystalline diamond: In vitro biocompatibility assessment by MG63 and human bone marrow cells cultures.

    Science.gov (United States)

    Amaral, M; Dias, A G; Gomes, P S; Lopes, M A; Silva, R F; Santos, J D; Fernandes, M H

    2008-10-01

    Nanocrystalline diamond (NCD) has a great potential for prosthetic implants coating. Nevertheless, its biocompatibility still has to be better understood. To do so, we employed several materials characterization techniques (SEM, AFM, micro-Raman spectroscopy) and cell culture assays using MG63 osteoblast-like and human bone marrow cells. Biochemical routines (MTT assays, Lowry's method, ALP activity) supported by SEM and confocal microscopy characterization were carried out. We used silicon nitride (Si3N4) substrates for NCD coatings based on a previous demonstration of the superior adhesion and tribological performance of these NCD coated ceramics. Results demonstrate an improved human osteoblast proliferation and the stimulation of differentiated markers, like ALP activity and matrix mineralization, compared with standard polystyrene tissue culture plates. The nanometric featuring of NCD, associated to its chemical affinity are key points for bone regeneration purposes. PMID:18085649

  19. Microfabrication of Bubbular Cavities in PDMS for Cell Sorting and Microcell Culture Applications

    Institute of Scientific and Technical Information of China (English)

    Ut-Binh T.Giang; Michael R.King; Lisa A.DeLouise

    2008-01-01

    We describe a novel technique, low surface energy Gas Expansion Molding (GEM), to fabricate microbubble arrays in polydimethylsiloxane (PDMS) which are incorporated into parallel plate flow chambers and tested in cell sorting and microcell culture applications. This architecture confers several operational advantages that distinguish this technology approach from currently used methods. Herein we describe the GEM process and the parameters that are used to control microbubble formation and a Vacuum-Assisted Coating (VAC) process developed to selectively and spatially alter the PDMS surface chemistry in the wells and on the microchannel surface. We describe results from microflow image visualization studies conducted to investigate fluid streams above and within microbubble wells and conclude with a discussion of cell culture studies in PDMS.

  20. A micro-Raman spectroscopic investigation of leukemic U-937 cells in aged cultures

    Science.gov (United States)

    Fazio, Enza; Trusso, Sebastiano; Franco, Domenico; Nicolò, Marco Sebastiano; Allegra, Alessandro; Neri, Fortunato; Musolino, Caterina; Guglielmino, Salvatore P. P.

    2016-04-01

    Recently it has been shown that micro-Raman spectroscopy combined with multivariate analysis is able to discriminate among different types of tissues and tumoral cells by the detection of significant alterations and/or reorganizations of complex biological molecules, such as nucleic acids, lipids and proteins. Moreover, its use, being in principle a non-invasive technique, appears an interesting clinical tool for the evaluation of the therapeutical effects and of the disease progression. In this work we analyzed molecular changes in aged cultures of leukemia model U937 cells with respect to fresh cultures of the same cell line. In fact, structural variations of individual neoplastic cells on aging may lead to a heterogeneous data set, therefore falsifying confidence intervals, increasing error levels of analysis and consequently limiting the use of Raman spectroscopy analysis. We found that the observed morphological changes of U937 cells corresponded to well defined modifications of the Raman contributions in selected spectral regions, where markers of specific functional groups, useful to characterize the cell state, are present. A detailed subcellular analysis showed a change in cellular organization as a function of time, and correlated to a significant increase of apoptosis levels. Besides the aforementioned study, Raman spectra were used as input for principal component analysis (PCA) in order to detect and classify spectral changes among U937 cells.

  1. Medium for development of bee cell cultures (Apis mellifera: Hymenoptera: Apidae)

    Science.gov (United States)

    A bee cell culture system was developed. A medium, WH2, for the production of cell cultures from hymenopteran species such as honey bee, Apis mellifera L. (Hymenoptera: Apidae) was developed. Multiple bee cell cultures were produced when using bee larvae and pupae as starting material and the modif...

  2. Culture of porcine hepatocytes or bile duct epithelial cells by inductive serum-free media

    Science.gov (United States)

    A serum-free, feeder-cell-dependent, selective culture system for the long-term culture of porcine hepatocytes or cholangiocytes was developed. Liver cells were isolated from 1 wk old pigs or young adult pigs (25 and 63 kg live weight) and were placed in primary culture on feeder-cell layers of mit...

  3. Human alveolar epithelial type II cells in primary culture.

    Science.gov (United States)

    Mao, Pu; Wu, Songling; Li, Jianchun; Fu, Wei; He, Weiqun; Liu, Xiaoqing; Slutsky, Arthur S; Zhang, Haibo; Li, Yimin

    2015-02-01

    Alveolar epithelial type II (AEII) cells are a key structure and defender in the lung but also are the targets in many lung diseases, including acute respiratory distress syndrome, ventilator-induced lung injury, and pulmonary fibrosis. We sought to establish an optimized method for high yielding and long maintenance of characteristics of primary human AEII cells to facilitate the investigation of the mechanisms of lung diseases at the cellular and molecular levels. Adult human peripheral normal lung tissues of oncologic patients undergoing lung resection were collected. The AEII cells were isolated and identified by the expression of pro-surfactant protein (SP)C, epithelial sodium channel (αENaC) and cytokeratin (CK)-8, the lamellar bodies specific for AEII cells, and confirmed by the histology using electron microscopy. The phenotype of AEII cells was characterized by the expression of surfactant proteins (SP-A, SP-B, SP-C, SP-D), CK-8, KL-6, αENaC, and aquaporin (AQP)-3, which was maintained over 20 days. The biological activity of the primary human AEII cells producing SP-C, cytokines, and intercellular adhesion molecule-1 was vigorous in response to stimulation with tumor necrosis factor-α. We have modified previous methods and optimized a method for isolation of high purity and long maintenance of the human AEII cell phenotype in primary culture. This method provides an important tool for studies aiming at elucidating the molecular mechanisms of lung diseases exclusively in AEII cells. PMID:25677546

  4. Reduction of fatal graft-versus-host disease by 3H--thymidine suicide of donor cells cultured with host cells

    International Nuclear Information System (INIS)

    The effect of the tritiated thymidine (3H-TdR) suicide technique on the ability of donor cells to induce fatal graft-versus-host disease (GVHD) was studied. C57BL/6 (H-2/sup b/) spleen cells were stimulated in vitro with irradiated BALB/c (H-2/sup d/) Moloney lymphoma cells in mixed culture and 3H-TdR of high-specific activity added to eliminate proliferating cells. The ability of such cells to induce fatal GVHD was assayed by injecting them i.v. into adult BALB/c mice immunosuppressed with cyclophosphamide (180 mg/kg). These cells induced fatal GVHD in fewer mice (52 percent) than did C57BL/6 cells cultured with BALB/c lymphoma cells but without 3H-TdR (87 percent) and C57BL/6 cells cultured with irradiated C57BL/6 cells with (95 percent) or without 3H-TdR (86 percent). Thus, the 3H-TdR suicide technique greatly diminished the ability of cells to induce lethal GVHD

  5. The Application of Dielectric Spectroscopy and Biocalorimetry for the Monitoring of Biomass in Immobilized Mammalian Cell Cultures

    Directory of Open Access Journals (Sweden)

    Harriet E. Cole

    2015-05-01

    Full Text Available The purpose of this study was to introduce dielectric spectroscopy and biocalorimetry as monitoring methods to follow immobilised Chinese Hamster Ovary (CHO cell culture development. The theory behind both monitoring techniques is explained and perfusion cultures are performed in a Reaction Calorimeter (eRC1 from Mettler Toledo as an application example. The findings of this work show that dielectric spectroscopy gives highly reliable information upon the viable cell density throughout the entire culture. On the other hand, the RC1 could only provide accurate data from day 5, when the cell density exceeded 4 × 106 vcells∙mL−1 (viable cell per mL working volume (WV. The method validation showed the limit of detection (LOD for 1.4 L cultures to be 8.86 × 106 vcells∙mL−1, a viable cell density commonly achieved in fed-batch and the early stages of a perfusion culture. This work suggests that biocalorimetry should be possible to implement at industrial scale to monitor CHO cell cultures.

  6. Use of immunofluorescence technique in cultured fibroblasts from Mediterranean cetaceans as new "in vitro" tool to investigate effects of environmental contaminants.

    Science.gov (United States)

    Marsili, Letizia; Casini, Silvia; Bucalossi, Daniela; Porcelloni, Serena; Maltese, Silvia; Fossi, Maria Cristina

    2008-07-01

    The aim of the present study was to propose the immunofluorescence technique in cultured fibroblasts from Mediterranean cetaceans as a new "in vitro" tool to explore the susceptibility of these marine mammals to different xenobiotic compounds. The cell lines were cultured from integument biopsies of free-ranging and stranded cetaceans (dead within 12h). Using the indirect immunofluorescence assay, we detected endogenous proteins induced by different contaminants. Here we present the method used for qualitative and quantitative evaluation of cytochromes P450 (CYP1A1 and CYP2B) induced by some POPs (DDTs and PCBs) and emerging contaminants (PBDEs) in fibroblast cell cultures of striped dolphin (Stenella coeruleoalba) and bottlenose dolphin (Tursiops truncatus). Immunofluorescence was quantified with a specially designed Olympus macro, DetectIntZ. A major result was the possibility of using this "in vitro" assay to quantify induction of endogenous proteins. PMID:18396327

  7. Controlled and reversible induction of differentiation and activation of adult human hepatocytes by a biphasic culture technique

    Institute of Scientific and Technical Information of China (English)

    Marcus K.H. Auth; Wolf-Otto Bechstein; Roman A. Blaheta; Kim A. Boost; Kerstin Leckel; Wolf-Dietrich Beecken; Tobias Engl; Dietger Jonas; Elsie Oppermann; Philip Hilgard; Bernd H. Markus

    2005-01-01

    AIM: Clinical application of human hepatocytes (HC) is hampered by the progressive loss of growth and differentiation in vitro. The object of the study was to evaluate the effect of a biphasic culture technique on expression and activation of growth factor receptors and differentiation of human adult HC.METHODS: Isolated HC were sequentially cultured in a hormone enriched differentiation medium (DM) containing nicotinamide, insulin, transferrin, selenium, and dexamethasone or activation medium (AM) containing hepatocyte growth factor (HGF), epidermal growth factor (EGF), andgranulocyte-macrophage colony-stimulating factor (GMCSF). Expression, distribution and activation of the HC receptors (MET and EGFR) and the pattern of characteristic cytokeratin (CK) filaments were measured by fluorometry, confocal microscopy and Western blotting.RESULTS: In the biphasic culture system, HC underwent repeated cycles of activation (characterized by expression and activation of growth factor receptors) and re-differentiation (illustrated by distribution of typical filaments CK-18 but low or absent expression of CK-19). In AM increased expression of MET and EGFR was associated with receptor translocation into the cytoplasm and induction of atypical CK-19. In DM low expression of MET and EGFR was localized on the cell membrane and CK-19 was reduced. Receptor phosphorylation required embedding of HC in collagen type Ⅰ gel.CONCLUSION: Control and reversible modulation of growth factor receptor activation of mature human HC can be accomplishedin vitro, when defined signals from the extracellular matrix and sequential growth stimuli are provided. The biphasic technique helps overcome dedifferentiation, which occurs during continuous stimulation by means of growth factors.

  8. Basis for cumulative cultural evolution in chimpanzees: social learning of a more efficient tool-use technique.

    Directory of Open Access Journals (Sweden)

    Shinya Yamamoto

    Full Text Available BACKGROUND: The evidence for culture in non-human animals has been growing incrementally over the past two decades. However, the ability for cumulative cultural evolution, with successive generations building on earlier achievements, in non-human animals remains debated. Faithful social learning of incremental improvements in technique is considered to be a defining feature of human culture, differentiating human from non-human cultures. This study presents the first experimental evidence for chimpanzees' social transmission of a more efficient tool-use technique invented by a conspecific group member. METHODOLOGY/PRINCIPAL FINDINGS: The chimpanzees were provided with a straw-tube, and spontaneously demonstrated two different techniques in obtaining juice through a small hole: "dipping" and "straw-sucking". Both the "dipping" and "straw-sucking" techniques depended on the use of the same tool (straw-tube for the same target (juice accessible from exactly the same location (small hole 1 cm in diameter. Therefore the difference between "dipping" and "straw-sucking" was only in "technique". Although the two techniques differed significantly in their efficiency, their cognitive and perceptuo-motor complexity were comparable. All five chimpanzees who initially performed the "dipping" technique switched to using the more efficient "straw-sucking" technique upon observing a conspecific or human demonstrate the more proficient alternate "straw-sucking" technique. CONCLUSIONS/SIGNIFICANCE: The social learning mechanism involved here was clearly not local or stimulus enhancement, but imitation or emulation of a tool-use technique. When there is no biologically relevant difference in cognitive or perceptuo-motor complexity between two techniques, and when chimpanzees are dissatisfied with their own technique, chimpanzees may socially learn an improved technique upon close observation of a proficient demonstrator. This study provides important insights into

  9. Bridging Mediterranean cultures in the IYS: A documentary exhibition on irrigation techniques in water scarcity conditions

    Science.gov (United States)

    Barontini, Stefano; Louki, Amina; Ben Slima, Zied; Ezzahra Ghaouch, Fatima; Labaran, Raisa; Raffelli, Giulia; Peli, Marco; Vitale, Nicola

    2015-04-01

    Brescia, an industrial city in Northern Italy, is now experiencing a crucial change in its traditional structure. In recent years in fact it has been elected as living and working seat by many foreigners and it is now one of the cities with the greatest percentage of migrants in the Country. This is an important challenge for the city and an opportunity to merge, compare and integrate different cultures to build its future. In this context some students of different Courses (engineering and medicine), belonging both to the Arabian and local community, met together and with researchers in the study team 'Al-B¯i r¯u n¯i , for culture, science and society'. The team aims at organising cultural events in which, starting from the figure of the Persian scientist Ab¯u Raih. ¯a n Al-B¯i r¯u n¯i (about 973, 1051), the contribution of the Arabian and Islamic culture to the development of the European one in the middle ages is investigated. Moving from the initial idea of the study team Al-B¯i r¯u n¯i and from the suggestions of the World Soil Day 2014 and of the International Year of Soils 2015, we built a documentary exhibition entitled 'Irrigation techniques in water scarcity conditions'. The exhibition, which stresses the importance of the irrigation techniques for the soil conservation, is focused on the idea of disseminating two main concepts, i.e. (1) the technological continuity of some water supply systems in countries, around the Mediterranean Sea, affected by similar conditions of water availability, and (2) the possibility of building environments where, due to severe or extreme climatic conditions, the sustainability is reached when the man lives in equilibrium with the nature. The exhibition, which is written in Italian and will move around in the city during all 2015, consists of about twenty posters organized into three main chapters, corresponding to three main classes of water supply systems which are common in most of the countries surrounding

  10. On-line Measurements and Control of Viable Cell Density in Cell Culture Manufacturing Processes using Radio-frequency Impedance.

    Science.gov (United States)

    Carvell, John P; Dowd, Jason E

    2006-03-01

    In this work, radio-frequency (RF) impedance is reviewed as a method for monitoring and controlling cell culture manufacturing processes. It is clear from the many publications cited that RF Impedance is regarded as an accurate and reliable method for measuring the live cell bio-volume both on-line and off-line and the technology is also sutable for animal cells in suspension, attached to micro-carriers or immobilized in fixed beds. In cGMP production, RF Impedance is being used in three main areas. Firstly, it is being used as a control instrument for maintaining consistent perfusion culture allowing the bioreactor to operate under optimum conditions for maximum production of recombinant proteins. In the second application it has not replaced traditional off-line live cell counting techniques but it is being used as an additional monitoring tool to check product conformance. Finally, RF Impedance is being used to monitor the concentration of live cells immobilized on micro-carriers or packed beds in cGMP processes where traditional off-line live cell counting methods are inaccurate or impossible to perform. PMID:19003069

  11. Establishment of automated culture system for murine induced pluripotent stem cells

    Directory of Open Access Journals (Sweden)

    Koike Hiroyuki

    2012-11-01

    Full Text Available Abstract Background Induced pluripotent stem (iPS cells can differentiate into any cell type, which makes them an attractive resource in fields such as regenerative medicine, drug screening, or in vitro toxicology. The most important prerequisite for these industrial applications is stable supply and uniform quality of iPS cells. Variation in quality largely results from differences in handling skills between operators in laboratories. To minimize these differences, establishment of an automated iPS cell culture system is necessary. Results We developed a standardized mouse iPS cell maintenance culture, using an automated cell culture system housed in a CO2 incubator commonly used in many laboratories. The iPS cells propagated in a chamber uniquely designed for automated culture and showed specific colony morphology, as for manual culture. A cell detachment device in the system passaged iPS cells automatically by dispersing colonies to single cells. In addition, iPS cells were passaged without any change in colony morphology or expression of undifferentiated stem cell markers during the 4 weeks of automated culture. Conclusions Our results show that use of this compact, automated cell culture system facilitates stable iPS cell culture without obvious effects on iPS cell pluripotency or colony-forming ability. The feasibility of iPS cell culture automation may greatly facilitate the use of this versatile cell source for a variety of biomedical applications.

  12. Mesh词表词汇实用例句:“细胞培养技术-Cell Culture Techniques”

    Institute of Scientific and Technical Information of China (English)

    2012-01-01

    例句:Six kinds of plant cell culture techniques,the important influence factor and its adjustment in the process of producing the plant secondary metabolites by plant cellculture,are elaborated in this paper,putting forward some rationalization suggestions for going deep into researches at the same time.

  13. Photocontrol of nitric oxide production in cell culture using a caged isoform selective inhibitor.

    Science.gov (United States)

    Perdicakis, Basil; Montgomery, Heather J; Abbott, Glenn L; Fishlock, Dan; Lajoie, Gilles A; Guillemette, J Guy; Jervis, Eric

    2005-01-01

    Over the past decade, multiphoton microscopy has progressed from a photonic novelty to a technique whose application is currently experiencing exponential growth in the biological sciences. A novel application of this technology with significant therapeutic potential is the control of drug activity by multiphoton photolysis of caged therapeutics. As an initial case study, the potent isoform selective inhibitor N-(3-(aminomethyl)benzyl) acetamidine (1400W) of inducible nitric oxide synthase (iNOS) has been conjugated to a caging molecule 6-bromo-7-hydroxy-4-hydroxyquinoline-2-ylmethyl acetyl ester (Bhc). Here we present the first report of a bulk therapeutic effect, inhibition of nitric oxide production, in mammalian cell culture by multiphoton photolysis of a caged drug, Bhc-1400W. Mouse macrophage RAW 264.7 cells induced with bacterial lipopolysaccharides to express iNOS were used to assess the therapeutic value of the conjugated inhibitor. Both 1400W and Bhc-1400W are stable in metabolically active cells and an optimal time interval for the photorelease of the inhibitor was determined. The ratios of the IC(50) values of Bhc-1400W over 1400W calculated in the presence of iNOS enzyme and in RAW 264.7 cell culture are 19 and 100, respectively, indicating that a broad therapeutic range exists in cell culture. Multiphoton uncaging protocols and therapeutic doses of inhibitors were not cytotoxic. Photocontrol of LPS induced nitric oxide production was achieved in mammalian cell culture using a single laser focal volume. This technology has the potential to control active drug concentrations in vivo, a lack of which is one of the main problems currently associated with systemic drug administration. PMID:15582451

  14. Three-dimensional Cell Culture Devices for Cancer Migration and Drug Testing

    Science.gov (United States)

    Ma, Liang

    Porous polymeric materials are widely used to mimic the extracellular matrix (ECM) environment for applications such as 3D cell culturing and tissue engineering. A series of comparative experiments on 3D cell cultures both in PLA porous scaffolds and alginate gels were conducted to create an in vitro tumor model. A novel 3D cell culture device based on porous polymeric material was developed to study cancer migration. Significant cell migration was observed through the porous channel within 1--2 weeks induced by 20% fetal bovine serum (FBS). A three-dimensional micro-scale perfusion-based two-chamber (3D-muPTC) tissue model system was developed to test the cytotoxicity of anticancer drugs by emulating liver metabolism effects in vitro. Hepatoma cells and glioblastoma multiforme (GBM) cancer cells were cultured in porous polymeric scaffolds in two separate chambers, representing the liver and tumor, respectively. The cytotoxic effect of temozolomide (TMZ) was first tested using this system. It was found that the GBM cells showed a much higher viability under the TMZ treatment with liver cells in the system, suggesting that the drug metabolism in liver is affecting the efficacy of the drug. The favorable metabolism effect of cytochrome P450 (CYP) was tested using a prodrug ifosfamide (IFO). Without the liver cells, IFO showed only slight toxicity to GBM cells. Moreover, it was shown that different expression levels of CYP 3A4, a major drug metabolizing enzyme, in liver cells caused significantly different levels of GBM cell viability. Simulation of the flow characteristics in the 3D-muPTC system was conducted using the finite-element analysis approach. The shear stress was predicted in the porous scaffolds under different flow rate conditions. The predicted shear stress effects agreed well with an experimental cell viability study. A low cost organic solvent free approach to fabricating tissue engineering scaffolds was developed by combining the twin-screw extrusion

  15. Cell Culture in Microgravity: Opening the Door to Space Cell Biology

    Science.gov (United States)

    Pellis, Neal R.; Dawson, David L. (Technical Monitor)

    1999-01-01

    Adaptational response of human cell populations to microgravity is investigated using simulation, short-term Shuttle experiments, and long-term microgravity. Simulation consists of a clinostatically-rotated cell culture system. The system is a horizontally-rotated cylinder completely filled with culture medium. Low speed rotation results in continuous-fall of the cells through the fluid medium. In this setting, cells: 1) aggregate, 2) propagate in three dimensions, 3) synthesize matrix, 4) differentiate, and 5) form sinusoids that facilitate mass transfer. Space cell culture is conducted in flight bioreactors and in static incubators. Cells grown in microgravity are: bovine cartilage, promyelocytic leukemia, kidney proximal tubule cells, adrenal medulla, breast and colon cancer, and endothelium. Cells were cultured in space to test specific hypotheses. Cartilage cells were used to determine structural differences in cartilage grown in space compared to ground-based bioreactors. Results from a 130-day experiment on Mir revealed that cartilage grown in space was substantially more compressible due to insufficient glycosaminoglycan in the matrix. Interestingly, earth-grown cartilage conformed better to the dimensions of the scaffolding material, while the Mir specimens were spherical. The other cell populations are currently being analyzed for cell surface properties, gene expression, and differentiation. Results suggest that some cells spontaneously differentiate in microgravity. Additionally, vast changes in gene expression may occur in response to microgravity. In conclusion, the transition to microgravity may constitute a physical perturbation in cells resulting in unique gene expressions, the consequences of which may be useful in tissue engineering, disease modeling, and space cell biology.

  16. Digital microfluidics for automated hanging drop cell spheroid culture.

    Science.gov (United States)

    Aijian, Andrew P; Garrell, Robin L

    2015-06-01

    Cell spheroids are multicellular aggregates, grown in vitro, that mimic the three-dimensional morphology of physiological tissues. Although there are numerous benefits to using spheroids in cell-based assays, the adoption of spheroids in routine biomedical research has been limited, in part, by the tedious workflow associated with spheroid formation and analysis. Here we describe a digital microfluidic platform that has been developed to automate liquid-handling protocols for the formation, maintenance, and analysis of multicellular spheroids in hanging drop culture. We show that droplets of liquid can be added to and extracted from through-holes, or "wells," and fabricated in the bottom plate of a digital microfluidic device, enabling the formation and assaying of hanging drops. Using this digital microfluidic platform, spheroids of mouse mesenchymal stem cells were formed and maintained in situ for 72 h, exhibiting good viability (>90%) and size uniformity (% coefficient of variation digital microfluidic platform provides a viable tool for automating cell spheroid culture and analysis. PMID:25510471

  17. Effects of methyl isocyanate on rat muscle cells in culture.

    Science.gov (United States)

    Anderson, D; Goyle, S; Phillips, B J; Tee, A; Beech, L; Butler, W H

    1988-04-01

    Since the Bhopal disaster, in which the causal agent was methyl isocyanate (MIC), exposed people have complained of various disorders including neuromuscular dysfunction. In an attempt to gain some information about the response of muscle tissue to MIC its effects were investigated in cells in culture isolated from muscle of 2 day old rats. After treatment with a range of MIC concentrations (0.025-0.5 microliter/5 ml culture) the total number of nuclei of the two main cell types (fibroblasts and myoblasts) and the number of nuclei in muscle fibres (myotubes) were recorded. At lower doses which had little effect on the total number of nuclei, the formation of muscle fibres--that is, fusion of muscle cells--was prevented as the proportion of nuclei in myotubes was decreased. At higher doses both cell types were killed. This would suggest either an effect on muscle differentiation or a selective toxicity towards myoblasts. The observations were supported by light and electron microscopy. PMID:3378004

  18. Thymic epithelial cells. I. Expression of strong suppressive (veto) activity in mouse thymic epithelial cell cultures

    DEFF Research Database (Denmark)

    Claesson, Mogens Helweg; Ropke, C

    1990-01-01

    We show that thymic epithelial cells grown under serum-free conditions in a chemically defined culture medium can act as veto cells in vitro. The veto activity of thymic epithelial cells results in inactivation of specific alloreactive cytotoxic T-cell precursors at the clonal level. It is...... concluded that the epithelial stromal cells of the thymus, by acting as veto cells, may be responsible for the negative intrathymic selection of self-reactive thymocytes leading to elimination of the vast majority of immature thymic lymphocytes....

  19. Cultured stem cells are sensitive to gravity changes

    Science.gov (United States)

    Buravkova, L. B.; Romanov, Yu. A.; Konstantinova, N. A.; Buravkov, S. V.; Gershovich, Yu. G.; Grivennikov, I. A.

    2008-09-01

    Stem and precursor cells play an important role in development and regeneration. The state of these cells is regulated by biochemical substances, mechanical stimuli and cellular interactions. To estimate gravity effects we used two types of cultured stem cells: human mesenchymal stromal cells (hMSCs) from bone marrow and mice embryonic stem (mESC) line R1. Gravity changes were simulated by long-term (4-7 days) slow clinorotation and leaded to decreased hMSC proliferation, changes of cell morphology and modified F-actin cytoskeleton. We did not find the shifts in cell phenotype except for decreased expression of HLA 1 and CD105 but excretion of IL-6 into medium increased significantly. Remodeling of cytoskeleton started after first 4 h and was similar to preapoptotic changes. This data suggested the modification in cell adhesion and possible commitment of hMSC. It was observed that expression of alkaline phosphatase by MSC in osteogenic medium was more intensive in control. On the contrary, clinorotation did not change formation of mESC colonies and increased proliferation activity in LIF+-medium. However, the number of embryonic bodies after clinorotation was less than in static control. It is suggested that ESCs kept the viability and proliferative potential but decreased the differentiation ability after changes in gravity stimulation.

  20. Sparse grid techniques for particle-in-cell schemes

    CERN Document Server

    Ricketson, Lee F

    2016-01-01

    We propose the use of sparse grids to accelerate particle-in-cell (PIC) schemes. By using the so-called `combination technique' from the sparse grids literature, we are able to dramatically increase the size of the spatial cells in multi-dimensional PIC schemes while paying only a slight penalty in grid-based error. The resulting increase in cell size allows us to reduce the statistical noise in the simulation without increasing total particle number. We present initial proof-of-principle results from test cases in two and three dimensions that demonstrate the new scheme's efficiency, both in terms of computation time and memory usage.

  1. Physical techniques for the study of exocytosis in isolated cells.

    Science.gov (United States)

    Henry, J P; Darchen, F; Cribier, S

    1998-01-01

    Membrane traffic is an important aspect of cell biology which implies shuttle vesicles and multiple binding/fusion events. In spite of rapid progress at the biochemical level, the mechanism of fusion is still not understood. A detailed physical description of the phenomenon is possible at the level of the plasma membrane where secretory vesicles fuse with the cell membrane, a process known as exocytosis. This process is specially active in neurons (release of neurotransmitter) and in endocrine cells (release of hormones), where exocytosis is tightly regulated. Among the biophysical techniques developed, cell membrane capacitance measurements by the technique of patch-clamp and amperometry of the oxidizable secretory products have resulted in interesting information. These techniques have described the initial fusion pore, its fluctuations, the efflux of material through the pore and its irreversible expansion. Optical techniques, using bioluminescent and fluorescent probes are also in progress. For instance, the dye FM 1-43 binds to but is not translocated through biological membranes and it has been used to measure membrane surface, as done by capacitance measurement. Evanescent wave fluorescence microscopy has been recently introduced to analyse the behaviour of secretory granules in the vicinity of the plasma membrane. PMID:9782378

  2. Total volume culture technique for the isolation of microorganisms from continuous ambulatory peritoneal dialysis patients with peritonitis.

    OpenAIRE

    Dawson, M S; Harford, A M; Garner, B K; Sica, D A; Landwehr, D M; Dalton, H P

    1985-01-01

    A total volume method of culturing dialysis fluid from continuous ambulatory peritoneal dialysis patients during episodes of peritonitis was developed. Concentrated culture media stored in small blood transfer bags were added directly to the drained dialysate exchange bags by the same technique used to carry out the dialysate exchange. The exchange bag with the added culture medium was incubated at 35 degrees C and observed for turbidity. Seventy-eight dialysis exchange bags from patients wit...

  3. Glycosylation-mediated phenylpropanoid partitioning in Populus tremuloides cell cultures

    Directory of Open Access Journals (Sweden)

    Babst Benjamin A

    2009-12-01

    Full Text Available Abstract Background Phenylpropanoid-derived phenolic glycosides (PGs and condensed tannins (CTs comprise large, multi-purpose non-structural carbon sinks in Populus. A negative correlation between PG and CT concentrations has been observed in several studies. However, the molecular mechanism underlying the relationship is not known. Results Populus cell cultures produce CTs but not PGs under normal conditions. Feeding salicyl alcohol resulted in accumulation of salicins, the simplest PG, in the cells, but not higher-order PGs. Salicin accrual reflected the stimulation of a glycosylation response which altered a number of metabolic activities. We utilized this suspension cell feeding system as a model for analyzing the possible role of glycosylation in regulating the metabolic competition between PG formation, CT synthesis and growth. Cells accumulated salicins in a dose-dependent manner following salicyl alcohol feeding. Higher feeding levels led to a decrease in cellular CT concentrations (at 5 or 10 mM, and a negative effect on cell growth (at 10 mM. The competition between salicin and CT formation was reciprocal, and depended on the metabolic status of the cells. We analyzed gene expression changes between controls and cells fed with 5 mM salicyl alcohol for 48 hr, a time point when salicin accumulation was near maximum and CT synthesis was reduced, with no effect on growth. Several stress-responsive genes were up-regulated, suggestive of a general stress response in the fed cells. Salicyl alcohol feeding also induced expression of genes associated with sucrose catabolism, glycolysis and the Krebs cycle. Transcript levels of phenylalanine ammonia lyase and most of the flavonoid pathway genes were reduced, consistent with down-regulated CT synthesis. Conclusions Exogenous salicyl alcohol was readily glycosylated in Populus cell cultures, a process that altered sugar utilization and phenolic partitioning in the cells. Using this system, we

  4. In vitro cell culture lethal dose submitted to gamma radiation

    International Nuclear Information System (INIS)

    The present study was designed to evaluate the in vitro effect of gamma radiation in cell culture of mouse connective tissue exposed to different doses of gamma radiation and under several conditions. The cell viability was analyzed by neutral red uptake methodology. This assay was developed for establish a methodology to be used in the future in the study of resveratrol radioprotection. Resveratrol (3,4',5- trihydroxystilbene), a phenolic phytoalexin that occurs naturally in some spermatophytes, such as grapevines, in response to injury as fungal infections and exposure to ultraviolet light. In the wines this compound is found at high levels and is considered one of the highest antioxidant constituents. The intense antioxidant potential of resveratrol provides many pharmacological activities including cardioprotection, chemoprevention and anti-tumor effects. Our results demonstrated that 60Co gamma radiation lethal dose (LD50) on NCTC clone 929 cells was about 340Gy. (author)

  5. Ribavirin Inhibits Parrot Bornavirus 4 Replication in Cell Culture.

    Science.gov (United States)

    Musser, Jeffrey M B; Heatley, J Jill; Koinis, Anastasia V; Suchodolski, Paulette F; Guo, Jianhua; Escandon, Paulina; Tizard, Ian R

    2015-01-01

    Parrot bornavirus 4 is an etiological agent of proventricular dilatation disease, a fatal neurologic and gastrointestinal disease of psittacines and other birds. We tested the ability of ribavirin, an antiviral nucleoside analog with antiviral activity against a range of RNA and DNA viruses, to inhibit parrot bornavirus 4 replication in duck embryonic fibroblast cells. Two analytical methods that evaluate different products of viral replication, indirect immunocytochemistry for viral specific nucleoprotein and qRT-PCR for viral specific phosphoprotein gene mRNA, were used. Ribavirin at concentrations between 2.5 and 25 μg/mL inhibited parrot bornavirus 4 replication, decreasing viral mRNA and viral protein load, in infected duck embryonic fibroblast cells. The addition of guanosine diminished the antiviral activity of ribavirin suggesting that one possible mechanism of action against parrot bornavirus 4 may likely be through inosine monophosphate dehydrogenase inhibition. This study demonstrates parrot bornavirus 4 susceptibility to ribavirin in cell culture. PMID:26222794

  6. In vitro cell culture lethal dose submitted to gamma radiation

    Energy Technology Data Exchange (ETDEWEB)

    Moreno, Carolina S.; Rogero, Sizue O.; Rogero, Jose Roberto [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)], e-mail: carolina_sm@hotmail.com; Ikeda, Tamiko I.; Cruz, Aurea S. [Instituto Adolfo Lutz, Sao Paulo, SP (Brazil)

    2009-07-01

    The present study was designed to evaluate the in vitro effect of gamma radiation in cell culture of mouse connective tissue exposed to different doses of gamma radiation and under several conditions. The cell viability was analyzed by neutral red uptake methodology. This assay was developed for establish a methodology to be used in the future in the study of resveratrol radioprotection. Resveratrol (3,4',5- trihydroxystilbene), a phenolic phytoalexin that occurs naturally in some spermatophytes, such as grapevines, in response to injury as fungal infections and exposure to ultraviolet light. In the wines this compound is found at high levels and is considered one of the highest antioxidant constituents. The intense antioxidant potential of resveratrol provides many pharmacological activities including cardioprotection, chemoprevention and anti-tumor effects. Our results demonstrated that {sup 60}Co gamma radiation lethal dose (LD50) on NCTC clone 929 cells was about 340Gy. (author)

  7. Sulphur XANES Analysis of Cultured Human Prostate Cancer Cells

    International Nuclear Information System (INIS)

    Prostate cancer is one of the most commonly diagnosed cancers in men throughout the world. It is believed that changes to the structure of protein binding sites, altering its metabolism, may play an important role in carcinogenesis. Sulphur, often present in binding sites, can influence such changes through its chemical speciation. Hence there is a need for precise investigation of coordination environment of sulphur. X-ray absorption near edge structure spectroscopy offers such possibility. Cell culture samples offer histologically well defined areas of good homogeneity, suitable for successful and reliable X-ray absorption near edge structure analysis. This paper presents sulphur speciation data collected from three different human prostate cancer cell lines (PC-3, LNCaP and DU-145). Sulphur X-ray absorption near edge structure analysis was performed on K-edge structure. The spectra of cells were compared with those of cancerous tissue and with organic substances as well as inorganic compounds. (authors)

  8. A non-invasive method for in situ quantification of subpopulation behaviour in mixed cell culture.

    Science.gov (United States)

    MacArthur, Ben D; Tare, Rahul S; Please, Colin P; Prescott, Philip; Oreffo, Richard O C

    2006-02-22

    Ongoing advances in quantitative molecular- and cellular-biology highlight the need for correspondingly quantitative methods in tissue-biology, in which the presence and activity of specific cell-subpopulations can be assessed in situ. However, many experimental techniques disturb the natural tissue balance, making it difficult to draw realistic conclusions concerning in situ cell behaviour. In this study, we present a widely applicable and minimally invasive method which combines fluorescence cell labelling, retrospective image analysis and mathematical data processing to detect the presence and activity of cell subpopulations, using adhesion patterns in STRO-1 immunoselected human mesenchymal populations and the homogeneous osteoblast-like MG63 continuous cell line as an illustration. Adhesion is considered on tissue culture plastic and fibronectin surfaces, using cell area as a readily obtainable and individual cell specific measure of spreading. The underlying statistical distributions of cell areas are investigated and mappings between distributions are examined using a combination of graphical and non-parametric statistical methods. We show that activity can be quantified in subpopulations as small as 1% by cell number, and outline behaviour of significant subpopulations in both STRO-1+/- fractions. This method has considerable potential to understand in situ cell behaviour and thus has wide applicability, for example in developmental biology and tissue engineering. PMID:16849218

  9. The Biological Study of the Cultured Human Lens Epithelial Cells in Vitro

    Institute of Scientific and Technical Information of China (English)

    1994-01-01

    The human lens epithelial cells (HLE) cultured in vitro was established in normal and cataractous lenses. The biological feature, histological characteristics and the ultrastructure of the cultured HLE cells were investigated. The results reveal that the proliferative capacity of the culutured HLE cells is reversely proportional to the donour age; the cultured HLE cells has the limited proliferative capacity in vitro. The relieve of the contact inhibition is the effective trigger of the HLE cell prolife...

  10. Bladder cancer cell in co-culture induces human stem cell differentiation to urothelial cells through paracrine FGF10 signaling

    OpenAIRE

    Chung, Seyung S.; Koh, Chester J.

    2013-01-01

    FGF10 is required for embryonic epidermal morphogenesis including brain development, lung morphogenesis, and initiation of limb bud formation. In this study, we investigated the role of FGF10 as a lead induction factor for stem cell differentiation toward urothelial cell. To this end, human multi-potent stem cell in vitro system was employed. Human amniotic fluid stem cells were co-cultured with immortalized bladder cancer lines to induce directed differentiation into urothelial cells. Urothe...

  11. Damaging effects of gliadin on three-dimensional cell culture model

    Institute of Scientific and Technical Information of China (English)

    Ersilia Dolfini; Maria Letizia Falini; Maria Teresa Bardella; Luca Elli; Leda Roncoroni; Barbara Costa; Maria Pia Colleoni; Vito Lorusso; Simona Ramponi; Paola Braidotti; Stefano Ferrero

    2005-01-01

    AIM: To evaluate the effects of gliadin on the oxidative environment in the"in vivo-like" model of a three-dimensional cell culture system.METHODS: LoVo cell line (intestinal adenocarcinoma)multicellular spheroids were treated with digested gliadin (with albumin used as a control). Spheroid volumes, cell viability and morphology, lactate dehydrogenase (LDH)release, content of reduced glutathione (GSH) and activity of GSH-related enzymes were examined. The data were statistically analyzed using the Student's t-test (P<0.05).was considered statistically significant.RESULTS: Gliadin reduced cell viability (from 20% to 60%)and led to morphological alterations characterized by apoptotic findings and cytoskeletal injuries. LDH activity increased. The content of GSH reduced (-20% vs controls),and activity of GSH-related enzymes was significantly inhibited.CONCLUSION: Gliadin treatment induces an imbalance in the antioxidative mechanism of cells cultured by the three-dimensional technique. This alteration may explain the cell damage directly caused by gliadin and the subsequent morphological abnormalities.

  12. Synthesis and characterization of biocompatible multicomponent polymer systems as supports for cell cultures

    International Nuclear Information System (INIS)

    Engineering living tissue for reconstructive surgery requires an appropriate cell source and optimal culture conditions, but also a suitable biodegradable scaffold as the basic elements. On the basis of the well known facts that scaffold chemistry and architecture can influence the fate and function of engrafted cells, a large number of polymers, as cell cultures supports, have been proposed. In this study, we report a synthesis, characterization and cell interactions with the following polymer systems: I. Poly[L- lactic acid / glycolic acid / poly(dimethylsiloxane)], copolymers; II. Poly(DL - lactic acid) / triblock PCL - PDMS - PCL copolymers; III. Blends of poly(DL - lactic - co - glycolic acid) and triblock PCL - PDMS - PCL copolymers. For the cell seeding experiments, Swiss 3T3 and/or L929 mouse fibroblasts were grown in RPMI 1640 and/or DMEM / F12 medium, and placed onto the bio polymer non porous or porous films, prepared using a particulate leaching technique. The amount of cells present on the surfaces of the scaffolds was quantified using a neutral red uptake assay. (Author)

  13. Extracellular mass transport considerations for space flight research concerning suspended and adherent in vitro cell cultures

    Science.gov (United States)

    Klaus, David M.; Benoit, Michael R.; Nelson, Emily S.; Hammond, Timmothy G.

    2004-01-01

    Conducting biological research in space requires consideration be given to isolating appropriate control parameters. For in vitro cell cultures, numerous environmental factors can adversely affect data interpretation. A biological response attributed to microgravity can, in theory, be explicitly correlated to a specific lack of weight or gravity-driven motion occurring to, within or around a cell. Weight can be broken down to include the formation of hydrostatic gradients, structural load (stress) or physical deformation (strain). Gravitationally induced motion within or near individual cells in a fluid includes sedimentation (or buoyancy) of the cell and associated shear forces, displacement of cytoskeleton or organelles, and factors associated with intra- or extracellular mass transport. Finally, and of particular importance for cell culture experiments, the collective effects of gravity must be considered for the overall system consisting of the cells, their environment and the device in which they are contained. This does not, however, rule out other confounding variables such as launch acceleration, on orbit vibration, transient acceleration impulses or radiation, which can be isolated using onboard centrifuges or vibration isolation techniques. A framework is offered for characterizing specific cause-and-effect relationships for gravity-dependent responses as a function of the above parameters.

  14. Comparison of various staining methods for the detection of Cryptosporidium in cell-free culture.

    Science.gov (United States)

    Boxell, Annika; Hijjawi, Nawal; Monis, Paul; Ryan, Una

    2008-09-01

    The complete development of Cryptosporidium in host cell-free medium first described in 2004, represented a significant advance that can facilitate many aspects of Cryptosporidium research. A current limitation of host cell-free cultivation is the difficulty involved in visualising the life-cycle stages as they are very small in size, morphologically difficult to identify and dispersed throughout the media. This is in contrast to conventional cell culture methods for Cryptosporidium, where it is possible to focus on the host cells and view the foci of infection on the host cells. In the present study, we compared three specific and three non-specific techniques for visualising Cryptosporidium parvum life-cycle stages in cell-free culture; antibody staining using anti-sporozoite and anti-oocyst wall antibodies (Sporo-Glo and Crypto Cel), fluorescent in-situ hybridization (FISH) using a Cryptosporidium specific rRNA oligonucleotide probe and the non-specific dyes; Texas Red, carboxyfluorescein diacetate succinimidyl ester (CFSE) and 4,6' diamino-2-phenylindole dihydrochloride (DAPI). Results revealed that a combination of Sporo-Glo and Crypto Cel staining resulted in easy and reliable identification of all life-cycle stages. PMID:18547565

  15. Best Viral Elution Method Available for Quantification of Enteroviruses in Sludge by Both Cell Culture and Reverse Transcription-PCR

    OpenAIRE

    Monpoeho, S.; Maul, A.; Mignotte-Cadiergues, B.; Schwartzbrod, L; Billaudel, S.; Ferré, V.

    2001-01-01

    The aim of this study was to select one or several virus extraction techniques that enable simultaneous detection of enterovirus genomes and infectious particles in different types of urban sludge. Eight techniques were compared by using 16 different liquid and solid sludge samples. The numbers of infectious enteroviruses in cell cultures were determined by using the most-probable-number method. The enterovirus genome was quantified by a single-tube reverse transcription-PCR using TaqMan tech...

  16. Optimization of Storage Temperature for Cultured ARPE-19 Cells

    Directory of Open Access Journals (Sweden)

    Lara Pasovic

    2013-01-01

    Full Text Available Purpose. The establishment of future retinal pigment epithelium (RPE replacement therapy is partly dependent on the availability of tissue-engineered RPE cells, which may be enhanced by the development of suitable storage methods for RPE. This study investigates the effect of different storage temperatures on the viability, morphology, and phenotype of cultured RPE. Methods. ARPE-19 cells were cultured under standard conditions and stored in HEPES-buffered MEM at nine temperatures (4°C, 8°C, 12°C, 16°C, 20°C, 24°C, 28°C, 32°C, and 37°C for seven days. Viability and phenotype were assessed by a microplate fluorometer and epifluorescence microscopy, while morphology was analyzed by scanning electron microscopy. Results. The percentage of viable cells preserved after storage was highest in the 16°C group (48.7%±9.8%; P<0.01 compared to 4°C, 8°C, and 24°C–37°C; P<0.05 compared to 12°C. Ultrastructure was best preserved at 12°C, 16°C, and 20°C. Expression of actin, ZO-1, PCNA, caspase-3, and RPE65 was maintained after storage at 16°C compared to control cells that were not stored. Conclusion. Out of nine temperatures tested between 4°C and 37°C, storage at 12°C, 16°C, and 20°C was optimal for maintenance of RPE cell viability, morphology, and phenotype. The preservation of RPE cells is critically dependent on storage temperature.

  17. The use of tissue culture techniques with irradiation to improve potato resistance to late blight

    International Nuclear Information System (INIS)

    A mutation breeding program was conducted to improve potato (Solanum tuberosum) resistance to late blight disease caused by Phytophthora infestans. In vitro cultured explants from potato cvs. Draga, Diamant, Spunta were irradiated with gamma ray doses 25, 30, and 35 Gy. Growing shoots were cut and re-cultured every 2 weeks until the 4th generation (MV4) to make sure no chimeral tissues still existed in the mutant material. Plantlets were subsequently propagated to obtain enough explants for in vitro selection pressure. Around 3000 plantlets from the three cultivars were subjected to selection pressure using co-culture technique. MV4 explants were incubated in jars, containing MS medium, with mycelia of P. infestans. Surviving plantlets were propagated and re-incubated with the pathogen for three consecutive generations. Resistant plantlets were acclimatized and transferred to pots and grown under glasshouse conditions. Plants were later inoculated, at the adult stage, with sporangial suspension. Cultivar Draga produced the highest number of resistant plants. Ten plants of Draga appeared to be resistant to late blight whereas only one plant from each of the other 2 cultivars was resistant. Mutant plants varied in number of produced minitubers from 13 to 70, Also, weight of these minitubers varied from less than 1 to 35 grams. Selected mutant lines will undergo further testing under field conditions for P. infestans resistance and other agronomic characteristics. (author)

  18. 紫背菜水培管理技术%Hydroponic Culture Techniques of Gynura Bicolor

    Institute of Scientific and Technical Information of China (English)

    罗思良; 潘廷由; 周连芳

    2012-01-01

    水培是无土栽培中的一种,即以营养液代替土壤,人为提供作物生长发育所需的养分、氧气、水分和热量。从育苗、消毒、定植、水肥调控、采收等方面介绍了紫背菜水培的管理技术,以期为蔬菜水培生产提供技术参考。%Hydroponic is a subset of soilless culture and is a method to artificially grow plants by nutrient solutions instead of soil. Hydroponics culture can provide the nutrient,oxygen, water and heat, which meet the require of plants' growth and development. The hydroponics culture techniques of Gynura bicolor were introduced from seedling,disinfecting,planting,water and fertilizer regulation harvesting,etc,so as to provide technical references for vegetable production by hydroponics.

  19. Mapping air pollution using Earth observation techniques for cultural heritage sites

    Science.gov (United States)

    Agapiou, Athos; Nisantzi, Argyro; Lysandrou, Vasiliki; Mamouri, Rodanthi; Alexakis, Dimitrios D.; Themistocleous, Kyriacos; Sarris, Apostolos; Hadjimitsis, Diofantos G.

    2013-08-01

    Air pollutants, together with climatic parameters, are of major importance for the deterioration of cultural heritage monuments. Atmospheric pollution is widely recognized as one of the major anthropogenic threats to architectural cultural heritage, in particular when associated with water absorption phenomena. Atmospheric particle deposition on surfaces of Monuments (of cultural heritage interest) may cause an aesthetic impact induced by a series of chemical reactions. Therefore there is a need for systematic monitoring and mapping of air pollution for areas where important archaeological sites and monuments are found. observation techniques, such as the use of satellite image for the retrieval of Aerosol Optical Thickness (AOT), are ideal for this purpose. In this paper, all important monuments of the Paphos District, listed by the Department of Antiquities of Cyprus, have been mapped using Geographical Information Systems. Several recent (2012) MODIS satellite images (both Aqua and Terra) have been used to extract the AOT values in this area. Multi-temporal analysis was performed to identify areas of high risk where AOT values are considered to be high. In situ observations have been also carried out to verify the results.

  20. Responsiveness of fetal rat brain cells to glia maturation factor during neoplastic transformation in cell culture

    DEFF Research Database (Denmark)

    Haugen, A; Laerum, O D; Bock, E

    1981-01-01

    The effect of partially purified extracts from adult pig brains containing a glia maturation protein factor (BE) has been investigated on neural cells during carcinogenesis. Pregnant BD IX-rats were given a single transplacental dose of the carcinogen ethylnitrosourea (EtNU) on the 18th day of...... gestation. The brains of the treated fetuses were transferred to cell culture and underwent neoplastic transformation with a characteristic sequence of phenotypic alterations which could be divided into five different stages. During the first 40 days after explantation (stage I & II) BE induced...... appreciable effect on GFA-content was seen any longer, although some few weakly GFA positive cells could be observed in all permanent cell lines. Fetal rat brain cells therefore seem to become less responsive to this differentiation inducer during neoplastic transformation in cell culture....