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Sample records for cell culture medium

  1. Isolated Cells of Porphyra yezoensis Cultured on Solid Medium

    Institute of Scientific and Technical Information of China (English)

    沈颂东; 戴继勋

    2001-01-01

    Vegetative cells of Porphyra yezoensis are isolated with sea snail enzyme and cultured on the solidified agar medium. The results of experiments show that the isolated cells can survive,divide and regenerate well on the medium solidified with agar. The first division on the solid medium starts after 7 days' culture, 4 days later than the liquid culture. The survival rate of isolated cells is 71.3% on the solid medium, lower than the 86.2% of that in seawater.Thalli, thalloids,conchocelis, spermatangia and multicellular masses are developed on the solid/medium in the first month, slowly but normally. Spermatangia sacs disappear within 4 weeks. Without adding nutrient liquid onto the surface of solid medium or injecting seawater under the agar layer in order to keep moisture, the thalli and cell groups release monospores to form new thalli instead of enlarging their areas after 5 weeks' culturing. Some monospores regenerate new thalli. Other monospores lose their pigments and minimize their volume and divide quickly to form light pink calli. After 16 weeks, numerous calli can be seen on the solid medium and after 24 weeks' culturing, almost only calli and conchocelis can be seen. If the calli are immersed in seawater, the monospores are released and may develop into young thallus.

  2. The Stimulatory Effect of Notochordal-Cell Conditioned Medium in a Nucleus Pulposus Explant Culture

    NARCIS (Netherlands)

    de Vries, Stefan; Doeselaar, Marina van; Meij, Björn; Tryfonidou, M; Ito, Keita

    2015-01-01

    OBJECTIVES: Notochordal cell-conditioned medium (NCCM) has previously shown to have a stimulatory effect on nucleus pulposus cells (NPCs) and bone marrow stromal cells (BMSCs) in alginate and pellet cultures. These culture methods provide a different environment than the nucleus pulposus (NP) tissue

  3. Flux characteristics of cell culture medium in rectangular microchannels.

    Science.gov (United States)

    Feng, Zhonggang; Fukuda, Shuhei; Yokoyama, Michio; Kitajima, Tatsuo; Nakamura, Takao; Umezu, Mitsuo

    2011-09-01

    Rectangular microchannels 50 μm high and 30, 40, 50, 60, or 70 μm wide were fabricated by adjusting the width of a gap cut in a polyethylene sheet 50 μm thick and sandwiching the sheet between an acrylic plate and a glass plate. Flux in the microchannels was measured under three different inner surface conditions: uncoated, albumin-coated, and confluent growth of rat fibroblasts on the bottom of the microchannels. The normalized flux in microchannels with cultured fibroblasts or albumin coating was significantly larger than that in the uncoated channels. The experimental data for all microchannels deviated from that predicted by classical hydrodynamic theory. At small aspect ratio the flux in the microchannels was larger than that predicted theoretically, whereas it became smaller at large aspect ratio. The aspect ratio rather than Reynolds number is the correct property for predicting the variation of the normalized friction factor. We postulate that two counteracting effects, rotation of large molecules and slip velocity at the corners of the microchannels, are responsible for the deviation. From these results we conclude that albumin coating should be carried out in the same way as when fabricating our integrating cell-culture system. The outcomes of this study are not only important for the design of our culture system, but also quite informative for general microfluidics.

  4. Aquatic flower-inspired cell culture platform with simplified medium exchange process for facilitating cell-surface interaction studies.

    Science.gov (United States)

    Hong, Hyeonjun; Park, Sung Jea; Han, Seon Jin; Lim, Jiwon; Kim, Dong Sung

    2016-02-01

    Establishing fundamentals for regulating cell behavior with engineered physical environments, such as topography and stiffness, requires a large number of cell culture experiments. However, cell culture experiments in cell-surface interaction studies are generally labor-intensive and time-consuming due to many experimental tasks, such as multiple fabrication processes in sample preparation and repetitive medium exchange in cell culture. In this work, a novel aquatic flower-inspired cell culture platform (AFIP) is presented. AFIP aims to facilitate the experiments on the cell-surface interaction studies, especially the medium exchange process. AFIP was devised to capture and dispense cell culture medium based on interactions between an elastic polymer substrate and a liquid medium. Thus, the medium exchange can be performed easily and without the need of other instruments, such as a vacuum suction and pipette. An appropriate design window of AFIP, based on scaling analysis, was identified to provide a criterion for achieving stability in medium exchange as well as various surface characteristics of the petal substrates. The developed AFIP, with physically engineered petal substrates, was also verified to exchange medium reliably and repeatedly. A closed structure capturing the medium was sustained stably during cell culture experiments. NIH3T3 proliferation results also demonstrated that AFIP can be applied to the cell-surface interaction studies as an alternative to the conventional method. PMID:26683462

  5. Application of Serum-Free Culture Medium for Preparation of A-NK Cells

    Institute of Scientific and Technical Information of China (English)

    Zhihua Wang; Zhibin Zhang; Hui Zhang; Yan Zhang

    2006-01-01

    To compare the differences between proliferation and cytotoxicity of adherent natural killer (A-NK) cells cultured with serum-free medium AIMV and standard serum-containing medium in vitro, and also observe the assisting effect of IL-12 on the activation and the morphology character of IL-2-treated A-NK cells, cellular proliferation was evaluated by MTT method in vitro. The morphology of the target cells killed by A-NK cells was observed through electroscope. All of the A-NK cells cultured in serum-free medium AIMV could rapidly proliferate and keep high cytotoxicity compared with that in standard serum-containing medium. A-NK cells activated by both moderate-dose IL-2 and IL-12 were superior to the high-dose IL-2-treated A-NK cells. These data indicated that serum-free medium AIMV could replace standard serum-containing medium for culturing A-NK cells, and moderate-dose IL-2 and IL-12 could reduce side effects caused by high-dose IL-2. The study provided a new experimental basis for experimental and clinical preparation of A-NK cells.

  6. Dissolved oxygen concentration in the medium during cell culture: Defects and improvements.

    Science.gov (United States)

    Zhang, Kuan; Zhao, Tong; Huang, Xin; He, Yunlin; Zhou, Yanzhao; Wu, Liying; Wu, Kuiwu; Fan, Ming; Zhu, Lingling

    2016-03-01

    In vitro cell culture has provided a useful model to study the effects of oxygen on cellular behavior. However, it remains unknown whether the in vitro operations themselves affect the medium oxygen levels and the living states of cells. In addition, a prevailing controversy is whether reactive oxygen species (ROS) production is induced by continuous hypoxia or reoxygenation. In this study, we have measured the effects of different types of cell culture containers and the oxygen environment where medium replacement takes place on the actual oxygen tension in the medium. We found that the deviations of oxygen concentrations in the medium are much greater in 25-cm(2) flasks than in 24-well plates and 35-mm dishes. The dissolved oxygen concentrations in the medium were increased after medium replacement in normoxia, but remained unchanged in glove boxes in which the oxygen tension remained at a low level (11.4, 5.7, and 0.5% O2 ). We also found that medium replacement in normoxia increased the number of ROS-positive cells and reduced the cell viability; meanwhile, medium replacement in a glove box did not produce the above effects. Therefore, we conclude that the use of 25-cm(2) flasks should be avoided and demonstrate that continuous hypoxia does not produce ROS, whereas the reoxygenation that occurs during the harvesting of cells leads to ROS and induces cell death.

  7. Enhanced growth medium and method for culturing human mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R. (7290 Sayre Dr., Oakland, CA 94611); Smith, Helene S. (5693 Cabot Dr., Oakland, CA 94611); Hackett, Adeline J. (82 Evergreen Dr., Orinda, CA 94563)

    1983-01-01

    Methods are disclosed for isolating and culturing human mammary epithelial cells of both normal and malignant origin. Tissue samples are digested with a mixture including the enzymes collagenase and hyaluronidase to produce clumps of cells substantially free from stroma and other undesired cellular material. Growing the clumps of cells in mass culture in an enriched medium containing particular growth factors allows for active cell proliferation and subculture. Clonal culture having plating efficiencies of up to 40% or greater may be obtained using individual cells derived from the mass culture by plating the cells on appropriate substrates in the enriched media. The clonal growth of cells so obtained is suitable for a quantitative assessment of the cytotoxicity of particular treatment. An exemplary assay for assessing the cytotoxicity of the drug adriamycin is presented.

  8. Effects of Supplementation of Various Medium Components on Chinese Hamster Ovary Cell Cultures Producing Recombinant Antibody

    OpenAIRE

    Kim, Do Yun; Lee, Joon Chul; Chang, Ho Nam; Oh, Duk Jae

    2005-01-01

    Thirteen vitamins, twenty amino acids, hormones, inorganic salts, and other chemical agents, which constitute typical serum-free media, were evaluated for the development of fortified medium to enhance cell growth and productivity of recombinant antibody in the cultures of the recombinant Chinese hamster ovary (rCHO) cells. Two different rCHO cell lines, rCHO-A producing recombinant antibodies against the human platelet and rCHO-B secreting recombinant antibodies against the S surface antigen...

  9. Silver nanoparticles inhibit fish gill cell proliferation in protein-free culture medium.

    Science.gov (United States)

    Yue, Yang; Behra, Renata; Sigg, Laura; Schirmer, Kristin

    2016-10-01

    While short-term exposures of vertebrate cells, such as from fish, can be performed in defined, serum-free media, long-term cultures generally require addition of growth factors and proteins, normally supplied with a serum supplement. However, proteins are known to alter nanoparticle properties by binding to nanoparticles. Therefore, in order to be able to study nanoparticle-cell interactions for extended periods, the rainbow trout (Oncorhynchus mykiss) gill cell line, RTgill-W1, was adapted to proliferate in a commercial, serum-free medium, InVitrus VP-6. The newly adapted cell strain was named RTgill-W1-pf (protein free). These cells proliferate at a speed similar to the RTgill-W1 cells cultured in a fully supplemented medium containing 5% fetal bovine serum. As well, they were successfully cryopreserved in liquid nitrogen and fully recovered after thawing. Yet, senescence set in after about 10 passages in InVitrus VP-6 medium, revealing that this medium cannot fully support long-term culture of the RTgill-W1 strain. The RTgill-W1-pf cell line was subsequently applied to investigate the effect of silver nanoparticles (AgNP) on cell proliferation over a period of 12 days. Indeed, cell proliferation was inhibited by 10 μM AgNP. This effect correlated with high levels of silver being associated with the cells. The new cell line, RTgill-W1-pf, can serve as a unique representation of the gill cell-environment interface, offering novel opportunities to study nanoparticle-cell interactions without serum protein interference. PMID:27030289

  10. Silver nanoparticles inhibit fish gill cell proliferation in protein-free culture medium.

    Science.gov (United States)

    Yue, Yang; Behra, Renata; Sigg, Laura; Schirmer, Kristin

    2016-10-01

    While short-term exposures of vertebrate cells, such as from fish, can be performed in defined, serum-free media, long-term cultures generally require addition of growth factors and proteins, normally supplied with a serum supplement. However, proteins are known to alter nanoparticle properties by binding to nanoparticles. Therefore, in order to be able to study nanoparticle-cell interactions for extended periods, the rainbow trout (Oncorhynchus mykiss) gill cell line, RTgill-W1, was adapted to proliferate in a commercial, serum-free medium, InVitrus VP-6. The newly adapted cell strain was named RTgill-W1-pf (protein free). These cells proliferate at a speed similar to the RTgill-W1 cells cultured in a fully supplemented medium containing 5% fetal bovine serum. As well, they were successfully cryopreserved in liquid nitrogen and fully recovered after thawing. Yet, senescence set in after about 10 passages in InVitrus VP-6 medium, revealing that this medium cannot fully support long-term culture of the RTgill-W1 strain. The RTgill-W1-pf cell line was subsequently applied to investigate the effect of silver nanoparticles (AgNP) on cell proliferation over a period of 12 days. Indeed, cell proliferation was inhibited by 10 μM AgNP. This effect correlated with high levels of silver being associated with the cells. The new cell line, RTgill-W1-pf, can serve as a unique representation of the gill cell-environment interface, offering novel opportunities to study nanoparticle-cell interactions without serum protein interference.

  11. Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells.

    Science.gov (United States)

    Martin, G R

    1981-12-01

    This report describes the establishment directly from normal preimplantation mouse embryos of a cell line that forms teratocarcinomas when injected into mice. The pluripotency of these embryonic stem cells was demonstrated conclusively by the observation that subclonal cultures, derived from isolated single cells, can differentiate into a wide variety of cell types. Such embryonic stem cells were isolated from inner cell masses of late blastocysts cultured in medium conditioned by an established teratocarcinoma stem cell line. This suggests that such conditioned medium might contain a growth factor that stimulates the proliferation or inhibits the differentiation of normal pluripotent embryonic cells, or both. This method of obtaining embryonic stem cells makes feasible the isolation of pluripotent cells lines from various types of noninbred embryo, including those carrying mutant genes. The availability of such cell lines should made possible new approaches to the study of early mammalian development.

  12. Stem cells from human exfoliated deciduous teeth differentiate toward neural cells in a medium dynamically cultured with Schwann cells in a series of polydimethylsiloxanes scaffolds

    Science.gov (United States)

    Su, Wen-Ta; Pan, Yu-Jing

    2016-08-01

    Objective. Schwann cells (SCs) are primary structural and functional cells in the peripheral nervous system. These cells play a crucial role in peripheral nerve regeneration by releasing neurotrophic factors. This study evaluated the neural differentiation potential effects of stem cells from human exfoliated deciduous teeth (SHEDs) in a rat Schwann cell (RSC) culture medium. Approach. SHEDs and RSCs were individually cultured on a polydimethylsiloxane (PDMS) scaffold, and the effects of the RSC medium on the SHEDs differentiation between static and dynamic cultures were compared. Main results. Results demonstrated that the SHED cells differentiated by the RSC cultured medium in the static culture formed neurospheres after 7 days at the earliest, and SHED cells formed neurospheres within 3 days in the dynamic culture. These results confirm that the RSC culture medium can induce neurospheres formation, the speed of formation and the number of neurospheres (19.16 folds high) in a dynamic culture was superior to the static culture for 3 days culture. The SHED-derived spheres were further incubated in the RSCs culture medium, these neurospheres continuously differentiated into neurons and neuroglial cells. Immunofluorescent staining and RT-PCR revealed nestin, β-III tubulin, GFAP, and γ-enolase of neural markers on the differentiated cells. Significance. These results indicated that the RSC culture medium can induce the neural differentiation of SHED cells, and can be used as a new therapeutic tool to repair nerve damage.

  13. Comparative evaluation of cytotoxicity of cadmium in rat liver cells cultured in serum-containing medium and commercially available serum-free medium.

    Science.gov (United States)

    Latinwo, Lekan M; Badisa, Veera L D; Odewumi, Caroline O; Ikediobi, Christopher O; Badisa, Ramesh B; Brooks-Walter, Alexis; Lambert, Ayuk-Takem T; Nwoga, Jude

    2008-07-01

    Cadmium (Cd) is an industrial pollutant and carcinogenic metal. Most in vitro Cd toxicity studies have been carried out in various cell lines cultured in 10% fetal bovine serum (FBS) containing medium. In this report, we compared the toxic effect of Cd (0-300 microM) on cell growth, total RNA, total proteins, and antioxidant enzymes in rat normal liver cells cultured in medium with 10% FBS or commercially available serum-free medium for 4 or 8 hours. With Cd concentration at above 100 microM, the total levels of RNA, protein and cell growth decreased in serum-containing medium, while their levels increased in serum-free medium compared to the controls. The glutathione peroxidase and glutathione reductase levels were lower in serum-free medium than in serum-containing medium, indicating less oxidative stress in cells grown in serum-free medium. These results clearly suggest that Cd showed higher toxicity to liver cells grown in serum-containing medium in comparison to commercially available serum-free medium. It is speculated that albumin and other substances present in commercial serum-free medium chelated Cd and thereby protected these cells against Cd toxicity. Even under in vivo conditions, cadmium enters into various organs after passing through blood which contains serum. Based on these studies, it appears that media containing serum may be ideal for in vivo toxicity correlation studies with animal cells.

  14. Growth factor-defined culture medium for human mesenchymal stem cells.

    Science.gov (United States)

    Mimura, Sumiyo; Kimura, Naohiro; Hirata, Mitsuhi; Tateyama, Daiki; Hayashida, Midori; Umezawa, Akihiro; Kohara, Arihiro; Nikawa, Hiroki; Okamoto, Tetsuji; Furue, Miho K

    2011-01-01

    Human bone marrow-derived mesenchymal stem cells (hMSCs) are potential cellular sources of therapeutic stem cells as they have the ability to proliferate and differentiate into a wide array of mesenchymal cell types such as osteoblasts, chondroblasts and adipocytes. hMSCs have been used clinically to treat patients with graft vs. host disease, osteogenesis imperfect, or alveolar cleft, suggesting that transplantation of hMSCs is comparatively safe as a stem cell-based therapy. However, conventional culture medium for hMSCs contains fetal bovine serum (FBS). In the present study, we developed a growth factor-defined, serum-free medium for culturing hMSCs. Under these conditions, TGF-beta1 promoted proliferation of hMSCs. The expanded hMSC population expressed the human pluripotency markers SSEA-3, -4, NANOG, OCT3/4 and SOX2. Furthermore, double positive cells for SSEA-3 and a mesenchymal cell marker, CD105, were detected in the population. The potential to differentiate into osteoblasts and adipocytes was confirmed. This work provides a useful tool to understand the basic biological properties of hMSCs in culture. PMID:21305471

  15. Human dental pulp stem cells cultured in serum-free supplemented medium

    Directory of Open Access Journals (Sweden)

    Virginie eBonnamain

    2013-12-01

    Full Text Available Growing evidence show that human dental pulp stem cells (DPSCs could provide a source of adult stem cells for the treatment of neurodegenerative pathologies. In this study, DPSCs were expanded and cultured with a protocol generally used for the culture of neural stem/progenitor cells.Methodology: DPSC cultures were established from third molars. The pulp tissue was enzymatically digested and cultured in serum-supplemented basal medium for 12 hours. Adherent (ADH and non-adherent (non-ADH cell populations were separated according to their differential adhesion to plastic and then cultured in serum-free defined N2 medium with epidermal growth factor (EGF and basic fibroblast growth factor (bFGF. Both ADH and non-ADH populations were analyzed by FACS and/or PCR.Results: FACS analysis of ADH-DPSCs revealed the expression of the mesenchymal cell marker CD90, the neuronal marker CD56, the transferrin receptor CD71, and the chemokine receptor CXCR3, whereas hematopoietic stem cells markers CD45, CD133 and CD34 were not expressed. ADH-DPSCs expressed transcripts coding for the Nestin gene, whereas expression levels of genes coding for the neuronal markers β-III tubulin and NF-M, and the oligodendrocyte marker PLP-1 were donor dependent. ADH-DPSCs did not express the transcripts for GFAP, an astrocyte marker. Cells of the non-ADH population that grew as spheroids expressed Nestin, β-III tubulin, NF-M and PLP-1 transcripts. DPSCs migrated out of the spheroids exhibited an odontoblast-like morphology and expressed a higher level of DSPP and osteocalcin transcripts than ADH-DPSCs. Conclusion: Collectively, these data indicate that human DPSCs can be expended and cultured in serum-free supplemented medium with EGF and bFGF. ADH-DPSCs and non-ADH populations contained neuronal and/or oligodendrocyte precursors at different stages of commitment and interestingly, cells from spheroid structures seem to be more engaged into the odontoblastic lineage than the

  16. Effects of medium nutrition on cell growth and isocamptothecin A and B production by suspension cell culture of Camptotheca acuminata

    Institute of Scientific and Technical Information of China (English)

    Zhang Dongyan; Yu Fang; Bai Fengwu; An Lijia

    2006-01-01

    The effects of initial sucrose concentration, nitrate to ammonium ratio, total N concentration and phosphate concentration in medium on cell growth and isocamptothecin A and B synthesis by suspension cell culture of Camptotheca acuminata were investigated in 250 mL shake flasks. 30 g L-1 sucrose concentration was beneficial to secondary metabolites synthesis. The cell growth and metabolites synthesis were also affected by the ratio of NO-3/NH+4, and nitrate was favourable for cell growth. The maximum dry weight was achieved when nitrate was used as the sole N source. The effect of total initial N on the cell cultures was also investigated with NO-3/NH+4 ratio of 1∶2. The final dry cell weight was similar throughout culture period and 50 mM initial N was favourable for secondary metabolite synthesis. 50 mM initial phosphate concentration facilitated both cell growth and secondary metabolites synthesis.

  17. Agglomeration, sedimentation, and cellular toxicity of alumina nanoparticles in cell culture medium

    International Nuclear Information System (INIS)

    The cytotoxicity of alumina nanoparticles (NPs) was investigated for a wide range of concentration (25–200 μg/mL) and incubation time (0–72 h) using floating cells (THP-1) and adherent cells (J774A.1, A549, and 293). Alumina NPs were gradually agglomerated over time although a significant portion of sedimentation occurred at the early stage within 6 h. A decrease of the viability was found in floating (THP-1) and adherent (J774A.1 and A549) cells in a dose-dependent manner. However, the time-dependent decrease in cell viability was observed only in adherent cells (J774A.1 and A549), which is predominantly related with the sedimentation of alumina NPs in cell culture medium. The uptake of alumina NPs in macrophages and an increased cell-to-cell adhesion in adherent cells were observed. There was no significant change in the viability of 293 cells. This in vitro test suggests that the agglomeration and sedimentation of alumina NPs affected cellular viability depending on cell types such as monocytes (THP-1), macrophages (J774A.1), lung carcinoma cells (A549), and embryonic kidney cells (293).

  18. HepG2 cells develop signs of riboflavin deficiency within four days of culture in riboflavin-deficient medium*

    OpenAIRE

    Werner, Ricarda; MANTHEY, KAROLINE C.; Griffin, Jacob B.; Zempleni, Janos

    2005-01-01

    Flavin mononucleotide and flavin adenine dinucleotide are essential coenzymes in redox reactions. For example, flavin adenine dinucleotide is a coenzyme for both glutathione reductase and enzymes that mediate the oxidative folding of secretory proteins. Here we investigated short-term effects of moderately riboflavin-deficient culture medium on flavin-related responses in HepG2 hepatocarcinoma cells. Cells were cultured in riboflavin-deficient (3.1 nmol/L) medium for up to six days; controls ...

  19. Culture medium modulates the behaviour of human dental pulp-derived cells: Technical Note

    Directory of Open Access Journals (Sweden)

    S Lopez-Cazaux

    2006-02-01

    Full Text Available In vitro approaches have extensively been developed to study reparative dentinogenesis. While dental pulp is a source of unidentified progenitors able to differentiate into odontoblast-like cells, we investigated the effect of two media; MEM (1.8mM Ca and 1mM Pi and RPMI 1640 (0.8mM Ca and 5mM Pi on the behaviour of human dental pulp cells. Our data indicate that MEM significantly increased cell proliferation and markedly enhanced the proportion of -smooth muscle actin positive cells, which represent a putative source of progenitors able to give rise to odontoblast-like cells. In addition, MEM strongly stimulated alkaline phosphatase activity and was found to induce expression of transcripts encoding dentin sialophosphoprotein, an odontoblastic marker, without affecting that of parathyroid hormone/parathyroid hormone related protein-receptor and osteonectin. In conclusion, these observations demonstrate that not only proliferation but also differentiation into odontoblast-like cells was induced by rich calcium and poor phosphate medium (MEM as compared to RPMI 1640. This study provides important data for the determination of the optimal culture conditions allowing odontoblast-like differentiation in human pulp cell culture.

  20. A chemically defined culture medium containing Rho kinase inhibitor Y-27632 for the fabrication of stratified squamous epithelial cell grafts

    Energy Technology Data Exchange (ETDEWEB)

    Aslanova, Afag [Department of Surgery, Institute of Gastroenterology, Tokyo Women' s Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666 (Japan); Institute of Advanced Biomedical Engineering and Science, Tokyo Women' s Medical University, TWIns, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666 (Japan); Takagi, Ryo; Yamato, Masayuki; Okano, Teruo [Institute of Advanced Biomedical Engineering and Science, Tokyo Women' s Medical University, TWIns, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666 (Japan); Yamamoto, Masakazu, E-mail: yamamoto.ige@twmu.ac.jp [Department of Surgery, Institute of Gastroenterology, Tokyo Women' s Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666 (Japan)

    2015-05-01

    With the development of a culture method for stratified squamous epithelial cells, tissue-engineered epithelial cell sheets have been successfully applied as clinical cell grafts. However, the implementation of these cell sheets without the use of any animal-derived materials is highly desirable. In this study, Rho-associated protein kinase inhibitor Y-27632 was used to develop a chemically defined culture medium for the fabrication of stratified epithelial cell grafts consisting of human epidermal and oral keratinocytes, and the proliferation activity, cell morphology, and gene expressions of the keratinocytes were analyzed. The results of a colorimetric assay indicated that Y-27632 significantly promoted the proliferation of the keratinocytes in culture media both with and without fetal bovine serum (FBS), although there were no indications of Y-27632 efficacy on cell morphology and stratification of the keratinocytes in culture medium without any animal-derived materials. The results of quantitative RT-PCR revealed that gene expressions correlated with cell adhesion, cell–cell junction, proliferation markers, and stem/progenitor markers in cultured keratinocytes were not strongly affected by the addition of Y-27632 to the culture medium. Moreover, gene expressions of differentiation markers in stratified keratinocytes cultured in medium without FBS were nearly identical to those of keratinocytes co-cultured with 3T3 feeder cells. Interestingly, the expressions of differentiation markers in cultured stratified keratinocytes were suppressed by FBS, whereas they were reconstructed by either co-culture of a 3T3 feeder layer or addition of Y-27632 into the culture medium containing FBS. These findings indicate that Y-27632 is a useful supplement for the development of a chemically defined culture medium for fabrication of stratified epithelial cell grafts for clinical applications for the purpose of developing the culture medium with a lower risk of pathogen

  1. A chemically defined culture medium containing Rho kinase inhibitor Y-27632 for the fabrication of stratified squamous epithelial cell grafts

    International Nuclear Information System (INIS)

    With the development of a culture method for stratified squamous epithelial cells, tissue-engineered epithelial cell sheets have been successfully applied as clinical cell grafts. However, the implementation of these cell sheets without the use of any animal-derived materials is highly desirable. In this study, Rho-associated protein kinase inhibitor Y-27632 was used to develop a chemically defined culture medium for the fabrication of stratified epithelial cell grafts consisting of human epidermal and oral keratinocytes, and the proliferation activity, cell morphology, and gene expressions of the keratinocytes were analyzed. The results of a colorimetric assay indicated that Y-27632 significantly promoted the proliferation of the keratinocytes in culture media both with and without fetal bovine serum (FBS), although there were no indications of Y-27632 efficacy on cell morphology and stratification of the keratinocytes in culture medium without any animal-derived materials. The results of quantitative RT-PCR revealed that gene expressions correlated with cell adhesion, cell–cell junction, proliferation markers, and stem/progenitor markers in cultured keratinocytes were not strongly affected by the addition of Y-27632 to the culture medium. Moreover, gene expressions of differentiation markers in stratified keratinocytes cultured in medium without FBS were nearly identical to those of keratinocytes co-cultured with 3T3 feeder cells. Interestingly, the expressions of differentiation markers in cultured stratified keratinocytes were suppressed by FBS, whereas they were reconstructed by either co-culture of a 3T3 feeder layer or addition of Y-27632 into the culture medium containing FBS. These findings indicate that Y-27632 is a useful supplement for the development of a chemically defined culture medium for fabrication of stratified epithelial cell grafts for clinical applications for the purpose of developing the culture medium with a lower risk of pathogen

  2. Calcification in human osteoblasts cultured in medium conditioned by the prostatic cancer cell line PC-3 and prostatic acid phosphatase.

    Science.gov (United States)

    Kimura, G; Sugisaki, Y; Masugi, Y; Nakazawa, N

    1992-01-01

    A medium that had been conditioned by PC-3 cells stimulated the calcification of a human osteoblastic cell line, Tak-10, in a nonmitogenic culture. The calcification of the osteoblasts was stimulated maximally at a 25% concentration of the conditioned medium. Calcification activity was markedly enhanced by the addition of both prostatic acid phosphatase (PAP) and its substrate, alpha-glycerophosphate, to the medium; however, PAP added alone did not enhance this activity. These results suggest that human prostatic carcinoma cells produce a factor that stimulates the calcification of the human osteoblasts. Results have also suggested that PAP is a requisite for osteogenesis provided that its substrates are abundant in the medium.

  3. Performance evaluation of CHO-K1 cell in culture medium supplemented with hemolymph

    Directory of Open Access Journals (Sweden)

    Tássia Raffoul

    2005-06-01

    Full Text Available The aim of this work was to evaluate the potential of hemolymph utilization as a culture medium supplement to cultivate the animal cell CHO-K1. For this purpose 1% v/v of hemolymph was added to DMEM medium containing 10% v/v of FBS and 1 or 4.5 g/L of glucose. The culture was grown in spinner flasks incubated in a 10% v/v CO2 environment, at 37ºC, with the Cytodex 1 microcarrier. Comparing the results obtained from the culture with hemolymph against those without hemolymph, a positive influence of the hemolymph was observed, as the experiment with hemolymph presented a 52% higher cell concentration and a higher productivity of up to 40%.Desenvolvimento de meios de cultura isentos de soro fetal bovino (SFB é uma das grandes prioridades de pesquisa em desenvolvimento de processos com célula animal. O objetivo do presente trabalho foi realizar uma análise do potencial de uso da hemolinfa como suplemento do meio utilizado no cultivo da célula animal ancorante CHO-K1. Para isso, foi adicionado 1% v/v de extrato de hemolinfa ao meio DMEM contendo 10% v/v de SFB e 1,0 ou 4,5 g/L de glicose. O cultivo foi realizado em frascos tipo spinner em um ambiente de 10% v/v de CO2, a 37ºC, utilizando o microcarregador Cytodex 1. Comparando os resultados obtidos no ensaio com hemolinfa com um sem hemolinfa pode-se notar uma influência positiva da hemolinfa no cultivo, já que o ensaio com hemolinfa apresentou uma concentração máxima de células 52% maior e uma produtividade máxima de até 40% maior.

  4. Design of serum-free medium for suspension culture of CHO cells on the basis of general commercial media.

    Science.gov (United States)

    Miki, Hideo; Takagi, Mutsumi

    2015-08-01

    The design of serum-free media for suspension culture of genetically engineered Chinese hamster ovary (CHO) cells using general commercial media as a basis was investigated. Subcultivation using a commercial serum-free medium containing insulin-like growth factor (IGF)-1 with or without FCS necessitated additives other than IGF-1 to compensate for the lack of FCS and improve cell growth. Suspension culture with media containing several combinations of growth factors suggested the effectiveness of addition of both IGF-1 and the lipid signaling molecule lysophosphatidic acid (LPA) for promoting cell growth. Subcultivation of CHO cells in suspension culture using the commercial serum-free medium EX-CELL™302, which contained an IGF-1 analog, supplemented with LPA resulted in gradually increasing specific growth rate comparable to the serum-containing medium and in almost the same high antibody production regardless of the number of generations. The culture with EX-CELL™302 supplemented with LPA in a jar fermentor with pH control at 6.9 showed an apparently higher cell growth rate than the cultures without pH control and with pH control at 6.8. The cell growth in the medium supplemented with aurintricarboxylic acid (ATA), which was much cheaper than IGF-1, in combination with LPA was synergistically promoted similarly to that in the medium supplemented with IGF-1 and LPA. In conclusion, the serum-free medium designed on the basis of general commercial media could support the growth of CHO cells and antibody production comparable to serum-containing medium in suspension culture. Moreover, the possibility of cost reduction by the substitution of IGF-1 with ATA was also shown.

  5. Optimization of the basal medium for improving production and secretion of taxanes from suspension cell culture of Taxus baccata L

    Directory of Open Access Journals (Sweden)

    Kajani Abolghasem

    2012-10-01

    Full Text Available Abstract Background and purpose of the study Taxol is one of the most effective anticancer drugs that isolated from Taxus sp. due to the slow growth of Taxus trees and low concentration of Taxol in the tissues, the biotechnological approaches especially plant cell culture have been considered to produce Taxol in commercial scale. Methods We investigated the effects of basal medium type used in culture media on production of Taxol and other taxane compounds from cell suspension culture of T. baccata L. Briefly, five commonly basal media including Gamborg, Murashige and Skoog, Woody Plant, Schenk and Hildebrandt, and Driver and Kuniyuki medium were used for preparing separate suspension culture media. The intra- and extra-cellular yields of taxanes were analyzed by using HPLC after 21 days period of culturing. Results The yields of taxanes were significantly different for the cultures prepared by different basal media. Moreover, the effects of basal medium on the yield of products differed for varius taxane compounds. Maximum yields of Baccatin III (10.03 mgl-1 and 10-deacetyl baccatin III (4.2 mgl-1 were achieved from the DKW basal media, but the yield of Taxol was maximum (16.58 mgl-1 in the WPM basal media. Furthermore, the secretion of taxanes from the cells into medium was also considerably affected by the type of basal medium. The maximum extra-cellular yield of Taxol (7.81 mgl-1, Baccatin III (5.0 mgl-1, and 10-deacetyl baccatin III (1.45 mgl-1 were also obtained by using DKW basal medium that were significantly higher than those obtained from other culture media.

  6. Optimization of the basal medium for improving production and secretion of taxanes from suspension cell culture of Taxus baccata L

    Directory of Open Access Journals (Sweden)

    Abolghasem Abbasi Kajani

    2012-10-01

    Full Text Available Background and purpose of the study Taxol is one of the most effective anticancer drugs that isolated from Taxus sp. due to the slow growth of Taxus trees and low concentration of Taxol in the tissues, the biotechnological approaches especially plant cell culture have been considered to produce Taxol in commercial scale.MethodsWe investigated the effects of basal medium type used in culture media on production of Taxol and other taxane compounds from cell suspension culture of T. baccata L. Briefly, five commonly basal media including Gamborg, Murashige and Skoog, Woody Plant, Schenk and Hildebrandt, and Driver and Kuniyuki medium were used for preparing separate suspension culture media. The intra- and extra-cellular yields of taxanes were analyzed by using HPLC after 21 days period of culturing.ResultsThe yields of taxanes were significantly different for the cultures prepared by different basal media. Moreover, the effects of basal medium on the yield of products differed for varius taxane compounds. Maximum yields of Baccatin III (10.03 mgl-1 and 10-deacetyl baccatin III (4.2 mgl-1 were achieved from the DKW basal media, but the yield of Taxol was maximum (16.58 mgl-1 in the WPM basal media. Furthermore, the secretion of taxanes from the cells into medium was also considerably affected by the type of basal medium. The maximum extra-cellular yield of Taxol (7.81 mgl-1, Baccatin III (5.0 mgl-1, and 10-deacetyl baccatin III (1.45 mgl-1 were also obtained by using DKW basal medium that were significantly higher than those obtained from other culture media.

  7. Kinetics and metabolic specificities of Vero cells in bioreactor cultures with serum-free medium

    OpenAIRE

    Quesney, Sébastien; Marc, Annie; Gerdil, Catherine; Gimenez, Cyrille; Marvel, Jacqueline; Richard, Yves; Meignier, Bernard

    2003-01-01

    The aim of this study was to understand the metabolism kinetics of Vero cells grown on microcarriers in bioreactors in serum-free medium (SFM). We sought to determine what nutrients are essential for Vero cells and how they are consumed. Contrary to glucose and to most of the amino acids, glutamine and serine were very quickly depleted in this medium and can be supposed to be responsible for cell apoptosis. Lactate and ammonium ions did not reach toxic levels for Vero cells. We payed more att...

  8. Optimisation of hybridoma cell growth and monoclonal antibody secretion in a chemically defined, serum- and protein-free culture medium.

    Science.gov (United States)

    Schneider, Y J

    1989-01-01

    Monoclonal antibodies (MAbs), for human use require chemical and biological purity. The best approach seems in vitro cultivation in a serum-, protein-free medium. A basal defined culture medium has been developed to sustain optimal hybridoma cell growth and MAb secretion. It consists of Iscove's Dulbecco's modified, Eagle's, Ham's F12 and NCTC 135 media in a 5:5:1 mixture (v/v/v), to which glucose is added to reach a final concentration of 25 mM, glutamine to 4-6 mM, 2-mercaptoethanol to 50 microM, Pluronic F68 to 0.01-0.1% (w/v), Hepes to 25 mM and NaHCO3 to 3 g/l. Hybridoma cells, derived from Sp 2/0 myeloma and secreting a MAb to a human milk fat globule membrane-associated high molecular weight glycoprotein, were cloned in this medium containing 1% (v/v) fetal calf serum and then sequentially adapted in serum-free medium further supplemented with transferrin and insulin, both at 10 micrograms/ml. Clones producing immunoreactive MAbs secrete a mean of 50 micrograms IgG/ml, i.e., ca. 80% of the concentration reached in Dulbecco's modified Eagle's medium containing 10% serum. When cells were cultured in spinner flasks with a semi-continuous mode of cultivation (with a daily removal of 20% of the volume and its replacement by fresh culture medium), in serum-free medium further supplemented with 10 nM estradiol, a mixture of trace elements and albumin (at 30 micrograms/ml) complexed to linoleic acid, MAb secretion reached 100 micrograms/ml and became equal or higher to that obtained in serum-containing medium. MAb secretion was not decreased and was even significantly increased during the growth phase, when transferrin was replaced by another iron source, i.e., ferric citrate at 500 microM associated with 20 microM ascorbic acid. Finally, deletion of insulin and of albumin-linoleic acid did not affect significantly cell density nor MAb secretion. In conclusion, it appears from this study that semi-continuous cultivation in spinner flasks of hybridoma cells, after

  9. Prototypical antipsychotic drugs protect hippocampal neuronal cultures against cell death induced by growth medium deprivation

    Directory of Open Access Journals (Sweden)

    Williams Sylvain

    2006-03-01

    Full Text Available Abstract Background Several clinical studies suggested that antipsychotic-based medications could ameliorate cognitive functions impaired in certain schizophrenic patients. Accordingly, we investigated the effects of various dopaminergic receptor antagonists – including atypical antipsychotics that are prescribed for the treatment of schizophrenia – in a model of toxicity using cultured hippocampal neurons, the hippocampus being a region of particular relevance to cognition. Results Hippocampal cell death induced by deprivation of growth medium constituents was strongly blocked by drugs including antipsychotics (10-10-10-6 M that display nM affinities for D2 and/or D4 receptors (clozapine, haloperidol, (±-sulpiride, domperidone, clozapine, risperidone, chlorpromazine, (+-butaclamol and L-741,742. These effects were shared by some caspases inhibitors and were not accompanied by inhibition of reactive oxygen species. In contrast, (--raclopride and remoxipride, two drugs that preferentially bind D2 over D4 receptors were ineffective, as well as the selective D3 receptor antagonist U 99194. Interestingly, (--raclopride (10-6 M was able to block the neuroprotective effect of the atypical antipsychotic clozapine (10-6 M. Conclusion Taken together, these data suggest that D2-like receptors, particularly the D4 subtype, mediate the neuroprotective effects of antipsychotic drugs possibly through a ROS-independent, caspase-dependent mechanism.

  10. Effect of Somatic Cell Types and Culture Medium on in vitro Maturation, Fertilization and Early Development Capability of Buffalo Oocytes

    Directory of Open Access Journals (Sweden)

    H. Jamil*, H. A. Samad, N. Rehman, Z. I. Qureshi and L. A. Lodhi

    2011-04-01

    Full Text Available This study was designed to evaluate the efficacy of different somatic cell types and media in supporting in vitro maturation (IVM, in vitro fertilization (IVF and early embryonic development competence of buffalo follicular oocytes. Cumulus oocyte complexes were collected for maturation from follicles (>6mm of buffalo ovaries collected at the local abattoir. Oocytes were co-cultured in tissue culture medium (TCM-199 with either granulosa cells, cumulus cells, or buffalo oviductal epithelial cells (BOEC @ 3x106 cells/ml or in TCM-199 without helper cells (control at 39°C and 5%CO2 in humidified air. Fresh semen was prepared in modified Ca++ free Tyrode medium. Fertilization was carried out in four types of media: i Tyrode lactate albumin pyruvate (TALP, ii TALP+BOEC, iii modified Ca++ free Tyrode and iv modified Ca++ free Tyrode+BOEC. Fertilized oocytes were cultured for early embryonic development in TCM-199 with and without BOEC. Higher maturation rates were observed in the granulosa (84.24% and cumulus cells (83.44% than BOEC co culture system (73.37%. Highest fertilization rate was obtained in modified Ca++ free Tyrode with BOEC co culture (70.42%, followed by modified Ca++ free Tyrode alone (63.77%, TALP with BOEC (36.92% and TALP alone (10.94%. Development of early embryos (8-cell stage improved in TCM-199 with BOEC co culture than TCM-199 alone. From the results of this study, it can be concluded that addition of somatic cells (granulosa cells, cumulus cells results in higher maturation rates of buffalo follicular oocytes than BOEC co culture system, while fertilization rate improved in modified Ca++ free Tyrode with and without BOEC. Addition of BOEC to TCM-199 improved the developmental capacity of early embryo.

  11. Production of imidazole alkaloids in cell cultures of jaborandi as affected by the medium pH.

    Science.gov (United States)

    Andreazza, N L; Abreu, I N; Sawaya, A C H F; Eberlin, M N; Mazzafera, P

    2009-04-01

    The effect of pH (from 4.8 to 9.8) on the production of pilosine and pilocarpine and on their partition between cell and medium was studied in two lineages (P and PP) of Pilocarpus microphyllus cell suspension cultures. Highest mass accumulation was observed at high pHs and both lineages produced pilocarpine while only lineage PP produced pilosine. Both alkaloids were released in the medium but higher accumulation occurred in the cells. The highest production of pilocarpine was at pH 8.8-9.8 in both cell lineages. Other imidazole alkaloids were also identified in both lineages. At all pHs tested, the pH in the media cultures tended to stabilize around 6 after 10-15 days of cultivation. NO3(-) and NH4+ variation in the media might partially explain the pH stabilization. PMID:19066732

  12. Search for Chemically Defined Culture Medium to Assist Initial Regeneration of Diseased Renal Parenchyma After Stem/Progenitor Cell Implantation

    OpenAIRE

    Minuth WW; Denk L; Gruber M

    2013-01-01

    Before an intended implantation stem/progenitor cells are usually kept in the beneficial atmosphere of a selected culture medium. However, after implantation the situation is drastically changing for them. Yet stem/progenitor cells must stand the harmful fluid environment within a diseased organ. In this coherence it is unknown, to which degree alterations in molecular composition of interstitial fluid can influence the initial regeneration of parenchyma. To obtain first ins...

  13. Medium Renewal Blocks Anti-Proliferative Effects of Metformin in Cultured MDA-MB-231 Breast Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Maruša Rajh

    Full Text Available Epidemiological studies indicate that metformin, a widely used type 2 diabetes drug, might reduce breast cancer risk and mortality in patients with type 2 diabetes. Metformin might protect against breast cancer indirectly by ameliorating systemic glucose homeostasis. Alternatively, it might target breast cancer cells directly. However, experiments using MDA-MB-231 cells, a standard in vitro breast cancer model, produced inconsistent results regarding effectiveness of metformin as a direct anti-cancer agent. Metformin treatments in cultured MDA-MB-231 cells are usually performed for 48-96 hours, but protocols describing renewal of cell culture medium during these prolonged treatments are rarely reported. We determined whether medium renewal protocol might alter sensitivity of MDA-MB-231 cells treated with metformin. Using the MTS assay, BrdU incorporation and Hoechst staining we found that treatment with metformin for 48-72 hours failed to suppress viability and proliferation of MDA-MB-231 cells if low-glucose (1 g/L medium was renewed every 24 hours. Conversely, metformin suppressed their viability and proliferation if medium was not renewed. Without renewal glucose concentration in the medium was reduced to 0.1 g/L in 72 hours, which likely explains increased sensitivity to metformin under these conditions. We also examined whether 2-deoxy-D-glucose (2-DG reduces resistance to metformin. In the presence of 2-DG metformin reduced viability and proliferation of MDA-MB-231 cells with or without medium renewal, thus demonstrating that 2-DG reduces their resistance to metformin. In sum, we show that medium renewal blocks anti-proliferative effects of metformin during prolonged treatments in low-glucose medium. Differences in medium renewal protocols during prolonged treatments might therefore lead to apparently inconsistent results as regards effectiveness of metformin as a direct anti-cancer agent. Finally, our results indicate that co-therapy with

  14. Medium Renewal Blocks Anti-Proliferative Effects of Metformin in Cultured MDA-MB-231 Breast Cancer Cells.

    Science.gov (United States)

    Rajh, Maruša; Dolinar, Klemen; Miš, Katarina; Pavlin, Mojca; Pirkmajer, Sergej

    2016-01-01

    Epidemiological studies indicate that metformin, a widely used type 2 diabetes drug, might reduce breast cancer risk and mortality in patients with type 2 diabetes. Metformin might protect against breast cancer indirectly by ameliorating systemic glucose homeostasis. Alternatively, it might target breast cancer cells directly. However, experiments using MDA-MB-231 cells, a standard in vitro breast cancer model, produced inconsistent results regarding effectiveness of metformin as a direct anti-cancer agent. Metformin treatments in cultured MDA-MB-231 cells are usually performed for 48-96 hours, but protocols describing renewal of cell culture medium during these prolonged treatments are rarely reported. We determined whether medium renewal protocol might alter sensitivity of MDA-MB-231 cells treated with metformin. Using the MTS assay, BrdU incorporation and Hoechst staining we found that treatment with metformin for 48-72 hours failed to suppress viability and proliferation of MDA-MB-231 cells if low-glucose (1 g/L) medium was renewed every 24 hours. Conversely, metformin suppressed their viability and proliferation if medium was not renewed. Without renewal glucose concentration in the medium was reduced to 0.1 g/L in 72 hours, which likely explains increased sensitivity to metformin under these conditions. We also examined whether 2-deoxy-D-glucose (2-DG) reduces resistance to metformin. In the presence of 2-DG metformin reduced viability and proliferation of MDA-MB-231 cells with or without medium renewal, thus demonstrating that 2-DG reduces their resistance to metformin. In sum, we show that medium renewal blocks anti-proliferative effects of metformin during prolonged treatments in low-glucose medium. Differences in medium renewal protocols during prolonged treatments might therefore lead to apparently inconsistent results as regards effectiveness of metformin as a direct anti-cancer agent. Finally, our results indicate that co-therapy with 2-DG and

  15. Hydrogen peroxide generation in caco-2 cell culture medium by addition of phenolic compounds: effect of ascorbic acid.

    Science.gov (United States)

    Roques, Sylvie Cambon; Landrault, Nicolas; Teissèdre, Pierre-Louis; Laurent, Caroline; Besançon, Pierre; Rouane, Jean-Max; Caporiccio, Bertrand

    2002-05-01

    Phenolic compounds have recently attracted special attention due to their beneficial health effects; their intestinal absorption and bioavailability need, therefore, to be investigated and Caco-2 cell culture model appeared as a promising tool. We have shown herein that the addition of a grape seed extract (GSE) to Dulbecco's modified Eagle's medium (DMEM) used for Caco-2 cell culture leads to a substantial loss of catechin, epicatechin and B2 and B3 dimers from GSE in the medium after 24 h and to a production of hydrogen peroxide (H2O2). When 1420 microM ascorbic acid is added to the DMEM, such H2O2 production was prevented. This hydrogen peroxide generation substantially involves inorganic salts from the DMEM. We recommend that ascorbic acid be added to circumvent such a risk. PMID:12150547

  16. Amperometric biosensor with nanostructured electrodes by using multi-alled carbon nanotubes for glucose detection in cell culture medium

    OpenAIRE

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

    2009-01-01

    The monitoring of metabolic compounds such as glucose is largely reported in literature. The applications of this type of analysis are mainly related to clinical purposes, e.g. in diabetes pathology, where a lot of studies are presented in literature. Recently, some authors presented studies about glucose and lactate detection in cell culture monitoring [1], [2]. A clear identification of medium compounds could be interesting for biologists and biotechnologists, since they may be identified a...

  17. Analysis of CHO cells metabolic redistribution in a glutamate-based defined medium in continuous culture.

    Science.gov (United States)

    Altamirano, C; Illanes, A; Casablancas, A; Gámez, X; Cairó, J J; Gòdia, C

    2001-01-01

    The effect of glutamine replacement by glutamate and the balance between glutamate and glucose metabolism on the redistribution of t-PA-producing recombinant CHO cells metabolism is studied in a series of glucose shift down and shift up experiments in continuous culture. These experiments reveal the existence of multiple steady states, and experimental data are used to perform metabolic flux analysis to gain a better insight into cellular metabolism and its redistribution. Regulation of glucose feed rate promotes a higher efficiency of glucose and nitrogen source utilization, with lower production of metabolic byproducts, but this reduces t-PA specific production rate. This reduction under glucose limitation can be attributed to the fact that the cells are forced to efficiently utilize the carbon and energy source for growth, impairing the production of dispensable metabolites. It is, therefore, the combination of growth rate and carbon and energy source availability that determines the level of t-PA production in continuous culture.

  18. Cell types and their status in Chlamydomonas-like algae (Chlorophyceae on agar medium culture

    Directory of Open Access Journals (Sweden)

    M.М. Pavlovska

    2014-04-01

    Full Text Available The classification of cell types under agar culture was proposed. Six cell morphotypes were allocated. The statuses were identified depending on the reduction of monade attributes of cells. The variants of transition from one cell morphotype to another under dissolving mucilage were shown. The monade, cocciod, palmeloid and gloeocysta morphotypes approximately equally represented in all clades. The asterococcus and mucogleocysta morphotypes presented only in Reinhardtinia аnd Oogamochlamydinia clades. Any morphotype isn’t typical for all clades of Chlamydomonas-like algae at once. The most of morphotypes numbers (5 from 6 are presented in Reinhardtinia clade. This demonstrates the diversity of the Reinhardtinia clade species. There are only one morphotype presented in Polytominia and Monadinia clades. There are four morphotypes presented in Oogamochlamydinia clade, three – in Moewusinia, two morphotypes – in Chloromonadinia.

  19. Improvement on light penetrability and microalgae biomass production by periodically pre-harvesting Chlorella vulgaris cells with culture medium recycling.

    Science.gov (United States)

    Huang, Yun; Sun, Yahui; Liao, Qiang; Fu, Qian; Xia, Ao; Zhu, Xun

    2016-09-01

    To improve light penetrability and biomass production in batch cultivation, a cultivation mode that periodically pre-harvesting partial microalgae cells from suspension with culture medium recycling was proposed. By daily pre-harvesting 30% microalgae cells from the suspension, the average light intensity in the photobioreactor (PBR) was enhanced by 27.05-122.06%, resulting in a 46.48% increase in total biomass production than that cultivated in batch cultivation without pre-harvesting under an incident light intensity of 160μmolm(-2)s(-1). Compared with the semi-continuous cultivation with 30% microalgae suspension daily replaced with equivalent volume of fresh medium, nutrients and water input was reduced by 60% in the proposed cultivation mode but with slightly decrease (12.82%) in biomass production. No additional nutrient was replenished when culture medium recycling. Furthermore, higher pre-harvesting ratios (40%, 60%) and lower pre-harvesting frequencies (every 2, 2.5days) were not advantageous for the pre-harvesting cultivation mode. PMID:27289058

  20. [Cell cultures].

    Science.gov (United States)

    Cipro, Simon; Groh, Tomáš

    2014-01-01

    Cell or tissue cultures (both terms are interchangeable) represent a complex process by which eukaryotic cells are maintained in vitro outside their natural environment. They have a broad usage covering not only scientific field but also diagnostic one since they represent the most important way of monoclonal antibodies production which are used for both diagnostic and therapeutic purposes. Cell cultures are also used as a "cultivation medium" in virology and for establishing proliferating cells in cytodiagnostics. They are well-established and easy-to-handle models in the area of research, e.g. as a precious source of nucleic acids or proteins. This paper briefly summarizes their importance and methods as well as the pitfalls of the cultivation and new trends in this field. PMID:24624984

  1. Adaptation of a Commonly Used, Chemically Defined Medium for Human Embryonic Stem Cells to Stable Isotope Labeling with Amino Acids in Cell Culture

    DEFF Research Database (Denmark)

    Liberski, A. R.; Al-Noubi, M. N.; Rahman, Z. H.;

    2013-01-01

    rarely employed in the context of complex culturing conditions as those required for human embryonic stem cells (hESCs). Classic hESC culture is based on the use of mouse embryonic fibroblasts (MEFs) as a feeder layer, and as a result, possible xenogeneic contamination, contribution of unlabeled amino......Metabolic labeling with stable isotopes is a prominent technique for comparative quantitative proteomics, and stable isotope labeling with amino acids in cell culture (SILAC) is the most commonly used approach. SILAC is, however, traditionally limited to simple tissue culture regimens and only...... developed by Ludwig et al. and commercially available as mTeSR1 [mTeSR1 is a trade mark of WiCell (Madison, WI) licensed to STEMCELL Technologies (Vancouver, Canada)]. This medium, together with adjustments to the culturing protocol, facilitates reproducible labeling that is easily scalable to the protein...

  2. Influence of Selenium Content in the Culture Medium on Protein Profile of Yeast Cells Candida utilis ATCC 9950

    Directory of Open Access Journals (Sweden)

    Marek Kieliszek

    2015-01-01

    Full Text Available Selenium is an essential trace element for human health and it has been recognized as a component of several selenoproteins with crucial biological functions. It has been identified as a component of active centers of many enzymes, as well as integral part of biologically active complexes. The aim of the study was to evaluate the protein content and amino acid profile of the protein of fodder yeast Candida utilis ATCC 9950 cultured in media control and experimental enriched selenium. Protein analysis was performed using SDS-PAGE method consisting of polyacrylamide gel electrophoresis in the presence of SDS. The highest contents of soluble protein (49,5 mg/g were found in yeast cells after 24-hour culture conducted in control (YPD medium. In the presence of selenium there were determined small amounts of protein content. With increasing time of yeast culture (to 72 hours the control and experimental media were reported to reduce soluble protein content. In electropherogram proteins from control cultures was observed the presence of 10 protein fractions, but in all the experimental cultures (containing 20, 30, and 40 mg/L selenium of 14 protein fractions. On the basis of the molecular weights of proteins, it can be concluded that they were among others: selenoprotein 15 kDa and selenoprotein 18 kDa.

  3. Effects of exogenous growth regulators on cell suspension culture of yin-hong grape (vitis vinifera l.) and establishment of the optimum medium

    International Nuclear Information System (INIS)

    Callus induced by stem of Yin-hong grape (Vitis vinifera L.) was used as materials and B5 medium as basic medium. The major growth parameters of cell suspension cultures with various levels of 1-Naphthaleneacetic acid (NAA) and 6-Benzyl aminopurine (6-BA) were investigated to provide a basis for the optimum medium of suspension cell cultures of Yin-hong grape regarding cell number, packed cell volume (PCV), dry cell weight (DCW), cell viability, and morphology. All data were analysed by of two-way analysis of variance (ANOVA). Results showed that the treatment of 6-BA and NAA would effect the cell growth dynamics, probably causing logarithmic phase in advance at higher levels of 6-BA. Different concentration of 6-BA and NAA had significant effects on cells number, PCV, DCW and viability (p<0.05), while no-significant effect was observed on the cells morphology. The optimum medium for suspension cell cultures of Yin-hong grape was identified as B5+1.5 mg/L6-BA+1.5 mg/LNAA+ 250 mg/L casein hydrolysate + 30 g/L sucrose. With the optimum medium, the maximum number of suspension cells after the logarithmic growth phase was 34.78 * 108 / mL, the highest cell viability reached 86.45%.; DCW reached 3.84 g/L and PCV reached 0.092 mL/mL after eight days cultivating. (author)

  4. Isomaltulose production using free cells: optimisation of a culture medium containing agricultural wastes and conversion in repeated-batch processes.

    Science.gov (United States)

    Kawaguti, Haroldo Y; Buzzato, Michele F; Sato, Hélia H

    2007-04-01

    The enzyme glucosyltransferase is an industrially important enzyme since it produces non-cariogenic isomaltulose (6-O-alpha-D-glucopyronosyl-1-6-D-fructofuranose) from sucrose by intramolecular transglucosylation. The experimental designs and response surface methodology (RSM) were applied for the optimisation of the nutrient concentrations in the culture medium for the production of glucosyltransferase by Erwinia sp. D12 in shaken flasks at 200 rpm and 30 degrees C. A statistical analysis of the results showed that, in the range studied, the factors had a significant effect (P sugar cane molasses (150 g l(-1)), corn steep liquor (20 g l(-1)), yeast extract Prodex Lac SD (15 g l(-1)) and K2HPO4 (0.5 g l(-1)) after 8 h at 30 degrees C. The production of cell biomass by the strain of Erwinia sp. D12 was carried out in a 6.6-l fermenter with a mixing rate of 200 rpm and an aeration rate of 1 vvm. Fermentation time, cellular growth, medium pH and glucosyltransferase production were observed. The greatest glucosyltransferase activity was 22.49 U/ml, obtained after 8 h of fermentation. The isomaltulose production from sucrose was performed using free Erwinia sp. D12 cells in a batch process using an orbital shaker. The influence of the parameters sucrose concentration, temperature, pH, and cell concentration on the conversion of sucrose into isomaltulose was studied. The free cells showed a high conversion rate of sucrose into isomaltulose using batch fermentation, obtaining an isomaltulose yield of 72.11% from sucrose solution 35% at 35 degrees C. PMID:17186209

  5. Size distribution of fullerenol nanoparticles in cell culture medium and their influence on antioxidative enzymes in Chinese hamster ovary cells

    Directory of Open Access Journals (Sweden)

    Srđenović Branislava U.

    2015-01-01

    Full Text Available Fullerenol (C60(OH24 nanoparticles (FNP have a significant role in biomedical research due to their numerous biological activities, some of which are cytoprotective and antioxidative properties. The aim of this study was to measure distribution of fullerenol nanoparticles and zeta potential in cell medium RPMI 1640 with 10% fetal bovine serum (FBS and to investigate the influence of FNP on Chinese hamster ovary cells (CHO-K1 survival, as well as to determine the activity of three antioxidative enzymes: superoxide-dismutase, glutathione-reductase and glutathione-S-transferase in mitomycin C-treated cell line. Our investigation implies that FNP, as a strong antioxidant, influence the cellular redox state and enzyme activities and thus may reduce cell proliferation, which confirms that FNP could be exploited for its use as a cytoprotective agent.[Projekat Ministarstva nauke Republike Srbije, br. III45005 i Pokrajinski Sekretarijat za nauku i tehnološki razvoj Vojvodine, grant number 114-451-2056/2011-01

  6. 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...

  7. Effects of nerve growth factor and heart cell conditioned medium on neurite regeneration of aged sympathetic neurons in culture.

    Science.gov (United States)

    Uchida, Y; Tomonaga, M

    1985-11-25

    The effects of nerve growth factor (NGF) and heart-cell-conditioned medium (HCM) on the neurite regeneration of aged sympathetic neurons were investigated in culture. Investigation of HCM was carried out by two different methods: one was the use of whole HCM on collagen substratum, which reflected component(s) effective in solution (HCM-S); the other was the use of polyornithine (PORN)-binding component(s) (P-HCM). Superior cervical ganglion neurons prepared from male mice from 6 to 30 months of age were cultured in MEM-10% FCS on collagen or gelatin-PORN substratum for 3 days. The number of neurons with neurites and the length of neurites were quantified as neurite production and elongation, respectively. Neuronal survival was not affected by addition of NGF, HCM-S or P-HCM. Neurite production of early adult neurons was enhanced by NGF, HCM-S or P-HCM. In contrast, neurite production of aged neurons was enhanced by only HCM-S, but not NGF or P-HCM. HCM-S did not promote neurite elongation in neurons at any age. Neurite elongation of early adult neurons was enhanced by NGF or P-HCM. Neurite elongation of aged neurons was enhanced by P-HCM. However, responsiveness of NGF for neurite elongation varied according to substrata. No age-related difference was found in neurite production and elongation in the absence of NGF, HCM-S or P-HCM. These results indicate that responsiveness of aged sympathetic neurons is various in different growth factors. PMID:3840716

  8. Cultivation and irradiation of human fibroblasts in a medium enriched with platelet lysate for obtaining feeder layer in epidermal cell culture

    International Nuclear Information System (INIS)

    For over 30 years, the use of culture medium, enriched with bovine serum, and murines fibroblasts, with the rate of proliferation controlled by irradiation or by share anticarcinogenic drugs, has been playing successfully its role in assisting in the development of keratinocytes in culture, for clinical purposes. However, currently there is a growing concern about the possibility of transmitting prions and animals viruses to transplanted patients. Taking into account this concern, the present work aims to cultivate human fibroblasts in a medium enriched with human platelets lysate and determine the irradiation dose of these cells, for obtaining feeder layer in epidermal cell culture. For carrying out the proposed objective, platelets lysis has standardized, this lysate was used for human fibroblasts cultivation and the irradiation dose enough to inhibit its duplication was evaluated. Human keratinocytes were cultivated in these feeder layers, in culture medium enriched with the lysate. With these results we conclude that the 10% platelets lysate promoted a better adhesion and proliferation of human fibroblasts and in all dose levels tested (60 to 300 Gy), these had their mitotic activity inactivated by ionizing irradiation, being that the feeder layers obtained with doses from 70 to 150 Gy were those that provided the best development of keratinocytes in medium containing 2.5% of human platelet lysate. Therefore, it was possible to standardize both the cultivation of human fibroblasts as its inactivation for use as feeder layer in culture of keratinocytes, so as to eliminate xenobiotics components. (author)

  9. 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

  10. Combined effects of low-level laser therapy and human bone marrow mesenchymal stem cell conditioned medium on viability of human dermal fibroblasts cultured in a high-glucose medium.

    Science.gov (United States)

    Hendudari, Farzane; Piryaei, Abbas; Hassani, Seyedeh-Nafiseh; Darbandi, Hasan; Bayat, Mohammad

    2016-05-01

    Low-level laser therapy (LLLT) exhibited biostimulatory effects on fibroblasts viability. Secretomes can be administered to culture mediums by using bone marrow mesenchymal stem cells conditioned medium (BM-MSCs CM). This study investigated the combined effects of LLLT and human bone marrow mesenchymal stem cell conditioned medium (hBM-MSCs CM) on the cellular viability of human dermal fibroblasts (HDFs), which was cultured in a high-glucose (HG) concentration medium. The HDFs were cultured either in a concentration of physiologic (normal) glucose (NG; 5.5 mM/l) or in HG media (15 mM/l) for 4 days. LLLT was performed with a continuous-wave helium-neon laser (632.8 nm, power density of 0.00185 W/cm(2) and energy densities of 0.5, 1, and 2 J/cm(2)). About 10 % of hBM-MSCs CM was added to the HG HDF culture medium. The viability of HDFs was evaluated using dimethylthiazol-diphenyltetrazolium bromide (MTT) assay. A significantly higher cell viability was observed when laser of either 0.5 or 1 J/cm(2) was used to treat HG HDFs, compared to the control groups. The cellular viability of HG-treated HDFs was significantly lower compared to the LLLT + HG HDFs, hBM-MSCs CM-treated HG HDFs, and LLLT + hBM-MSCs CM-treated HG HDFs. In conclusion, hBM-MSCs CM or LLLT alone increased the survival of HG HDFs cells. However, the combination of hBM-MSCs CM and LLLT improved these results in comparison to the conditioned medium. PMID:26984346

  11. α-transforming growth factor secreted by untransformed bovine anterior pituitary cells in culture. I. Purification from conditioned medium

    International Nuclear Information System (INIS)

    A 6-kDa α-transforming growth factor (TGF) was purified 100,000-fold to homogeneity from the culture fluid conditioned by normal bovine anterior pituitary-derived cells. Initial purification of the acid-soluble TGF from concentrated conditioned medium was achieved by Bio-Gel P-60 gel filtration (apparent molecular mass of 9 kDa). After the Bio-Gel step, three different steps of reverse-phase fast-protein liquid chromatography on the same Pharmacia C18 column, using linear acetonitrile gradients, gave complete purification. The ion-pairing agents used in the three consecutive steps were: 0.1% trifluoroacetic acid, 0.13% heptafluorobutyric acid, and again, 0.1% trifluoroacetic acid at a shallower gradient. Homogeneity was confirmed by reverse-phase high performance liquid chromatography, and by polyacrylamide gel electrophoresis, where TGF visualization was facilitated by autoradiography of 125I-TGF. The 125I-TGF bound to epidermal growth factor (EGF) receptors and after elution ran identically to the starting material. The molecular mass of TGF is 6 kDa by polyacrylamide gel electrophoresis and 6.6 kDa by amino acid analysis. The amino acid composition of bovine TGF is similar to that of rat or human αTGF and distinct from epidermal growth factor. Colony-stimulating activity was lost after purification, but the TGF retained its ability to stimulate thymidine uptake by quiescent cells. This mitogenic activity could be blocked completely by anti-EGF-receptor monoclonal antibodies, indicating that the activity was mediated through the EGF-receptor

  12. A Newly Defined and Xeno-Free Culture Medium Supports Every-Other-Day Medium Replacement in the Generation and Long-Term Cultivation of Human Pluripotent Stem Cells.

    Science.gov (United States)

    Ahmadian Baghbaderani, Behnam; Tian, Xinghui; Scotty Cadet, Jean; Shah, Kevan; Walde, Amy; Tran, Huan; Kovarcik, Don Paul; Clarke, Diana; Fellner, Thomas

    2016-01-01

    Human pluripotent stem cells (hPSCs) present an unprecedented opportunity to advance human health by offering an alternative and renewable cell resource for cellular therapeutics and regenerative medicine. The present demand for high quality hPSCs for use in both research and clinical studies underscores the need to develop technologies that will simplify the cultivation process and control variability. Here we describe the development of a robust, defined and xeno-free hPSC medium that supports reliable propagation of hPSCs and generation of human induced pluripotent stem cells (hiPSCs) from multiple somatic cell types; long-term serial subculturing of hPSCs with every-other-day (EOD) medium replacement; and banking fully characterized hPSCs. The hPSCs cultured in this medium for over 40 passages are genetically stable, retain high expression levels of the pluripotency markers TRA-1-60, TRA-1-81, Oct-3/4 and SSEA-4, and readily differentiate into ectoderm, mesoderm and endoderm. Importantly, the medium plays an integral role in establishing a cGMP-compliant process for the manufacturing of hiPSCs that can be used for generation of clinically relevant cell types for cell replacement therapy applications. PMID:27606941

  13. Induction of E-cadherin+ human amniotic fluid cell differentiation into oocyte-like cells via culture in medium supplemented with follicular fluid.

    Science.gov (United States)

    Liu, Te; Huang, Yongyi; Bu, Yanzhen; Zhao, Yanhui; Zou, Gang; Liu, Zhixue

    2014-07-01

    Pluripotent human amniotic fluid cells (HuAFCs) can differentiate into various types of somatic cell in vitro. However, their differentiation into oocyte-like cells has never been described to the best of our knowledge. In the present study, differentiation of E-cadherin+ and E-cadherin- HuAFC sub-populations into oocyte-like cells was induced via culture in medium containing bovine follicular fluid and β-mercaptoethanol. The E-cadherin+ HuAFCs expressed DAZL highly. Post-induction, cells with an oocyte-like phenotype were found among the E-cadherin+ HuAFCs, expressing markers specific to germ cells and oocytes (VASA, ZP3 and GDF9) and meiosis (DMC1 and SCP3). When specific small interfering RNA (siRNA) was used to suppress E-cadherin in the E-cadherin+ HuAFCs, the levels of DAZL expression were reduced. Post-induction, the morphology of the siRNA‑E‑cadherin HuAFCs was poorer and the expression levels of germ cell-specific markers were lower compared with those of the siRNA-mock HuAFCs. Therefore, E-cadherin+ HuAFCs could be more easily induced to differentiate into oocyte-like cells by bovine follicular fluid and β-mercaptoethanol. In addition, the E-cadherin+ HuAFCs exhibited potential characteristics of DAZL protein expression, and thus it was conjectured that bovine follicular fluid acts on DAZL protein and promotes E-cadherin+ HuAFC differentiation into oocyte-like cells.

  14. Development and characterization of a clinically compliant xeno-free culture medium in good manufacturing practice for human multipotent mesenchymal stem cells.

    Science.gov (United States)

    Chase, Lucas G; Yang, Sufang; Zachar, Vladimir; Yang, Zheng; Lakshmipathy, Uma; Bradford, Jolene; Boucher, Shayne E; Vemuri, Mohan C

    2012-10-01

    Human multipotent mesenchymal stem cell (MSC) therapies are currently being tested in clinical trials for Crohn's disease, multiple sclerosis, graft-versus-host disease, type 1 diabetes, bone fractures, cartilage damage, and cardiac diseases. Despite remarkable progress in clinical trials, most applications still use traditional culture media containing fetal bovine serum or serum-free media that contain serum albumin, insulin, and transferrin. The ill-defined and variable nature of traditional culture media remains a challenge and has created a need for better defined xeno-free culture media to meet the regulatory and long-term safety requirements for cell-based therapies. We developed and tested a serum-free and xeno-free culture medium (SFM-XF) using human bone marrow- and adipose-derived MSCs by investigating primary cell isolation, multiple passage expansion, mesoderm differentiation, cellular phenotype, and gene expression analysis, which are critical for complying with translation to cell therapy. Human MSCs expanded in SFM-XF showed continual propagation, with an expected phenotype and differentiation potential to adipogenic, chondrogenic, and osteogenic lineages similar to that of MSCs expanded in traditional serum-containing culture medium (SCM). To monitor global gene expression, the transcriptomes of bone marrow-derived MSCs expanded in SFM-XF and SCM were compared, revealing relatively similar expression profiles. In addition, the SFM-XF supported the isolation and propagation of human MSCs from primary human marrow aspirates, ensuring that these methods and reagents are compatible for translation to therapy. The SFM-XF culture system allows better expansion and multipotentiality of MSCs and serves as a preferred alternative to serum-containing media for the production of large scale, functionally competent MSCs for future clinical applications.

  15. Melatonin affects voltage-dependent calcium and potassium currents in MCF-7 cell line cultured either in growth or differentiation medium.

    Science.gov (United States)

    Squecco, Roberta; Tani, Alessia; Zecchi-Orlandini, Sandra; Formigli, Lucia; Francini, Fabio

    2015-07-01

    Big efforts have been dedicated up to now to identify novel targets for cancer treatment. The peculiar biophysical profile and the atypical ionic channels activity shown by diverse types of human cancers suggest that ion channels may be possible targets in cancer therapy. Earlier studies have shown that melatonin exerts an oncostatic action on different tumors. In particular, it was shown that melatonin was able to inhibit growth/viability and proliferation, to reduce the invasiveness and metastatic properties of human estrogen-sensitive breast adenocarcinoma MCF-7 cell line cultured in growth medium, with substantial impairments of epidermal growth factor (EGF) and Notch-1-mediated signaling. The purpose of this work was to evaluate on MCF-7 cells the possible effects of melatonin on the biophysical features known to have a role in proliferation and differentiation, by using the patch-clamp technique. Our results show that in cells cultured in growth as well as in differentiation medium melatonin caused a hyperpolarization of resting membrane potential paralleled by significant changes of the inward Ca(2+) currents (T- and L-type), outward delayed rectifier K(+) currents and cell capacitance. All these effects are involved in MCF-7 growth and differentiation. These findings strongly suggest that melatonin, acting as a modulator of different voltage-dependent ion channels, might be considered a new promising tool for specifically disrupting cell viability and differentiation pathways in tumour cells with possible beneficial effects on cancer therapy. PMID:25843408

  16. 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...

  17. 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 tissue like structuring of the cultures. However, as this chapter illustrates, many issues remain to be identified regarding perfusion cell culture such as design, material choice and how to use these systems before they will be widespread amongst biomedical researchers....

  18. Secretion of nerve growth factor, brain-derived neurotrophic factor, and glial cell-line derived neurotrophic factor in co-culture of four cell types in cerebrospinal fluid-containing medium

    Institute of Scientific and Technical Information of China (English)

    Sanjiang Feng; Minghua Zhuang; Rui Wu

    2012-01-01

    The present study co-cultured human embryonic olfactory ensheathing cells, human Schwann cells, human amniotic epithelial cells and human vascular endothelial cells in complete culture medium- containing cerebrospinal fluid. Enzyme linked immunosorbent assay was used to detect nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor secretion in the supernatant of co-cultured cells. Results showed that the number of all cell types reached a peak at 7–10 days, and the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor peaked at 9 days. Levels of secreted nerve growth factor were four-fold higher than brain-derived neurotrophic factor, which was three-fold higher than glial cell line-derived neurotrophic factor. Increasing concentrations of cerebrospinal fluid (10%, 20% and 30%) in the growth medium caused a decrease of neurotrophic factor secretion. Results indicated co-culture of human embryonic olfactory ensheathing cells, human Schwann cells, human amniotic epithelial cells and human vascular endothelial cells improved the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor. The reduction of cerebrospinal fluid extravasation at the transplant site after spinal cord injury is beneficial for the survival and secretion of neurotrophic factors from transplanted cells.

  19. Hydrogen photoproduction in green algae Chlamydomonas reinhardtii sustainable over 2 weeks with the original cell culture without supply of fresh cells nor exchange of the whole culture medium.

    Science.gov (United States)

    Yagi, Takafumi; Yamashita, Kyohei; Okada, Norihide; Isono, Takumi; Momose, Daisuke; Mineki, Shigeru; Tokunaga, Eiji

    2016-07-01

    Unicellular green algae Chlamydomonas reinhardtii are known to make hydrogen photoproduction under the anaerobic condition with water molecules as the hydrogen source. Since the hydrogen photoproduction occurs for a cell to circumvent crisis of its survival, it is only temporary. It is a challenge to realize persistent hydrogen production because the cells must withstand stressful conditions to survive with alternation of generations in the cell culture. In this paper, we have found a simple and cost-effective method to sustain the hydrogen production over 14 days in the original culture, without supply of fresh cells nor exchange of the culture medium. This is achieved for the cells under hydrogen production in a sulfur-deprived culture solution on the {anaerobic, intense light} condition in a desiccator, by periodically providing a short period of the recovery time (2 h) with a small amount of TAP(+S) supplied outside of the desiccator. As this operation is repeated, the response time of transition into hydrogen production (preparation time) is shortened and the rate of hydrogen production (build up time) is increased. The optimum states of these properties favorable to the hydrogen production are attained in a few days and stably sustained for more than 10 days. Since generations are alternated during this consecutive hydrogen production experiment, it is suggested that the improved hydrogen production properties are inherited to next generations without genetic mutation. The properties are reset only when the cells are placed on the {sulfur-sufficient, aerobic, moderate light} conditions for a long time (more than 1 day at least). PMID:27083446

  20. Purification and characterization of three chitinases and one beta-1,3-glucanase accumulating in the medium of cell suspension cultures of barley (Hordeum vulgare L.)

    DEFF Research Database (Denmark)

    Kragh, K.M.; Jacobsen, S.; Dalgaard Mikkelsen, J.;

    1991-01-01

    Three basic chitinases and one basic beta-1,3-glucanase were secreted into the medium when embryogenic cell suspensions of barley (Hordeum vulgare L.) cv. 'Igri' were cultured as undifferentiated aggregates in the presence of 2,4-D. The enzymes were purified by affinity and ion exchange...... chromatography. Two of the chitinases were identified as the previously described endochitinases T and C from barley grain. The third and novel chitinase, designated K, was the major basic chitinase in the medium constituting 4% of the soluble protein. Chitinase K was found to be a 33-kDa endochitinase with p......I at 8.7. Further analysis showed that this enzyme is also expressed in barley grain. The amino acid composition and five partial amino acid sequences covering 93 residues of chitinase K were determined. A high similarity was found between chitinase K and barley chitinase T and C as well as basic...

  1. Effect of different surfactants in biorelevant medium on the secretion of a lipophilic compound in lipoproteins using Caco-2 cell culture

    DEFF Research Database (Denmark)

    Karpf, Ditte M; Holm, René; Garafalo, Carole;

    2006-01-01

    The impact of a pharmaceutical relevant metabolizable, ionic surfactant or two synthetic, nonionic surfactants on the absorption and lipoprotein incorporation of a lipophilic drug, retinol, was studied in the Caco-2 cell culture. Filter-grown monolayers of Caco-2 cells were incubated for 20 h...... with (3)H-retinol and (14)C-oleic acid and with increasing concentrations of lyso-phosphatidylcholine (lyso-PC), Cremophor RH40, or Tween 80. The concentration of (3)H-retinol and (14)C-lipid was measured in the apical, intracellular, and basolateral compartments. The basolateral medium...... was ultracentrifugated into different lipoprotein classes and their (3)H-retinol and (14)C-lipid concentrations were determined. The cells incubated with lyso-PC and Tween 80 increased the incorporation of (3)H-retinol and (14)C-lipid into chylomicrons and very low density lipoproteins (VLDL). The explored surfactants...

  2. Ultrastructure of Single Cells, Callus-like and Monosore-like Cells in Porphyra yezoensis Ueda on Semi solid Culture Medium

    Institute of Scientific and Technical Information of China (English)

    梅俊学; 沈颂东; 姜明; 费修绠

    2003-01-01

    It had been demonstrated that individual cells or protoplasts isolated from Porphyra thallus by enzyme could develop into normal leafy thalli in the same way as monospores, and that isolated cells develop in different way in liquid and on semi solid media. The authors observed the ultrastructure of isolated vegetative cells cultured on semi solid media and compared them with those of monospores and isolated cells cultured in liquid media. The results showed that subcellular structures were quite different among cells in different conditions. In their development, isolated cells on semi solid media did not show the characteristic subcellular feature of monospore formation, such as production of fibrous vesicles. Callus like cells formed on semi solid media underwent a distinctive modification in cellular organization. They developed characteristic cell inclusions and a special 2 layer cell covering. Golgi bodies, ER, starch grains, mitochondria. Vacuoles were not commonly found in them.

  3. Development of a vinasse culture medium for plant tissue culture

    International Nuclear Information System (INIS)

    Vinasse is the main pollutant (effluent) obtained from the distillation of sugarcane in the production of fuel alcohol. However, this residue is rich in nutrients that are required by plants. We developed a new culture medium using vinasse for the In vitro propagation of an orchid. The vinasse was treated (decanted and filtered), and the nutrients were determined and quantified. Different formulations using vinasse were tested for an In vitro culture. The vinasse dilutions demonstrated a good buffering effect. The ideal vinasse dilution for media formulation was 2.5%. The best KC formulations with vinasse were KCV1 and KCV5. Compared to KC medium, these formulations demonstrated similar results for In vitro multiplication, with the exception of protocorm-like body number, which was inferior in the vinasse formulations. Conversely, for In vitro elongation and rooting, these vinasse media were superior to KC medium. KC medium promotes a low rooting rate (8%) compared to 68 and 100% obtained by KCV1 and KCV5, respectively. Moreover, plantlets cultured on KC medium become protocorm-like body clusters, which impeded the acclimatization of these explants. Plantlets elongated and rooted on KCV1 and KCV5 were successfully acclimatized with a 91% survival rate for both KC vinasse formulations. This study shows the great potential of this technology as a rational alternative to vinasse disposal and adds value to what is currently considered a waste product. (author)

  4. 21 CFR 866.2330 - Enriched culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Enriched culture medium. 866.2330 Section 866.2330...) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2330 Enriched culture medium. (a) Identification. An enriched culture medium is a device that consists primarily of liquid...

  5. 21 CFR 866.2360 - Selective culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Selective culture medium. 866.2360 Section 866...) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2360 Selective culture medium. (a) Identification. A selective culture medium is a device that consists primarily of liquid...

  6. 21 CFR 866.2390 - Transport culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Transport culture medium. 866.2390 Section 866...) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2390 Transport culture medium. (a) Identification. A transport culture medium is a device that consists of a semisolid,...

  7. 21 CFR 866.2350 - Microbiological assay culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Microbiological assay culture medium. 866.2350... Microbiological assay culture medium. (a) Identification. A microbiological assay culture medium is a device that... (general controls). The device is exempt from the premarket notification procedures in subpart E of...

  8. Application of near-infrared (NIR) spectroscopy for screening of raw materials used in the cell culture medium for the production of a recombinant therapeutic protein.

    Science.gov (United States)

    Kirdar, Alime Ozlem; Chen, Guoxiang; Weidner, James; Rathore, Anurag S

    2010-01-01

    Control of raw materials based on an understanding of their impact on product attributes has been identified as a key aspect of developing a control strategy in the Quality by Design (QbD) paradigm. This article presents a case study involving use of a combined approach of Near-infrared (NIR) spectroscopy and Multivariate Data Analysis (MVDA) for screening of lots of basal medium powders based on their impact on process performance and product attributes. These lots had identical composition as per the supplier and were manufactured at different scales using an identical process. The NIR/MVDA analysis, combined with further investigation at the supplier site, concluded that grouping of medium components during the milling and blending process varied with the scale of production and media type. As a result, uniformity of blending, impurity levels, chemical compatibility, and/or heat sensitivity during the milling process for batches of large-scale media powder were deemed to be the source of variation as detected by NIR spectra. This variability in the raw materials was enough to cause unacceptably large variability in the performance of the cell culture step and impact the attributes of the resulting product. A combined NIR/MVDA approach made it possible to finger print the raw materials and distinguish between good and poor performing media lots. PMID:19938040

  9. Buffering the pH of the culture medium does not extend yeast replicative lifespan

    OpenAIRE

    Wasko, Brian M.; Carr, Daniel T; Herman Tung; Ha Doan; Nathan Schurman; Neault, Jillian R; Joey Feng; Janet Lee; Ben Zipkin; Jacob Mouser; Edward Oudanonh; Tina Nguyen; Torin Stetina; Anna Shemorry; Mekedes Lemma

    2013-01-01

    During chronological aging of budding yeast cells, the culture medium can become acidified, and this acidification limits cell survival.  As a consequence, buffering the culture medium to pH 6 significantly extends chronological life span under standard conditions in synthetic medium.  In this study, we assessed whether a similar process occurs during replicative aging of yeast cells.  We find no evidence that buffering the pH of the culture medium to pH levels either higher or lower than the...

  10. Medium Renewal Blocks Anti-Proliferative Effects of Metformin in Cultured MDA-MB-231 Breast Cancer Cells

    OpenAIRE

    Maruša Rajh; Klemen Dolinar; Katarina Miš; Mojca Pavlin; Sergej Pirkmajer

    2016-01-01

    Epidemiological studies indicate that metformin, a widely used type 2 diabetes drug, might reduce breast cancer risk and mortality in patients with type 2 diabetes. Metformin might protect against breast cancer indirectly by ameliorating systemic glucose homeostasis. Alternatively, it might target breast cancer cells directly. However, experiments using MDA-MB-231 cells, a standard in vitro breast cancer model, produced inconsistent results regarding effectiveness of metformin as a direct ant...

  11. INNOVATIVE CULTURE IN SMALL AND MEDIUM ENTERPRISES

    Directory of Open Access Journals (Sweden)

    Aluisio Broering Mambrini

    2011-10-01

    Full Text Available In the last two decades, innovation has been a key driver of economic growth. Innovation is closely related to creating value and generating wealth through successful service to consumer needs. Thus, it is not necessarily restricted to the use of new knowledge generated from research, but on the development of new products or services that are obtained with creative use of knowledge, new or already known. This study aimed to identify management practices that promote a culture of innovation in small and medium enterprises and analyze how they contribute to the innovative capacity of these companies. The research method was the multiple case study with six small and medium businesses that have at least one case of significant innovation in its history. The main results showed that amongst the practices are: a performance in highly specialized niches and deep focus on customer needs; b strong investment and incorporation of new knowledge outside the company (open innovation; c speed and agility in the absorption and deployment of new knowledge and technologies; d retention of employees; e acting as an integrator combining diverse knowledge and technologies; f the information management of the knowledge acquired by the company; g little concern to patent the technology; h flexibility and informal, fluid and open communication between employees of the company that promotes agility in management and i the management of partnerships across the value chain, including the functional areas.

  12. Micropropagation of orchid hybrids in knudson culture medium with addiction of vitamins of ms culture medium, benzilaminopurine and activated charcoal

    Directory of Open Access Journals (Sweden)

    Fabíola Villa

    2014-02-01

    Full Text Available The genera Cattleya and Brassavola, naturally occurring in Brazil, are widely used in hybridization to obtain better shape of the lip in the hybrids. This demand creates the need to develop more effective propagation to meet the market and contributing to avoid the extinction of these plants. This study aimed to test BAP concentrations, activated charcoal, culture medium Knudson and culture medium MS’s vitamins in subculture of Brassocattleya Pastoral x Laeliocattleya Amber Glow orchids plants. There were used seedlings from in vitro germination, with 1-1.5 cm length, undergoing standardization in the culture medium Knudson for three months. The experiment were performed using concentrations of culture medium Knudson (0%, 50%, 100%, 150% and 200%, combined with concentrations of culture medium MS’s vitamins (0%, 50%, 100% and 200% and BAP concentrations (0; 1.0; 2.0 and 4.0 mg L-1 versus activated charcoal (0, 200, 400 and 600 mg L-1, incorporated in a culture medium Knudson. In each flask of 250 mL containing approximately 50 mL of culture medium, four seedlings were placed under aseptic conditions. It was concluded that can use 129% of Knudson, supplemented with 137.5 mg L-1 activated charcoal, 104% of vitamin MS medium without addition of BAP as the best composition for culture medium of hybrid orchid in vitro propagation. The Knudson culture medium does not promote increase in the root system.

  13. Immunohistochemical expression of stem cell, endothelial cell, and chemosensitivity markers in primary glioma spheroids cultured in serum-containing and serum-free medium

    DEFF Research Database (Denmark)

    Christensen, Karina; Aaberg-Jessen, Charlotte; Andersen, Claus;

    2010-01-01

    To investigate the influence of serum-free medium (SFM) supplemented with epidermal growth factor and basic fibroblast growth factor compared with conventional serum-containing medium (SCM) on the phenotype of organotypic primary spheroids from seven gliomas.......To investigate the influence of serum-free medium (SFM) supplemented with epidermal growth factor and basic fibroblast growth factor compared with conventional serum-containing medium (SCM) on the phenotype of organotypic primary spheroids from seven gliomas....

  14. Identification of candidate agents active against N. ceranae infection in honey bees: establishment of a medium throughput screening assay based on N. ceranae infected cultured cells.

    Directory of Open Access Journals (Sweden)

    Sebastian Gisder

    Full Text Available Many flowering plants in both natural ecosytems and agriculture are dependent on insect pollination for fruit set and seed production. Managed honey bees (Apis mellifera and wild bees are key pollinators providing this indispensable eco- and agrosystem service. Like all other organisms, bees are attacked by numerous pathogens and parasites. Nosema apis is a honey bee pathogenic microsporidium which is widely distributed in honey bee populations without causing much harm. Its congener Nosema ceranae was originally described as pathogen of the Eastern honey bee (Apis cerana but jumped host from A. cerana to A. mellifera about 20 years ago and spilled over from A. mellifera to Bombus spp. quite recently. N. ceranae is now considered a deadly emerging parasite of both Western honey bees and bumblebees. Hence, novel and sustainable treatment strategies against N. ceranae are urgently needed to protect honey and wild bees. We here present the development of an in vitro medium throughput screening assay for the identification of candidate agents active against N. ceranae infections. This novel assay is based on our recently developed cell culture model for N. ceranae and coupled with an RT-PCR-ELISA protocol for quantification of N. ceranae in infected cells. The assay has been adapted to the 96-well microplate format to allow automated analysis. Several substances with known (fumagillin or presumed (surfactin or no (paromomycin activity against N. ceranae were tested as well as substances for which no data concerning N. ceranae inhibition existed. While fumagillin and two nitroimidazoles (metronidazole, tinidazole totally inhibited N. ceranae proliferation, all other test substances were inactive. In summary, the assay proved suitable for substance screening and demonstrated the activity of two synthetic antibiotics against N. ceranae.

  15. Identification of candidate agents active against N. ceranae infection in honey bees: establishment of a medium throughput screening assay based on N. ceranae infected cultured cells.

    Science.gov (United States)

    Gisder, Sebastian; Genersch, Elke

    2015-01-01

    Many flowering plants in both natural ecosytems and agriculture are dependent on insect pollination for fruit set and seed production. Managed honey bees (Apis mellifera) and wild bees are key pollinators providing this indispensable eco- and agrosystem service. Like all other organisms, bees are attacked by numerous pathogens and parasites. Nosema apis is a honey bee pathogenic microsporidium which is widely distributed in honey bee populations without causing much harm. Its congener Nosema ceranae was originally described as pathogen of the Eastern honey bee (Apis cerana) but jumped host from A. cerana to A. mellifera about 20 years ago and spilled over from A. mellifera to Bombus spp. quite recently. N. ceranae is now considered a deadly emerging parasite of both Western honey bees and bumblebees. Hence, novel and sustainable treatment strategies against N. ceranae are urgently needed to protect honey and wild bees. We here present the development of an in vitro medium throughput screening assay for the identification of candidate agents active against N. ceranae infections. This novel assay is based on our recently developed cell culture model for N. ceranae and coupled with an RT-PCR-ELISA protocol for quantification of N. ceranae in infected cells. The assay has been adapted to the 96-well microplate format to allow automated analysis. Several substances with known (fumagillin) or presumed (surfactin) or no (paromomycin) activity against N. ceranae were tested as well as substances for which no data concerning N. ceranae inhibition existed. While fumagillin and two nitroimidazoles (metronidazole, tinidazole) totally inhibited N. ceranae proliferation, all other test substances were inactive. In summary, the assay proved suitable for substance screening and demonstrated the activity of two synthetic antibiotics against N. ceranae.

  16. Development of a selective culture medium for Fusarium moniliforme.

    Science.gov (United States)

    Castellá, G; Bragulat, M R; Rubiales, M V; Cabañes, F J

    1997-12-01

    Nash and Snyder medium and malachite green agar 2.5 ppm medium, a new selective culture medium designed in our laboratory, were challenged with pure cultures of Fusarium moniliforme strains and two different mixed-conidium suspensions, which included rapidly spreading fungi, for their utility in the isolation and enumeration of F. moniliforme. From the results of this comparative study, malachite green agar 2.5 ppm allowed only the selective growth of F. moniliforme whereas Nash and Snyder medium allowed both the growth of F. moniliforme and other species not belonging to Fusarium spp. The enumeration of F. moniliforme propagules was similar in both culture media.

  17. Concise review: no breakthroughs for human mesenchymal and embryonic stem cell culture: conditioned medium, feeder layer, or feeder-free; medium with fetal calf serum, human serum, or enriched plasma; serum-free, serum replacement nonconditioned medium, or ad hoc formula? All that glitters is not gold!

    Science.gov (United States)

    Mannello, Ferdinando; Tonti, Gaetana A

    2007-07-01

    The choice of an optimal strategy of stem cell culture is at the moment an impossible task, and the elaboration of a culture medium adapted to the production of embryonic and adult mesenchymal stem cells for the clinical application of cell therapy remains a crucial matter. To make an informed choice, it is crucial to not underestimate the theoretical health risk of using xenogenic compounds, to limit the immunological reactions once stem cells are transplanted, to not overestimate the controversial results obtained with human serum, plasma, and blood derivatives, as well as to carefully examine the pros and cons of serum-free and ad hoc formulation strategies; besides that, to also maintain multipotentiality, self-renewal, and transplantability. The extent to which we are able to achieve effective cell therapies will depend on assimilating a rapidly developing base of scientific knowledge with the practical considerations of design, delivery, and host response. Although clinical studies have already started, many questions remain unsolved, and concomitantly even more evidence on suitable and safe off-the-shelf products (mainly xeno-free) for embryonic and mesenchymal stem cells is cropping up, even though there should be no rush to enter the clinical stage while the underlying basic research is still not so solid; this solely will lead to high-quality translational research, without making blunders stemming from the assumption that all that glitters is not gold. Disclosure of potential conflicts of interest is found at the end of this article. PMID:17395775

  18. Concise review: no breakthroughs for human mesenchymal and embryonic stem cell culture: conditioned medium, feeder layer, or feeder-free; medium with fetal calf serum, human serum, or enriched plasma; serum-free, serum replacement nonconditioned medium, or ad hoc formula? All that glitters is not gold!

    Science.gov (United States)

    Mannello, Ferdinando; Tonti, Gaetana A

    2007-07-01

    The choice of an optimal strategy of stem cell culture is at the moment an impossible task, and the elaboration of a culture medium adapted to the production of embryonic and adult mesenchymal stem cells for the clinical application of cell therapy remains a crucial matter. To make an informed choice, it is crucial to not underestimate the theoretical health risk of using xenogenic compounds, to limit the immunological reactions once stem cells are transplanted, to not overestimate the controversial results obtained with human serum, plasma, and blood derivatives, as well as to carefully examine the pros and cons of serum-free and ad hoc formulation strategies; besides that, to also maintain multipotentiality, self-renewal, and transplantability. The extent to which we are able to achieve effective cell therapies will depend on assimilating a rapidly developing base of scientific knowledge with the practical considerations of design, delivery, and host response. Although clinical studies have already started, many questions remain unsolved, and concomitantly even more evidence on suitable and safe off-the-shelf products (mainly xeno-free) for embryonic and mesenchymal stem cells is cropping up, even though there should be no rush to enter the clinical stage while the underlying basic research is still not so solid; this solely will lead to high-quality translational research, without making blunders stemming from the assumption that all that glitters is not gold. Disclosure of potential conflicts of interest is found at the end of this article.

  19. Determinant factors for hybridmyeloma culture (H9r9 in the yield of anti - D – impact of medium (IMDM and RPMI, serum (FCS and cell density

    Directory of Open Access Journals (Sweden)

    Sathya R

    2008-12-01

    empirical informations of H9r9 cell metabolism to achieve optimization. Keywords: Hybridoma (H9r9, Foetal calf serum (FCS, Anti – D, Apoptosis, IMDM/RPMI Medium Received: 12 January 2009 / Received in revised form: 6 February 2009, Accepted: 25 February 2009, Published online: 28 February 2009

  20. 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.

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

    KAUST Repository

    Hadj-Romdhane, F.

    2013-03-01

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

  2. 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...

  3. Organizational Culture Assessment of Small & Medium-Sized Enterprises

    OpenAIRE

    Arshad Zaheer; Kashif ur Rehman; Abrar Ahmad

    2006-01-01

    This paper is an ethnographic study defining and assessing the organizational culture exhibited by Small and Medium Enterprises (SMEs). It primarily focuses on four cultural categories: clan, adhocracy, hierarchy and market-driven. These conceptual domains have been examined by the Organizational Culture Assessment Instrument. Results from a sample of 162 SMEs in the Rawalpindi/Islamabad area indicate that SME culture lacks creativity, innovation, freedom and risk taking. SMEs are not looking...

  4. 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...

  5. [Effects of beryllium chloride on cultured cells].

    Science.gov (United States)

    Sakaguchi, T; Sakaguchi, S; Nakamura, I; Kagami, M

    1984-05-01

    The effects of beryllium on cultured cells were investigated. Three cell-lines (HeLa-S3, Vero, HEL-R66) were used in these experiments and they were cultured in Eagle's MEM plus 5 or 10% FBS (Fetal Bovine Serum) containing beryllium in various concentrations. HeLa cells or Vero cells were able to grow in the medium with 10 micrograms Be/ml (1.1 mM). On the other hand, the growth of HEL cells were strongly inhibited, even when cultured in the medium with 1 microgram Be/ml (1.1 X 10(-1) mM) and the number of living cells showed markedly low level as compared to that of the control samples cultured in the medium without beryllium. The cytotoxic effects of beryllium on these cells, which were cultured for three days in the medium with beryllium, were observed. None of cytotoxic effects were found on HeLa cells cultured with 0.5 micrograms/ml (5.5 X 10(-2) mM) and on Vero cells cultured with 0.05 micrograms Be/ml (5.5 X 10(-3) mM), while HEL cells received cytotoxic effects even when cultured in the medium containing 0.05 micrograms Be/ml (5.5 X 10(-3) mM), and these effects on the cells appeared strong when cultured in the medium without FBS. It was revealed from these experiments that HEL cells are very sensitive in terms of toxic effects of beryllium. Therefore, there cells can be used for the toxicological study on low level concentrations of the metal.

  6. INNOVATIVE CULTURE IN SMALL AND MEDIUM ENTERPRISES

    OpenAIRE

    Aluisio Broering Mambrini; Seiji Cintho; Erni Dattein Dattein; Jorge Antonio Arias Medina; Emerson Antonio Maccari

    2011-01-01

    In the last two decades, innovation has been a key driver of economic growth. Innovation is closely related to creating value and generating wealth through successful service to consumer needs. Thus, it is not necessarily restricted to the use of new knowledge generated from research, but on the development of new products or services that are obtained with creative use of knowledge, new or already known. This study aimed to identify management practices that promote a culture of innovation i...

  7. Evaluation of primary and secondary production using wastewater as a culture medium.

    Science.gov (United States)

    Nandini, S; Ramírez-García, Pedro; Sarma, S S S

    2010-10-01

    The ability of rotifers and cladocerans to convert primary to secondary production in wastewaters was tested. Scenedesmus acutus was cultured on Bold's (defined) medium, wastewater from the tertiary phase of water treatment and a mixture of both. The algal growth rates (µ) ranged from 0.4 to 0.7 day⁻¹, being highest in defined medium. The demographic characteristics of Brachionus rubens and Moina macrocopa were tested using algae at a density of 1.0 x 10⁶ cells mL⁻¹. Into each test jar, we introduced 20 neonates (Moina (0.28-0.57 day⁻¹). Brachionus had significantly higher growth rates on algae cultured on Bold medium than on treated wastewater while Moina grew significantly better on Scenedesmus cultured on Bold medium or a mixture of treated wastewater and Bold medium than on treated wastewater alone. PMID:19748945

  8. Architecture of Institution & Home. Architecture as Cultural Medium

    OpenAIRE

    Robinson, J W

    2004-01-01

    This dissertation addresses how architecture functions as a cultural medium. It does so by by investigating how the architecture of institution and home each construct and support different cultural practices. By studying the design of ordinary settings in terms of how qualitative differences in architectural environments affect those who use them, this study exemplifies architectural research directed toward constructive cultural change. In the United States, ordinary people often describe h...

  9. Direct coating of culture medium from cells secreting classical swine fever virus E2 antigen on ELISA plates for detection of E2-specific antibodies.

    Science.gov (United States)

    Cheng, Ta-Chun; Pan, Chu-Hsiang; Chen, Chien-Shu; Chuang, Kuo-Hsiang; Chuang, Chih-Hung; Huang, Chien-Chaio; Chu, Yu-Yi; Yang, Ya-Chun; Chu, Pei-Yu; Kao, Chien-Han; Hsieh, Yuan-Chin; Cheng, Tian-Lu

    2015-07-01

    The envelope glycoprotein E2 of classical swine fever virus (CSFV) is widely used as a marker for measuring vaccine efficacy and antibody titer. The glycosylation profile of E2 may affect the immunogenicity of the vaccine and the timing of re-vaccination. In this study, a human embryonic kidney cell line was used to secrete fully-glycosylated CSFV E2, which was then coated onto ELISA plates without purification or adjustment. The resulting E2-secreting medium-direct-coating (E2-mDc) ELISA was successfully used to measure anti-E2 antibody titers in vaccinated and field pig sera samples. Compared with a virus neutralization test (as standard), the E2-mDc ELISA was found to be more accurate (90%) than a commercial CSFV antibody diagnostic kit (62%). In conclusion, the mammalian cell-secreted antigen can provide cheap, accurate and effective assays for vaccine efficacy and disease diagnoses. PMID:25975854

  10. Inulinase from Kluyveromyces marxianus: culture medium composition and enzyme extraction

    Directory of Open Access Journals (Sweden)

    A. PESSOA JR

    1999-09-01

    Full Text Available K. marxianus DSM 70106 was cultivated for inulinase production in a medium containing 2.0 g/L of yeast extract, 5.0 g/L of peptone and salts. The addition of corn steep liquor did not increase enzyme production. Inulin, as the main carbon source, afforded higher inulinase production than glucose, fructose, sucrose, lactose, maltose and starch. Glucose, fructose and sucrose reduced enzyme production by 46, 58 and 71%, respectively. By using the best culture medium enzyme activity remained stable for 22 months at 4oC; while at -18oC it decreased by 10%. Maximal activity was found in the pH range of 3.5 to 5.0 and at temperatures from 50 to 60oC. Flocculation was used for cell separation. Shifting the pH was more efficient than using polyelectrolytes, CaCl2, bentonite and Fe2O3. Recovery of inulinase by AOT(sodium di-2-ethylhexyl sulfosuccinate-reversed micelles yielded up to ~20%.

  11. 脐带间充质干细胞无血清和含胎牛血清培养体系比较%Comparison of the Umbilical Cord Mesenchymal Stem Cells Cultured by Serum-free Medium and Fetal Bovine Serum-contained Medium

    Institute of Scientific and Technical Information of China (English)

    陈林; 张坤; 董伟; 杨珂; 艾辉; 陈禄华

    2014-01-01

    为了比较在无血清与含胎牛血清培养基中人脐带间充质干细胞的增殖能力、生物学特性、分化潜能,将脐带华通氏胶剥出剪碎,分别在无血清与含胎牛血清培养基中培养扩增脐带间充质干细胞。用MTT法检测细胞增殖活性,并绘制生长曲线;在倒置显微镜下对细胞形态进行观察;利用流式细胞仪检测鉴定其表面标记CD73,CD90,CD105,CD14,CD34,CD45,CD79a及HLA-DR的表达;将成骨及成脂诱导液培养1~2周后观察其细胞形态学变化,用碱性磷酸酶染色鉴定其成骨情况,油红O染色鉴定其成脂情况。结果表明:无血清与含胎牛血清培养基所培养的人脐带间充质干细胞在细胞形态、增殖活性、表面标记和分化潜能相似;但在无血清培养基成分中无动物源蛋白引入,是更理想的人脐带间充质干细胞培养基种类。%To compare the differences of proliferation,biological characteristics,differentiation poten-tial of serum-free and fetal bovine serum-contained medium cultured human umbilical cord-derived mesenchymal stem cells.Separated Wharton’s jelly from human umbilical cord,cultured umbilical cord mesenchymal stem cells in serum-free medium and serum-supplied medium.Detected the cells’ proliferation rate by MTT method and get the cell growth curve ;observe the cell morphology by mi-croscope;Use flow cytometer to identify surface markers CD73,CD90,CD105,CD14,CD34, CD45 ,CD79 a,and HLA-DR.We observe cells change of shape after cells cultured by Osteogenesis Differentiation and Adiposeness Differentiation medium.Use alkaline phosphatase and oil red O staining methods to ensure cells’differentiation.The cell morphology,proliferation activity,surfacemarkers and differentiation potential of human umbilical cord mesenchymal stem cells cultured in ser-um-free medium are similar with which cultured in FBS-supplied medium .The composition of the ser-um-free medium are

  12. Optimization of ferric chloride concentration and pH to improve both cell growth and flocculation in Chlorella vulgaris cultures. Application to medium reuse in an integrated continuous culture bioprocess.

    Science.gov (United States)

    Lecina, Martí; Nadal, Gisela; Solà, Carles; Prat, Jordi; Cairó, Jordi J

    2016-09-01

    Combined effect of ferric chloride and pH on Chlorella vulgaris growth and flocculation were optimized using DoE. Afterwards, an integrated bioprocess for microalgae cultivation and harvesting conceived as a sole step was run in continuous operation mode. Microalgae concentration in a 2L-photobioreactor was about 0.5gL(-1) and the efficiency of flocculation in the coupled sedimentation tank was about 95%. Dewatered microalgae reached a biomass concentrations increase about 50-fold, whereas it was only about 0.02gL(-1) in the clarified medium. Then, the reuse of the clarified medium recovered was further evaluated. The clarified medium was reused without any further nutrient supplementation, whereas a second round of medium reuse was performed after supplementation of main nutrients (phosphate-sulfate-nitrate), micronutrients and ferric chloride. The medium reuse strategy did not affect cell growth and flocculation. Consequently, the reuse of medium reduces the nutrients requirements and the demand for water, and therefore the production costs should be reduced accordingly. PMID:27240237

  13. Optimization of ferric chloride concentration and pH to improve both cell growth and flocculation in Chlorella vulgaris cultures. Application to medium reuse in an integrated continuous culture bioprocess.

    Science.gov (United States)

    Lecina, Martí; Nadal, Gisela; Solà, Carles; Prat, Jordi; Cairó, Jordi J

    2016-09-01

    Combined effect of ferric chloride and pH on Chlorella vulgaris growth and flocculation were optimized using DoE. Afterwards, an integrated bioprocess for microalgae cultivation and harvesting conceived as a sole step was run in continuous operation mode. Microalgae concentration in a 2L-photobioreactor was about 0.5gL(-1) and the efficiency of flocculation in the coupled sedimentation tank was about 95%. Dewatered microalgae reached a biomass concentrations increase about 50-fold, whereas it was only about 0.02gL(-1) in the clarified medium. Then, the reuse of the clarified medium recovered was further evaluated. The clarified medium was reused without any further nutrient supplementation, whereas a second round of medium reuse was performed after supplementation of main nutrients (phosphate-sulfate-nitrate), micronutrients and ferric chloride. The medium reuse strategy did not affect cell growth and flocculation. Consequently, the reuse of medium reduces the nutrients requirements and the demand for water, and therefore the production costs should be reduced accordingly.

  14. The Modified Culture Medium and Special Method to Increase the Derivation of Human Embryonic Stem Cells%提高人胚胎干细胞建系率的优选培养体系与方法

    Institute of Scientific and Technical Information of China (English)

    骆玉梅; 范勇; 陈欣洁; 李少英; 郑育红; 徐霞; 孙筱放

    2012-01-01

    目的:建立一种从废弃胚胎中提高囊胚形成率和质量的培养体系,寻找多种促进内细胞团(ICM)数目增多、贴壁、增值的方法,提高人胚胎干细胞(human embryonic stem cell,hESC)建系效率,建立人胚胎干细胞库.方法:将179枚IVFDay3废弃的胚胎放入优选培养体系中培养(G2.5培养液中添加10%人血清蛋白,人白细胞抑制生长因子(hLIF),碱性成纤维细胞生长因子(bFGF)).到Day7将形成的囊胚全部用机械法分离ICM,接种于丝裂霉素C灭活处理的原代小鼠胚胎成纤维细胞(MEF)上,培养8-9天,每4-5天传代1次.结果:优选培养体系的囊胚形成率为29.1%(52/179),其中A级囊胚形成率为11.2%(20/179),50个ICM贴壁生长,20个出现克隆形态,成功建立11株hESC(FY-hES-11至FY-hES-21).11株hESC均具有共同的多能性生物学特性.结论:优选培养体系可以明显提高囊胚形成的质量,促进ICM的增值,纯熟的机械切割法可以避免损伤ICM并提高其贴壁率,原代灭活的MEF饲养层可以明显促进细胞增殖.%Objective: To establish a modified culture medium to improve the blastocyst formation rate and quality from the discarded embryos, and find good methods to promote the number of inner cell mass (ICM) increase, adherence and growth, so as to increase the derivation of human embryonic stem cells (human embryonic stem cell, hESC ) and build hESC bank. Methods: We collected 179 Day3 discarded embryos into the modified culture medium (G2.S medium supplemented with 10% human serum albumin, human leukocyte inhibiting growth factor (hLIF), basic fibroblast growth factor (bFGF)). At Day7 separated ICM from the blastocyst and seeded on the mouse embryonic fibroblasts (MEF) of passage 0 which have been inactivated by mitomycin C. The cells were cultured 8 to 9 days with hESC medium and passaged every 4 to 5 days. Results: The data showed that the formation of blastocyst in the modified culture medium was 29.1% (52/179), and

  15. 21 CFR 866.2300 - Multipurpose culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Multipurpose culture medium. 866.2300 Section 866.2300 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2300 Multipurpose...

  16. 21 CFR 866.2320 - Differential culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Differential culture medium. 866.2320 Section 866.2320 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices § 866.2320 Differential...

  17. Culture medium for amylase production by toxigenic fungi

    Directory of Open Access Journals (Sweden)

    Figueira Edson Luiz Zangrando

    2000-01-01

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

  18. 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)

  19. Development of a rapid lateral flow immunoassay test for detection of exosomes previously enriched from cell culture medium and body fluids.

    Science.gov (United States)

    Oliveira-Rodríguez, Myriam; López-Cobo, Sheila; Reyburn, Hugh T; Costa-García, Agustín; López-Martín, Soraya; Yáñez-Mó, María; Cernuda-Morollón, Eva; Paschen, Annette; Valés-Gómez, Mar; Blanco-López, Maria Carmen

    2016-01-01

    Exosomes are cell-secreted nanovesicles (40-200 nm) that represent a rich source of novel biomarkers in the diagnosis and prognosis of certain diseases. Despite the increasingly recognized relevance of these vesicles as biomarkers, their detection has been limited due in part to current technical challenges in the rapid isolation and analysis of exosomes. The complexity of the development of analytical platforms relies on the heterogeneous composition of the exosome membrane. One of the most attractive tests is the inmunochromatographic strips, which allow rapid detection by unskilled operators. We have successfully developed a novel lateral flow immunoassay (LFIA) for the detection of exosomes based on the use of tetraspanins as targets. We have applied this platform for the detection of exosomes purified from different sources: cell culture supernatants, human plasma and urine. As proof of concept, we explored the analytical potential of this LFIA platform to accurately quantify exosomes purified from a human metastatic melanoma cell line. The one-step assay can be completed in 15 min, with a limit of detection of 8.54×10(5) exosomes/µL when a blend of anti-CD9 and anti-CD81 were selected as capture antibodies and anti-CD63 labelled with gold nanoparticles as detection antibody. Based on our results, this platform could be well suited to be used as a rapid exosome quantification tool, with promising diagnostic applications, bearing in mind that the detection of exosomes from different sources may require adaptation of the analytical settings to their specific composition. PMID:27527605

  20. Development of a rapid lateral flow immunoassay test for detection of exosomes previously enriched from cell culture medium and body fluids

    Directory of Open Access Journals (Sweden)

    Myriam Oliveira-Rodríguez

    2016-08-01

    Full Text Available Exosomes are cell-secreted nanovesicles (40–200 nm that represent a rich source of novel biomarkers in the diagnosis and prognosis of certain diseases. Despite the increasingly recognized relevance of these vesicles as biomarkers, their detection has been limited due in part to current technical challenges in the rapid isolation and analysis of exosomes. The complexity of the development of analytical platforms relies on the heterogeneous composition of the exosome membrane. One of the most attractive tests is the inmunochromatographic strips, which allow rapid detection by unskilled operators. We have successfully developed a novel lateral flow immunoassay (LFIA for the detection of exosomes based on the use of tetraspanins as targets. We have applied this platform for the detection of exosomes purified from different sources: cell culture supernatants, human plasma and urine. As proof of concept, we explored the analytical potential of this LFIA platform to accurately quantify exosomes purified from a human metastatic melanoma cell line. The one-step assay can be completed in 15 min, with a limit of detection of 8.54×105 exosomes/µL when a blend of anti-CD9 and anti-CD81 were selected as capture antibodies and anti-CD63 labelled with gold nanoparticles as detection antibody. Based on our results, this platform could be well suited to be used as a rapid exosome quantification tool, with promising diagnostic applications, bearing in mind that the detection of exosomes from different sources may require adaptation of the analytical settings to their specific composition.

  1. Selection of osteoprogenitors from the jaw periosteum by a specific animal-free culture medium.

    Directory of Open Access Journals (Sweden)

    Dorothea Alexander

    Full Text Available The goal of our research work is to establish mesenchymal osteoprogenitors derived from human jaw periosteum for tissue engineering applications in oral and maxillofacial surgery. For future autologous and/or allogeneic transplantations, some issues must be addressed. On the one hand, animal-free culture conditions have yet to be established. On the other hand, attempts should be undertaken to shorten the in vitro culturing process efficiently. The aim of the present study is to compare and analyze the phenotype of osteoprogenitors from the jaw periosteum under normal FCS-containing and animal-free culture conditions. Therefore, we analyzed the proliferation rates of MesenCult-XF medium (MC- in comparison to DMEM-cultured JPCs. Whereas jaw periosteal cells (JPCs show relatively slow proliferation rates and a fibroblastoid shape under DMEM culture conditions, MC-cultured JPCs diminished their cell size significantly and proliferated rapidly. By live-monitoring measurements of adhesion and proliferation, we made an interesting observation: whereas the proliferation of the MSCA-1(+ subpopulation and the unseparated cell fraction were favored by the animal-free culture medium, the proliferation of the MSCA-1(- subpopulation seemed to be repressed under these conditions. The alkaline phosphatase expression pattern showed similar results under both culture conditions. Comparison of the mineralization capacity revealed an earlier formation of calcium-phosphate precipitates under MC culture conditions; however, the mineralization capacity of the DMEM-cultured cells seemed to be higher. We conclude that the tested animal-free medium is suitable for the in vitro expansion and even for the specific selection of osteoprogenitor cells derived from the jaw periosteum.

  2. Promising medium duration multiple resistant rice culture CB 05 022

    Directory of Open Access Journals (Sweden)

    R. Pushpam,S. Robin, S.Rajeswari, K. Mohanasundaram and K. Thiagarajan

    2010-07-01

    Full Text Available CB 05022, the medium duration rice culture with the parentage of CO 43/ ADT 39 was developed at Department of Rice,TNAU, Coimbatore. The culture was evaluated at station trials and it gave 21.8% increased yield over ADT (R 46. Thisculture was evaluated under AICRIP in irrigated transplanted low land ecosystem as IET 20884 for two years 2006-2008.Under IVT-IM the culture recorded a mean grain yield of 5485 kg/ha which was 21, 22% increase over the checks Jayaand Triguna respectively. Again the culture was tested under IVT-L during kharif 2009 wherein it recorded a mean grainyield of 5551 kg/ha which was 11.4% increase over Savitri. In the region wise performance, this culture ranked first inEastern region and fifth in Western region. In overall basis, it ranked second among all entries. The culture was alsotested under Disease Screening Nursery during 2009 and it showed multiple disease resistance again leaf blast, neck blastand brown spot. The culture is having good quality characteristics of intermediate amylose and GT, soft GC, high milling(70.7% and head rice recovery (60.6. The culture is now under third year of testing under AICRIP. Owing to highyielding, multiple resistance to major diseases, this culture has great scope for exploitation under our varietalimprovement programmes.

  3. 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.

  4. Fish stem cell cultures.

    Science.gov (United States)

    Hong, Ni; Li, Zhendong; Hong, Yunhan

    2011-04-13

    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.

  5. Evaluation of Endotoxin in Culture Medium for Human in vitro Fertilization

    Institute of Scientific and Technical Information of China (English)

    Jing LI; Wei-jie ZHU; Wen-hong ZHANG

    2005-01-01

    Objective To determine whether the presence of bacterial endotoxin in the commercial culture media utilized for human in vitro fertilization (IVF), and evaluate the difference in detecting endotoxin in culture medium between the human sperm motility assay and the 2-cell mouse embryo assay.Methods Thirty-six batches of culture media commonly used in IVF laboratories from 3 manufacturers were determined for the presence ofendotoxin before using the medium for the assisted reproductive programs (group A). After being used, 25 specimens among above media were also tested (group B). The chromogenic limulus amoebocyte lysate (LAL) test was used for quantification the content of endotoxin. In addition, the human sperm motility assay was compared with the 2-cell mouse embryo assay to evaluate the difference in detecting endotoxin in culture medium.Results Endotoxin was not detected in group A. However, 2 samples were positive in group B. Sperm did not show significant change in motility in group A during 24 h of incubation when compared with the control (P>0.05). However, in group A the 2-cell embryo development to blastocyst was suppressed in 3 batches of media.Conclusions Regular screening of each batch of culture medium should be performed if possible although there was no evidence of endotoxin contamination in commercially prepared pre-tested media. Culture environment should be stringently controlled in case the medium is polluted. The sensitivity of the sperm motility assay was lower than that of the mouse embryo assay for detecting low levels of endotoxin or toxic compounds in the medium.

  6. Effects of Proteins from Culture Medium on Surface Property of Silanes- Functionalized Magnetic Nanoparticles

    Directory of Open Access Journals (Sweden)

    Chen ZP

    2008-01-01

    Full Text Available Abstract Monodisperse magnetic nanoparticles (MNPs were synthesized by thermal decomposition of iron-oleate and functionalized with silanes bearing various functional groups such as amino group (NH2, short-chain poly(ethylene glycol (PEG, and carboxylic group (COOH. Then, silanes-functionalized magnetic nanoparticles (silanes-MNPs were incubated in cell culture medium plus fetal calf serum to investigate the effects of proteins from culture medium on surface property of MNPs. Zeta potential measurements showed that although surface charges of silanes-MNPs were different, they exhibited negative charges at neutral pH and approximate isoelectric points after they were incubated in cell culture medium. The reason was that silanes-MNPs could easily adsorb proteins from culture medium via non-covalent binding, resulting in the formation of protein-silanes-MNPs conjugates. Moreover, silanes-MNPs with various functional groups had different adsorption capacity to proteins, as confirmed by Coomassie blue fast staining method. The in vitro cell experiments showed that protein-silanes-MNPs had higher cellular uptake by cancer cells than silanes-MNPs.

  7. Production of Normal Mammalian Organ Culture Using a Medium Containing Mem-Alpha, Leibovitz L 15, Glucose Galactose Fructose

    Science.gov (United States)

    Goodwin, Thomas J. (Inventor); Wolf, David A. (Inventor); Spaulding, Glenn F. (Inventor); Prewett, Tacey L. (Inventor)

    1999-01-01

    Normal mammalian tissue and the culturing process has been developed for the three groups of organ, structural and blood tissue. The cells are grown in vitro under micro- gravity culture conditions and form three dimensional cells aggregates with normal cell function. The microgravity culture conditions may be microgravity or simulated microgravity created in a horizontal rotating wall culture vessel. The medium used for culturing the cells, especially a mixture of epithelial and mesenchymal cells contains a mixture of Mem-alpha and Leibovits L15 supplemented with glucose, galactose and fructose.

  8. Aleuria aurantia - indole metabolites of fruit bodies, mycelial culture and culture medium

    Directory of Open Access Journals (Sweden)

    Janina Węgiel

    2014-08-01

    Full Text Available The aim of present study was to investigate and compare indole metabolites of fruit bodies, mycelium cultivated in vitro and culture medium of the fungus Aleuria aurantia (Fr. Fuck. By use of a number of chromatographic and spectroscopic methods several indole metabolites have been detected and identified among other the 3-indolebutyric acid was produced and extracted to the culture medium. Furthermore 3-indoleatonitrile and tryptophane degradative products have been found both in fruit bodies and mycelium.

  9. Aleuria aurantia - indole metabolites of fruit bodies, mycelial culture and culture medium

    OpenAIRE

    Janina Węgiel; Stanisław Kohlmünzer

    2014-01-01

    The aim of present study was to investigate and compare indole metabolites of fruit bodies, mycelium cultivated in vitro and culture medium of the fungus Aleuria aurantia (Fr.) Fuck. By use of a number of chromatographic and spectroscopic methods several indole metabolites have been detected and identified among other the 3-indolebutyric acid was produced and extracted to the culture medium. Furthermore 3-indoleatonitrile and tryptophane degradative products have been found both in fruit bodi...

  10. Sugarcane Bagasse: A Potential Medium for Fungal Cultures

    OpenAIRE

    Arushdeep Sidana; Umar Farooq

    2014-01-01

    Worldwide, sugarcane industries produce tons of sugarcane bagasse as residual/waste material. This residual material is rich in complex lignocellulosic substances and may be used as a low cost carbon and energy source for the growth of fungal species. The present work was aimed at designing a sugarcane waste-based medium as a substitute for expensive commercial media for growing fungal cultures. Eight species of fungi, namely, Aspergillus niger, Candida albicans, Saccharomyces cerevisiae, Fus...

  11. 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

  12. Cell survival of glioblastoma grown in medium containing hydrogen peroxide and/or nitrite, or in plasma-activated medium.

    Science.gov (United States)

    Kurake, Naoyuki; Tanaka, Hiromasa; Ishikawa, Kenji; Kondo, Takashi; Sekine, Makoto; Nakamura, Kae; Kajiyama, Hiroaki; Kikkawa, Fumitaka; Mizuno, Masaaki; Hori, Masaru

    2016-09-01

    Non-equilibrium atmospheric pressure plasmas generate a high electron density (on the order of 10(16) electrons per cm(-3)) using Ar gas. Culture medium in air at room temperature was plasma-irradiated for several hundred seconds. Tens of micromolar hydrogen peroxide (H2O2) and millimolar levels of nitrous ion (NO2(-)) were detected in the plasma-irradiated culture medium (plasma activated medium; PAM) and selectively induced the apoptotic death of glioblastoma tumor cells, but did not kill normal mammary epithelial cells. A similar antitumor effect was induced by spiking the medium with comparable concentrations of H2O2 and NO2(-). The PAM remained still a somewhat difference that it should also be assessed for understanding other latent mechanisms. PMID:26820218

  13. Estrogen and phenol red free medium for osteoblast culture: study of the mineralization ability.

    Science.gov (United States)

    de Faria, A N; Zancanela, D C; Ramos, A P; Torqueti, M R; Ciancaglini, P

    2016-08-01

    To design an estrogen and phenol red free medium for cell culture and check its effectiveness and safety on osteoblast growth it is necessary to maintain the estrogen receptors free for tests. For this purpose, we tested some modifications of the traditional culture media: estrogen depleted fetal bovine serum; estrogen charcoal stripped fetal bovine serum and phenol red free α-MEM. The aim of this work is to examine the effects of its depletion in the proliferation, differentiation, and toxicity of mesenchymal stromal cells differentiated into osteoblasts to obtain an effective interference free culture medium for in vitro studies, focused on non-previously studied estrogen receptors. We performed viability tests using the following techniques: MTT, alkaline phosphatase specific activity, formation of mineralized matrix by Alizarin technique and analysis of SEM/EDX of mineralized nodules. The results showed that the culture media with estrogen free α-MEM + phenol red free α-MEM did not impact viability, alkaline phosphatase activity and mineralization of the osteoblasts culture compared to control. In addition, its nodules possess Ca/P ratio similar to hydroxyapatite nodules on the 14th and 21st day. In conclusion, the modified culture medium with phenol red free α-MEM with estrogen depleted fetal bovine serum can be safely used in experiments where the estrogen receptors need to be free. PMID:25634598

  14. Development and optimisation of a procedure for the production of Parapoxvirus ovis by large-scale microcarrier cell culture in a non-animal, non-human and non-plant-derived medium.

    Science.gov (United States)

    Pohlscheidt, M; Langer, U; Minuth, T; Bödeker, B; Apeler, H; Hörlein, H-D; Paulsen, D; Rübsamen-Waigmann, H; Henzler, H-J; Reichl, U

    2008-03-17

    For the production of a chemically inactivated Parapoxvirus ovis (PPVO), an adherent bovine kidney cell line was cultivated on Cytodex-3 microcarriers in suspension culture. The inactivated and purified virus particles have shown immune modulatory activity in several animal models. PPVO was produced by a biphasic batch process at the 3.5 and 10 L scale. Aeration was realised by bubble-free membrane oxygenation via a tube stator with a central two-blade anchor impeller. In order to increase efficiency, process robustness and safety, the established process was optimised. The cell line was adapted to a protein-free medium (except recombinant insulin) in order to increase biosafety. A scale up to a 50 L pilot plant with direct cell expansion was performed successfully. In parallel, the biphasic batch process was optimised with special emphasis on different operating conditions (cell number, Multiplicity of Infection (MOI), etc.) and process management (fed-batch, dialysis, etc.). The quality and concentration of the purified virus particles was assessed by quantitative electron microscopy, residual host cell protein and DNA-content and, finally, biologic activity in a transgenic mouse model. This integrated approach led to a new, safe, robust and highly productive large-scale production process, called "Volume-Expanded-Fed" Batch with cell densities up to 6-7e06 cells/mL. By subsequent dilution of infected cells into the next process scale, an increase in total productivity by a factor of 40 (related to an established biphasic batch process) was achieved. PMID:18295380

  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. Effects of glucose concentration, medium osmotic pressure and light intensity on the growth of Marchantia paleacea var. diptera cells in photomixotrophic culture; Hikari kongo eiyo baiyo ni okeru futaba zenigoke saibo no zoshoku ni oyobosu glucose nodo, baichi shintoatsu to hikari kyodo no koka

    Energy Technology Data Exchange (ETDEWEB)

    Kinooka, M.; Miyaoka, M.; Taya, M.; Tone, S. [Osaka University, Osaka (Japan); Ono, K. [Kumamoto University, Kumamoto (Japan)

    1997-07-10

    Batch cultures of photomixotrophic cells of Marchantia paleacea var. diptera were conducted at 25degC under the initial conditions of glucose concentration of 20 or 40 kg/m{sup 3}, medium osmotic pressure of 520 or 830 kPa and mean incident light intensity of 0, 13 or 28 W/m{sup 2}. The glucose concentration, osmotic pressure and mean light intensity in medium declined with increasing cell mass concentration, so that they became limiting factors against the cell growth during the cultures. A kinetic expression for the cell growth was presented by considering the effects of three limiting factors, and applied to the data obtained from the batch cultures. The calculation results fitted closely with the experimental data in the cultures. 8 refs., 2 tabs., 1 tab.

  17. Cell Culturing of Cytoskeleton

    Science.gov (United States)

    2004-01-01

    Biomedical research offers hope for a variety of medical problems, from diabetes to the replacement of damaged bone and tissues. Bioreactors, which are used to grow cells and tissue cultures, play a major role in such research and production efforts. Cell culturing, such as this bone cell culture, is an important part of biomedical research. The BioDyn payload includes a tissue engineering investigation. The commercial affiliate, Millenium Biologix, Inc. has been conducting bone implant experiments to better understand how synthetic bone can be used to treat bone-related illnesses and bone damaged in accidents. On STS-95, the BioDyn payload will include a bone cell culture aimed to help develop this commercial synthetic bone product. Millenium Biologix, Inc. is exploring the potential for making human bone implantable materials by seeding its proprietary artificial scaffold material with human bone cells. The product of this tissue engineering experiment using the Bioprocessing Modules (BPMs) on STS-95 is space-grown bone implants, which could have potential for dental implants, long bone grafts, and coating for orthopedic implants such as hip replacements.

  18. 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

  19. Dense cultures of Neisseria gonorrhoeae in liquid medium.

    Science.gov (United States)

    Brookes, R; Hedén, C G

    1967-03-01

    Cultivation of Neisseria gonorrhoeae was effected in a conical glass culture vessel surrounded by a constant-temperature water jacket, and with facilities for stirring, aeration, and pH measurement and control. With the use of an aerated peptone-based medium, containing polypropylene glycol to prevent foam build-up, the yields obtained over the pH range from 5.8 to 7.4 were determined. The greatest yield was obtained at pH 6.4 when the dry weight was 1.5 g/liter. At pH 7.2 to 7.6, lysis was extensive.

  20. 21 CFR 866.1700 - Culture medium for antimicrobial susceptibility tests.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Culture medium for antimicrobial susceptibility tests. 866.1700 Section 866.1700 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND....1700 Culture medium for antimicrobial susceptibility tests. (a) Identification. A culture medium...

  1. Influence of Culture Conditions and Medium Composition on the Production of Cellulose by Shiga Toxin-Producing Escherichia coli Cells

    OpenAIRE

    Yoo, Byong Kwon; Chen, Jinru

    2009-01-01

    Culture conditions favoring cellulose production by Shiga toxin-producing Escherichia coli included a 28°C incubation temperature, an aerobic atmosphere, and the presence of 2% ethanol in Luria-Bertani no-salt agar with pH 6.0 and a water activity of 0.99. These findings will assist in formulating microbiological media useful for cellulose and biofilm research.

  2. High Pdr12 levels in spoilage yeast (Saccharomyces cerevisiae) correlate directly with sorbic acid levels in the culture medium but are not sufficient to provide cells with acquired resistance to the food preservative.

    Science.gov (United States)

    Papadimitriou, Minas N B; Resende, Catarina; Kuchler, Karl; Brul, Stanley

    2007-01-25

    Sorbic acid is a commonly used food preservative against yeast and fungal food spoilage. Understanding its effect on the molecular physiology of yeast cells will allow the food industry to develop knowledge-based strategies to make more optimal use of its preservative action. Here we show that the yeast membrane protein Pdr12, previously shown to be prominently involved in sorbic acid resistance development in laboratory strains, was strongly induced by the presence of sorbic acid in the culture medium in Saccharomyces strains isolated from spoiled foods. Induction of Pdr12 expression was seen both under laboratory conditions and upon growth in a commercial soft drink. Induction was rapid and maintained for the duration of the stress. No Pdr12-like protein induction was seen in Zygosaccharomyces bailii or Zygosaccharomyces lentus, two well-known beverages spoilage organisms. Finally, unexpectedly, our studies showed for the first time that pre-inducing Pdr12p to maximal levels by subjecting cells to a mild sorbic acid stress did not lead to cells with an acquired resistance. Neither more rapid growth in the presence of the acid nor growth at higher sorbic acid concentrations at a given environmental pH was observed. Thus we have shown that while important in resistance development against sorbic acid, by itself induction of the pump is not sufficient to acquire resistance to the preservative.

  3. In vitro differentiation of mouse embryonic stem cells into inner ear hair cell-like cells using stromal cell conditioned medium

    OpenAIRE

    Ouji, Y; Ishizaka, S.; NAKAMURA-UCHIYAMA, F; Yoshikawa, M

    2012-01-01

    Hearing loss is mainly caused by loss of sensory hair cells (HCs) in the organ of Corti or cochlea. Although embryonic stem (ES) cells are a promising source for cell therapy, little is known about the efficient generation of HC-like cells from ES cells. In the present study, we developed a single-medium culture method for growing embryoid bodies (EBs), in which conditioned medium (CM) from cultures of ST2 stromal cells (ST2-CM) was used for 14-day cultures of 4-day EBs. At the end of the 14-...

  4. 不同因子影响骨髓间充质干细胞的多向分化%Different ingredients of cell culture medium influence multi-differentiation of bone marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    栗扬阳; 赵庆华

    2013-01-01

      背景:在组织工程领域,关于骨髓间充质干细胞定向诱导分化的研究越来越多,但是细胞培养基中不同成分会对骨髓间充质干细胞的体外增殖分化产生影响。目的:针对培养基中不同因子对骨髓间充质干细胞定向诱导分化的作用加以综述。方法:第一作者应用计算机检索1998年1月至2012年4月Pubmed数据库及万方数据库。检索英文关键词为“bone marrow mesenchymal stromal cel s, cel culture medium, differentiation”,中文关键词为“骨髓间充质干细胞,细胞培养,定向诱导分化”,纳入有关不同因子对骨髓间充质干细胞向成骨细胞、软骨细胞和脂肪细胞定向诱导分化作用的文献,排除重复研究。结果与结论:计算机初检共得到184篇文献,根据纳入排除标准,对其中30篇文献进行综述。大体说来,骨髓间充质干细胞向成骨分化的主要因子有地塞米松、转化生长因子、维生素 C、维生素 D3、β-甘油磷酸钠及乙烯雌酚等;向软骨分化的主要因子有地塞米松、转化生长因子、维生素C、胰岛素样生长因子及成纤维细胞生长因子等;向脂肪细胞转化的主要因子有地塞米松、3-异丁基-1-甲基黄嘌呤、胰岛素和消炎痛等,但是其中一些因子的作用机制及不良反应还不明确,需要进一步的研究与验证。同时,骨髓间充质干细胞在骨髓中的含量较低,不同的分离方法会导致不同的分离率,因此如何选取一种分离率高的分离方法仍有待研究。%  BACKGROUND: In the field of tissue engineering, there are an increasing number of studies describing oriented differentiation of bone marrow mesenchymal stem cel s. Different ingredients of culture medium produce effects on in vitro proliferation and differentiation of bone marrow mesenchymal stem cel s. OBJECTIVE: To summarize the effects of different ingredients of cel culture

  5. 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

    A streamlined, simple technique for primary cell culture from E17 rat tissue is presented. In an attempt to standardize culturing methods for all neuronal cell types in the embryo, we evaluated a commercial medium without serum and used similar times for trypsinization and tested different surfaces...... for plating. In 1 day, using one method and a single medium, it is possible to produce robust E17 cultures of dorsal root ganglia (DRG), cerebellum, and enteric plexi. Allowing the endogenous glial cells to repopulate the cultures saves time compared with existing techniques, in which glial cells are added...... to cultures first treated with antimitotic agents. It also ensures that all the cells present in vivo will be present in the culture. Myelination commences after approximately 2 weeks in culture for dissociated DRG and 3-4 weeks in cerebellar cultures. In enteric cultures, glial wrapping of the enteric...

  6. Dynamic culture improves cell reprogramming efficiency.

    Science.gov (United States)

    Sia, Junren; Sun, Raymond; Chu, Julia; Li, Song

    2016-06-01

    Cell reprogramming to pluripotency is an inefficient process and various approaches have been devised to improve the yield of induced pluripotent stem cells. However, the effect of biophysical factors on cell reprogramming is not well understood. Here we showed that, for the first time, dynamic culture with orbital shaking significantly improved the reprogramming efficiency in adherent cells. Manipulating the viscosity of the culture medium suggested that the improved efficiency is mainly attributed to convective mixing rather than hydrodynamic shear stress. Temporal studies demonstrated that the enhancement of reprogramming efficiency required the dynamic culture in the middle but not early phase. In the early phase, fibroblasts had a high proliferation rate, but as the culture became over-confluent in the middle phase, expression of p57 was upregulated to inhibit cell proliferation and consequently, cell reprogramming. Subjecting the over confluent culture to orbital shaking prevented the upregulation of p57, thus improving reprogramming efficiency. Seeding cells at low densities to avoid over-confluency resulted in a lower efficiency, and optimal reprogramming efficiency was attained at a high seeding density with dynamic culture. Our findings provide insight into the underlying mechanisms of how dynamic culture condition regulate cell reprogramming, and will have broad impact on cell engineering for regenerative medicine and disease modeling.

  7. Development of a rapid lateral flow immunoassay test for detection of exosomes previously enriched from cell culture medium and body fluids

    OpenAIRE

    Oliveira-Rodríguez, Myriam; López-Cobo, Sheila; Hugh T. Reyburn; López-Martín, Soraya; Yáñez Mo, María; Cernuda-Morollón, Eva; Paschen, Annette; Valés-Gómez, Mar; Blanco-López, Maria Carmen; Costa-García, Agustín

    2016-01-01

    Exosomes are cell-secreted nanovesicles (40–200 nm) that represent a rich source of novel biomarkers in the diagnosis and prognosis of certain diseases. Despite the increasingly recognized relevance of these vesicles as biomarkers, their detection has been limited due in part to current technical challenges in the rapid isolation and analysis of exosomes. The complexity of the development of analytical platforms relies on the heterogeneous composition of the exosome membrane. One of the most ...

  8. How do culture media influence in vitro perivascular cell behavior?

    Science.gov (United States)

    Huber, Birgit; Volz, Ann-Cathrin; Kluger, Petra Juliane

    2015-12-01

    Perivascular cells are multilineage cells located around the vessel wall and important for wall stabilization. In this study, we evaluated a stem cell media and a perivascular cell-specific media for the culture of primary perivascular cells regarding their cell morphology, doubling time, stem cell properties, and expression of cell type-specific markers. When the two cell culture media were compared to each other, perivascular cells cultured in the stem cell medium had a more elongated morphology and a faster doubling rate and cells cultured in the pericyte medium had a more typical morphology, with several filopodia, and a slower doubling rate. To evaluate stem cell properties, perivascular cells, CD146(-) cells, and mesenchymal stem cells (MSCs) were differentiated into the adipogenic, osteogenic, and chondrogenic lineages. It was seen that perivascular cells, as well as CD146(-) cells and MSCs, cultured in stem cell medium showed greater differentiation than cells cultured in pericyte-specific medium. The expression of pericyte-specific markers CD146, neural/glial antigen 2 (NG2), platelet-derived growth factor receptor-β (PDGFR-β), myosin, and α-smooth muscle actin (α-SMA) could be found in both pericyte cultures, as well as to varying amounts in CD146(-) cells, MSCs, and endothelial cells. The here presented work shows that perivascular cells can adapt to their in vitro environment and cell culture conditions influence cell functionality, such as doubling rate or differentiation behavior. Pericyte-specific markers were shown to be expressed also from cells other than perivascular cells. We can further conclude that CD146(+) perivascular cells are inhomogeneous cell population probably containing stem cell subpopulations, which are located perivascular around capillaries.

  9. Optimizing culture medium for meristem tissue culture of several Saccharum species and commercial hybrids

    Science.gov (United States)

    The optimal range of medium nutrients and plant growth regulators (PGR) was investigated for in vitro culture of diverse sugarcane species and cultivars. Macro-nutrients, nitrogen (N), phosphorous (P) and potassium (K), were essential for growth of leaf primordia. Although the best concentration of ...

  10. Growth regulation of cultured human nevus cells.

    Science.gov (United States)

    Mancianti, M L; Györfi, T; Shih, I M; Valyi-Nagy, I; Levengood, G; Menssen, H D; Halpern, A C; Elder, D E; Herlyn, M

    1993-03-01

    Cells isolated from congenital melanocytic nevi and cultured in vitro have growth characteristics that resemble their premalignant stage in situ. A serum-free, chemically defined medium has been developed that allows continuous growth of established nevus cultures for up to several months. Like primary melanoma cells, nevus cells in high-calcium-containing W489 medium require insulin for growth. In contrast to melanoma cells, nevus cells in serum-free medium require the presence of alpha-melanocyte-stimulating hormone, which enhanced intracellular levels of cyclic adenosine monophosphate. In contrast to the requirements of normal human melanocytes from newborn foreskin, congenital nevus cells grow with less dependency on basic fibroblast growth factor (bFGF). Nevus cultures contain bFGF-like activity, and they express bFGF mRNA. Nevic cells of compound nevi also express bFGF mRNA in situ but only in the junctional areas. These results indicate that bFGF plays an important growth regulatory role for nevus cells in vitro and in vivo. PMID:8440904

  11. 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.

  12. Participation of cob tissue in the transport of medium components into maize kernels cultured in vitro

    International Nuclear Information System (INIS)

    Maize (Zea mays L.) kernels cultured in vitro while still attached to cob pieces have been used as a model system to study the physiology of kernel development. In this study, the role of the cob tissue in uptake of medium components into kernels was examined. Cob tissue was essential for in vitro kernel growth, and better growth occurred with larger cob/kernel ratios. A symplastically transported fluorescent dye readily permeated the endosperm when supplied in the medium, while an apoplastic dye did not. Slicing the cob tissue to disrupt vascular connections, but not apoplastic continuity, greatly reduced [14C]sucrose uptake into kernels. [14C]Sucrose uptake by cob and kernel tissue was reduced 31% and 68%, respectively, by 5 mM PCMBS. L-[14C]glucose was absorbed much more slowly than D-[14C]glucose. These and other results indicate that phloem loading of sugars occurs in the cob tissue. Passage of medium components through the symplast cob tissue may be a prerequisite for uptake into the kernel. Simple diffusion from the medium to the kernels is unlikely. Therefore, the ability of substances to be transported into cob tissue cells should be considered in formulating culture medium

  13. Differentiation of bone marrow mesenchymal stem cells induced by myocardial medium under hypoxic conditions

    Institute of Scientific and Technical Information of China (English)

    Xiao-jie XIE; Jian-an WANG; Jiang CAO; Xing ZHANG

    2006-01-01

    Aim: To explore whether bone marrow mesenchymal stem cells (MSC) can differentiate into myocardial-like cells induced by myocardial medium, especially the hypoxia/reoxygenation-conditioned medium of cardiomyocytes. Methods: Myocardial cells obtained from neonatal Sprague-Dawley rat ventricles were isolated and cultured in vitro and a hypoxia reoxygenation model was established. The MSC isolated from adult Sprague-Dawley rats were purified and then incubated with 3 different mediums: medium A - the conditioned medium of normal cardiomyocytes; medium B - the conditioned medium of cardiomyocytes after hypoxia reoxygenation; and the control medium - ordinary medium. The expressions of the cardiac myosin heavy chain (MHC), troponin T(TnT) and connexin 43 were investigated in the MSC after 24 h, 48 h and 72 h cultivation, respectively. Results: The MSC expressed MHC and TnT when incubated with the conditioned medium of cardiomyocytes after hypoxia reoxygenation, but did not express connexin 43. None of MHC, TnT and connexin 43 was detected in the MSC incubated with the conditioned medium of normal cardiomyocytes. Conclusion: The results indicate for the first time that myocardial medium for hypoxic preconditioning can induce MSC differentiation into myocardial-like cells.

  14. M8-An effective medium for anther culture of indica rice

    Institute of Scientific and Technical Information of China (English)

    MEIChuansheng; ZHANGJinyu; WuGuangnan

    1992-01-01

    A new dedifferentiation medium (MS) was developed, which greatly improved the efficiency of anther culture of indica rive. The percentage of green plantlets for anthers inoculated on M8 medium was 40% higher than that on N6 medium in 6 cultivars and it was 2.6%, on average, on M8 medium in more than 20 cultivars and lines,

  15. Cultivation and irradiation of human fibroblasts in a medium enriched with platelet lysate for obtaining feeder layer in epidermal cell culture; Cultivo e irradiacao de fibroblastos humanos em meio enriquecido com lisado de plaquetas para obtencao de camada de sustentacao em culturas de celulas da epiderme

    Energy Technology Data Exchange (ETDEWEB)

    Yoshito, Daniele

    2011-07-01

    For over 30 years, the use of culture medium, enriched with bovine serum, and murines fibroblasts, with the rate of proliferation controlled by irradiation or by share anticarcinogenic drugs, has been playing successfully its role in assisting in the development of keratinocytes in culture, for clinical purposes. However, currently there is a growing concern about the possibility of transmitting prions and animals viruses to transplanted patients. Taking into account this concern, the present work aims to cultivate human fibroblasts in a medium enriched with human platelets lysate and determine the irradiation dose of these cells, for obtaining feeder layer in epidermal cell culture. For carrying out the proposed objective, platelets lysis has standardized, this lysate was used for human fibroblasts cultivation and the irradiation dose enough to inhibit its duplication was evaluated. Human keratinocytes were cultivated in these feeder layers, in culture medium enriched with the lysate. With these results we conclude that the 10% platelets lysate promoted a better adhesion and proliferation of human fibroblasts and in all dose levels tested (60 to 300 Gy), these had their mitotic activity inactivated by ionizing irradiation, being that the feeder layers obtained with doses from 70 to 150 Gy were those that provided the best development of keratinocytes in medium containing 2.5% of human platelet lysate. Therefore, it was possible to standardize both the cultivation of human fibroblasts as its inactivation for use as feeder layer in culture of keratinocytes, so as to eliminate xenobiotics components. (author)

  16. An alternative to evaluate the efficiency of in vitro culture medium using a logistic regression model

    Directory of Open Access Journals (Sweden)

    Daniel Furtado Ferreira

    2003-01-01

    Full Text Available The evaluation of a culture medium for the in vitro culture of a species is performed using its physical and/or chemical properties. However, the analysis of the experimental results makes it possible to evaluate its quality. In this sense, this work presents an alternative using a logistic model to evaluate the culture medium to be used in vitro. The probabilities provided by this model will be used as a medium evaluator index. The importance of this index is based on the formalization of a statistical criterion for the selection of the adequate culture medium to be used on in vitro culture without excluding its physical and/or chemical properties. To demonstrate this procedure, an experiment determining the ideal medium for the in vitro culture of primary explants of Ipeca [Psychotria ipecacuanha (Brot. Stokes] was evaluated. The differentiation of the culture medium was based on the presence and absence of the growth regulator BAP (6-benzilaminopurine. A logistic model was adjusted as a function of the weight of fresh and dry matter. Minimum, medium and maximum probabilities obtained with this model showed that the culture medium containing BAP was the most adequate for the explant growth. Due to the high discriminative power of these mediums, detected by the model, their use is recommended as an alternative to select culture medium for similar experiments.

  17. Therapeutic potential of mesenchymal stromal cells and MSC conditioned medium in Amyotrophic Lateral Sclerosis (ALS--in vitro evidence from primary motor neuron cultures, NSC-34 cells, astrocytes and microglia.

    Directory of Open Access Journals (Sweden)

    Hui Sun

    Full Text Available Administration of mesenchymal stromal cells (MSC improves functional outcome in the SOD1G93A mouse model of the degenerative motor neuron disorder amyotrophic lateral sclerosis (ALS as well as in models of other neurological disorders. We have now investigated the effect of the interaction between MSC and motor neurons (derived from both non-transgenic and mutant SOD1G93A transgenic mice, NSC-34 cells and glial cells (astrocytes, microglia (derived again from both non-transgenic and mutant SOD1G93A ALS transgenic mice in vitro. In primary motor neurons, NSC-34 cells and astrocytes, MSC conditioned medium (MSC CM attenuated staurosporine (STS - induced apoptosis in a concentration-dependent manner. Studying MSC CM-induced expression of neurotrophic factors in astrocytes and NSC-34 cells, we found that glial cell line-derived neurotrophic factor (GDNF and ciliary neurotrophic factor (CNTF gene expression in astrocytes were significantly enhanced by MSC CM, with differential responses of non-transgenic and mutant astrocytes. Expression of Vascular Endothelial Growth Factor (VEGF in NSC-34 cells was significantly upregulated upon MSC CM-treatment. MSC CM significantly reduced the expression of the cytokines TNFα and IL-6 and iNOS both in transgenic and non-transgenic astrocytes. Gene expression of the neuroprotective chemokine Fractalkine (CX3CL1 was also upregulated in mutant SOD1G93A transgenic astrocytes by MSC CM treatment. Correspondingly, MSC CM increased the respective receptor, CX3CR1, in mutant SOD1G93A transgenic microglia. Our data demonstrate that MSC modulate motor neuronal and glial response to apoptosis and inflammation. MSC therefore represent an interesting candidate for further preclinical and clinical evaluation in ALS.

  18. 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 c...

  19. Effect of Conditioned Medium and Bone Marrow Stem Cell Lysate on the Course of Acetaminophen-Induced Liver Failure.

    Science.gov (United States)

    Khubutiya, M Sh; Temnov, A A; Vagabov, V A; Sklifas, A N; Rogov, K A; Zhgutov, Yu A

    2015-05-01

    A composition containing culture medium conditioned by mesenchymal stem cells and mesenchymal stem cell lysate improves biochemical parameters, reduces inflammation, and stimulates regenerative processes in the liver. PMID:26033600

  20. Ca2+ enrichment in culture medium potentiates effect of oligonucleotides.

    Science.gov (United States)

    Hori, Shin-Ichiro; Yamamoto, Tsuyoshi; Waki, Reiko; Wada, Shunsuke; Wada, Fumito; Noda, Mio; Obika, Satoshi

    2015-10-30

    Antisense and RNAi-related oligonucleotides have gained attention as laboratory tools and therapeutic agents based on their ability to manipulate biological events in vitro and in vivo. We show that Ca(2+) enrichment of medium (CEM) potentiates the in vitro activity of multiple types of oligonucleotides, independent of their net charge and modifications, in various cells. In addition, CEM reflects in vivo silencing activity more consistently than conventional transfection methods. Microscopic analysis reveals that CEM provides a subcellular localization pattern of oligonucleotides resembling that obtained by unassisted transfection, but with quantitative improvement. Highly monodispersed nanoparticles ~100 nm in size are found in Ca(2+)-enriched serum-containing medium regardless of the presence or absence of oligonucleotides. Transmission electron microscopy analysis reveals that the 100-nm particles are in fact an ensemble of much smaller nanoparticles (ϕ ∼ 15 nm). The presence of these nanoparticles is critical for the efficient uptake of various oligonucleotides. In contrast, CEM is ineffective for plasmids, which are readily transfected via the conventional calcium phosphate method. Collectively, CEM enables a more accurate prediction of the systemic activity of therapeutic oligonucleotides, while enhancing the broad usability of oligonucleotides in the laboratory. PMID:26101258

  1. Ca2+ enrichment in culture medium potentiates effect of oligonucleotides.

    Science.gov (United States)

    Hori, Shin-Ichiro; Yamamoto, Tsuyoshi; Waki, Reiko; Wada, Shunsuke; Wada, Fumito; Noda, Mio; Obika, Satoshi

    2015-10-30

    Antisense and RNAi-related oligonucleotides have gained attention as laboratory tools and therapeutic agents based on their ability to manipulate biological events in vitro and in vivo. We show that Ca(2+) enrichment of medium (CEM) potentiates the in vitro activity of multiple types of oligonucleotides, independent of their net charge and modifications, in various cells. In addition, CEM reflects in vivo silencing activity more consistently than conventional transfection methods. Microscopic analysis reveals that CEM provides a subcellular localization pattern of oligonucleotides resembling that obtained by unassisted transfection, but with quantitative improvement. Highly monodispersed nanoparticles ~100 nm in size are found in Ca(2+)-enriched serum-containing medium regardless of the presence or absence of oligonucleotides. Transmission electron microscopy analysis reveals that the 100-nm particles are in fact an ensemble of much smaller nanoparticles (ϕ ∼ 15 nm). The presence of these nanoparticles is critical for the efficient uptake of various oligonucleotides. In contrast, CEM is ineffective for plasmids, which are readily transfected via the conventional calcium phosphate method. Collectively, CEM enables a more accurate prediction of the systemic activity of therapeutic oligonucleotides, while enhancing the broad usability of oligonucleotides in the laboratory.

  2. Initiate and Maintain Cavitation by Combining High Amplitude Bursts and Continuous Ultrasound Exposure in Culture Medium

    Science.gov (United States)

    Mestas, J.-L.; Alberti, L.; Chesnais, S.; Lafon, C.; Blay, J.-Y.; Cathignol, D.

    2006-05-01

    Ultrasound can produce a variety of nonthermal bioeffects via acoustic cavitation. Most studies on cavitation effects pointed on the difficulty of initiating and controlling the cavitation phenomenon. Our objective is to obtain reproducible viability and transfection rate in the case of the application of a continuous low intensity ultrasound exposure. We propose to initiate and maintain cavitation in the medium by combining a continuous ultrasound exposure with periodical high amplitude bursts. Cells were exposed to ultrasound (444.5 kHz) transmitted through the bottom of twelve-well culture plates containing prostatic cells (AT2, 2.5 106 cells/mL), the plasmid DsRed in transfection case (200 μg/mL) and culture media. The cavitation effects were evaluated on the cell viability and transfection, determined 0 to 3 days after exposure by a flow cytometer (FACScan; total counted events: 10 000). Bursts of 1.73 W/cm2 intensity level had no effect on cells when their duration was lower than 100 ms and their frequency lower than 4 bursts/min. When combined with continuous exposure, only one burst of 1.73 W/cm2 intensity level and 50 ms duration was sufficient to activate the cavitation phenomenon in the medium.

  3. The effect of chemically defined medium on spontaneous calcium signaling of in situ chondrocytes during long-term culture.

    Science.gov (United States)

    Zhou, Yilu; Park, Miri; Cheung, Enoch; Wang, Liyun; Lu, X Lucas

    2015-04-13

    Chemically defined serum-free medium has been shown to better maintain the mechanical integrity of articular cartilage explants than serum-supplemented medium during long-term in vitro culture, but little is known about its effect on cellular mechanisms. We hypothesized that the chemically defined culture medium could regulate the spontaneous calcium signaling of in situ chondrocytes, which may modulate the cellular metabolic activities. Bovine cartilage explants were cultured in chemically defined serum-free or serum-supplemented medium for four weeks. The spontaneous intracellular calcium ([Ca(2+)]i) signaling of in situ chondrocytes was longitudinally measured together along with the biomechanical properties of the explants. The spontaneous [Ca(2+)]i oscillations in chondrocytes were enhanced at the initial exposure of serum-supplemented medium, but were significantly dampened afterwards. In contrast, cartilage explants in chemically defined medium preserved the level of calcium signaling, and showed more responsive cells with higher and more frequent [Ca(2+)]i peaks throughout the four week culture in comparison to those in serum medium. Regardless of the culture medium that the explants were exposed, a positive correlation was detected between the [Ca(2+)]i responsive rate and the stiffness of cartilage (Spearman's rank correlation coefficient=0.762). A stable pattern of [Ca(2+)]i peaks was revealed for each chondrocyte, i.e., the spatiotemporal features of [Ca(2+)]i peaks from a cell were highly consistent during the observation period (15 min). This study showed that the beneficial effect of chemically defined culture of cartilage explants is associated with the spontaneous [Ca(2+)]i signaling of chondrocytes in cartilage.

  4. Cell culture's spider silk road.

    Science.gov (United States)

    Perkel, Jeffrey

    2014-06-01

    A number of synthetic and natural materials have been tried in cell culture and tissue engineering applications in recent years. Now Jeffrey Perkel takes a look at one new culture component that might surprise you-spider silk.

  5. Efficient Differentiation of Embryonic Stem Cells into Neurons in Glial Cell-conditioned Medium under Attaching Conditions

    Institute of Scientific and Technical Information of China (English)

    Hai-Bin TIAN; Zeng-Liang BAI; Hong WANG; Jian-Quan CHEN; Guo-Xiang CHENG

    2005-01-01

    Embryonic stem (ES) cells can differentiate into neurons in vitro, which provides hope for the treatment of some neurodegenerative diseases through cell transplantation. However, it remains a challenge to efficiently induce ES cells to differentiate into neurons. Here, we show that murine ES cells can efficiently differentiate into neurons when cultured in glial cell- conditioned medium (GCM) under attaching conditions without the formation of embryoid bodies. In comparison with murine embryonic fibroblast-conditioned medium, we found that GCM has a positive effect on limiting the generation of non-neuronal cells, such as astrocytes. In addition, compared with suspension conditions, attaching conditions delay the differentiation process of ES cells.

  6. 条件培养液对Wistar大鼠海马神经干细胞单细胞克隆增殖的影响%Effect of conditioned culture medium on the proliferation of neural stem cell single cell clone in hippocampus of Wistar rat

    Institute of Scientific and Technical Information of China (English)

    佟雷; 王振宇; 季丽莉; 佟晓杰

    2011-01-01

    Objective To study the effect of conditioned culture medium (CM) on the proliferation of neural stem cells (NSCs) single cell clone in hippecampus of Wistar rat. Methods The serum-free medium was used to culture NSCs isolated from hippocampus of rats, the conditioned-medium was collected for further use. Cells clusters from passage 4 were performed for mono-cloning and divided into control (sentra-free medium) and CM groups. The proliferation of NSCs was evaluated by MTT. Immunocytochemistry for nestin, neuron specific enolase (NSE), glial fibrillary acidic protein (GFAP) and galactocerebroside (Galc) was used to identify monoclonal cells, neurons, astrocytes and oligodendrocytes respectively.Results The cultured monoclonal cells expressed nestin, and expressed NSE, GFAP and Galc one week after induced differentiation. The proliferation of NSCs was faster in CM group than in the control group. Conclusions CM can enhance the proliferation of the single cell clone of NSCs single cell clone in hippocampus of Wistar rat.%目的 探讨神经干细胞(NSCs)条件培养液对其单细胞克隆增殖的影响.方法 采用无血清培养法体外培养Wistar大鼠海马神经干细胞并进行传代培养,然后收集神经干细胞条件培养液.第四代细胞进行单细胞克隆,并将单细胞克隆细胞分为条件培养液组、无血清培养液对照组.通过MTT去比较两组细胞的增殖情况.单克隆培养细胞行巢蛋白(nestin)免疫细胞化学染色,诱导分化后细胞行神经元特异性烯醇化酶(NSE )、胶质原纤维酸性蛋白(GFAP)、半乳糖脑苷脂(Galc)免疫细胞化学染色.结果 单克隆培养后克隆球表达nestin,诱导分化后细胞表达NSE、GFAP、Galc,条件培养液组细胞增殖速度高于对照组.结论 条件培养液能提高Wistar大鼠海马神经干细胞单细胞克隆的增殖速度.

  7. 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...

  8. 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...

  9. Evaluation of a New Culture Medium for Borrelia burgdorferi

    OpenAIRE

    Marques, Adriana R.; Stock, Frida; Gill, Vee

    2000-01-01

    We evaluated the new MPM medium for the growth of Borrelia burgdorferi. All 18 blood samples from 17 patients with Lyme disease were negative. Growth studies showed that by day 4, most organisms in MPM were not viable. Our results reinforce the use of BSK medium as the primary choice for growing B. burgdorferi.

  10. Differentiation of rat oligodendrocyte precursor cells in chemical conditional medium in vitro

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Objective: To investigate in vitro differentiation of oligodendrocyte precursor cells (OPCs) into mature oligodendrocytes in chemical conditional medium. Methods: The mixed glial cells from cerebral cortices of 48-hour-old Sprague-Dawley (SD) rats were cultured in vitro. The OPCs were separated by shaking procedure around 9-10 d in the primary culture. Then the isolated OPCs were further transferred into the chemical conditional medium for cell differentiation. The pattern of OPCs maturation in vitro was continuously observed with contrast phase microscopy and mature oligodendrocytes were further identified by immunocytochemical assays. Results: OPCs grew well when co-cultured with glial cells and distinct cellular stratification formed about 9-10 d in the primary culture, which indicated the appropriate opportunity for the separation of OPCs. Following cultured in the chemical conditional medium, the OPCs progressively differentiated into the mature oligodendrocytes. These mature oligodendrocytes were also immunostained with the oligodendrocyte lineage-specific antibody, Oligo2. Conclusion: The OPCs isolated from the cerebral cortices of neonatal SD rats can progressively differentiate into mature oligodendrocytes in the chemical conditional medium in vitro.

  11. Culture medium optimization for pigment production with RSM method

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Response surface methodology was used to optimize a medium for a red-pigmented marine bacterium S-9801 strain (Flavobacterium sp.). In the first optimization step the influence of yeast extract, peptone, glucose and sodium chloride on pigment production was evaluated using a fractional factorial design. Pigment production was positively influenced by glucose and sodium chloride while other components had no significant effect. In the second step the path of steepest ascent was used to approach the optimal region of the medium composition. In the third step the optimal concentration of glucose and sodium chloride was determined by a central composite design and response analysis. The optimized medium allowed pigment production (A 535~650) to be increased from 0.137 to0.559, being 320% higher than the original medium.

  12. Submerged monoxenic culture medium development for Heterorhabditis bacteriophora and its symbiotic bacterium Photorhabdus luminescens: protein sources.

    Science.gov (United States)

    Cho, Chun-Hwi; Whang, Kyung Sook; Gaugler, Randy; Yoo, Sun Kyun

    2011-08-01

    Most medium formulations for improving culture of entomopathogenic nematodes (EPN) based on protein sources have used enriched media like animal feed such as dried egg yolk, lactalbumin, and liver extract, among other ingredients. Most results, however, showed unstable yields and longer production time. Many of the results do not show the detailed parameters of fermentation. Soy flour, cotton seed flour, corn gluten meal, casein powder, soytone, peptone, casein hydrolysates, and lactalbumin hydrolysate as protein sources were tested to determine the source to support optimal symbiotic bacteria and nematode growth. The protein hydrolysates selected did not improve bacterial cell mass compared with the yeast extract control, but soy flour was the best, showing 75.1% recovery and producing more bacterial cell number (1.4×10⁹/ml) than all other sources. The highest yield (1.85×10⁵ IJs/ml), yield coefficient (1.67×10⁶ IJs/g medium), and productivity (1.32×10⁷ IJs/l/day) were also achieved at enriched medium with soybean protein. PMID:21876379

  13. Three-dimensional cell culturing by magnetic levitation.

    Science.gov (United States)

    Haisler, William L; Timm, David M; Gage, Jacob A; Tseng, Hubert; Killian, T C; Souza, Glauco R

    2013-10-01

    Recently, biomedical research has moved toward cell culture in three dimensions to better recapitulate native cellular environments. This protocol describes one method for 3D culture, the magnetic levitation method (MLM), in which cells bind with a magnetic nanoparticle assembly overnight to render them magnetic. When resuspended in medium, an external magnetic field levitates and concentrates cells at the air-liquid interface, where they aggregate to form larger 3D cultures. The resulting cultures are dense, can synthesize extracellular matrix (ECM) and can be analyzed similarly to the other culture systems using techniques such as immunohistochemical analysis (IHC), western blotting and other biochemical assays. This protocol details the MLM and other associated techniques (cell culture, imaging and IHC) adapted for the MLM. The MLM requires 45 min of working time over 2 d to create 3D cultures that can be cultured in the long term (>7 d). PMID:24030442

  14. [Method of producing fusarin C in perlite-liquid culture medium].

    Science.gov (United States)

    Li, G; Li, M; Ma, J

    1992-02-01

    For researching the biosynthesis labelled Fusarin C(Fc) by Fuscarium moniliforme, a more quick and convenient method of Fusarin C production and purification were established, and a good liquid culture medium consisted of different kinds organic matters (hydroxy proline, sucrose and glycerin), inorganic salts and perlite replaced corn grit medium. The perlite-liquid culture medium inoculated with the strain of F. moniliforme yields 936mg Fc/kg organic matter with in 14 days of incubation at 28 degrees C. As compared with the corn grit medium, the amount of Fc from perlite-liquid medium was more than that from corn grit medium (831mg Fc/kg corn grit). In all experiments both thin-layer chromatography and high-pressure liquid chromatography were used to confirm the presence of Fc. parameters which were important for the optimal biosynthesis of Fc included hydroxy proline and sucrose concentrations, incubated time/temperature and amount of perlite. The 40g of sucrose/L liquid culture was optimal concentration for Fusarin C production. Of three contained N-matter tested, hydroxy proline was the best sources of N-atom for Fusarin C. Under the absence of hydroxy proline, the Fc wasn't synthesized in perlite-liquid culture medium by F. moniliforme. A culture time/temperature study of Fc production was done, and the optimal Fc amounts was synthesized after incubation for 14 days at 28 degrees C on perlite-liquid culture medium.

  15. Maintenance and neuronal cell differentiation of neural stem cells C17.2 correlated to medium availability sets design criteria in microfluidic systems.

    Directory of Open Access Journals (Sweden)

    Bu Wang

    Full Text Available BACKGROUND: Neural stem cells (NSCs play an important role in developing potential cell-based therapeutics for neurodegenerative disease. Microfluidics has proven a powerful tool in mechanistic studies of NSC differentiation. However, NSCs are prone to differentiate when the nutrients are limited, which occurs unfavorable by fast medium consumption in miniaturized culture environment. For mechanistic studies of NSCs in microfluidics, it is vital that neuronal cell differentiation is triggered by controlled factors only. Thus, we studied the correlation between available cell medium and spontaneous neuronal cell differentiation of C17.2 NSCs in standard culture medium, and proposed the necessary microfluidic design criteria to prevent undesirable cell phenotype changes. METHODOLOGY/PRINCIPAL FINDINGS: A series of microchannels with specific geometric parameters were designed to provide different amount of medium to the cells over time. A medium factor (MF, defined as the volume of stem cell culture medium divided by total number of cells at seeding and number of hours between medium replacement successfully correlated the amount of medium available to each cell averaged over time to neuronal cell differentiation. MF smaller than 8.3×10(4 µm3/cell⋅hour produced significant neuronal cell differentiation marked by cell morphological change and significantly more cells with positive β-tubulin-III and MAP2 staining than the control. When MF was equal or greater than 8.3×10(4 µm3/cell⋅hour, minimal spontaneous neuronal cell differentiation happened relative to the control. MF had minimal relation with the average neurite length. SIGNIFICANCE: MFs can be controlled easily to maintain the stem cell status of C17.2 NSCs or to induce spontaneous neuronal cell differentiation in standard stem cell culture medium. This finding is useful in designing microfluidic culture platforms for controllable NSC maintenance and differentiation. This study also

  16. Dependence of synchronized bursting activity on medium stirring and the perfusion rate in a cultured network of neurons

    Science.gov (United States)

    Heo, Ryoun; Kim, Hyun; Lee, Kyoung J.

    2016-05-01

    A cultured network of neurons coupled with a multi-electrode-array (MEA) recording system has been a useful platform for investigating various issues in neuroscience and engineering. The neural activity supported by the system can be sensitive to environmental fluctuations, for example, in the medium's nutrient composition, ph, and temperature, and to mechanical disturbances, yet this issue has not been the subject. Especially, a normal practice in maintaining neuronal cell cultures involves an intermittent sequence of medium exchanges, typically at a time interval of a few days, and one such sudden medium exchange is unavoidably accompanied by many unintended disturbances. Here, based on a quantitative time-series analysis of synchronized bursting events, we explicitly demonstrate that such a medium exchange can, indeed, bring a huge change in the existing neural activity. Subsequently, we develop a medium perfusion-stirring system and an ideal protocol that can be used in conjunction with a MEA recording system, providing long-term stability. Specifically, we systematically evaluate the effects of medium stirring and perfusion rates. Unexpectedly, even some vigorous mechanical agitations do not have any impacts on neural activity. On the other hand, too much replenishment ( e.g., 1.8 ml/day for a 1.8-ml dish) of neurobasal medium results in an excitotoxicity.

  17. [In vitro cultivation of dendritic cells with serum-free medium].

    Science.gov (United States)

    Zhang, Huai-Dong; Song, Zhen-Lan; Li, Wei-Ping

    2006-10-01

    This study was aimed to investigate the protocol in vitro to incubate the dendritic cell (DC) derived from peripheral blood monocytes using serum-free medium X-VIVO 20. Peripheral blood monocytes from healthy donors were treated with 100 ng/ml GM-CSF and 500 U/ml IL-4, respectively. After cultivation for 6 days, they were treated with 100 ng/ml calcium ionophore A23187. After cultivation for 24 hours the cellular morphology was observed under invert microscope, the surface markers were analyzed by flow cytometry, the proliferation of allogenetic T cells was detected by MTT colorimetry, the specific cytotoxicity of T cells primed with DC was examined by MTT assay. The results showed that in all three groups with serum-free, fetal calf serum (FCS) and human AB serum mediums, cells displayed characteristic morphological features of DC. Simultaneously CD14 expression was decreased, and CD83, HLA-DR and CDw123 expression were increased on these cells. In addition, DCs cultured with these methods could evidently stimulate the proliferation of allogenetic T cell. As compared with the two controls of serum containing groups, the cultured cells in the serum-free groups showed almost the same allo-stimulatory capability and cellular morphology and surface markers, and T lymphocytes primed with the culture-derived DC exhibited the similar killing activity to K562 (P > 0.05). It is concluded that there is no significance in DC numbers, morphology, epitope and ability to stimulate the proliferation of allogenetic T cells between DC induced by serum-free X-VIVO 20 medium and DC induced by serum-contained medium. DC cultured and induced by serum-free medium is worth using in practice widely.

  18. RSW Mixed Element Cell-Centered Medium Mesh

    Data.gov (United States)

    National Aeronautics and Space Administration — This RSW gridset is designed as the medium size mixed element grid for use with cell-centered unstructured meshes. UG3 : Grid File Name = rsw_med_mixedcc.b8.ugrid...

  19. RSW Fully Tet Medium Cell-Centered Mesh

    Data.gov (United States)

    National Aeronautics and Space Administration — This is the RSW Fully tetrahedral medium cell-centered unstructured grid with a viscous wall. UG3 : Grid File Name = rsw_med_tetcc.b8.ugrid UG3 : Quad Surface...

  20. Selective buffered charcoal-yeast extract medium for isolation of nocardiae from mixed cultures.

    OpenAIRE

    Garrett, M. A.; Holmes, H T; Nolte, F S

    1992-01-01

    The recovery of Nocardia species from mixed cultures is facilitated by use of a selective medium. We show that buffered charcoal-yeast extract medium with polymyxin, anisomycin, and vancomycin can be used for the selective isolation of nocardiae from contaminated specimens.

  1. The serial cultivation of suspended BHK-21/13 cells in serum-free Waymouth medium.

    Science.gov (United States)

    Guskey, L E; Jenkin, H M

    1976-01-01

    A simple medium system was developed to obtain growth of BHK-21 cells in shaker cultures in the absence of serum. These cells have now undergone over 80 serial passages in serum-free Waymouth medium and have been recovered from the frozen state after storage for over 1 month in medium containing 10% dimethyl sulfoxide (DMSO) and 1% bovine serum albumin (BSA). Various amounts of exogenous lipid in the form of sodium oleate were added to cultures of cells growing in serum-free Waymouth medium. Concentrations of 10-50 mug of sodium oleate/ml had no detrimental effects on the cells as measured by trypan blue uptake. Furthermore, the cells were serially passed ten times in the presence of 10 mug sodium oleate/ml. Depletion of calf serum from the growth medium and addition of known quantities of lipids to the system provides a means of revealing subtle changes in lipid synthesis and lipid turnover during cellular growth. PMID:1250851

  2. [Effect of calcium on medium alkalinization induced by salicylic acid in Salvia miltiorrhiza suspension cultures].

    Science.gov (United States)

    Liu, Liancheng; Wang, Cong; Dong, Juan'e; Su, Hui; Zhuo, Zequn; Xue, Yaxin

    2013-07-01

    We studied medium alkalinization in Salvia miltiorrhiza suspension cultures treated with salicylic acid and the effect of Ca2+ in this process through application of calcium channel antagonists (Verapamil, LaCl3, LiCl, 2-APB) and ionophore A23187. The results show that salicylic acid could induce significant medium alkalinization in S. miltiorrhiza culture. Verapamil and LaCl3 or LiCl and 2-APB, two different groups of calcium channel antagonist, significantly inhibited the medium alkalinization induced by salicylic acid. However, the suppression effect of verapamil or LaCl3 on medium alkalinization induced by salicylic acid was higher than that of LiCl or 2-APB. When two types of calcium channel inhibitor (LaCl3 and 2-APB) were used together, the medium alkalinization induced by salicylic acid was completely suppressed and even reduced the pH in medium. On the other hand, A23187 could promote the medium alkalinization. Based on the results above, we speculated that salicylic acid could induce significant medium alkalinization in S. miltiorrhiza culture, depending on the calcium from both extracell and intracell. Moreover, calcium from extracell plays a more dominant role in this process. Reveal of relationship in this research between Ca2+ and medium alkalinization can provide theory evidence for mechanism of the plant secondary metabolism.

  3. Bicarbonate Plays a Critical Role in the Generation of Cytotoxicity during SIN-1 Decomposition in Culture Medium

    Directory of Open Access Journals (Sweden)

    Kyo Shirai

    2012-01-01

    Full Text Available 3-Morpholinosydnonimine (SIN-1 is used as a donor of peroxynitrite (ONOO− in various studies. We demonstrated, however, that, the cell-culture medium remains cytotoxic to PC12 cells even after almost complete SIN-1 decomposition, suggesting that reaction product(s in the medium, rather than ONOO−, exert cytotoxic effects. Here, we clarified that significant cytotoxicity persists after SIN-1 decomposes in bicarbonate, a component of the culture medium, but not in NaOH. Cytotoxic SIN-1-decomposed bicarbonate, which lacks both oxidizing and nitrosating activities, degrades to innocuous state over time. The extent of SIN-1 cytotoxicity, irrespective of its fresh or decomposed state, appears to depend on the total number of initial SIN-1 molecules per cell, rather than its concentration, and involves oxidative/nitrosative stress-related cell damage. These results suggest that, despite its low abundance, the bicarbonate-dependent cytotoxic substance that accumulates in the medium during SIN-1 breakdown is the cytotoxic entity of SIN-1.

  4. Desarrollo de embriones de bovino obtenidos por fecundación in vitro cultivados con células oviductales o medio condicionado y transferidos a hembras receptoras Bovine embryo development produced by in vitro fertilization cultured with oviductal cell or conditioned medium and transfer to recipients

    Directory of Open Access Journals (Sweden)

    M RATTO

    1999-01-01

    Full Text Available Se comparó el desarrollo in vitro de ovocitos obtenidos de ovarios de vaca de matadero, madurados, fecundados y cultivados in vitro bajo dos sistemas. Los ovocitos fueron cultivados en un medio de maduración a 39 °C, 5 % de CO2 y humedad relativa de 95 % durante 22 horas. Posteriormente, fueron incubados con espermatozoides seleccionados a través de una gradiente discontinua de Percoll. La tasa de maduración nuclear y fecundación fueron de 93,7 % (74/79 y 76,9 % (50/65 respectivamente. Un total de 252 ovocitos fecundados fueron cultivados in vitro. El porcentaje de desarrollo in vitro a las 2 días post-inseminación (embriones de 4-8 células fue de 62,7 % (64/102 para los cigotos cultivados con células oviductales y de 67 % (100/150 para los cultivados en medio condicionado (P0,05. El porcentaje de desarrollo de mórulas fue de 17,6 % (18/102 para los cigotos cultivados con células oviductales y de 13,3 % (20/150 para los cultivados con medio condicionado (P0,05. Se obtuvo una tasa de desarrollo del 15,7 % (16/102 de blastocistos para aquellos cigotos cultivados con células oviductales. No se obtuvo blastocistos a partir de cigotos cultivados en medio condicionado. Cuatro blastocistos fueron transferidos a dos hembras receptoras. A los 42 y 57 días se encontró la presencia de un feto en cada hembraThe in vitro development of matured and fertilized bovine oocytes was compared between two culture systems. Oocytes were collected by aspiration of follicles of 3-8 mm in diameter using an 18g needle. After morphological selection the oocytes were incubated at 39 0C, 5 % C02 y 95 % relative humidity, during 22 hours. Afterwards, oocytes were incubated with spermatozoa selected by Percoll gradient system. The rate of nuclear maturation and fertilization was 93,7 % (74/79 and 76,9 % (50/65, respectively. A total of 252 zygotes were cultured, 102 with oviductal cells and 150 in conditioned medium. The in vitro development on day 2 of culture

  5. Serum-free culture conditions for serial subculture of undifferentiated PC12 cells.

    Science.gov (United States)

    Ohnuma, Kiyoshi; Hayashi, Yohei; Furue, Miho; Kaneko, Kunihiko; Asashima, Makoto

    2006-03-15

    PC12 cells, a widely used model neuronal cell line, are usually cultured in serum-supplemented medium. This report describes a serum-free medium for the culture of PC12 cells. PC12 cells grown in the two media types had similar growth rates and released dopamine in response to high potassium-induced calcium elevation. However, the levels of dopamine and of dopamine release in cells cultured in the serum-free medium were less than 10% of that in cells cultured in serum-supplemented medium. Dopamine levels recovered within 10 days if cells were returned to serum-supplemented medium, but dopamine release could not be recovered. Nerve growth factor (NGF) induced similar responses in PC12 cells cultured in both media, including phosphorylation of extracellular signal-regulated protein kinases and neurite extension. Transferrin was necessary for survival of neurite-bearing PC12 cells subcultured in serum-free medium and insulin promoted the cells proliferation. Ten days culture with NGF produced a similar increase in neurofilament expression and acetylcholinesterase activity in both media. These results suggest that PC12 in the hormonally defined serum-free media are qualitatively the same as those cultured in serum-supplemented media, and therefore this new culture protocol should enable more precise studies of PC12 cells culture in the absence of confounding unknown factors.

  6. Improved elastase production by Bacillus sp.EL31410--further optimization and kinetics studies of culture medium for batch fermentation

    Institute of Scientific and Technical Information of China (English)

    何国庆; 陈启和; 琚晓捷; 石乃冬

    2004-01-01

    An efficient culture medium producing a bacterial elastase with high yields was developed further following preliminary studies by means of response surface method. Central composite design (CCD) and response surface methodology were applied to optimize the medium constituents. A central composite design was used to explain the combined effect of three medium constituents, viz, glucose, K2HPO4, MgSO4@7H2O. The strain produced more elastase in the completely optimized medium, as compared with the partially optimized medium. The fitted model of the second model, as per RSM,showed that glucose was 7.4 g/100 ml, casein 1.13 g/100 ml, corn steep flour 0.616 g/100 ml, K2HPO4 0.206 g/100 ml and MgSO4@7H2O 0.034 g/100 ml. The fermentation kinetics of these two culture media in the flask experiments were analyzed. It was found that the highest elastase productivity occurred at 54 hours. Higher glucose concentration had inhibitory effect on elastase production. At the same time, we observed that the glucose consumption rate was slow in the completely optimized medium, which can explain the lag period of the highest elastase production. Some metal ions and surfactant additives also affected elastase production and cell growth.

  7. USE OF SODIUM HIPOCHLORITE IN STERILIZATION OF CULTURE MEDIUM FOR MULTIPLICATION OF Eucalyptus pellita L.

    Directory of Open Access Journals (Sweden)

    Silvio Lopes Teixeira

    2009-10-01

    Full Text Available Lately it has been observed a great interest in the research area of plant tissue culture in discovering new alternatives leading to cost reduction of the plants produced in commercial laboratories, in order to turn this alternative of plant propagation more economical. A potentially promising alternative for this reduction of costs, but which has not been receiving the due attention, is the possibility of substituting the autoclaving technique to a more economical one. With this purpose, two tests were carried out, using a new protocol of medium preparation, which consisted of the chemical sterilization of all the utensils used in the preparation and packaging of the culture medium as well, associated to the addition of the sterilizing agent to the medium, in different concentrations. The objective of the first test was to observe the influence of different concentrations of NaClO added to the culture medium, on its sterilization. The second test aimed at verifying the reaction of the Eucalyptus pellita tissues to different concentrations of NaClO in the culture medium. The addition of NaClO to the culture medium, equal or higher than 0.0005% in the fist test and of 0.005% in the second one, allowed complete sterilization of the medium, without observing any damage to the Eucalyptus pellita tissues, even when they were grown on culture medium containing up to 0.009%, the maximum concentration tried. The results showed the viability of eliminating the autoclave for the sterilization of culture media.

  8. Development of an Integrated Microfluidic Perfusion Cell Culture System for Real-Time Microscopic Observation of Biological Cells

    OpenAIRE

    Chih-Chin Oh-Yang; Min-Hsien Wu; Jr-Lung Lin; Shih-Siou Wang

    2011-01-01

    This study reports an integrated microfluidic perfusion cell culture system consisting of a microfluidic cell culture chip, and an indium tin oxide (ITO) glass-based microheater chip for micro-scale perfusion cell culture, and its real-time microscopic observation. The system features in maintaining both uniform, and stable chemical or thermal environments, and providing a backflow-free medium pumping, and a precise thermal control functions. In this work, the performance of the medium pumpin...

  9. Evaluation of the effects of a plasma activated medium on cancer cells

    Science.gov (United States)

    Mohades, S.; Laroussi, M.; Sears, J.; Barekzi, N.; Razavi, H.

    2015-12-01

    The interaction of low temperature plasma with liquids is a relevant topic of study to the field of plasma medicine. This is because cells and tissues are normally surrounded or covered by biological fluids. Therefore, the chemistry induced by the plasma in the aqueous state becomes crucial and usually dictates the biological outcomes. This process became even more important after the discovery that plasma activated media can be useful in killing various cancer cell lines. Here, we report on the measurements of concentrations of hydrogen peroxide, a species known to have strong biological effects, produced by application of plasma to a minimum essential culture medium. The activated medium is then used to treat SCaBER cancer cells. Results indicate that the plasma activated medium can kill the cancer cells in a dose dependent manner, retain its killing effect for several hours, and is as effective as apoptosis inducing drugs.

  10. Evaluation of the effects of a plasma activated medium on cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Mohades, S.; Laroussi, M., E-mail: mlarouss@odu.edu; Sears, J.; Barekzi, N.; Razavi, H. [Plasma Engineering and Medicine Institute, Old Dominion University, Norfolk, Virginia 23529 (United States)

    2015-12-15

    The interaction of low temperature plasma with liquids is a relevant topic of study to the field of plasma medicine. This is because cells and tissues are normally surrounded or covered by biological fluids. Therefore, the chemistry induced by the plasma in the aqueous state becomes crucial and usually dictates the biological outcomes. This process became even more important after the discovery that plasma activated media can be useful in killing various cancer cell lines. Here, we report on the measurements of concentrations of hydrogen peroxide, a species known to have strong biological effects, produced by application of plasma to a minimum essential culture medium. The activated medium is then used to treat SCaBER cancer cells. Results indicate that the plasma activated medium can kill the cancer cells in a dose dependent manner, retain its killing effect for several hours, and is as effective as apoptosis inducing drugs.

  11. Human adipose derived stroma/stem cells grow in serum-free medium as floating spheres.

    Science.gov (United States)

    Dromard, C; Bourin, P; André, M; De Barros, S; Casteilla, L; Planat-Benard, V

    2011-04-01

    With the goal of obtaining clinically safe human adipose-derived stroma/stem cells (ASC) and eliminating the use of serum, we have developed a new culture system that allows the expansion of ASC as spheres in a defined medium. These spheres can be passaged several times. They are not only aggregated cells but rather originate from single cells as clonal spheres can be obtained after seeding at very low density and reform clonal spheres after dissociation. These spheres can also revert to monolayer growth when plated in medium containing human plasma and even generate fibroblast-like colonies (CFU-f). Under several differentiation-specific media, spheres-derived ASC maintain their capacity to differentiate into osteoblasts, endothelial cells and adipocytes. These results indicate that human ASC can be maintained in a serum-free 3D culture system, which is of great interest for the expansion in bioreactors of autologous ASC and their use in clinical trials.

  12. 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...

  13. Enhancement of the thermal transport in a culture medium with Au nanoparticles

    Science.gov (United States)

    Jiménez-Pérez, J. L.; Fuentes, R. Gutierrez; Alvarado, E. Maldonado; Ramón-Gallegos, E.; Cruz-Orea, A.; Tánori-Cordova, J.; Mendoza-Alvarez, J. G.

    2008-11-01

    In this work, it is reported the gold nanoparticles synthesis, their characterization, and their application to the enhancement of the thermal transport in a cellular culture medium. The Au nanoparticles (NPs), with average size of 10 nm, contained into a culture medium (DMEM (1)/F12(1)) (CM) increased considerably the heat transfer in the medium. Thermal lens spectrometry (TLS) was used to measure the thermal diffusivity of the nanofluids. The characteristic time constant of the transient thermal lens was obtained by fitting the theoretical expression, for transient thermal lens, to the experimental data. Our results show that the thermal diffusivity of the culture medium is highly sensitive to the Au nanoparticle concentration and size. The ability to modify the thermal properties to nanometer scale becomes very important in medical applications as in the case of cancer treatment by using photodynamic therapy (PDT). A complementary study with UV-vis and TEM techniques was performed to characterize the Au nanoparticles.

  14. 兔脂肪干细胞在含有转化因子和转铁蛋白培养基中向软骨细胞的分化%Chondrogenic differentiation of rabbit adipose-derived stem cells in culture medium containing transforming factors and transferrin

    Institute of Scientific and Technical Information of China (English)

    杨国庆; 王兆杰; 安荣泽; 赵俊延; 齐新文

    2011-01-01

    BACKGROUND: The choice of seed cells is a key factor to the cartilage tissue engineering.OBJECTIVE: To explore the chondrogenic potential of adipose-derived stem cells (ADSCs) in culture medium containingtransforming factors and transferring.METHODS: ADSCs were obtained from the subcutaneous adipose tissue of 6-month-old New Zealand white rabbits'neckbackby mechanical digestion and enzyme digestion, and then cultured and amplified in vitro. The adhesion and growth of cells wereobserved using inverted phase contrast microscope. Two weeks after the cells were cultured in chondrogenic medium, collagentype II expression was detected by immunohistochemistry staining.RESULTS AND CONCLUSION: The primary stem cells isolated from adipose tissue adhered on the plate in 24 hours, andreached 90% confluence in single layers after 96 hours. The cartilage nodules were observed at 7 days after ADSCs werecultured under chondrogenic medium, while collagen type II was positive 14 days later. ADSCs of rabbits can expresschondrogenic phenotype when they were cultured in chondrogenic medium, which proves the ADSCs can be one of the optionsfor cartilage tissue engineering.%背景:成软骨的种子细胞的选择是软骨组织工程研究中的关键因素.目的:观察脂肪干细胞在含有转化因子和转铁蛋白诱导条件下向软骨细胞分化的能力.方法:取新西兰大白兔颈背部脂肪组织,机械分离及酶消化法获得脂肪干细胞,显微镜下观察细胞黏附及生长情况;加入含有转化因子β1和转铁蛋白的诱导培养基培养2周后以免疫组化方法检测Ⅱ型胶原的表达.结果与结论:从兔脂肪组织中分离出的干细胞原代培养时24 h贴壁,96 h后达80%融合;于软骨诱导培养基内向软骨诱导7 d后形成软骨结节,14 d后Ⅱ型胶原免疫组化阳性.结果初步表明兔脂肪干细胞经诱导后可以向软骨细胞分化.

  15. Optimization of an effective growth medium for culturing probiotic bacteria for applications in strict vegetarian food products

    Directory of Open Access Journals (Sweden)

    Manju Pathak

    2012-10-01

    Full Text Available Background: This study aimed to modify de Man Rogosa Sharpe culture medium (termed MRS for selective cultivation of probiotics strain for the consumption by the strictly vegetarian human population. Vegetarian probiotic foods by definition must be free from all animal-derived ingredients. This not only includes the product ingredients but the probiotic inoculum as well. Probiotic starter cultures are traditionally grown and stored in media containing milk or meatderived ingredients. The presence of these ingredients makes the probiotic cell concentrates unsuitable for use in vegetarian products and thus creates the need for a growth medium which isfree from animal-derived ingredients. Present study investigated the growth of a strain of Lactobacillus lactis in MRS. The present invention relates in general to a bacterial culture media,and more specifically a complex microbial culture media, based on plant seed powder extract in place of animal extract for probiotic bacterial growth.Methods: Lactobacillus lactis, a probiotic, was grown in standard MRS culture medium as well as in our various test media (TM containing various vegetal source in place of beef extract, yeast extract and peptone as in case of MRS. The inoculated culture mediums were incubated at 37C for 72 hours and growth of probiotic is recorded at regular intervals. The growth was recorded as Colony Forming Units (CFUs.Results: The best growth of probiotic is observed in TM 2. TM 2 is the leguminous seed extract. Starter culture mediums for probiotics or other bacteria primarily contain protein from animal source. The possibility of using vegetal protein from TM 2 extract in place of peptones and meat extract for the nitrogen supplementation of culture media for the growth of lactic acid bacteria has been demonstrated.Functional Foods in Health and Disease 2012, 2(10:369-378 Conclusion: The absolute vegetarian culture medium containing TM 2 is better than standard MRS for the

  16. Evaluation on Sensitivity of the Human Sperm Motility Assay for Detecting Endotoxin in Culture Medium

    Institute of Scientific and Technical Information of China (English)

    Wei-jie ZHU; Jing LI; Wen-hong ZHANG; Kang-shou YAO

    2003-01-01

    Objective To investigate the sensitivity of the human sperm motility assay for detecting endotoxin in culture medium Materials & Methods Motile sperm were separated and exposed to different concentrations of endotoxin (0.5 ng/mL, 1 ng/mL, 10 ng/mL, 1 000 ng/mL, 10 000 ng/mL, and 50 000 ng/mL), and sperm motility was determined after incubation. Effects of endotoxin on sperm motility in media without albumin were also examined. In addition, at the same concentrations of endotoxin (0.5 ng/mL, 1 ng/mL, and 10 ng/mL), the sensitivity of the human sperm motility assay was compared to those of 1-cell and 2-cell mouse embryo bioassays.Results At levels of 0.5 ng/mL~1 000 ng/mL endotoxin in media with 2 mg/mL albumin, sperm did not show significant change in motility during 24 h of incubation when compared with the control (P>0.05). However, the sperm motility was significantly inhibited at endotoxin dosages of 10 000 and 50 000 ng/mL. In the absence of albumin supplementation, at endotoxin levels of 50 000 ng/mL, and 1 000 ng/mL, there was a marked decrease in sperm motility compared with the control after 2 h or 8 h of incubation, respectively (P<0.01). In media containing 0.5 ng/mL and 1 ng/mL endotoxin, 1-cell and 2-cell mouse embryos had significantly reduced developmental rates in all developmental stages, and at the level of 10 ng/mL, the development of the embryos was arrested.Conclusion The human sperm motility assay could detect high levels of endotoxin in culture medium but its sensitivity to endotoxin would be inferior to that of the 1-cell or 2-cell mouse embryo bioassay. In the absence of albumin supplementation, the sensitivity of the sperm motility assay could be improved.

  17. Modified PEHPS Medium as an Alternative for the In Vitro Culture of Giardia lamblia

    Science.gov (United States)

    Vargas-Villarreal, Javier; Mata-Cárdenas, Benito D.; Hernández-García, Magda E.; Garza-González, Jesús N.; De La Garza-Salinas, Laura H.; González-Salazar, Francisco

    2014-01-01

    Commercial culture media present interlot variations in biological activity. We have previously designed a homemade and economic culture medium, PEHPS medium, for the axenic cultivation of Entamoeba histolytica and Trichomonas vaginalis. Trophozoites of amoebae and trichomonads grow well in this medium. Furthermore, the medium is stable for several months when stored frozen or refrigerated. The objective of this work was to modify PEHPS medium to support the in vitro growth of Giardia lamblia. Inocula of 5 × 103 trophozoites/mL of G. lamblia were incubated at 36.5°C in modified PEHPS or TYI-S-33 medium. Then, the growths of the three Giardia strains in both media were compared. The logarithmic growth phase lasted 72 h; the mean yield of the strains ranged from 10.06 to 11.43 × 105Giardia trophozoites/mL, and the range of duplication time in the three strains was from 5.67 to 6.06 in modified PEHPS medium. These growth characteristics were not significantly different from those obtained with TYI-S-33 medium. We conclude that modified PEHPS medium might be used for the axenic cultivation of G. lamblia. PMID:24982905

  18. Modified PEHPS Medium as an Alternative for the In Vitro Culture of Giardia lamblia

    Directory of Open Access Journals (Sweden)

    Javier Vargas-Villarreal

    2014-01-01

    Full Text Available Commercial culture media present interlot variations in biological activity. We have previously designed a homemade and economic culture medium, PEHPS medium, for the axenic cultivation of Entamoeba histolytica and Trichomonas vaginalis. Trophozoites of amoebae and trichomonads grow well in this medium. Furthermore, the medium is stable for several months when stored frozen or refrigerated. The objective of this work was to modify PEHPS medium to support the in vitro growth of Giardia lamblia. Inocula of 5 × 103 trophozoites/mL of G. lamblia were incubated at 36.5°C in modified PEHPS or TYI-S-33 medium. Then, the growths of the three Giardia strains in both media were compared. The logarithmic growth phase lasted 72 h; the mean yield of the strains ranged from 10.06 to 11.43 × 105Giardia trophozoites/mL, and the range of duplication time in the three strains was from 5.67 to 6.06 in modified PEHPS medium. These growth characteristics were not significantly different from those obtained with TYI-S-33 medium. We conclude that modified PEHPS medium might be used for the axenic cultivation of G. lamblia.

  19. 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

  20. Acute toxicity bioassays using Daphnia magna Straus (Cladocera, Daphniidae) maintained in a modified culture medium

    OpenAIRE

    Mónica Núñez; Jasmin Hurtado

    2013-01-01

    Daphnia magna is a test organism used in ecotoxicological assays of freshwater; however, traditional culture systems for this organism could result expensive, for that the aim of this research was to developed a new economic culture medium. With this purpose, 10 strains of D. magna were isolated, their population development was evaluated by total count of organisms and pregnant females using 3 different culture media: (A) alfalfa juice, (B) solved yeast and (C) a mixture of alfalfa juice plu...

  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. Effect of medium osmotic potential on callus induction and shoot regeneration in flax anther culture.

    Science.gov (United States)

    Chen, Yurong; Dribnenki, Paul

    2004-11-01

    Development of an efficient and cost-effective doubled haploid production system in flax (Linum usitatissimum L.) is the prerequisite for the application of doubled haploid technology in a practical breeding program. Pre-culture of anthers on a medium containing 15% sucrose for 2-7 days before transfer to the same medium containing 6% sucrose for a total of 28 days culture period significantly increased shoot regeneration for all four genotypes evaluated. Moreover, pre-culture of anthers on medium containing 15% sucrose for 2-7 days was sufficient to dramatically reduce the frequency of shoot regeneration from somatic tissues and thereby to increase the frequency of microspore-derived plants in flax anther culture. Furthermore, replacing 15% sucrose with 6% sucrose and 9% polyethylene glycol (PEG), or 3% sucrose and 12% PEG, in pre-culture medium did not significantly affect callus induction and shoot regeneration. The results indicate that sucrose may act as carbon/energy source as well as an osmotic regulator in flax anther culture. Sucrose as an osmotic regulator may be replaced by a non-metabolizable osmoticum: PEG. The implication of this study in flax anther culture and breeding is discussed. PMID:15235814

  3. Mesenchymal Progenitor Cells: Tissue Origin, Isolation and Culture.

    Science.gov (United States)

    Bourin, Philippe; Gadelorge, Mélanie; Peyrafitte, Julie-Anne; Fleury-Cappellesso, Sandrine; Gomez, Marilyn; Rage, Christine; Sensebé, Luc

    2008-01-01

    SUMMARY: Since the pioneering work of Alexander Friedenstein on multipotent mesenchymal stromal cells (MSCs), a tremendous amount of work has been done to isolate, characterize and culture such cells. Assay of colony forming unit-fibroblasts (CFU-Fs), the hallmark of MSCs, is used to estimate their frequency in tissue. MSCs are adherent cells, so they are easy to isolate, and they show contact inhibition. Thus, several parameters must be taken into account for culture: cell density, number of passages, culture medium, and growth factors used. The purity of the initial material is not a limiting parameter. Similar but not identical cell populations are found in almost all mammal or human tissues. MSCs seem to be very abundant in adipose tissue but at low frequency in blood from umbilical cord or in adult tissue. The culture conditions are very similar, whatever the source of cells. Because of their favorable properties, MSCs are very promising tools for regenerative medicine.

  4. Induction of mitosis in the cultured rabbit lens initiated by the addition of insulin to medium KEI-4

    Energy Technology Data Exchange (ETDEWEB)

    Reddan, J.R.; Unakar, N.J.; Harding, C.V.; Bagchi, M.; Saldana, G.

    1975-01-01

    The epithelium of lenses cultured in KEI-4, a completely defined medium formulated with specific reference to the biochemistry and physiology of the rabbit lens, exhibits a pattern of cell division similar to that noted for the organ in situ. Initial fluctuations in mitotic activity occurred in the area of the germinative zone during the first 24 hr of culture. Mitosis decreased at 1 hr, was extremely low at 3 hr and returned to values comparable for lens in vivo by 22 hr. The precipitous drop in mitosis noted at 3 hr is in part attributable to the isolation of the lens from adjoining tissue. The addition of insulin to KEI-4 triggers a parasynchronous burst of DNA synthesis throughout the central lens epithelium. The activation requires the intact hormone; neither proinsulin nor the A and/or B chains of insulin, nor glucagon nor zinc chloride can initiate mitosis. The gamma-globulin-rich fraction of rabbit serum can also stimulate mitosis. The addition of dibutyryl adenosine 3':5' cyclic monophosphate (DBeAMP) plus theophylline to KEI-4-insulin inhibits mitosis and prevents the cells from entering the synthetic phase of the cell cycle. Theophylline alone or DBeAMP alone brings about a 90 percent reduction in the insulin-induced mitotic responses. Lenses exposed to insulin show a marked increase in RNA synthesis and also exhibit an increased binding of tritiated actinomycin D at 1 and 3 hr of culture relative to KEI-4 controls. The hormone apparently activates the genome including those genes governing cell division. The system is amenable for long-term culture of the mammalian lens and since the constituents of the medium are known it should be possible to determine the factor(s) in the medium which, in conjunction with insulin, are needed for the induction of cell division.

  5. STUDIES REGARDING CULTURE MEDIUM INFLUENCE ON IN VITRO REGENERATION FROM WHEAT IMATURE EMBRYOS

    Directory of Open Access Journals (Sweden)

    M. DANCI

    2013-12-01

    Full Text Available Surnamed „embryos’ saving method”, embryos culture is an in vitro technique used for over half of the century for saving the distant hybridization products, that would have degenerate in other conditions. Immature embryos culture is used for initiation of in vitro cultures imposed by the impossibility of using other explants for some of the plant species. Wheat is one of the crops that immature embryos culture technique is suitable for. This methods principle is based on aseptic embryos excision and their inoculation to an adequate culture medium. In vitro culture results depend in a greater manner of the basic culture medium and the hormonal balance used. Immature embryos isolated from two Romanian wheat cultivars – Dropia and Lovrin 41 – were inoculated for callus production on two types of basic media added with 2,4 D. The selected calluses were transferred on MS basic medium and several parameters were registered. Both cultivars emphasized a good callusing capacity, in a different percentage depending on the culture media used, such as 71,09 – 94,45%.. big differences between the cultivars regarding embriogenic callus frequency, shooting callus frequency and regenerated plants percentage were registered.

  6. 干酪乳杆菌DI-1发酵产胞壁蛋白酶的培养基优化%Optimization of Culture Medium for the Production of Cell Envelope Proteinase by Lactobacillus Casei DI-1

    Institute of Scientific and Technical Information of China (English)

    吴振; 潘道东

    2013-01-01

    采用Plackett-Burman和Box-Behnken方案,对影响干酪乳杆菌DI-1产胞壁蛋白酶(CEP)的MRS培养基成分进行筛选,结果表明,当在MRS培养基中添加磷酸氢二钾2.78 g/L,柠檬酸二铵2.92 g/L和乙酸钠4.36g/L时,CEP活性显著提高,此时酶活性为11.36U/mL,比基本MRS培养基提高了63.68%.在氨基酸和鸭肉水解肽添加试验中,当基础培养基中分别添加0.1 g/L的精氨酸、甘氨酸、异亮氨酸、丝氨酸、丙氨酸和谷氨酰胺时,CEP的活性从6.94 U/mL提高到11.11 U/mL;当添加木瓜风味复合蛋白酶水解鸭肉蛋白多肽时,CEP活力提高到13.89 U/mL.%In the present study, Plackett-Burman and Box-Behnken design were adopted to analyze the basal MRS medium components that significantly influenced the CEP production. K2HPO4, diamine hydrogen citrate and CH3C00Na were screened as the three main factors for effection the production of cell envelope proteinase (CEP) of Lactobacillus ca-sei DI-1. The optimized concentrations of K2HPO4, diamine hydrogen citrate and CH3CO0Na in the media were 2.78, 2.92 and 4.36 g/L, respectively. The maximum activity of CEP in the media was 11.36 U/mL, which was 63.68% higher than that of basal MRS medium. Furthermore, the CEP activity increased from 6.94 to 11.11 U/mL when adding 0.10 g/L of L-arginine, glycine, L-isoleucine, L-serine, L-alanine and L-glutamine to the MRS medium respectively, and the CEP activity increased from 6.94 to 13.89 U/mL when adding peptides from the duck protein hydrolysate produced by papaya-flavour compund enzyme.

  7. 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.

  8. 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

  9. 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)

  10. In vitro culture medium (IVC) supplementation with sericin improves developmental competence of ovine zygotes.

    Science.gov (United States)

    Aghaz, Faranak; Hajarian, Hadi; KaramiShabankareh, Hamed

    2016-03-01

    This study was carried out to investigate the effects of supplementation of potassium simplex optimized medium (KSOM-aa) with various sericin concentrations (0, 0.1, 0.5, 1 and 2.5%) on ovine zygotes. The results indicate that the supplementation of oocyte in vitro culture medium with optimal concentration of sericin (0.1 and 0.5%) may have beneficial effects on developmental competence of in vitro-derived ovine embryos. PMID:26952758

  11. 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.

  12. Study on Cell Suspension Culture of Floribunda Rose

    Institute of Scientific and Technical Information of China (English)

    ZHANG Chun'ai; WANG Jingang; FAN Jinping; GONG Shufang; CHE Daidi

    2008-01-01

    Friable callus was induced when immature seeds of floribunda rose were inoculated on MS medium supplemented with 2,4-D 3.0 mg-L-1.When transfered onto subculture media,fi-iable callus developed into embryogenic callus,which was used to establish cell suspension lines.Cell suspensions had to be subcultured at a interval of 4-5 days at the first several culture cycles.The best subculturing cycle for the stable cell suspensions was 8-10 days.The best inoculum quantity was 1 mL PCV(Packed Cell Volume) per 40 mL culture fluid.

  13. 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.

  14. Fermentation profile of Saccharomyces cerevisiae and Candida tropicalis as starter cultures on barley malt medium.

    Science.gov (United States)

    Alloue-Boraud, Wazé Aimée Mireille; N'Guessan, Kouadio Florent; Djeni, N'Dédé Théodore; Hiligsmann, Serge; Djè, Koffi Marcellin; Delvigne, Franck

    2015-08-01

    Saccharomyces cerevisiae C8-5 and Candida tropicalis F0-5 isolated from traditional sorghum beer were tested for kinetic parameters on barley malt extract, YPD (863 medium) and for alcohol production. The results showed that C. tropicalis has the highest maximum growth rate and the lowest doubling time. Values were 0.22 and 0.32 h(-1) for maximum growth rate, 3 h 09 min and 2 h 09 min for doubling time respectively on barley malt extract and YPD. On contrary, glucose consumption was the fastest with S. cerevisiae (-0.36 and -0.722 g/l/h respectively on barley malt extract and YPD). When these two yeasts were used as starters in pure culture and co-culture at proportion of 1:1 and 2:1 (cell/cell) for barley malt extract fermentation, we noticed that maltose content increased first from 12.12 g/l to 13.62-16.46 g/l and then decreased. The highest increase was obtained with starter C. tropicalis + S. cerevisiae 2:1. On contrary, glucose content decreased throughout all the fermentation process. For all the starters used, the major part of the ethanol was produced at 16 h of fermentation. Values obtained in the final beers were 11.4, 11.6, 10.4 and 10.9 g/l for fermentation conducted with S. cerevisiae, C. tropicalis, C. tropicalis + S. cerevisiae 1:1 and C. tropicalis + S. cerevisiae 2:1. Cell viability measurement during the fermentation by using flow cytometry revealed that the lowest mean channel fluorescence for FL3 (yeast rate of death) was obtained with C. tropicalis + S. cerevisiae 2:1 after 48 h of fermentation. PMID:26243947

  15. Enrichment of breast cancer stem cells using a keratinocyte serum-free medium

    Institute of Scientific and Technical Information of China (English)

    LIU Zhen-zhen; CHEN Ping; LU Zhen-duo; CUI Shu-de; DONG Zi-ming

    2011-01-01

    Background Keratinocyte serum-free medium (K-SFM) is a defined medium used to support the growth of primary keratinocytes and embryonic stem cell. The aim of this research was to optimize enrichment of breast cancer stem cells (CSCs) using K-SFM.Methods A K-SFM was used to enrich CSCs from two breast cancer cell lines and a primary culture of breast cancer.RPMI-1640 supplemented with 10% fetal calf serum (FCS) was used as a control. CSCs were identified with flow cytometry using CD44+/CD24-as molecular markers. The expression of a variety of CSC markers (Oct-4, ABCG2, Nanog,N-cadherin, and E-cadherin) was analyzed with real-time PCR.Results Much higher percentage of CSCs was achieved with K-SFM: 17.3% for MCF-7 cells, 17.4% for SKBR-3, and 20.0% for primary breast cancer culture. Less than 1% CSC was achieved using RPMI-1640 supplemented with 10% FCS. In comparison to the CSCs obtained with RPMI-1640, CSCs in the K-SFM expressed higher levels of Oct-4,ABCG2, Nanog and N-cadherin, and lower level of E-cadherin.Conclusion K-SFM is an optimal culture medium to maintain and to enrich breast CSCs.

  16. Does Embryo Culture Medium Influence the Health and Development of Children Born after In Vitro Fertilization?

    Science.gov (United States)

    Bouillon, Céline; Léandri, Roger; Desch, Laurent; Ernst, Alexandra; Bruno, Céline; Cerf, Charline; Chiron, Alexandra; Souchay, Céline; Burguet, Antoine; Jimenez, Clément; Sagot, Paul; Fauque, Patricia

    2016-01-01

    In animal studies, extensive data revealed the influence of culture medium on embryonic development, foetal growth and the behaviour of offspring. However, this impact has never been investigated in humans. For the first time, we investigated in depth the effects of embryo culture media on health, growth and development of infants conceived by In Vitro Fertilization until the age of 5 years old. This single-centre cohort study was based on an earlier randomized study. During six months, in vitro fertilization attempts (No. 371) were randomized according to two media (Single Step Medium--SSM group) or Global medium (Global group). This randomized study was stopped prematurely as significantly lower pregnancy and implantation rates were observed in the SSM group. Singletons (No. 73) conceived in the randomized study were included (42 for Global and 31 for SSM). The medical data for gestational, neonatal and early childhood periods were extracted from medical records and parental interviews (256 variables recorded). The developmental profiles of the children in eight domains (social, self-help, gross motor, fine motor, expressive language, language comprehension, letter knowledge and number knowledge--270 items) were compared in relation to the culture medium. The delivery rate was significantly lower in the SSM group than in the Global group (pchildren of the Global group were less likely than those of the SSM group to show developmental problems (p = 0.002), irrespective of the different domains. In conclusion, our findings showed that the embryo culture medium may have an impact on further development.

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

    DEFF Research Database (Denmark)

    Engvild, Kjeld Christensen

    1974-01-01

    Suspension cultures of Datura innoxia Mill, were successfully grown on a modified Murashige and Skoog medium with 2,4–D, NAA or BAP as growth substances, provided the micronutrient levels were reduced to 1/10. Normal amounts of micronutrients were toxic. Attempts to identify the toxic elements did...... 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 above 500...

  18. Effect of Sodium Chloride on Growth of Staphylococcus aureus in Synthetic Tissue Culture Medium

    OpenAIRE

    出来尾, 哲; 地土井, 襄爾

    1983-01-01

    Staphylococcus aureus Smith was cultured in a synthetic tissue culture medium containing various concentrations of sodium chloride. The growth was at a higher level in the media containing sodium chloride of concentrations of 2.0 and 4.0% than in those of less than 2.0% and more than 4.0%. The result suggests that the bacteria grow more vigorously in the fairly high concentrations of sodium chloride than in the physiological concentration of it, in the synthetic medium. It may be considered t...

  19. [Culture medium based on biogas slurry and breeding of oil Chlorella].

    Science.gov (United States)

    Zhao, Feng-Min; Mei, Shuai; Cao, You-Fu; Ding, Jin-Feng; Xu, Jia-Jie; Li, Shu-Jun

    2014-06-01

    The oil chlorella cultivation and biogas slurry treatment were combined. The biogas slurry provided water and nutrient for growing chlorella, at the same time, harmless treatment of biogas slurry was realized. This paper cultivated 4 species of oil chlorella in the mixed medium of biogas slurry and green algae medium (the volume ratios were 1 : 9, 1 : 3, 1 : 1 and 3 : 1, respectively), and compared their oil productivity to select the best oil chlorella species and the optimal culture medium. The results showed that, the combination of medium and chlorella species to reach the highest oil productivity was a volume ratio of 1 : 3 and the chlorella species BJ05, and the oil productivity of chlorella BJ05 was 9.20 mg x (L x d)(-1), higher than that in green algae medium [8.66 mg x (L x d)(-1)]. In mixed medium with a volume ratio of 1:3, the effect of adding different nutrients into the green algae medium on the oil productivity was examined, and the results showed that, sodium carbonate and citric acid had no negative effect on the oil productivity of chlorella BJ05. in the absence of sodium carbonate and citric acid, the oil productivity of chlorella BJ05 was 9.36 mg x (L x d)(-1), and the removal of COD (chemical oxygen demand), total nitrogen, total phosphorus and ammonia nitrogen rates were 59%, 75%, 61% and 100%, respectively. Deficiency in other nutrients had negative effect on the oil productivity. Therefore, the culture medium was further optimized to the mixed medium of biogas slurry and green algae medium with a volume ratio of 1 : 3 and without addition of sodium carbonate and citric acid.

  20. Quantitative Characterization of the Growth of Deinococcus geothermalis DSM-11302: Effect of Inoculum Size, Growth Medium and Culture Conditions

    Directory of Open Access Journals (Sweden)

    Julie Bornot

    2015-08-01

    Full Text Available Due to their remarkable resistance to extreme conditions, Deinococcaceae strains are of great interest to biotechnological prospects. However, the physiology of the extremophile strain Deinococcus geothermalis has scarcely been studied and is not well understood. The physiological behaviour was then studied in well-controlled conditions in flask and bioreactor cultures. The growth of D. geothermalis type strains was compared. Among the strains tested, the strain from the German Collection of Microorganisms (Deutsche Sammlung von Mikroorganismen DSM DSM-11302 was found to give the highest biomass concentration and growth rate: in a complex medium with glucose, the growth rate reached 0.75 h−1 at 45 °C. Yeast extract concentration in the medium had significant constitutive and catalytic effects. Furthermore, the results showed that the physiological descriptors were not affected by the inoculum preparation steps. A batch culture of D. geothermalis DSM-11302 on defined medium was carried out: cells grew exponentially with a maximal growth rate of 0.28 h−1 and D. geothermalis DSM-11302 biomass reached 1.4 g·L−1 in 20 h. Then, 1.4 gDryCellWeight of biomass (X was obtained from 5.6 g glucose (Glc consumed as carbon source, corresponding to a yield of 0.3 CmolX·CmolGlc−1; cell specific oxygen uptake and carbon dioxide production rates reached 216 and 226 mmol.CmolX−1·h−1, respectively, and the respiratory quotient (QR value varied from 1.1 to 1.7. This is the first time that kinetic parameters and yields are reported for D. geothermalis DSM-11302 grown on a mineral medium in well-controlled batch culture.

  1. A GROWTH INHIBITOR SECRETED FROM CULTURED RABBIT SMOOTHMUSCLE CELLS IS DISTINCT FROM TGF-β

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To ascertain whether the growth inhibitor in conditioned medium from cultured rabbit arte rial cells is distinct from TGF-β. Methods Rabbit aortic smooth muscle ceils were grown from explained segments of the aorta. Conditioned medium from cultured rabbit aortic smooth muscle ceils and anti-TGF-β were employed in this study. Smooth muscle cell proliferation was measured by XTT detection (Boehringer Mannheim). Results Acidified conditioned medium from smooth muscle ceils had significantly stronger effects of growth inhibition than controls, and anti-TGF-β did not affect the growth inhibitory effect of conditioned medium from cultured rabbit arterial smooth muscle cells. Conclusion The growth inhibiting substance in conditioned medium from cultured rabbit aortic smooth muscle cells is distinct from TGF-β.

  2. Does Embryo Culture Medium Influence the Health and Development of Children Born after In Vitro Fertilization?

    Directory of Open Access Journals (Sweden)

    Céline Bouillon

    Full Text Available In animal studies, extensive data revealed the influence of culture medium on embryonic development, foetal growth and the behaviour of offspring. However, this impact has never been investigated in humans. For the first time, we investigated in depth the effects of embryo culture media on health, growth and development of infants conceived by In Vitro Fertilization until the age of 5 years old. This single-centre cohort study was based on an earlier randomized study. During six months, in vitro fertilization attempts (No. 371 were randomized according to two media (Single Step Medium--SSM group or Global medium (Global group. This randomized study was stopped prematurely as significantly lower pregnancy and implantation rates were observed in the SSM group. Singletons (No. 73 conceived in the randomized study were included (42 for Global and 31 for SSM. The medical data for gestational, neonatal and early childhood periods were extracted from medical records and parental interviews (256 variables recorded. The developmental profiles of the children in eight domains (social, self-help, gross motor, fine motor, expressive language, language comprehension, letter knowledge and number knowledge--270 items were compared in relation to the culture medium. The delivery rate was significantly lower in the SSM group than in the Global group (p<0.05. The culture medium had no significant effect on birthweight, risk of malformation (minor and major, growth and the frequency of medical concerns. However, the children of the Global group were less likely than those of the SSM group to show developmental problems (p = 0.002, irrespective of the different domains. In conclusion, our findings showed that the embryo culture medium may have an impact on further development.

  3. Culture materials affect ex vivo expansion of hematopoietic progenitor cells.

    Science.gov (United States)

    LaIuppa, J A; McAdams, T A; Papoutsakis, E T; Miller, W M

    1997-09-01

    Ex vivo expansion of hematopoietic cells is important for applications such as cancer treatment, gene therapy, and transfusion medicine. While cell culture systems are widely used to evaluate the biocompatibility of materials for implantation, the ability of materials to support proliferation of primary human cells in cultures for reinfusion into patients has not been addressed. We screened a variety of commercially available polymer (15 types), metal (four types), and glass substrates for their ability to support expansion of hematopoietic cells when cultured under conditions that would be encountered in a clinical setting. Cultures of peripheral blood (PB) CD34+ cells and mononuclear cells (MNC) were evaluated for expansion of total cells and colony-forming unit-granulocyte monocyte (CFU-GM; progenitors committed to the granulocyte and/or monocyte lineage). Human hematopoietic cultures in serum-free medium were found to be extremely sensitive to the substrate material. The only materials tested that supported expansion at or near the levels of polystyrene were tissue culture polystyrene, Teflon perfluoroalkoxy, Teflon fluorinated ethylene propylene, cellulose acetate, titanium, new polycarbonate, and new polymethylpentene. MNC were less sensitive to the substrate materials than the primitive CD34+ progenitors, although similar trends were seen for expansion of the two cell populations on the substrates tested. CFU-GM expansion was more sensitive to substrate materials than was total cell expansion. The detrimental effects of a number of the materials on hematopoietic cultures appear to be caused by protein adsorption and/or leaching of toxins. Factors such as cleaning, sterilization, and reuse significantly affected the performance of some materials as culture substrates. We also used PB CD34+ cell cultures to examine the biocompatibility of gas-permeable cell culture and blood storage bags and several types of tubing commonly used with biomedical equipment

  4. Implementing an English-Medium Course on Japanese Language and Culture

    OpenAIRE

    Schnickel, Jacob; Yamauchi, Hiroyuki; Fukushima, Takenobu

    2015-01-01

    This paper presents an overview of a semester-long English-medium course on Japanese language and culture offered to exchange students studying at a women’s university in Tokyo, Japan. The paper describes participating students’ backgrounds, course content, methods of evaluation, and learner outcomes. In addition, feedback from students is included, as are instructors’ ideas for future versions of the course.

  5. Emerging Culture of English-Medium Instruction in Korea: Experiences of Korean and International Students

    Science.gov (United States)

    Kim, Jeongyeon; Tatar, Bradley; Choi, Jinsook

    2014-01-01

    This study aims to contrastively examine Korean and international students' experiences of taking subject courses at a Korean university. Focusing on the viewpoints of the students, rather than central authorities, we attempt to reveal how language use and cultural factors are interpenetrated in the praxis of English-medium instruction (EMI).…

  6. 21 CFR 866.2410 - Culture medium for pathogenic Neisseria spp.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Culture medium for pathogenic Neisseria spp. 866.2410 Section 866.2410 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Microbiology Devices §...

  7. Effect of genotype, Culture medium and cold pretreatment on another culture of wheat (T. aestivum L.) mutant lines

    International Nuclear Information System (INIS)

    This research was conducted in Tissue Culture Laboratory of the Nuclear Research Center for Agriculture and Medicine in the Atomic Energy Organization of Iran in 1996-1997. The objective was to investigate the effect of genotype, culture medium and cold pretreatment in calli production from anthers. Anthers of four mutant lines obtained from three cvs, (Omid, Tabassi and Roshan) were produced in the Nuclear Agriculture Center and two check cvs, (Omid and Tabassi) were cultured in two media PII and modified C17. Some of the spikes before plating were kept in dark at 4-5 deg C for one week. The result indicated that the genotype of donar plants had a significant effect on the calli and plants formation. Cold pre-treatment and medium type had no effect on the calli formation, but interaction between the two factors was very significant, and the best result was obtained when anthers were cultured in modified C17 medium without cold pre-treatment

  8. Cell Culture, Technology: Enhancing the Culture of Diagnosing Human Diseases.

    Science.gov (United States)

    Hudu, Shuaibu Abdullahi; Alshrari, Ahmed Subeh; Syahida, Ahmad; Sekawi, Zamberi

    2016-03-01

    Cell culture involves a complex of processes of cell isolation from their natural environment (in vivo) and subsequent growth in a controlled environmental artificial condition (in vitro). Cells from specific tissues or organs are cultured as short term or established cell lines which are widely used for research and diagnosis, most specially in the aspect of viral infection, because pathogenic viral isolation depends on the availability of permissible cell cultures. Cell culture provides the required setting for the detection and identification of numerous pathogens of humans, which is achieved via virus isolation in the cell culture as the "gold standard" for virus discovery. In this review, we summarized the views of researchers on the current role of cell culture technology in the diagnosis of human diseases. The technological advancement of recent years, starting with monoclonal antibody development to molecular techniques, provides an important approach for detecting presence of viral infection. They are also used as a baseline for establishing rapid tests for newly discovered pathogens. A combination of virus isolation in cell culture and molecular methods is still critical in identifying viruses that were previously unrecognized. Therefore, cell culture should be considered as a fundamental procedure in identifying suspected infectious viral agent.

  9. 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

  10. The culture of human embryonic stem cells in microchannel perfusion bioreactors

    Science.gov (United States)

    Korin, Natanel; Bransky, Avishay; Dinnar, Uri; Levenberg, Shulamit

    2007-12-01

    The culture of human Embryonic Stem (ES) cells in microchannel bioreactors can be highly beneficial for ES cell biology studies and ES tissue engineering applications. In the present study we examine the use of Human Foreskin Fibroblasts (HFF) cells as feeder cells for human ES culture in a microchannel perfusion bioreactor. PDMS microchannels (depth:130 micron) were fabricated using conventional soft-lithography techniques. The channels were sterilized, coated with a human fibronectin solution and seeded with cells. Following a period of static incubation, culture medium was perfused through the channels at various flow rates and cell growth was monitored throughout the culture process. Mass transport and fluid mechanics models were used to evaluate the culture conditions (shear stress, oxygen levels within the micro-bioreactor as a function of the medium flow rate. The conditions for successful long-term culture (>7 days) of HFF under flow were established. Experiments with human embryonic stem cells cultured in microchannels show that the conditions essential to co-culture human ES cell on HFF cells under perfusion differ from the conditions necessary for HFF cell culture. Human ES cells were found to be highly sensitive to flow and culture conditions and did not grow under flow rates which were suitable for HFF long-term culture. Successful culture of undifferentiated human ES cell colonies in a perfusion micro-bioreactor is a basic step towards utilizing microfluidic techniques to explore stem cell biology.

  11. Effects of sciatic-conditioned medium on neonatal rat retinal cells in vitro

    Directory of Open Access Journals (Sweden)

    Torres P.M.M.

    1998-01-01

    Full Text Available Schwann cells produce and release trophic factors that induce the regeneration and survival of neurons following lesions in the peripheral nerves. In the present study we examined the in vitro ability of developing rat retinal cells to respond to factors released from fragments of sciatic nerve. Treatment of neonatal rat retinal cells with sciatic-conditioned medium (SCM for 48 h induced an increase of 92.5 ± 8.8% (N = 7 for each group in the amount of total protein. SCM increased cell adhesion, neuronal survival and glial cell proliferation as evaluated by morphological criteria. This effect was completely blocked by 2.5 µM chelerythrine chloride, an inhibitor of protein kinase C (PKC. These data indicate that PKC activation is involved in the effect of SCM on retinal cells and demonstrate that fragments of sciatic nerve release trophic factors having a remarkable effect on neonatal rat retinal cells in culture.

  12. Systematic evaluation of sericin protein as a substitute for fetal bovine serum in cell culture.

    Science.gov (United States)

    Liu, Liyuan; Wang, Jinhuan; Duan, Shengchang; Chen, Lei; Xiang, Hui; Dong, Yang; Wang, Wen

    2016-01-01

    Fetal bovine serum (FBS) shows obvious deficiencies in cell culture, such as low batch to batch consistency, adventitious biological contaminant risk, and high cost, which severely limit the development of the cell culture industry. Sericin protein derived from the silkworm cocoon has become increasingly popular due to its diverse and beneficial cell culture characteristics. However, systematic evaluation of sericin as a substitute for FBS in cell culture medium remains limited. In this study, we conducted cellular morphological, physiological, and transcriptomic evaluation on three widely used mammalian cells. Compared with cells cultured in the control, those cultured in sericin-substitute medium showed similar cellular morphology, similar or higher cellular overall survival, lower population doubling time (PDT), and a higher percentage of S-phase with similar G2/G1 ratio, indicating comparable or better cell growth and proliferation. At the transcriptomic level, differentially expressed genes between cells in the two media were mainly enriched in function and biological processes related to cell growth and proliferation, reflecting that genes were activated to facilitate cell growth and proliferation. The results of this study suggest that cells cultured in sericin-substituted medium perform as well as, or even better than, those cultured in FBS-containing medium. PMID:27531556

  13. Effects of bovine serum proteins in culture medium on post-warming survival of bovine blastocysts developed in vitro.

    Science.gov (United States)

    Ohboshi, S; Etoh, T; Sakamoto, K; Fujihara, N; Yoshida, T; Tomogane, H

    1997-04-15

    Experiments were conducted to investigate the factors affecting the survival of bovine blastocysts produced in vitro after cryopreservation by vitrification. Zygotes were obtained by in vitro maturation and fertilization of oocytes. Embryos used in this study were developed in vitro at Day 7 and 8 (Day 0 = insemination day) in modified synthetic oviduct fluid medium supplemented with calf serum or BSA. Embryos were cryopreserved in a two-step protocol consisting of exposure to 10% ethylene glycol for 5 min, followed by the original vitrification solution (designated as VS) consisting of 40% (v/v) ethylene glycol, 6% (w/v) polyethylene glycol and 0.5 M sucrose in phosphate-buffered saline for 1 min. After warming, embryos were cultured in modified TCM-199 for an in vitro survival assay. The highest survival rate was obtained from the warmed embryos developed at Day 7 in medium supplemented with BSA (82.6%), and there were significant differences between results with calf scrum and BSA treatment (42.4 and 70.7%, respectively; P < 0.01). However, there were no significant differences in the cell numbers of embryos among the treatments. These results suggest that the survival of embryos developed in medium with BSA is superior to that of embryos developed in medium containing calf serum, although the cell numbers of the embryos developed under both media were similar. PMID:16728072

  14. Large scale production of Blackleg vaccine by fermenter and enriched culture medium in Iran

    Directory of Open Access Journals (Sweden)

    Pilehchian Langroudi, R.

    2012-06-01

    Full Text Available In all biological systems growth is defined as increase of chemical compounds. Bacteria can achieve to balanced growth if they are growing in a medium, which are completely adapted to it. Clostridium chauvoei, (Clostridium feseri is an anaerobic, spore forming, motile, and polymorph bacteria, which its size varies from 0.5-1 to 3-8 micron and could be observed as individual bacterium, diplo, and rarely streptococcus. Blackleg is a fatal disease of young cattle. It produces an acute local infection, and the resulting blood poisoning leads to rapid death. Clostridium chauvoei and, less frequently, Clostridium septicum are the most commonly responsible organisms. Vaccination is the only effective means for controlling of blackleg disease. Several kinds of vaccine are available commercially. It is 4 decades that blackleg vaccine is produced in Razi institute and because of enhanced demand of country, decision was made to improve the production procedure of this vaccine using large-scale fermenter, so the aim of this study was adaptation of Clostridium chauvoei to growth and proliferation in fermenter for preparation of a potent vaccine. Accordingly attempts were made to prepare and formulate the ingredients in order to obtain high yield of Clostridium chauvoei in culture medium by fermenter. All experiments were done in two sets: A-growth in glass bottles using conventional culture medium and B-growth in fermenter using conventional culture medium similar to A and also enriched culture medium. Results showed high yield of Clostridium chauvoei suspension in fermenter after 10 hours, using enriched culture medium (more than 1,480,000,000 organisms/ml, but no significant changes was obtained in glass bottles conditions comparing to the fermenter conditions. The safety and potency of the prepared vaccine was determined in sheep and guinea pigs according to British pharmacopoeia (veterinary with satisfactory results. Since this research has been

  15. 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.

  16. 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.

  17. Dynamized preparations in cell culture.

    Science.gov (United States)

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

    2009-06-01

    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 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. PMID:18955237

  18. Inorganic ions in the medium modify tropane alkaloids and riboflavin output in Hyoscyamus niger root cultures

    Directory of Open Access Journals (Sweden)

    Katrin Pudersell

    2012-01-01

    Full Text Available Context: Hyoscyamus niger L. (Solanaceae roots are rich of tropane alkaloids, such as hyoscyamine and scopolamine are used as the source of raw material for the pharmaceutical industry. Aims: The aim of the present study was to investigate the effect of calcium, magnesium, and iron ions on the production of tropane alkaloids and excretion of riboflavin in H. niger root cultures. Materials and Methods: The calcium, magnesium, or iron enriched/deprived Murashige and Skoog (MS growth medium were used for culture of H. niger root tissues. The secondary metabolites were quantified using high performance liquid chromatography with ultraviolet detector (HPLC-UV and fluorimetry techniques. Results: An increased calcium content in the medium unidirectionally reduced hyoscyamine, while increasing scopolamine production with only a moderate impact on riboflavin excretion. Manipulations with magnesium and iron contents in the medium resulted in divergent changes in hyoscyamine, scopolamine, and riboflavin concentrations. Conclusions: Our results show that increased calcium ion content in the Murashige and Skoog medium may be used for the intensification of the scopolamine production in H. niger root cultures.

  19. Technology for cell cycle research with unstressed steady-state cultures

    OpenAIRE

    Lebleu, Valerie S.; Thornton, Maureen; Gonda, Steven R.; Helmstetter, Charles E.

    2006-01-01

    A culture system for performing cell cycle analyses on cells in undisturbed steady-state populations was designed and tested. In this system, newborn cells are shed continuously from an immobilized, perfused culture rotating about the horizontal axis. As a result of this arrangement, the number of newborn cells released into the effluent medium each generation is identical to the number of cells residing in the immobilized population, indicating that one of the two new daughter cells is shed ...

  20. Does Embryo Culture Medium Influence the Health and Development of Children Born after In Vitro Fertilization?

    Science.gov (United States)

    Bouillon, Céline; Léandri, Roger; Desch, Laurent; Ernst, Alexandra; Bruno, Céline; Cerf, Charline; Chiron, Alexandra; Souchay, Céline; Burguet, Antoine; Jimenez, Clément; Sagot, Paul; Fauque, Patricia

    2016-01-01

    In animal studies, extensive data revealed the influence of culture medium on embryonic development, foetal growth and the behaviour of offspring. However, this impact has never been investigated in humans. For the first time, we investigated in depth the effects of embryo culture media on health, growth and development of infants conceived by In Vitro Fertilization until the age of 5 years old. This single-centre cohort study was based on an earlier randomized study. During six months, in vitro fertilization attempts (No. 371) were randomized according to two media (Single Step Medium--SSM group) or Global medium (Global group). This randomized study was stopped prematurely as significantly lower pregnancy and implantation rates were observed in the SSM group. Singletons (No. 73) conceived in the randomized study were included (42 for Global and 31 for SSM). The medical data for gestational, neonatal and early childhood periods were extracted from medical records and parental interviews (256 variables recorded). The developmental profiles of the children in eight domains (social, self-help, gross motor, fine motor, expressive language, language comprehension, letter knowledge and number knowledge--270 items) were compared in relation to the culture medium. The delivery rate was significantly lower in the SSM group than in the Global group (peffect on birthweight, risk of malformation (minor and major), growth and the frequency of medical concerns. However, the children of the Global group were less likely than those of the SSM group to show developmental problems (p = 0.002), irrespective of the different domains. In conclusion, our findings showed that the embryo culture medium may have an impact on further development. PMID:27008092

  1. 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.

  2. Atrazine and its metabolites degradation in mineral salts medium and soil using an enrichment culture.

    Science.gov (United States)

    Kumar, Anup; Singh, Neera

    2016-03-01

    An atrazine-degrading enrichment culture was used to study degradation of atrazine metabolites viz. hydroxyatrazine, deethylatrazine, and deisopropylatrazine in mineral salts medium. Results suggested that the enrichment culture was able to degrade only hydroxyatrazine, and it was used as the sole source of carbon and nitrogen. Hydroxyatrazine degradation slowed down when sucrose and/or ammonium hydrogen phosphate were supplemented as the additional sources of carbon and nitrogen, respectively. The enrichment culture could degrade high concentrations of atrazine (up to 110 μg/mL) in mineral salts medium, and neutral pH was optimum for atrazine degradation. Further, except in an acidic soil, enrichment culture was able to degrade atrazine in three soil types having different physico-chemical properties. Raising the pH of acidic soil to neutral or alkaline enabled the enrichment culture to degrade atrazine suggesting that acidic pH inhibited atrazine-degrading ability. The study suggested that the enrichment culture can be successfully utilized to achieve complete degradation of atrazine and its persistent metabolite hydroxyatrazine in the contaminated soil and water.

  3. 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)

  4. Fucose content of monoclonal antibodies can be controlled by culture medium osmolality for high antibody-dependent cellular cytotoxicity

    OpenAIRE

    Konno, Yoshinobu; Kobayashi, Yuki; Takahashi, Ken; TAKAHASHI, Eiji; Sakae, Shinji; Wakitani, Masako; Yamano, Kazuya; Suzawa, Toshiyuki; Yano, Keiichi; Ohta, Toshio; Koike, Masamichi; Wakamatsu, Kaori; Hosoi, Shinji

    2011-01-01

    Antibody-dependent cellular cytotoxicity (ADCC) is dependent on the fucose content of oligosaccharides bound to monoclonal antibodies (MAbs). As MAbs with a low fucose content exhibit high ADCC activity, it is important to control the defucosylation levels (deFuc%) of MAbs and to analyze the factors that affect deFuc%. In this study, we observed that the deFuc% was inversely related to culture medium osmolality for MAbs produced in the rat hybridoma cell line YB2/0, with r2 values as high as ...

  5. Cultured 3T3L1 adipocytes dispose of excess medium glucose as lactate under abundant oxygen availability

    OpenAIRE

    Sabater Martínez, David; Arriarán, Sofía; Romero Romero, María del Mar; Agnelli, Silvia; Fernández López, José Antonio; Remesar Betlloch, Xavier; Alemany, Marià

    2014-01-01

    White adipose tissue (WAT) produces lactate in significant amount from circulating glucose, especially in obesity;Under normoxia, 3T3L1 cells secrete large quantities of lactate to the medium, again at the expense of glucose and proportionally to its levels. Most of the glucose was converted to lactate with only part of it being used to synthesize fat. Cultured adipocytes were largely anaerobic, but this was not a Warburg-like process. It is speculated that the massive production of lactate, ...

  6. Properties of ras-amplified recombinant BHK-21 cells in protein-free culture

    OpenAIRE

    Inoue, Yuichi; Kawamoto, Seiji; Shoji, Masahiro; Hashizume, Shuichi; Teruya, Kiichiro; Katakura, Yoshinori; Shirahata, Sanetaka

    2000-01-01

    We compared serum and protein-free cultures ofa ras-amplified recombinant BHK-21 cell line(ras-rBHK-IgG), which hyperproduces a lungcancer specific recombinant human monoclonal antibody. Ras-rBHK-IgG cells were shown to grow well, evenin protein-free medium and to be morphologicallysimilar to cells cultured in serum containing medium. However, the growth rate of ras-rBHK-IgG cellswas considerably slower in protein-free medium, whichresults in a longer maintenance period compared with cells cu...

  7. Variation of Spirulina maxima biomass production in different depths of urea-used culture medium

    Directory of Open Access Journals (Sweden)

    Md-Abu Affan

    2015-01-01

    Full Text Available Fewer studies have assessed the outdoor cultivation of Spirulina maxima compared with S. platensis, although the protein content of S. maxima is higher than S. platensis. Spirulina growth medium requires an increased amount of NaHCO3, Na2CO3, and NaNO3, which increases the production cost. Therefore, the current study used a low-cost but high-efficiency biomass production medium (Medium M-19 after testing 33 different media. The medium depth of 25 cm (group A was sub-divided into A1 (50% cover with a black curtain (PolyMax, 12 oz ultra-blackout, A2 (25% cover, and A3 (no cover. Similarly the medium depths of 30 and 35 cm were categorized as groups B (B1, B2, and B3 and C (C1, C2, and C3, respectively, and the effects of depth and surface light availability on growth and biomass production were assessed. The highest biomass production was 2.05 g L-1 in group A2, which was significantly higher (p < 0.05 than that in all other groups and sub-groups. Spirulina maxima died in B1 and C1 on the fifth day of culture. The biochemical composition of the biomass obtained from A2 cultures, including protein, carbohydrate, lipid, moisture, and ash, was 56.59%, 14.42%, 0.94%, 5.03%, and 23.02%, respectively. Therefore, S. maxima could be grown outdoors with the highest efficiency in urea-enriched medium at a 25-cm medium depth with 25% surface cover or uncovered.

  8. Optimizing a culture medium for biomass and phenolic compounds production using Ganoderma lucidum

    Directory of Open Access Journals (Sweden)

    Carlos Andrés Zárate-Chaves

    2013-01-01

    Full Text Available The present work was aimed at optimizing a culture medium for biomass production and phenolic compounds by using Ganoderma lucidum. The culture was optimized in two stages; a Plackett-Burman design was used in the first one for identifying key components in the medium and a central composite design was used in the second one for optimizing their concentration. Both responses (biomass and phenolic compounds were simultaneously optimized by the latter methodology regarding desirability, and the optimal concentrations obtained were 50.00 g/L sucrose, 13.29 g/L yeast extract and 2.99 g/L olive oil. Maximum biomass production identified in these optimal conditions was 9.5 g/L and that for phenolic compounds was 0.0452 g/L, this being 100% better than that obtained in the media usually used in the laboratory. Similar patterns regarding chemical characterization and biological activity towards Aspergillus sp., from both fruiting body and mycelium-derived secondary metabolites and extracts obtained in the proposed medium were observed. It was shown that such statistical methodologies are useful for optimizing fermentation and, in the specific case of G. lucidum, optimizing processes for its production and its metabolites in submerged culture as an alternative to traditional culture.

  9. A microdroplet cell culture based high frequency somatic embryogenesis system for pigeonpea, Cajanus cajan (L.) Millsp.

    Science.gov (United States)

    Kumar, Nagan Udhaya; Gnanaraj, Muniraj; Sindhujaa, Vajravel; Viji, Maluventhen; Manoharan, Kumariah

    2015-09-01

    A protocol for high frequency production of somatic embryos was worked out in pigeonpea, Cajanus cajan (L.) Millsp. The protocol involved sequential employment of embryogenic callus cultures, low density cell suspension cultures and a novel microdroplet cell culture system. The microdroplet cell cultures involved culture of a single cell in 10 μI of Murashige and Skoog's medium supplemented with phytohormones, growth factors and phospholipid precursors. By employing the microdroplet cell cultures, single cells in isolation were grown into cell clones which developed somatic embryos. Further, 2,4-dichlorophenoxyacetic acid, kinetin, polyethylene glycol, putrescine, spermine, spermidine, choline chloride, ethanolamine and LiCl were supplemented to the low density cell suspension cultures and microdroplet cell cultures to screen for their cell division and somatic embryogenesis activity. Incubation of callus or the inoculum employed for low density cell suspension cultures and microdroplet cell cultures with polyethylene glycol was found critical for induction of somatic embryogenesis. Somatic embryogenesis at a frequency of 1.19, 3.16 and 6.51 per 10(6) cells was achieved in the callus, low density cell suspension cultures and microdroplet cell cultures, respectively. Advantages of employing microdroplet cell cultures for high frequency production of somatic embryos and its application in genetic transformation protocols are discussed. PMID:26548080

  10. Evaluation of new transport medium for detection of herpes simplex virus by culture and direct enzyme-linked immunosorbent assay.

    Science.gov (United States)

    Ogburn, J R; Hoffpauir, J T; Cole, E; Hood, K; Michael, D; Nguyen, T; Raden, S; Raju, B; Reisinger, V; Oefinger, P E

    1994-12-01

    The transport medium Multi-Microbe Media (M4) was evaluated prospectively by culture and direct enzyme-linked immunosorbent assay (ELISA) for detection of herpes simplex virus from 473 specimens. In addition, 377 specimens in Bartels Viral Transport Medium were evaluated. By using culture as a "gold standard," the ELISA sensitivity was approximately 85%, while the specificities exceeded 96% for both media.

  11. 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.

  12. Variations of secretome profiles according to conditioned medium preparation: The example of human mesenchymal stem cell-derived adipocytes.

    Science.gov (United States)

    Clabaut, Aline; Grare, Céline; Léger, Thibaut; Hardouin, Pierre; Broux, Odile

    2015-10-01

    One challenging point in analyzing cellular secretome collected as conditioned medium is cross-contamination by cell culture media components, especially bovine serum proteins. A common approach for serum removal is to wash the cells, an alternative is to grow cells using serum-free conditions. Given that the sample processing may influence the phenotype of cells and thus the secretome, it is important to establish the optimal protocol for each cell type. In this study, we compared two methods for preparing conditioned medium from human adipocytes derived from mesenchymal stem cells. Cells were either washed twice with PBS or cultured the last four days of differentiation in serum-free adipogenic medium. Gene expression of the cells was evaluated by using real-time PCR and 1D LC-MS/MS was used to compare secreted proteins present in the culture supernatants. Surprisingly, results showed significant differences in gene expression patterns of the cells and in protein content of the conditioned media and suggested that PBS washes induced severe modifications of the phenotype of cells and thus changes in protein secretion profiles. These data emphasize the significant variations in protein species related to cell manipulations and underline the importance of procedure optimization prior to any proteomic investigation. PMID:26105977

  13. METHYLCELLULOSE CELL-CULTURE AS A NEW CYTOTOXICITY TEST SYSTEM FOR BIOMATERIALS

    NARCIS (Netherlands)

    VANLUYN, MJA; VANWACHEM, PB; NIEUWENHUIS, P; DAMINK, LO; TENHOOPEN, H; FEIJEN, J

    1991-01-01

    The cytotoxicity of biomaterials can be tested in vitro using various culture systems. Liquid culture systems may detect cytotoxicity of a material either by culture of cells with extracts or with the material itself. In the latter instance, renewing the medium will remove possible released cytotoxi

  14. Optimization of Buffalo (Bubalus bubalis Embryonic Stem Cell Culture System

    Directory of Open Access Journals (Sweden)

    Mohammad Zandi

    2015-07-01

    Full Text Available Objective: In order to retain an undifferentiated pluripotent state, embryonic stem (ES cells have to be cultured on feeder cell layers. However, use of feeder layers limits stem cell research, since experimental data may result from a combined ES cell and feeder cell response to various stimuli. Materials and Methods: In this experimental study, a buffalo ES cell line was established from in vitro derived blastocysts and characterized by the Alkaline phosphatase (AP and immunoflourescence staining of various pluripotency markers. We examined the effect of various factors like fibroblast growth factor 2 (FGF-2, leukemia inhibitory factor (LIF and Y-27632 to support the growth and maintenance of bubaline ES cells on gelatin coated dishes, in order to establish feeder free culture systems. We also analyzed the effect of feeder-conditioned media on stem cell growth in gelatin based cultures both in the presence as well as in the absence of the growth factors. Results: The results showed that Y-27632, in the presence of FGF-2 and LIF, resulted in higher colony growth and increased expression of Nanog gene. Feeder-Conditioned Medium resulted in a significant increase in growth of buffalo ES cells on gelatin coated plates, however, feeder layer based cultures produced better results than gelatin based cultures. Feeder layers from buffalo fetal fibroblast cells can support buffalo ES cells for more than two years. Conclusion: We developed a feeder free culture system that can maintain buffalo ES cells in the short term, as well as feeder layer based culture that can support the long term maintenance of buffalo ES cells.

  15. Optimization of Culturing Condition and Medium Composition for the Production of Alginate Lyase by a Marine Vibrio sp. YKW-34

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Carbohydrases secreted by marine Vibrio sp. YKW-34 with strong Laminaria cell wall degrading ability were screened,and among them alginate lyase was found to be dominant. The effects of medium composition and culturing condition on the production of alginate lyase by marine Vibrio sp. YKW-34 in flask were investigated in this study. In the culture medium of marine broth, no alginate lyase was produced. The activity of the alginate lyase, after being induced, reached 5 UmL-1. The best inoculum volume and inoculum age were 10% and 12 h, respectively. The optimal temperature for alginate lyase production was 25℃. The fermentation medium was composed of 0.5% of Laminaria powder and 0.2% of KNO3 with an initial acidity of pH 8.0. Alginate could induce alginate lyase production but not as efficiently as Laminaria powder did. The addition of fucoidan, cellulose and glucose had negative effect on the alginate lyase production. Other kinds of nitrogen sources, such as yeast extract, beef extract and peptone, had positive effect on the growth of the microorganism and negative effect on alginate lyase production. In addition, the time course of alginate lyase production under the optimized condition was described. The optimal harvest time was 48 h.

  16. Optimization of culturing condition and medium composition for the production of alginate lyase by a marine Vibrio sp. YKW-34

    Science.gov (United States)

    Fu, Xiaoting; Lin, Hong; Kim, Sang Moo

    2008-02-01

    Carbohydrases secreted by marine Vibrio sp. YKW-34 with strong Laminaria cell wall degrading ability were screened, and among them alginate lyase was found to be dominant. The effects of medium composition and culturing condition on the production of alginate lyase by marine Vibrio sp. YKW-34 in flask were investigated in this study. In the culture medium of marine broth, no alginate lyase was produced. The activity of the alginate lyase, after being induced, reached 5 UmL-1. The best inoculum volume and inoculum age were 10% and 12 h, respectively. The optimal temperature for alginate lyase production was 25°C. The fermentation medium was composed of 0.5% of Laminaria powder and 0.2% of KNO3 with an initial acidity of pH 8.0. Alginate could induce alginate lyase production but not as efficiently as Laminaria powder did. The addition of fucoidan, cellulose and glucose had negative effect on the alginate lyase production. Other kinds of nitrogen sources, such as yeast extract, beef extract and peptone, had positive effect on the growth of the microorganism and negative effect on alginate lyase production. In addition, the time course of alginate lyase production under the optimized condition was described. The optimal harvest time was 48 h.

  17. 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

  18. Effect of anti-Mullerian hormone in culture medium on quality of mouse oocytes matured in vitro.

    Directory of Open Access Journals (Sweden)

    Yihui Zhang

    Full Text Available Anti-mullerian hormone (AMH is thought to reflect the growth of follicles and the ovarian function. However, the role of AMH in culture medium during in vitro maturation (IVM on oocyte quality and subsequent development potential is unclear. The objective of this study is to investigate the effect of recombinant human AMH (rh-AMH supplemented into IVM medium on oocyte quality. Cumulus-oocyte complexes (COCs were obtained from ICR mice and cultured in vitro with the different concentrations (0-1,000 ng/ml of rh-AMH. Following 16-18 h of culture, quantitative PCR and ELISA were performed to analyze GDF9 and BMP15 mRNA expression and protein production from the oocytes. Subsequently, in vitro fertilization (IVF and early embryonic development were employed to further evaluate the quality of in vitro matured oocytes. The results showed that AMH was only expressed in cumulus cells but not in the oocytes. However, AMH most specific receptor, AMHR-II, was expressed in both oocytes and cumulus cells. The levels of GDF9 and BMP15 expression and blastocyst formation rate were significantly increased (p<0.05 when the IVM medium was supplemented with 100 ng/ml of rh-AMH. With AdH1-SiRNA/AMH for knocking down of AMH expression during IVM significantly reduced (p<0.05 the levels of GDF9 and BMP15 expression and blastocysts formation rate. These results suggest that AHM improves oocytes quality by up-regulating GDF9 and BMP15 expressions during IVM.

  19. Culturing Selenastrum capricornutum (Chlorophyta) in a synthetic algal nutrient medium with defined mineral particulates

    Science.gov (United States)

    Kuwabara, J.S.; Davis, J.A.; Chang, Cecily C.Y.

    1985-01-01

    Algal nutrient studies in chemically-defined media typically employ a synthetic chelator to prevent iron hydroxide precipitation. Micronutrient-particulate interactions may, however, significantly affect chemical speciation and hence biovailability of these nutrients in natural waters. A technique is described by which Selenastrum capricornutum Printz (Chlorophyta) may be cultured in a medium where trace metal speciation (except iron) is controlled, not by organic chelation, but by sorption onto titanium dioxide. Application of this culturing protocol in conjunction with results from sorption studies of nutrient ions on mineral particles provides a means of studying biological impacts of sorptive processes in aquatic environments. ?? 1985 Dr W. Junk Publishers.

  20. Maintenance of mesenchymal stem cells culture due to the cells with reduced attachment rate

    Directory of Open Access Journals (Sweden)

    Shuvalova N. S.

    2013-01-01

    Full Text Available Aim. The classic detachment techniques lead to changes in cells properties. We offer a simple method of cultivating the population of cells that avoided an influence on the surface structures. Methods. Mesenchymal stem cells (MSC from human umbilical cord matrix were obtained and cultivated in standard conditions. While substituting the culture media by a fresh portion, the conditioned culture medium, where the cells were maintained for three days, was transferred to other culture flacks with addition of serum and growth factors. Results. In the flacks, one day after medium transfer, we observed attached cells with typical MSC morphology. The cultures originated from these cells had the same rate of surface markers expression and clonogenic potential as those replated by standard methods. Conclusions. MSC culture, derived by preserving the cells with reduced attachment ability, actually has the properties of «parent» passage. Using this method with accepted techniques of cells reseeding would allow maintaining the cells that avoided an impact on the cell surface proteins.

  1. Effects of different extracts of Eucalyptus camaldulensis on Trichomonas vaginalis parasite in culture medium

    OpenAIRE

    Solmaz Hassani; Gholamreza Asghari; Hossseinali Yousefi; Afsaneh Kazemian; Mahmood Rafieiean; Hossein Yousofi Darani

    2013-01-01

    Background: Trichomonas vaginalis is considered one of the main causes of vulvovaginitis in women. Metronidazole with vast side effects is now the drug of choice for treatment of this infection. In an attempt to find an alternative drug, the effect of Eucalyptus camaldulensis on this parasite was shown in previous studies. In this investigation, the effect of different extracts of this plant on T. vaginalis in culture medium has been investigated. Materials and Methods: Five different ext...

  2. PROPAGATION OF Portulaca oleracea L. IN LIQUID MEDIUM: IMPLICATIONS OF PLANT GROWTH REGULATORS IN CULTURE

    Directory of Open Access Journals (Sweden)

    Mahipal S. Shekhawat

    2015-02-01

    Full Text Available Portulaca oleracea L. is a medicinal plant, growing in warm and moist regions of north hemisphere of the world. A protocol for in vitro propagation using nodal shoot segments as explants has been outlined. The percent shoot response with shoot induction rate, 6.4 ± 0.7 shoots per explant, was achieved when cultured on agar-gelled Murashige and Skoog (MS medium containing 2.0 mg/L of BAP (6-benzylaminopurine. The cultures were amplified by passages on MS medium with 1.0 mg/L each BAP and kinetin (Kn. The best shoot amplification (37.5±0.9 shoots per vessel was achieved by subculturing of in vitro regenerated shoot clumps on liquid MS medium. Shoots regenerated in vitro by both the processes were rooted on ½ strength of MS medium + 2.0 mg/L of indole-3 butyric acid (IBA. Ninety six percent of the shoots rooted in vitro. The in vitro rooted plantlets were hardened under different regimes of temperature and humidity in the greenhouse. The hardened plantlets were transferred to mixture of soil and manure in polybags.

  3. Adapting the Medium: Dynamics of Intermedial Adaptation in Contemporary Japanese Popular Visual Culture

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    Pusztai Beáta

    2015-08-01

    Full Text Available With respect to adaptation studies, contemporary Japanese popular culture signifies a unique case, as different types of media (be those textual, auditive, visual or audio-visual are tightly intertwined through the “recycling” of successful characters and stories. As a result, a neatly woven net of intermedial adaptations has been formed - the core of this complex system being the manga-anime-live-action film “adaptational triangle.” On the one hand, the paper addresses the interplay of the various factors by which the very existence of this network is made possible, such as the distinctive cultural attitude to “originality,” the structure of the comics, animation and film industries, and finally, the role of fictitious genealogies of both traditional and contemporary media in the negotiation of national identity. On the other hand, the essay also considers some of the most significant thematic, narrative, and stylistic effects this close interconnectedness has on the individual medium. Special attention is being paid to the nascent trend of merging the adaptive medium with that of the original story (viewing adaptation as integration, apparent in contemporary manga-based live- action comedies, as the extreme case of intermedial adaptation. That is, when the aim of the adaptational process is no longer the transposition of the story but the adaptation (i.e. the incorporation of the medium itself- elevating certain medium-specific devices into transmedial phenomena.

  4. RNA-seq transcriptome analysis of a Pseudomonas strain with diversified catalytic properties growth under different culture medium.

    Science.gov (United States)

    Yang, Jia-Wei; Zheng, Dai-Jun; Cui, Bao-Dong; Yang, Min; Chen, Yong-Zheng

    2016-08-01

    Biocatalysis is an emerging strategy for the production of enantio-pure organic molecules. However, lacking of commercially available enzymes restricts the widespread application of biocatalysis. In this study, we report a Pseudomonas strain which exhibited versatile oxidation activity to synthesize chiral sulfoxides when growing under M9-toluene medium and reduction activity to synthesize chiral alcohols when on Luria-Bertani (LB) medium, respectively. Further comparative transcriptome analysis on samples from these two cultural conditions has identified 1038 differentially expressed genes (DEG). Gene Ontology (GO) enrichment and KEGG pathways analysis demonstrate significant changes in protein synthesis, energy metabolism, and biosynthesis of metabolites when cells cultured under different conditions. We have identified eight candidate enzymes from this bacterial which may have the potential to be used for synthesis of chiral alcohol and sulfoxide chemicals. This work provides insights into the mechanism of diversity in catalytic properties of this Pseudomonas strain growth with different cultural conditions, as well as candidate enzymes for further biocatalysis of enantiomerically pure molecules and pharmaceuticals. PMID:27061463

  5. Use of BBL CHROMagar MRSA Medium for Identification of Methicillin-Resistant Staphylococcus aureus Directly from Blood Cultures

    OpenAIRE

    Pape, John; Wadlin, Jill; Nachamkin, Irving

    2006-01-01

    We evaluated the ability of BBL CHROMagar MRSA medium (Becton Dickinson, Sparks, MD) to identify methicillin-resistant Staphylococcus aureus (MRSA) directly upon subculture from positive blood culture bottles. There were 124 MRSA isolates recovered from blood cultures in the study. BBL CHROMagar MRSA medium was highly sensitive (97.6% [121/124] at 18 to 24 h of incubation and 100% [124/124] at 48 h) and 99.9% specific for identifying MRSA from positive blood cultures.

  6. 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.

  7. Auxin requirements of sycamore cells in suspension culture.

    Science.gov (United States)

    Moloney, M M; Hall, J F; Robinson, G M; Elliott, M C

    1983-04-01

    Sycamore (Acer pseudoplatanus L.) cell suspension cultures (strain OS) require 2,4-dichlorophenoxyacetic acid (2,4-D) in their culture medium for normal growth. If the 2,4-D is omitted, rates of cell division are dramatically reduced and cell lysis may occur. Despite this ;auxin requirement,' it has been shown by gas chromatography-mass spectrometry that the cells synthesize indol-3yl-acetic acid (IAA). Changes in free 2,4-D and IAA in the cells during a culture passage have been monitored.There is a rapid uptake of 2,4-D by the cells during the lag phase leading to a maximum concentration per cell (125 nanograms per 10(6) cells) on day 2 followed by a decline to 45 nanograms per 10(6) cells by day 9 (middle of linear phase). The initial concentration of IAA (0.08 nanograms per 10(6) cells) rises slowly to a peak of 1.4 nanograms per 10(6) cells by day 9 then decreases rapidly to 0.2 nanograms per 10(6) cells by day 15 (early declining phase) and 0.08 nanograms per 10(6) cells by day 23 (early stationary phase).

  8. Derivation of Myogenic Progenitors Directly From Human Pluripotent Stem Cells Using a Sphere-Based Culture

    OpenAIRE

    Hosoyama, Tohru; McGivern, Jered V.; Van Dyke, Jonathan M.; Allison D Ebert; Suzuki, Masatoshi

    2014-01-01

    The authors present a novel protocol for deriving myogenic progenitors from human embryonic stem cells and induced pluripotent stem cells using free-floating spherical culture. Results show that sphere-based cultures of human pluripotent stem cells, expanded in medium containing high concentrations of fibroblast growth factor and epidermal growth factor, can propagate myogenic progenitors from human embryonic stem cells and healthy and disease-specific induced pluripotent stem cells.

  9. A Preliminary Observation on the Development of Mouse Embryos Co-cultured with Human Oviductal Tissue or Conditioned Medium in Vitro

    Institute of Scientific and Technical Information of China (English)

    钟瑜; 张春雪; 潘善培

    1994-01-01

    The Present investigation has been carried out to examine the effect of human oviductal tissue co-culture system on the development of mouse embryos in vitro.Two-cell embryos collected from superovulated mouse were co-cultured with human oviductal tissue suspended in Ham'd F10+10%Fetal Calf Serum(F10 FCS),or in oviductal tissue conditioned medium and F10FCS as control.The results showed that the proportion developed into blastocyst,proportion of hatched and the velocity of cmbryo development were higher in both tissue co-culture and conditioned medium as compared with F10 FCS control.Furthermore,the velocity and percentage of embryomic devetopmem were higher in co-culture with ampullary tissue or its conditioned medium than that of isthmus,the effects of co-culture and conditioned medium on embryo development had no significant difference.All the embryos obtained from two co-culture systems could cleave normally,This experimental observation indicated that human oviductal epithelium might secrete some factors to promote the embryonic development in vitro.

  10. Optimization of culture conditions and medium composition for the marine algicidal bacterium Alteromonas sp. DH46 by uniform design

    Science.gov (United States)

    Lin, Jing; Zheng, Wei; Tian, Yun; Wang, Guizhong; Zheng, Tianling

    2013-09-01

    Harmful algal blooms (HABs) have led to extensive ecological and environmental issues and huge economic losses. Various HAB control techniques have been developed, and biological methods have been paid more attention. Algicidal bacteria is a general designation for bacteria which inhibit algal growth in a direct or indirect manner, and kill or damage the algal cells. A metabolite which is strongly toxic to the dinoflagellate Alexandrium tamarense was produced by strain DH46 of the alga-lysing bacterium Alteromonas sp. The culture conditions were optimized using a single-factor test method. Factors including carbon source, nitrogen source, temperature, initial pH value, rotational speed and salinity were studied. The results showed that the cultivation of the bacteria at 28°C and 180 r min-1 with initial pH 7 and 30 salt contcentration favored both the cell growth and the lysing effect of strain DH46. The optimal medium composition for strain DH46 was determined by means of uniform design experimentation, and the most important components influencing the cell density were tryptone, yeast extract, soluble starch, NaNO3 and MgSO4. When the following culture medium was used (tryptone 14.0g, yeast extract 1.63g, soluble starch 5.0 g, NaNO3 1.6 g, MgSO4 2.3 g in 1L), the largest bacterial dry weight (7.36 g L-1) was obtained, which was an enhancement of 107% compared to the initial medium; and the algal lysis rate was as high as 98.4% which increased nearly 10% after optimization.

  11. Optimization of Culture Conditions and Medium Composition for the Marine Algicidal Bacterium Alteromonas sp.DH46 by Uniform Design

    Institute of Scientific and Technical Information of China (English)

    LIN Jing; ZHENG Wei; TIAN Yun; WANG Guizhong; ZHENG Tianling

    2013-01-01

    Harmful algal blooms (HABs) have led to extensive ecological and environmental issues and huge economic losses.Various HAB control techniques have been developed,and biological methods have been paid more attention.Algicidal bacteria is a general designation for bacteria which inhibit algal growth in a direct or indirect manner,and kill or damage the algal cells.A metabolite which is strongly toxic to the dinoflagellate Alexandrium tamarense was produced by strain DH46 of the alga-lysing bacterium Alteromonas sp.The culture conditions were optimized using a single-factor test method.Factors including carbon source,nitrogen source,temperature,initial pH value,rotational speed and salinity were studied.The results showed that the cultivation of the bacteria at 28℃ and 180r min-1 with initial pH 7 and 30 salt contcentration favored both the cell growth and the lysing effect of strain DH46.The optimal medium composition for strain DH46 was determined by means of uniform design experimentation,and the most important components influencing the cell density were tryptone,yeast extract,soluble starch,NaNO3 and MgSO4.When the following culture medium was used (tryptone 14.0g,yeast extract 1.63g,soluble starch 5.0g,NaNO3 1.6g,MgSO4 2.3 g in 1L),the largest bacterial dry weight (7.36gL-1) was obtained,which was an enhancement of 107% compared to the initial medium; and the algal lysis rate was as high as 98.4% which increased nearly 10% after optimization.

  12. Enrichment of prostate cancer stem cells from primary prostate cancer cultures of biopsy samples

    OpenAIRE

    Wang, Shunqi; Huang, Shengsong; Zhao, Xin; Zhang, Qimin; Wu, Min; Sun, Feng; Han, Gang; Wu, Denglong

    2013-01-01

    This study was to enrich prostate cancer stem cells (PrCSC) from primary prostate cancer cultures (PPrCC). Primary prostate cancer cells were amplified in keratinocyte serum-free medium with epidermal growth factor (EGF) and bovine pituitary extract (BPE), supplemented with leukemia inhibitory factor (LIF), stem cell factor (SCF) and cholera toxin. After amplification, cells were transferred into ultra-low attachment dishes with serum-free DMEM/F12 medium, supplemented with EGF, basic fibrobl...

  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. Culture conditions affect cardiac differentiation potential of human pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Marisa Ojala

    Full Text Available Human pluripotent stem cells (hPSCs, including human embryonic stem cells (hESCs and human induced pluripotent stem cells (hiPSCs, are capable of differentiating into any cell type in the human body and thus can be used in studies of early human development, as cell models for different diseases and eventually also in regenerative medicine applications. Since the first derivation of hESCs in 1998, a variety of culture conditions have been described for the undifferentiated growth of hPSCs. In this study, we cultured both hESCs and hiPSCs in three different culture conditions: on mouse embryonic fibroblast (MEF and SNL feeder cell layers together with conventional stem cell culture medium containing knockout serum replacement and basic fibroblast growth factor (bFGF, as well as on a Matrigel matrix in mTeSR1 medium. hPSC lines were subjected to cardiac differentiation in mouse visceral endodermal-like (END-2 co-cultures and the cardiac differentiation efficiency was determined by counting both the beating areas and Troponin T positive cells, as well as studying the expression of OCT-3/4, mesodermal Brachyury T and NKX2.5 and endodermal SOX-17 at various time points during END-2 differentiation by q-RT-PCR analysis. The most efficient cardiac differentiation was observed with hPSCs cultured on MEF or SNL feeder cell layers in stem cell culture medium and the least efficient cardiac differentiation was observed on a Matrigel matrix in mTeSR1 medium. Further, hPSCs cultured on a Matrigel matrix in mTeSR1 medium were found to be more committed to neural lineage than hPSCs cultured on MEF or SNL feeder cell layers. In conclusion, culture conditions have a major impact on the propensity of the hPSCs to differentiate into a cardiac lineage.

  15. Modeling of cell culture damage and recovery leads to increased antibody and biomass productivity in CHO cell cultures.

    Science.gov (United States)

    Naderi, Saeideh; Nikdel, Ali; Meshram, Mukesh; McConkey, Brendan; Ingalls, Brian; Budman, Hector; Scharer, Jeno

    2014-09-01

    The development of an efficient and productive cell-culture process requires a deep understanding of intracellular mechanisms and extracellular conditions for optimal product synthesis. Mathematical modeling provides an effective strategy to predict, control, and optimize cell performance under a range of culture conditions. In this study, a mathematical model is proposed for the investigation of cell damage of a Chinese hamster ovary cell culture secreting recombinant anti-RhD monoclonal antibody (mAb). Irreversible cell damage was found to be correlated with a reduction in pH. This irreversible damage to cellular function is described mathematically by a Tessier-based model, in which the actively growing fraction of cells is dependent on an intracellular metabolic product acting as a growth inhibitor. To further verify the model, an offline model-based optimization of mAb production in the cell culture was carried out, with the goal of minimizing cell damage and thereby enhancing productivity through intermittent refreshment of the culture medium. An experimental implementation of this model-based strategy resulted in a doubling of the yield as compared to the batch operation and the resulting biomass and productivity profiles agreed with the model predictions.

  16. 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

  17. Lack of evidence of chalone activity in used medium and extract of JB-1 tumor cells in vitro. A flow cytometry study.

    Science.gov (United States)

    Naeser, F K; Barfod, N M; Bichel, P

    1978-02-14

    In cell-free mouse ascites fluid from the JB-1 ascites tumor in the plateau phase of growth low-molecular chalone substances have been found which reversibly and specifically arrest JB-1 cells in the G1 and G2 phase of the cell cycle. The aim of this study was to investigate whether chalones were involved in the regulation of in vitro growth of JB-1 tumor cells. Used medium and cell extract from confluent, stationary JB-1 cell cultures were investigated for proliferation-inhibitory properties. JB-1 cells from stationary cultures were explanted in test cultures and the traverse of cells through the S phase was investigated by means of flow cytometry (FCM). Inhibition--expressed as a delay of the traverse of cells through the S phase--was not observed when a surplus of used medium, concentrated and fractionated used medium or concentrated and fractionated cell extract from JB-1 cells in vitro was added to test cultures. On the contrary, used medium and concentrated and fractionated used medium stimulated growth. Thus, no involvement of chalones in the growth regulation of JB-1 tumor cells in vitro was detected.

  18. 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...... and introducing the observed difference between batches as one of the factors in an expanded three-dimensional ANOVA, we were able to overcome an otherwisecrucial lack of sufficiently reproducible duplicate values. We could thereby show that the effect of changing the apical medium was much more marked when...... the radioactive lipid precursors were added on the apical, rather than on the basolateral, side. Theinsert cell cultures were obviously polarized. We argue that it is not reasonable to reject troublesome experimental results, when we do not know a priori that something went wrong. The ANOVA is a very useful...

  19. Gas-permeable lifecell tissue culture flasks give improved growth of Helicobacter pylori in a liquid medium.

    OpenAIRE

    Secker, D A; Tompkins, D S; Alderson, G

    1991-01-01

    Experiments were conducted to investigate the microaerobic culture of Helicobacter pylori in a liquid medium by using gas-permeable Lifecell tissue culture flasks. Growth in Lifecell tissue culture flasks was 1.2 to 1.6 log units greater than that in glass control bottles. These results were comparable to those reported by the use of gyrated media.

  20. Preparation and Primary Culture of Liver Cells Isolated from Adult Rats by Dispase Perfusion

    Directory of Open Access Journals (Sweden)

    Wahid,Syarifuddin

    1984-06-01

    Full Text Available The dispase perfusion technique was used to isolate liver cells from adult rats. The optimum conditions for obtaining many isolated liver cells with high viability were an enzyme concentration of 2000 U/ml, a pH of 7.5 and a perfusion time of 20 min. The population of isolated liver cells prepared with dispase consisted of 43.6% cells with diameters less than 20 micron and 56.4% cells with diameters above 20 micron. The isolated liver cells were cultured in basal culture medium either supplemented with or without dexamethasone (1 X 10(-5M and insulin (10 micrograms/ml. The addition of hormones to the culture medium improved the attachment efficiency of the isolated liver cells and delayed the disappearance of mature hepatocytes. Epithelial-like clear cells proliferated early in primary culture even in the presence of hormones. Therefore, functioning mature hepatocytes and proliferating epithelial-like clear cells coexisted well in the hormone-containing medium. Furthermore, the number of cultured cells reached a maximal level earlier in the presence of hormones than in the absence of hormones. The level of TAT activity in primary cultured cells was higher up to 3 days after inoculation in the presence of hormones than in their absence. No difference between G6Pase activity in primary cultured cells in the presence of hormones and that in the absence of hormones was found.

  1. Organizational Culture and Open Innovation Performance in Small and Medium-sized Enterprises (SMEs in Poland

    Directory of Open Access Journals (Sweden)

    Mazur Jolanta

    2016-09-01

    Full Text Available This study investigates the links between organizational culture, the use of open innovation sources and the performance of SMEs. The main hypothesis of the study is that a special type of organizational culture (termed innovative culture, which fosters creativity, learning and inter-employee cooperation – will correspond with a greater scope of open innovation sources and higher levels of innovative, operational and financial performance. The study was based on a representative CATI survey of 473 SMEs operating in manufacturing and services industries in Poland. Our statistical analysis relied on building and testing structural equation model with the AMOS software. The findings confirmed a positive association between innovative culture and the scope of open sources of innovation. However, innovative culture had no direct effect on the percentage of sales from new and modified products, which is often used as a metric of innovativeness, but did show a positive influence on an index of operational performance and ROI. Such statistical patterns suggest that fostering innovative culture is beneficial to a company, though probably not through an increased number of product innovations, but rather via process, administrative and marketing innovations, as well as other gains in efficiency attained due to more streamlined employee cooperation and knowledge exchange. The study adds to the existing body of knowledge in management science by providing a better understanding of mechanisms underlying innovative culture’s impacts on open innovation practices and metrics of operational and financial performance in the context of small and medium enterprises.

  2. 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

  3. Disposable Bioreactors for Plant Micropropagation and Mass Plant Cell Culture

    Science.gov (United States)

    Ducos, Jean-Paul; Terrier, Bénédicte; Courtois, Didier

    Different types of bioreactors are used at Nestlé R&D Centre - Tours for mass propagation of selected plant varieties by somatic embryogenesis and for large scale culture of plants cells to produce metabolites or recombinant proteins. Recent studies have been directed to cut down the production costs of these two processes by developing disposable cell culture systems. Vegetative propagation of elite plant varieties is achieved through somatic embryogenesis in liquid medium. A pilot scale process has recently been set up for the industrial propagation of Coffea canephora (Robusta coffee). The current production capacity is 3.0 million embryos per year. The pre-germination of the embryos was previously conducted by temporary immersion in liquid medium in 10-L glass bioreactors. An improved process has been developed using a 10-L disposable bioreactor consisting of a bag containing a rigid plastic box ('Box-in-Bag' bioreactor), insuring, amongst other advantages, a higher light transmittance to the biomass due to its horizontal design. For large scale cell culture, two novel flexible plastic-based disposable bioreactors have been developed from 10 to 100 L working volumes, validated with several plant species ('Wave and Undertow' and 'Slug Bubble' bioreactors). The advantages and the limits of these new types of bioreactor are discussed, based mainly on our own experience on coffee somatic embryogenesis and mass cell culture of soya and tobacco.

  4. 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)

  5. Somatic Embryogenesis from Cell Suspension Cultures of Aspen Clone

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Suspension cultures initiated from callus derived from petiole explants of aspen hybrid (Populus tremuloides × P.tremula) produced somatic embryos. Callus was induced on a MS medium supplemented with 5 mg·L-1 2,4-D and 0.05 mg·L-1 zeatin under light conditions. Embryogenic calli were obtained when a subsequent subculture of calli was suspended in the same basal medium with 10 mg·L-1 2,4-D. The highest number of globular embryos were induced from embryogenic calli by cell suspension culture in a MS liquid medium supplemented with 10 mg·L-1 2,4-D. Genotype and 2,4-D concentration were vital to the induction of embryogenic calli producing competent cells. Embryogenic calli for each genotype were heterogeneous. Green calli with gel-like consistency could yield more competent cells than light yellow embryogenic calli. However, some globular embryos broke into slices and some developed abnormally after one month of culture under the same or other hormonal conditions.

  6. The effect of medium composition on interleukin-2 production by murine EL-4 thymoma cells

    OpenAIRE

    Galesi A. L. L.; Tamashiro W. M. S. C.; Moraes A. M.

    2004-01-01

    Due to the role of interleukin-2 (IL-2) in the mediation of immune response, this cytokine has been used in the treatment of some types of cancer and infectious diseases. However, relatively high levels of this cytokine are required to achieve significant activity. The aim of this work was to study a culture medium composition designed to increase the production of IL-2 by suspended murine EL-4 cells. The cultivations were carried out aiming at producing IL-2 in stirred bioreactors. The effec...

  7. 筋脉通含药血清降低高糖培养大鼠雪旺细胞活性氧水平及PARP-1蛋白表达%Effects of Chinese herbal medicine Jinmaitong-containing serum on the ROS level and expression of PARP-1 of rat Schwann cells cultured in high-glucose medium

    Institute of Scientific and Technical Information of China (English)

    石玥; 梁晓春; 张宏; 王普艳; 赵丽

    2012-01-01

    Objective To investigate the effects of medicated serum prepared by administration of Jinmaitong (JMT) , a compound Chinese herbal medicine, on oxidative damage and poly ADP-ribose polymerase-1 (PARP-1) of Schwann cells cultured in high-glucose medium. Methods SD rats were divided into normal control group ( distilled water) ,JMT group and vitamin C group to prepare medicated serum. Bilateral sciatic nerves of new born SD rats were used to separate Schwann cells. Schwann cells cultured in high-glucose medium were divided into high glucose group (50 mmol/L glucose medium,JMT group( JMT-medica-ted serum) and vitamin C( VC) group (VC-medicated serum). Schwann cells cultured in DMEM were used as the normal control. After48 h culturing, the level of ROS was measured by confocal laser scanning microscope with 2',7'-dichlorofluorescein( DCF) as a molecular probe and the expression of PARP-1 protein was detected by Western blot. Results 1) Compared with high glucose group, the fluorescence intensities of ROS-DEC in Schwann cells cultured in JMT and VC groups were weaker significantly (P <0. 01). There were no significant differences between these two treated groups. 2) Compared with high glucose group,the expression of PARP-1 (89 ku) in Schwann cells cultrued in JMT group decreased significantly (P <0. 01). The expression of JMT group was also much lower than that of VC group (P <0. 01). Conclusions The medicated serum of JMT down-regulates the expression of ROS and PARP-1 of Schwann cells cultured in high glucose medium and reduces the oxidative DNA damage.%目的 研究筋脉通含药血清对体外高糖培养雪旺细胞活性氧(ROS)水平及多聚(ADP-核糖)聚合酶-1(PARP-1)蛋白表达的影响.方法 30只雄性SD大鼠随机分为3组,分别灌胃筋脉通(JMT)、维生素C(VC)或蒸馏水制备含药血清和对照血清.取新出生大鼠的双侧坐骨神经用于制备雪旺细胞,分为高糖组、JMT组(加入筋脉通含药血清)、VC组(加入维生

  8. Optimization of the medium perfusion rate in a packed-bed bioreactor charged with CHO cells.

    Science.gov (United States)

    Meuwly, F; von Stockar, U; Kadouri, A

    2004-09-01

    In the present study, the optimal medium perfusion rate to be used for the continuous culture of a recombinant CHO cell line in a packed-bed bioreactor made of Fibra-Cel((R)) disk carriers was determined. A first-generation process had originally been designed with a high perfusion rate, in order to rapidly produce material for pre-clinical and early clinical trials. It was originally operated with a perfusion of 2.6 vvd during production phase in order to supply the high cell density (2.5x10(7) cell ml(-1) of packed-bed) with sufficient fresh medium. In order to improve the economics of this process, a reduction of the medium perfusion rate by -25% and -50% was investigated at small-scale. The best option was then implemented at pilot scale in order to further produce material for clinical trials with an improved second-generation process. With a -25% reduction of the perfusion rate, the volumetric productivity was maintained compared to the first-generation process, but a -30% loss of productivity was obtained when the medium perfusion rate was further reduced to -50% of its original level. The protein quality under reduced perfusion rate conditions was analyzed for purity, N-glycan sialylation level, abundance of dimers or aggregates, and showed that the quality of the final drug substance was comparable to that obtained in reference conditions. Finally, a reduction of -25% medium perfusion was implemented at pilot scale in the second-generation process, which enabled to maintain the same productivity and the same quality of the molecule, while reducing costs of media, material and manpower of the production process. For industrial applications, it is recommended to test whether and how far the perfusion rate can be decreased during the production phase - provided that the product is not sensitive to residence time - with the benefits of reduced cost of goods and to simplify manufacturing operations. PMID:19003257

  9. Timescale of silver nanoparticle transformation in neural cell cultures impacts measured cell response

    Energy Technology Data Exchange (ETDEWEB)

    Hume, Stephanie L.; Chiaramonti, Ann N.; Rice, Katherine P.; Schwindt, Rani K. [National Institute of Standards and Technology (NIST), Applied Chemicals and Materials Division (United States); MacCuspie, Robert I. [National Institute of Standards and Technology (NIST), Materials Measurement Science Division (United States); Jeerage, Kavita M., E-mail: jeerage@boulder.nist.gov [National Institute of Standards and Technology (NIST), Applied Chemicals and Materials Division (United States)

    2015-07-15

    Both serum protein concentration and ionic strength are important factors in nanoparticle transformation within cell culture environments. However, silver nanoparticles are not routinely tracked at their working concentration in the specific medium used for in vitro toxicology studies. Here we evaluated the transformation of electrostatically stabilized citrate nanoparticles (C-AgNPs) and sterically stabilized polyvinylpyrrolidone nanoparticles (PVP-AgNPs) in a low-serum (∼ 0.2 mg/mL bovine serum albumin) culture medium, while measuring the response of rat cortex neural progenitor cells, which differentiate in this culture environment. After 24 h, silver nanoparticles at concentrations up to 10 µg/mL did not affect adenosine triphosphate levels, whereas silver ions decreased adenosine triphosphate levels at concentrations of 1.1 µg/mL or higher. After 240 h, both silver nanoparticles, as well as silver ion, unambiguously decreased adenosine triphosphate levels at concentrations of 1 and 1.1 µg/mL, respectively, suggesting particle dissolution. Particle transformation was investigated in 1:10 diluted, 1:2 diluted, or undiluted differentiation medium, all having an identical protein concentration, to separate the effect of serum protein stabilization from ionic strength destabilization. Transmission electron microscopy images indicated that particles in 1:10 medium were not surrounded by proteins, whereas particles became clustered within a non-crystalline protein matrix after 24 h in 1:2 medium and at 0 h in undiluted medium. Despite evidence for a protein corona, particles were rapidly destabilized by high ionic strength media. Polyvinylpyrrolidone increased the stability of singly dispersed particles compared to citrate ligands; however, differences were negligible after 4 h in 1:2 medium or after 1 h in undiluted medium. Thus low-serum culture environments do not provide sufficient colloidal stability for long-term toxicology studies with citrate

  10. Timescale of silver nanoparticle transformation in neural cell cultures impacts measured cell response

    International Nuclear Information System (INIS)

    Both serum protein concentration and ionic strength are important factors in nanoparticle transformation within cell culture environments. However, silver nanoparticles are not routinely tracked at their working concentration in the specific medium used for in vitro toxicology studies. Here we evaluated the transformation of electrostatically stabilized citrate nanoparticles (C-AgNPs) and sterically stabilized polyvinylpyrrolidone nanoparticles (PVP-AgNPs) in a low-serum (∼ 0.2 mg/mL bovine serum albumin) culture medium, while measuring the response of rat cortex neural progenitor cells, which differentiate in this culture environment. After 24 h, silver nanoparticles at concentrations up to 10 µg/mL did not affect adenosine triphosphate levels, whereas silver ions decreased adenosine triphosphate levels at concentrations of 1.1 µg/mL or higher. After 240 h, both silver nanoparticles, as well as silver ion, unambiguously decreased adenosine triphosphate levels at concentrations of 1 and 1.1 µg/mL, respectively, suggesting particle dissolution. Particle transformation was investigated in 1:10 diluted, 1:2 diluted, or undiluted differentiation medium, all having an identical protein concentration, to separate the effect of serum protein stabilization from ionic strength destabilization. Transmission electron microscopy images indicated that particles in 1:10 medium were not surrounded by proteins, whereas particles became clustered within a non-crystalline protein matrix after 24 h in 1:2 medium and at 0 h in undiluted medium. Despite evidence for a protein corona, particles were rapidly destabilized by high ionic strength media. Polyvinylpyrrolidone increased the stability of singly dispersed particles compared to citrate ligands; however, differences were negligible after 4 h in 1:2 medium or after 1 h in undiluted medium. Thus low-serum culture environments do not provide sufficient colloidal stability for long-term toxicology studies with citrate

  11. Application of a serum-free medium for the growth of Vero cells and the production of reovirus.

    Science.gov (United States)

    Butler, M; Burgener, A; Patrick, M; Berry, M; Moffatt, D; Huzel, N; Barnabé, N; Coombs, K

    2000-01-01

    Two strains of reovirus (serotype 1 Lang/TIL and serotype 3 Dearing/T3D) were propagated in Vero cells grown in stationary or agitated cultures in a serum-free medium, M-VSFM. Solid microcarriers (Cytodex-1) were used to support cell growth in agitated cultures with a normal doubling time of 25 h. Cell yields of 1 x 10(6) cells/mL were obtained from an inoculum of 2 x 10(5) cells/mL in 4 days in microcarrier cultures. The growth profile and cell yield was not significantly different from serum-supplemented cultures. The virus titer increased by 3-4 orders of magnitude over a culture period of 150 h. The maximum virus titer in stationary cultures reached >1 x 10(9) pfu/mL for both strains of reovirus in M-VSFM. M-VSFM also supported high viral yields in microcarrier cultures. Both the specific productivity and final viral yield was higher in M-VSFM than serum-supplemented cultures. The high viral productivity suggests that this is a suitable system for the production of reovirus as an oncolytic agent for human therapeutic use. PMID:11027181

  12. 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 conclu....... 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....

  13. Acute toxicity bioassays using Daphnia magna Straus (Cladocera, Daphniidae maintained in a modified culture medium

    Directory of Open Access Journals (Sweden)

    Mónica Núñez

    2013-05-01

    Full Text Available Daphnia magna is a test organism used in ecotoxicological assays of freshwater; however, traditional culture systems for this organism could result expensive, for that the aim of this research was to developed a new economic culture medium. With this purpose, 10 strains of D. magna were isolated, their population development was evaluated by total count of organisms and pregnant females using 3 different culture media: (A alfalfa juice, (B solved yeast and (C a mixture of alfalfa juice plus solved yeast. Successful development of 4 strains was observed in the A medium, but the same strains failed to survive in the B and the C media. The 24h and 48h EC50 average values in acute ecotoxicological assays with potassium dicromate were 0,4045 mg/L ± 0,0389 and 0,1857 mg/L ± 0,0072 respectively. Also, acute ecotoxicological assays with these 4 strains were performed using potassium cyanide, which is a toxic reactive frequently used in mining operations. In this case 24h EC50 value was 1,5388 mg/L ± 0,1146 and 48h EC50 values were 0,6359 mg/L ± 0,0516. 48h EC50 values were lower than the cyanide permissible effluent values established by the Energy and Mining Authority.

  14. Optimization of production of Brucella abortus S19 culture in bioreactor using soyabean casein digest medium.

    Science.gov (United States)

    Kamaraj, Govindasamy; Rajendra, Lingala; Shankar, Chinchkar Ramachandra; Srinivasan, Villuppanoor Alwar

    2010-10-01

    A method of cultivating Brucella abortus S19 culture in bioreactor was attempted using three different media. Culture conditions in bioreactor were optimized by varying agitation and aeration parameters. Varying the aeration ranging from 0.5 vvm to 0.8 vvm and agitation rate ranging from 250 rpm to 400 rpm during bacterial growth was found to yield highest viable count within 48 hours of culture period. A count of > 1 x 10(11) CFU per ml within 48 to 60 hours post seeding was obtained consistently in all five consecutive batches (P > 0.05) with 6 x 10(11) CFU per ml being the maximum yield when the organism is grown in soyabean casein digest medium. B. abortus S19 maintained its smooth characteristics throughout its growth in bioreactor. The vaccine prepared with soyabean casein digest medium was found to be potent and safe with a protective index of 3.33 in mice. The vaccine was tested in 10 cattle calves of 3 to 13 months age and all the vaccinated animals were seropositive on 28, 60, 90, 120 and 150 days post-vaccination when analyzed by fluorescence polarization assay (FPA). PMID:21213590

  15. Development of an alternative culture medium for the selective enumeration of Lactobacillus casei in fermented milk.

    Science.gov (United States)

    Colombo, Monique; de Oliveira, Aline Evelyn Zimmermann; de Carvalho, Antonio Fernandes; Nero, Luís Augusto

    2014-05-01

    Monitoring the populations of probiotic strains of the species Lactobacillus casei in food is required by food industries in order to assure that a minimum concentration of these organisms will be ingested by consumers. In this context, Petrifilm™ AC plates can be used along with selective culture media to allow the enumeration of specific groups of lactic acid bacteria. The present study aimed to assess chemical substances as selective agents for Lb. casei in order to propose a selective culture medium to be used with Petrifilm™ AC plates as an alternative protocol for the enumeration of probiotic strains of this species in fermented milk. Twenty-six probiotic and starter cultures (including six strains of Lb. casei) were plated on de Man Rogosa and Sharpe (MRS) agar with distinct concentrations of nalidixic acid, bile, lithium chloride, metronidazole, sodium propionate, and vancomycin. Vancomycin at 10 mg/L demonstrated selective activity for Lb. casei. In addition, 2,3,5-triphenyltetrazolium chlorine was identified as a compound that did not inhibit Lb. casei, and Petrifilm™ AC plates used with MRS and vancomycin at 10 mg/L (MRS-V) demonstrated more colonies of this organism when incubated under anaerobic conditions than aerobic conditions. Acidophilus milk and yoghurt were prepared, added to Lb. casei strains, and stored at 4 °C. Lb. casei populations were monitored using MRS-V and MRTLV by conventional plating and associated with Petrifilm™ AC plates. All correlation indices between counts obtained by conventional plating and Petrifilm™ AC were significant (p enumeration of Lb. casei strains in fermented milk. However, the selective potential of this culture medium must be evaluated considering the specific strains of Lb. casei and the starter cultures inoculated in the fermented milk that requires monitoring.

  16. Effect and Mechanism of Kisspeptin-10 on Progesterone Secretion in Granulosa Cells of Chicken F1 Follicles Cultured in Serum-Free Medium%Kisspeptin-10对无血清培养鸡F1级卵泡颗粒细胞孕酮分泌的影响及机制研究

    Institute of Scientific and Technical Information of China (English)

    肖蕴祺; 黄炎冰; 吴晶; 倪迎冬; 赵茹茜

    2012-01-01

    [目的]研究kisspeptin-10对无血清培养的鸡卵泡颗粒细胞孕酮分泌的作用及可能机制.[方法]选取200日龄处于产蛋高峰期的伊莎蛋鸡,于排卵前剖腹收集F1卵泡,分离纯化颗粒细胞,于含胎牛血清培养基中培养24 h,换成无血清培养基稳定培养24 h后,用不同浓度的Kp-10、U73122、EGTA等单独或共同处理细胞,收集细胞上清,放射免疫学方法检测培养基中孕酮含量.[结果](1)通过免疫细胞化学方法,显示颗粒细胞上有Kp-10免疫活性样物质的阳性表达;(2)Kp-10处理可显著增加无血清培养的颗粒细胞的活力,100 nmol·L-1时可显著促进孕酮的分泌(P<0.05);(3)与对照组相比,2μmol·L-1 U73122(磷脂酶C抑制剂)对孕酮的分泌无显著影响(P>0.05),而0.5、2μmol·L-1 U73122能显著降低Kp-10对孕酮分泌的促进作用(P<0.05); (4)钙离子阻断剂Verapamil( 1-100 μmol·L-1)呈剂量依赖性降低孕酮的分泌,于100μmol·L-1时达显著降低水平(P<0.05);与1 μ mol·L-1Verapamil单独处理组相比,100 nmol·L-1 Kp-10与1μmol·L-1 Verapamil同时处理可显著增加孕酮的分泌,而与10、100 μ mol·L-1Verapamil共同处理不能逆转其对孕酮分泌的抑制作用(P>0.05),且胞内钙离子含量与上清液中孕酮分泌水平相一致; (5)与100 nmol·L-1 Kp-10处理组相比,1和5mmol·L-1 EGTA共同处理时孕酮分泌显著降低,当再加入1.5 mmol·L-1 Ca2+时孕酮分泌量显著增加.[结论]Kp-10促进体外培养的鸡卵泡颗粒细胞孕酮的分泌,其机制可能与胞内Ca2+信号通路有关.%[Objective] The objective of this study was to investigate the effect and mechanism of Kisspeptin-10 (Kp-10) on progesterone (P4) secretion in cultured granulose cells of chicken follicles. [Method] Two-hundred-day ISA laying hens were sacrificed before the expected time to ovulation. The follicles (Fi) were collected and the granulosa cells were isolated and cultured in serum medium

  17. Crustacean peptidergic neurons in culture show immediate outgrowth in simple medium.

    Science.gov (United States)

    Cooke, I; Graf, R; Grau, S; Haylett, B; Meyers, D; Ruben, P

    1989-01-01

    The survival and outgrowth of neurons in culture has usually required conditioning factors. We now report that crustacean neurons, taken from the peptidergic neurosecretory system of the eyestalk of crabs (Cardisoma carnifex) and lobsters (Panulirus marginatus), show immediate outgrowth, sustained for a week or more, in defined medium as simple as physiological saline with glucose and glutamine. The neurons show peptide hormone immunoreactivity that is prominent at growth cones, exhibit differences in form correlated with their immunoreactivity, release peptides to the medium, and have voltage-dependent currents, including a well-sustained Ca current. Cd blocks secretion, growth, and the Ca current. Peptidergic secretory neurons may be able to utilize existing membrane from their store of granules and already active synthetic, transport, and secretory mechanisms for immediate outgrowth.

  18. Resource efficiency and culture--workplace training for small and medium-sized enterprises.

    Science.gov (United States)

    Bliesner, Anna; Liedtke, Christa; Rohn, Holger

    2014-05-15

    Although there are already some qualification offers available for enterprises to support resource efficiency innovations, the high potentials that can be identified especially for small and medium sized enterprises (SMEs) have not been activated until now. As successful change lies in the hands of humans, the main aim of vocational education has to be the promotion of organisational and cultural changes in the enterprises. As there is already a small but increasing number of enterprises that perform very well in resource efficiency innovations one question arises: What are typical characteristics of those enterprises? Leaning on a good-practice approach, the project "ResourceCulture" is going to prove or falsify the hypothesis that enterprises being successful with resource efficiency innovations have a specific culture of trust, which substantially contributes to innovation processes, or even initially enables them. Detailed empirical field research will light up which correlations between resource efficiency, innovation and cultures of trust can be found and will offer important aspects for the improvement of management instruments and qualification concepts for workplace training. The project seizes qualification needs that were likewise mentioned by enterprises and consultants, regarding the implementation of resource efficiency. This article - based on first empirical field research results - derives preliminary indications for the design of the qualification module for the target groups resource efficiency consultants and managers. On this basis and in order to implement "ResourceCulture" conceptual and methodological starting points for workplace training are outlined.

  19. 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...

  20. 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

  1. Data in support of the mutagenic potential of the isoflavone irilone in cultured V79 cells

    OpenAIRE

    Anne Scheffler; Annette E. Albrecht; Esch, Harald L.; Leane Lehmann

    2015-01-01

    The isoflavone irilone is found in human plasma after ingestion of red clover-based dietary supplements, but information allowing safety assessment is rare. Here, data in support of the mutagenic potential of irilone in cultured V79 cells [1] are presented. These data include (i) a quantitative assessment of irilone in the culture medium during the cell culture experiments, (ii) changes in the mutation spectrum in cDNA of the hypoxanthine-guanine phosphoribosyltransferase locus of irilone-tre...

  2. Condition medium of HepG-2 cells induces the transdifferentiation of human umbilical cord mesenchymal stem cells into cancerous mesenchymal stem cells.

    Science.gov (United States)

    Yang, Juan; Miao, Yinglei; Chang, Yefei; Zhang, Fan; Wang, Yubo; Zheng, Sheng

    2016-01-01

    This study aimed to investigate the transdifferentiation of human umbilical cord mesenchymal stem cells (hUCMSCs) into cancer-associated mesenchymal stem cells (CA-MSCs) after incubation with condition medium (CM) from liver cancer HepG-2 cells, and the biobehaviors (proliferation and migration) of these CA-MSCs were further evaluated. The supernatant of HepG-2 cells was collected and mixed with equal volume of low glucose DMEM. The resultant medium was used to treat hUCMSCs for 48 h. The expression of CA-MSCs related proteins and miR-221 was detected in cells. The supernatant of induced hUCMSCs was mixed with equal volume of high glucose DMEM, and the resultant medium was used treat HepG-2 cells for 48 h and the proliferation and migration of HepG-2 cells were evaluated. Moreover, HepG-2 cells were co-cultured with hUCMSCs and then the proliferation and migration of HepG-2 cells were assessed. After incubation with the supernatant from HepG-2 cells, hUCMSCs showed significantly elevated expression of vimentin, fibroblast activation protein (FAP) and miR-221. The supernatant of induced hUCMSCs was able to significantly increase the proliferation and migration of HepG-2 cells. Following co-culture, the proliferation and migration of HepG-2 cells increased dramatically. These findings suggest that the supernatant of HepG-2 cells is able to induce the phenotype of CA-MSCs and the supernatant of CA-MSCs may promote the proliferation and migration of HepG-2 cells. These findings provide experimental evidence for the cellular remodeling in tumor microenvironment and the safety of clinical use of hUCMSCs. PMID:27648133

  3. Withaferin A from cell cultures of Withania somnifera

    Directory of Open Access Journals (Sweden)

    Ciddi Veeresham

    2006-01-01

    Full Text Available Suspension cultures of Withania somnifera cells were established and shown to produce withaferin A. The identification of withaferin A was done by TLC, UV absorption, HPLC and electron spray mass spectroscopy. These cultures could be strongly elicited by exposure to salacin. Addition of salacin at the concentration of 750 µM to the cultures in production medium enhanced production levels of withaferin A to 25±2.9 mg/l compared to 0.47±0.03 mg/l in unelicited controls. This report is the first to demonstrate withaferin A production in plant suspension cultures and provides prerequisites for commercial scale, controlled production of withaferin A.

  4. Lipids shed into the culture medium by trypomastigotes of Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Agusti Rosalia

    2000-01-01

    Full Text Available Trypomastigote forms of Trypanosoma cruzi were metabolically labeled with [14C]-ethanolamine and [3H]-palmitic acid. Lipids shed to the culture medium were analyzed and compared with the parasite components. Phosphatidylcholine and lysophosphatidylcholine accounted for 53% of the total incorporated precursor. Interestingly, phosphatidylethanolamine and its lyso derivative lysophosphatidylethanolamine, although present in significant amounts in the parasites, could not be detected in the shed material. Shed lipids were highly enriched in the desaturated fatty acids C16:1 and C18:1 when compared to the total fatty acid pool isolated from the parasites.

  5. 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.

  6. Legionella pneumophila Arthritis: use of medium specific for Mycobacteria for isolation of L. pneumophila in culture of articular fluid specimens.

    Science.gov (United States)

    Bemer, Pascale; Leautez, Sophie; Ninin, Emmanuelle; Jarraud, Sophie; Raffi, François; Drugeon, Henri

    2002-07-01

    We report the first case, to our knowledge, of acute purulent arthritis due to Legionella pneumophila in an immunosuppressed patient. L. pneumophila was isolated from samples of blood and articular fluid cultured with use of medium specific for mycobacteria (Bactec 13A medium).

  7. Culture of hormone-dependent functional epithelial cells from rat thyroids.

    OpenAIRE

    Ambesi-Impiombato, F S; Parks, L A; Coon, H G

    1980-01-01

    Primary cultures of rat thyroid cells were made in medium supplemented with 0.1--0.5% calf serum and containing six hormones or growth factors: insulin, thyrotropin, transferrin, hydrocortisone, somatostatin, and glycyl-L-histidyl-L-lysine acetate. The FRTL strain was purified by successive colonial isolations and was found to maintain highly differentiated features (secretion into the culture medium of physiological amounts of thyroglobulin and concentration of iodide by 100-fold). The FRTL ...

  8. Autofluorescence of viable cultured mammalian cells.

    Science.gov (United States)

    Aubin, J E

    1979-01-01

    The autofluorescence other than intrinsic protein emission of viable cultured mammalian cells has been investigated. The fluorescence was found to originate in discrete cytoplasmic vesicle-like regions and to be absent from the nucleus. Excitation and emission spectra of viable cells revealed at least two distinct fluorescent species. Comparison of cell spectra with spectra of known cellular metabolites suggested that most, if not all, of the fluorescence arises from intracellular nicotinamide adenine dinucleotide (NADH) and riboflavin and flavin coenzymes. Various changes in culture conditions did not affect the observed autofluorescence intensity. A multiparameter flow system (MACCS) was used to compare the fluorescence intensities of numerous cultured mammalian cells.

  9. Detection of microorganisms in culture medium through the neutron radiographic technique

    International Nuclear Information System (INIS)

    The study aims to obtain a more effective and faster method for the detection of bacteria in several culture media, such as potable water and blood. After the process growth in the culture medium, separation and suspension in buffer solution based in boron, the bacteria are deposited in track detectors that are submitted to thermal neutron beams (neutron flux: 2,2 x 105 n.cm-2.s-1), resulting from the channel J-9 of the Argonauta research reactor, from the Nuclear Engineering Institute, IEN/CNEN. The latent tracks arisen from the alpha particles proceeding from the reaction B (η, α) Li and, after having been revealed, are analyzed by an optical microscope that allows to detect the existence of the bacteria. Afterwards, they were analyzed in a nanoscope which helps the identification of the tracks of the alpha particles. (author)

  10. Influence of culture medium growth variables on Ganoderma lucidum exopolysaccharides structural features.

    Science.gov (United States)

    Fraga, Irene; Coutinho, João; Bezerra, Rui M; Dias, Albino A; Marques, Guilhermina; Nunes, Fernando M

    2014-10-13

    In this work the effect of carbon and nitrogen levels and initial pH of the wheat extract culture medium of submerged culture of Ganoderma lucidum on the amount, purity and structural features of exopolysaccharides (EPS) were studied. A low peptone level (1.65 g L(-1)) favored mycelium biomass, EPS purity, but a higher supply of peptone (4.80 g L(-1)) is needed for maximum EPS production. The carbohydrate composition of the EPS and structural features also changed significantly according to the different growing conditions, being observed significant differences in the (1 → 3)/(1 → 4)-Glcp ratio and also on the branching degree of EPS. As the biological activities of EPS are highly dependent on the polysaccharide structural features, this variability can have implications on the EPS biological activities, but can also be used advantageously to produce tailor made polysaccharides with specific applications.

  11. In Vitro Selection of Peanut Somatic Embryos on Medium Containing Culture Filtrate of Sclerotium rolfsii and Plantlet Regeneration

    Directory of Open Access Journals (Sweden)

    YUSNITA

    2005-06-01

    Full Text Available Attempts to identify somaclonal variants of peanut with resistance to Sclerotium stem rot disease due to infection of S. rolfsii were conducted. The objectives of this study were to develop in vitro selection method using culture filtrates of S. rolfsii, identify culture filtrate-insensitive somatic embryo (SE of peanut after in vitro selection and regenerate peanut R0 lines originated from culture filtrate-insensitive SE. To achieve these objectives, peanut embryogenic tissues were cultured on selective medium containing various concentrations of S. rolfsii culture filtrates and sublethal concentration of the filtrates. Medium containing sublethal level of S. rolfsii culture filtrates was used to identify culture filtrate-insensitive SE of peanut. Subsequently, the selected SEs were germinated, plantlets were regenerated and preliminary tested against S. rolfsii. Results of the experiments showed that addition of S. rolfsii culture filtrates into medium for inducing peanut somatic embryos drastically reduced their growth and proliferation. S. rolfsii culture filtrates at 10% concentration has significantly reduced the number of proliferated SE per explant. However, sublethal level was achieved at 30% of culture filtrates concentration. Responses of five peanut cultivars against 30% of culture filtrates were similar, indicating they were similar in their susceptibility against S. rolfsii. A number of culture filtrate-insensitive SE were identified after culturing 1500 clumps of embryogenic tissue of peanut cv. Kelinci for three consecutive passages on medium containing 30% of culture filtrates. Germination of selected SE and regeneration of plantlet from culture filtrate-insensitive SE resulted in 50 peanut R0 lines. These lines have been grown in the plastic house and produced normal seeds for further evaluation. Results of S. rolfsii inoculation indicated the existence of chimera for insensitivity against S. rolfsii.

  12. Effects of sinusoidal endothelial cell conditioned medium on the expressionof connective tissue growth factor in rat hepatic stellate cells

    Institute of Scientific and Technical Information of China (English)

    Xiao Jing Liu; Fang Liu; Wen Jun Xiao; Ming Hui Huang; Song Min Huang; Yi Ping Wang

    2000-01-01

    AIM To investigate the effects of sinusoidal endothelial cell (SEC) conditioned medium on the expression ofconnective tissue growth factor (CTGF) in rat hepatic stellate cells (HSC).METHODS By in situ collagenase perfusion and two-step Percoll gradient centrifugation, SECs wereisolated and cultured from normally and CCl4-treated Wistar rats, and the SEC conditioned media werecollected. HSCs were prepared from Wistar rats by in situ perfusion and single-step Nycodenz gradient, andwere cultured with SEC conditioned media. Expression of CTGF in HSC was assessed using reversetranscription-polymerase chain reaction (RT-PCR).RESULTS Expression of CTGF was not found in freshly isolated HSC and in primary culture of HSC onday 4 with SEC conditioned media from normal rats, but was present in primary culture of HSC on day 4 withSEC conditioned media from CCl4-induced liver fibrosis rats. Expression of CTGF was observed in culture-activated HSCs, and the effect of SEC conditioned media from CCl4-induced liver fibrosis rats on theexpression of CTGF gene in activated HSCs was not significant.CONCLUSION Expression of CTGF might be relative to the activation of HSC and the liver fibrogenesis,and damaged SECs play a very important role in the early stage of activation of HSC.

  13. 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

  14. Hemolymph analysis and evaluation of newly formulated media for culture of shrimp cells (Penaeus stylirostris).

    Science.gov (United States)

    Shimizu, C; Shike, H; Klimpel, K R; Burns, J C

    2001-06-01

    Creation of a shrimp cell line has been an elusive goal. This failure may be due to the composition of the cell culture medium, which may be inadequate to support primary cultured cells. Shrimp hemolymph should contain the nutritional components needed to support cell growth and division. We report here the comprehensive biochemical analysis of hemolymph from the blue shrimp, Penaeus stylirostris (Litopenaeus stylirostris) (see Holthuis, L. B. Shrimps and prawns of the world, in: FAO species catalog. Vol. 1. Rome: Food and Agriculture Organization of the United Nations; 1980), for free amino acids (FAAs), carbohydrates, electrolytes, metals, pH, and osmolality. Levels of hemolymph components were compared to 2xL-15 with 20% fetal bovine serum, a commonly used culture medium for crustacean cells. The FAAs, taurine and proline, and the metals, strontium and zinc, were significantly higher in hemolymph than in the 2 x L-15 medium. In contrast, other FAAs were up to 50 times higher in the 2 x L-15 medium than in the hemolymph. To mimic more closely the hemolymph composition, we created two new media based on either the 0.2 x L-15 or the M199 medium. We compared the microscopic appearance of cells cultured in these media and evaluated deoxyribonucleic acid (DNA) and protein synthesis by 3H-thymidine uptake and 35S-methionine uptake assays. The ovary cells of P. stylirostris cultured in either of the new media formed monolayers, while the cells cultured in 2 x L-15 medium did not. Despite these differences, there was no evidence of sustained DNA or protein synthesis with any of the media. Future studies to establish a shrimp cell line should focus on analysis of the cell cycle and on overcoming the molecular blocks to cell division. PMID:11515962

  15. Development of an Integrated Microfluidic Perfusion Cell Culture System for Real-Time Microscopic Observation of Biological Cells

    Directory of Open Access Journals (Sweden)

    Chih-Chin Oh-Yang

    2011-08-01

    Full Text Available This study reports an integrated microfluidic perfusion cell culture system consisting of a microfluidic cell culture chip, and an indium tin oxide (ITO glass-based microheater chip for micro-scale perfusion cell culture, and its real-time microscopic observation. The system features in maintaining both uniform, and stable chemical or thermal environments, and providing a backflow-free medium pumping, and a precise thermal control functions. In this work, the performance of the medium pumping scheme, and the ITO glass microheater were experimentally evaluated. Results show that the medium delivery mechanism was able to provide pumping rates ranging from 15.4 to 120.0 μL·min−1. In addition, numerical simulation and experimental evaluation were conducted to verify that the ITO glass microheater was capable of providing a spatially uniform thermal environment, and precise temperature control with a mild variation of ±0.3 °C. Furthermore, a perfusion cell culture was successfully demonstrated, showing the cultured cells were kept at high cell viability of 95 ± 2%. In the process, the cultured chondrocytes can be clearly visualized microscopically. As a whole, the proposed cell culture system has paved an alternative route to carry out real-time microscopic observation of biological cells in a simple, user-friendly, and low cost manner.

  16. Cultivation and identification of human colon cancer stem-like cells in the serum-free culture medium in vitro%结肠癌干细胞样细胞的无血清法培养及特性鉴定

    Institute of Scientific and Technical Information of China (English)

    杨婷; 马磊; 李盛; 管向东

    2015-01-01

    Objective To generate suspended spheres ,Human colon cancer cell line SW620 cells were acclimated gradually in the free serum culture medium ,1 week later ,a great quantity of spheres were obtained for the further research .Methods SW620 cells and spheres were incubated with the following Abs :anti‐SSEA‐4 Ab and anti‐TRA‐1‐60 Ab and detected with confocal microscopy . SW620 cells and spheres were collected and stained by PI ,Cell cycle distribution was then acquired by flow cytometry .SW620 cells and spheres were transferred into Adipocyte Differentiation Medium and cultured for 7 d and then lipid droplets were counted . SW620 cells and sphere cells were injected subcutaneously into the lateral root of one posterior limb of a nude mouse ,tumor volume was calculated .Results Spheres formed when grown under serum‐free conditions in 7 days ,and the ability of SW620 cells to form spheres was sustained .A few of SW620 adherent cells be cultured in DF+10% FBS clearly showed the expression of pluripotent stem cell markers Ssea‐4(25 .739 ± 7 .62)% ,Tra‐1‐60(27 .742 ± 4 .311)% ,but almost all spheres expressed the 2 markers and there was a significant difference between the two groups in the expression ratio (P<0 .05) .It had been shown that the spheres exhibited a relatively high proportion of cells in G0/G1 phase(84 .19 ± 2 .52)% and low proportion in S phase(7 .18 ± 1 .35)% ;when com‐pared to SW620 cells ,G0/G1 phase(63 .02 ± 6 .73)% ,S phase(20 .89 ± 3 .84)% (P<0 .05) .Adipocytes drops from spheres and ad‐herent cells were calculated under microscope after been cultured in adipocytes liquid for 7 d .The number of adipocytes drops in spheres was (583 .80 ± 77 .69) ,but(169 .20 ± 26 .43)in SW620 adherent(P<0 .05) .2 × 104 sphere cells injected subcutaneously into the lateral root of one posterior limb of a nude mouse fromed transplantation tumor after 4 weeks ,and the tumor size was (2 279 .98 ± 346 .27) mm3 ;2 × 105 adherent cells

  17. Improved elastase production by Bacillus sp. EL31410—further optimization and kinetics studies of culture medium for batch fermentation

    Institute of Scientific and Technical Information of China (English)

    何国庆; 陈启和; 琚晓捷; 石乃冬

    2004-01-01

    An efficient culture medium producing a bacterial elastase with high yields was developed further following preliminary studies by means of response surface method. Central composite design (CCD) and response surface methodology were applied to optimize the medium constituents. A central composite design was used to explain the combined effect of three medium constituents, viz, glucose, K2HPO4, MgSO4·7H2O. The strain produced more elastase in the completely optimized medium, as compared with the partially optimized medium. The fitted model of the second model, as per RSM, showed that glucose was 7.4 g/100 ml, casein 1.13 g/100 ml, corn steep flour 0.616 g/100 ml, KEHPO4 0.206 g/100 ml and MgSO4·7H2O 0.034 g/100 ml. The fermentation kinetics of these two culture media in the flask experiments were analyzed. It was found that the highest elastase productivity occurred at 54 hours. Higher glucose concentration had inhibitory effect on elastase production. At the same time, we observed that the glucose consumption rate was slow in the completely optimized medium, which can explain the lag period of the highest elastase production. Some metal ions and surfactant additives also affected elastase production and cell growth.

  18. Therapy of Chronic Cardiosclerosis in WAG Rats Using Cultures of Cardiovascular Cells Enriched with Cardiac Stem Cell.

    Science.gov (United States)

    Chepeleva, E V; Pavlova, S V; Malakhova, A A; Milevskaya, E A; Rusakova, Ya L; Podkhvatilina, N A; Sergeevichev, D S; Pokushalov, E A; Karaskov, A M; Sukhikh, G T; Zakiyan, S M

    2015-11-01

    We developed a protocol for preparing cardiac cell culture from rat heart enriched with regional stem cells based on clonogenic properties and proliferation in culture in a medium with low serum content. Experiments on WAG rats with experimental ischemic myocardial damage showed that implantation of autologous regional stem cells into the left ventricle reduced the volume of cicatricial tissue, promoted angiogenesis in the damaged zone, and prevented the risk of heart failure development.

  19. Treatment of Mycoplasma Contamination in Cell Cultures with Plasmocin

    Directory of Open Access Journals (Sweden)

    Cord C. Uphoff

    2012-01-01

    Full Text Available A high percentage of cell lines are chronically infected with various mycoplasma species. The addition of antibiotics that are particularly effective against these contaminants to the culture medium during a limited period of time is a simple, inexpensive, and very practical approach for decontaminating cell cultures. Here, we examined the effectiveness of the new antimycoplasma compound Plasmocin that has been employed routinely to cleanse chronically infected cell lines. In a first round of treatment 45 out of 58 (78% mycoplasma-positive cell lines could be cured. In a second attempt using back-up cryopreserved original cells, four additional cell lines were cured; thus, the overall cure rate was 84%. Even if the mycoplasma contamination was not eradicated by Plasmocin, the parallel treatment with several other antibiotics (Baytril, BM-Cyclin, Ciprobay, MRA, or MycoZap led to the cure of all 58 cell lines. The successful decontamination was permanent as mycoplasmas were no longer detected at day +14 posttreatment and at later time points as examined by PCR which is the most sensitive and specific mycoplasma detection method. Collectively, our results highlight certain antibiotics as effective antimycoplasma reagents and support the therapeutic rationale for their use in the eradication of this notorious cell culture contaminant.

  20. PENICILLIN-STREPTOMYCIN IN THE CULTURE MEDIUM DURING IN VITRO MATURATION (IVM OF BOVINE OOCYTES AFFECTS NUCLEAR MATURATION AND SUBSEQUENT EMBRYO DEVELOPMENT

    Directory of Open Access Journals (Sweden)

    SHIRAZI A

    2001-01-01

    Full Text Available Introduction: Standard concentrations of antibiotics in culture media are thought to have no detectable toxic effects on the cultured cells. However, since antibiotics are biologically active substances, the possibility that they interfere to some extent with cellular processes occurring in the cultured cells can not always be totally excluded. This study, therefore, was conducted to assess whether the presence of penicllin-streptomycin (pen-strep during in vitro maturation (IVM of bovine cumulus oocyte complexes (COCs affect nuclear and cytoplasmic maturation and subsequent embryo development. Materials and Methods: Bovine COCs were matured at 39oC in a humidified atmosphere with 5 % CO2 in air for 24 h in: 1- culture medium M 199 supplemented with 10 % FCS (Fetal calf serum, 0.05 IU/ml rhFSH (recombinant human FSH and 100 units penicillin and 100 ?g streptomycin/ ml. 2- culture medium M 199 without FCS and rhFSH in the presence of pen-strep. Cultures without antibiotics served as control. Six series of experiments, each consisted of at least 3 replicates, were performed. Results: In vitro maturation in the presence of pen-strep in culture medium supplemented with FCS and rhFSH significantly (P<0.05 increased the percentage of MII oocytes, however, when the COCs were divided, on the basis of appearance of the cumulus investment, into bright and dark groups, this effect was less obvious in both types of COCs, 76% vs 72% in bright COCs (P= 0.149 or 83% vs 80% in dark COCs (P=0.296 in treated and control groups respectively. The percentage of oocytes with type III of cortical granules (CGs distribution was not affected in the presence of pen-strep. The COCs expansion after IVM was not affected by the presence of antibiotics in culture medium. The subsequent embryo development of IVM/IVF produced ova, which were exposed to pen-strep during IVM, was significantly (P<0.05 decreased with respect to blastocyst formation by day 9. In vitro maturation in

  1. Accumulation of {sup 137}Cs in trefoil (leaf and stem), ``Mitsuba``, Cryptotaenia japonica Hassk, immersed in hydroponic culture medium

    Energy Technology Data Exchange (ETDEWEB)

    Motegi, Misako; Miyake, Sadaaki; Ohsawa, Takashi; Nakazawa, Kiyoaki [Saitama Institute of Public Health, Urawa (Japan); Izumo, Yoshiro

    1998-11-01

    Accumulation of {sup 137}Cs in trefoil (leaf and stem), ``Mitsuba``, Cryptotaenia japonica Hassk, with or without root was investigated to prepare higher radioactive plant in hydroponic culture medium (140-150 Bq/ml). It was found that {sup 137}Cs concentration in plant tissue was increased with time, as high as 1.6 times of that in the culture medium after 4 days. On the other hand, {sup 137}Cs concentration was affected by carrier element (Cs>6 ppm) and coexistent elements in the medium. Radioactivity of the plant after 4 days was shown to be sufficient for successive experiments. (author)

  2. Cultured 3T3L1 adipocytes dispose of excess medium glucose as lactate under abundant oxygen availability

    Science.gov (United States)

    Sabater, David; Arriarán, Sofía; Romero, María Del Mar; Agnelli, Silvia; Remesar, Xavier; Fernández-López, José Antonio; Alemany, Marià

    2014-01-01

    White adipose tissue (WAT) produces lactate in significant amount from circulating glucose, especially in obesity;Under normoxia, 3T3L1 cells secrete large quantities of lactate to the medium, again at the expense of glucose and proportionally to its levels. Most of the glucose was converted to lactate with only part of it being used to synthesize fat. Cultured adipocytes were largely anaerobic, but this was not a Warburg-like process. It is speculated that the massive production of lactate, is a process of defense of the adipocyte, used to dispose of excess glucose. This way, the adipocyte exports glucose carbon (and reduces the problem of excess substrate availability) to the liver, but the process may be also a mechanism of short-term control of hyperglycemia. The in vivo data obtained from adipose tissue of male rats agree with this interpretation.

  3. Comparison of phenotype and functions of dendritic cells in culture medium supplemented with different serum or plasma%不同来源血清或血浆培养条件下的树突状细胞表型和功能的比较

    Institute of Scientific and Technical Information of China (English)

    易辉君; 卢光琇

    2012-01-01

    目的:比较在添加100 mL/L胎牛血清(FBS)、20mL/L人AB血清和10 mL/L供者自体血浆3种不同培养条件下树突状细胞(DC)的表型和功能,为临床应用DC找到安全且有效的培养方法.方法:将外周血单个核细胞( PBMCs)分别在添加100 mL/L FBS,20 mL/L人AB血清,10 mL/L供者自体血浆培养条件下诱导成DC,在培养的第8天,利用流式细胞术(FCM)检测3种培养条件下DC的得率、纯度、表型和抗原捕获能力;用MTT的方法检测DC刺激同种异体淋巴细胞增殖的能力.结果:在DC集落数量、DC得率、DC纯度、抗原捕获能力和刺激淋巴细胞增殖的能力上,100 mL/L FBS培养组优于20 mL/L AB血清和10 mL/L自体血浆组,3组之间DC得率差异比较具有统计学的意义(P<0.05),而其他比较都无统计学的意义(P>0.05).结论:利用供者自体血浆培养DC是一种安全有效的培养方法,符合细胞治疗规范,可替代异种血清或同种异体血清,成为临床级别DC培养的选择.%AIM: To find a safe and effective method of culturing dendritic cells (DCs) for clinical application by comparing the phenotype and functions of DCs in different culture mediums which are supplemented with 100 mL/L fetal bovine serum (FBS), 20 mL/L human AB serum and 10 mL/L autologous plasma. METHODS; Peripheral blood mononuclear cells (PBMCs) were induced to DCs in 100 ml/L FBS, 20 mL/L human AB serum, and 10 mL/L autologous plasma, respectively. On day 8, the purity, phenotype and antigen uptake capability of DCs were examined by flow cytometry. The mixed lymphocyte reaction (MLR) was detected by MTT assay. RESULTS; Focused on DC aggregates, DC yield, DC purity, antigen uptake capability and ability of stimulating lymphocyte proliferation, the experiments showed that those of 100 mL/L FBS culture group were better than those of 20 mL/L human AB serum and 10 mL/L autologous plasma culture groups. There was no significant difference among these three culture groups

  4. Testicular cell-conditioned medium supports embryonic stem cell differentiation toward germ lineage and to spermatocyte- and oocyte-like cells.

    Science.gov (United States)

    Shah, Syed M; Saini, Neha; Singh, Manoj K; Manik, Radheysham; Singla, Suresh K; Palta, Prabhat; Chauhan, Manmohan S

    2016-08-01

    Testicular cells are believed to secrete various growth factors that activate signaling pathways finally leading to gametogenesis. In vitro gametogenesis is an obscure but paramountly important task primarily because of paucity of the precursor cells and first trimester gonadal tissues. To overcome these limitations for development of in vitro gametes, the present study was designed to induce differentiation of buffalo embryonic stem (ES) cells into germ lineage cells on stimulation by testicular cell-conditioned medium (TCM), on the basis of the assumption that ES cells have the intrinsic property to differentiate into any cell type and TCM would provide the necessary growth factors for differentiation toward germ cell lineage. For this purpose, buffalo ES cells were differentiated as embryoid bodies (EB) in floating cultures and as monolayer adherent cultures in different doses (10%, 20%, and 40%) of TCM for different culture intervals (4, 8, and 14 days), to identify the optimum dose-and-time period. We observed that 40% TCM dose induces highest expression of primordial germ cell-specific (DAZL, VASA, and PLZF), meiotic (SYCP3, MLH1, TNP1/2, and PRM2), spermatocyte-specific (BOULE and TEKT1), and oocyte-specific genes (GDF9 and ZP2/3) for a culture period of 14 days under both floating and adherent differentiation. Immunocytochemical analysis of EBs and adherent cultures revealed presence of primordial germ cell markers (c-KIT, DAZL, and VASA), meiotic markers (SYCP3, MLH1 and PROTAMINE1), spermatocyte markers (ACROSIN and HAPRIN), and oocyte markers (GDF9 and ZP4), indicating progression into post-meiotic gametogenesis. The detection of germ cell-specific proteins in Day 14 EBs like VASA, GDF9, and ZP4 by Western blotting further confirmed germ lineage differentiation. The significantly lower (P propagation, augmentation of reproductive performance in poor breeding buffalo species, and as a model for understanding human germ cell formation. PMID:27056417

  5. A simple method for the measurement of labelled compound incorporation into cells in culture.

    Science.gov (United States)

    Suplisson, A; Boissel, J P

    1976-02-10

    A simple method for the measurement of labelled compound incorporation into cells in layer culture was developed. Compared to other methods it proves to spare time and to be more sensitive owing to the fact that cells are not detached from the culture vials until the end of the manipulation as these are dissolved in the scintillation medium together with the cells just before counting.

  6. Suitability of human mesenchymal stem cells for gene therapy depends on the expansion medium

    International Nuclear Information System (INIS)

    Great hope is set in the use of mesenchymal stem cells for gene therapy and regenerative medicine. Since the frequency of this subpopulation of stem cells in bone marrow is low, mesenchymal stem cells are expanded ex vivo and manipulated prior to experimental or clinical use. Different methods for isolation and expansion are available, but the particular effect on the stem cell character is unclear. While the isolation of mesenchymal stem cells by density centrifugation followed by selection of the plastic adherent fraction is frequently used, the composition of expansion media differs. Thus, in the present study we cultured mesenchymal stem cells isolated from five healthy young volunteers in three widely used expansion media and performed a detailed analysis of the effect on morphology, proliferation, clonogenicity, passaging, differentiation and senescence. By this way we clearly show that the type of expansion medium used determines the stem cell character and time of senescence which is critical for future gene therapeutic and regenerative approaches using mesenchymal stem cells

  7. Stimulation of DNA synthesis in cultured rat alveolar type II cells

    Energy Technology Data Exchange (ETDEWEB)

    Leslie, C.C.; McCormick-Shannon, K.; Robinson, P.C.; Mason, R.J.

    1985-01-01

    Restoration of the alveolar epithelium after injury is thought to be dependent on the proliferation of alveolar type II cells. To understand the factors that may be involved in promoting type II cell proliferation in vivo, we determined the effect of potential mitogens and culture substrata on DNA synthesis in rat alveolar type II cells in primary culture. Type II cells cultured in basal medium containing 10% fetal bovine serum (FBS) exhibited essentially no DNA synthesis. Factors that stimulated /sup 3/H-thymidine incorporation included cholera toxin, epidermal growth factor, and rat serum. The greatest degree of stimulation was achieved by plating type II cells on an extracellular matrix prepared from bovine corneal endothelial cells and then by culturing the pneumocytes in medium containing rat serum, cholera toxin, insulin, and epidermal growth factor. Under conditions of stimulation of /sup 3/H-thymidine incorporation there was an increased DNA content per culture dish but no increase in cell number. The ability of various culture conditions to promote DNA synthesis in type II cells was verified by autoradiography. Type II cells were identified by the presence of cytoplasmic inclusions, which were visualized by tannic acid staining before autoradiography. These results demonstrate the importance of soluble factors and culture substratum in stimulating DNA synthesis in rat alveolar type II cells in primary culture.

  8. Methods for Maintaining Insect Cell Cultures

    OpenAIRE

    Lynn, Dwight E.

    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.

  9. Exposure to Chlorinated Biphenyls Causes Polymorphonucleocytes to Induce Progenitor Cell Toxicity in Culture

    Directory of Open Access Journals (Sweden)

    Tanika V. Martin

    2006-03-01

    Full Text Available Progenitor cells (PC are the precursors for many developmental structures and are sensitive to a variety of toxic agents including the environmental contaminants, polychlorinated biphenyls (PCBs. The mechanism(s that contributes to the development of PCB-induced progenitor cell-related fetotoxicities are not completely understood. However, several studies have demonstrated an important role for neutrophils (polymorphonucleocytes in the development of PCB induced toxicities. Our recent findings have indicated that conditioned medium collected from PC (CMPC exposed to a single dose of the PCB mixture, Aroclor 1248, can activate isolated neutrophil populations. Because of our recent findings, this study was conducted to determine if conditioned medium from PC treated with a PCB mixture causes neutrophils to injure PC in culture. Isolated PC were cultured and treated with different concentrations of Aroclor 1248 for 24 hours. The resulting PC-derived conditioned media was collected and its affect on neutrophil activity was analyzed. Conditioned medium from PC treated with Aroclor 1248 was chemotactic for neutrophils. The conditioned medium from Aroclor 1248 treated-PC also stimulated neutrophils to release super oxide anion, cathepsin G and elastase into culture medium. Furthermore, the conditioned medium from Aroclor 1248 treated- PC was able to stimulate neutrophils to cause progenitor cell toxicity in co-cultures. The conditioned medium from Aroclor 1248 treated-PC was not toxic to individual neutrophil cultures or PC cultures. Moreover, the addition of a protease inhibitor to the co-cultures containing neutrophils and PC, afforded protection against neutrophil-induced cytotoxicity of PC. These data suggest that a PCB mixture can cause progenitor cells to produce a factor(s that activates neutrophils and stimulates them to damage PC populations in culture.

  10. Improved culture-based isolation of differentiating endothelial progenitor cells from mouse bone marrow mononuclear cells.

    Directory of Open Access Journals (Sweden)

    Haruki Sekiguchi

    Full Text Available Numerous endothelial progenitor cell (EPC-related investigations have been performed in mouse experiments. However, defined characteristics of mouse cultured EPC have not been examined. We focused on fast versus slow adherent cell population in bone marrow mononuclear cells (BMMNCs in culture and examined their characteristics. After 24 h-culture of BMMNCs, attached (AT cells and floating (FL cells were further cultured in endothelial differentiation medium separately. Immunological and molecular analyses exhibited more endothelial-like and less monocyte/macrophage-like characteristics in FL cells compared with AT cells. FL cells formed thick/stable tube and hypoxia or shear stress overload further enhanced these endothelial-like features with increased angiogenic cytokine/growth factor mRNA expressions. Finally, FL cells exhibited therapeutic potential in a mouse myocardial infarction model showing the specific local recruitment to ischemic border zone and tissue preservation. These findings suggest that slow adherent (FL but not fast attached (AT BMMNCs in culture are EPC-rich population in mouse.

  11. 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.

  12. Comparative study on the stem cell phenotypes of C6 cells under different culture conditions

    Institute of Scientific and Technical Information of China (English)

    ZHANG Suo-jun; YE Fei; XIE Rui-fan; HU Feng; WANG Bao-feng; WAN Feng; GUO Dong-sheng; LEI Ting

    2011-01-01

    Background Glioma stem cell (GSC) hypothesis posits that a subpopulation of cells within gliomas have true clonogenic and tumorigenic potential. Significantly, a more controversial correlate to GSC is that cells in different culture conditions might display distinct stem cell properties. Considering these possibilities, we applied an approach comparing stem cell characteristics of C6 glioma cells under different culture conditions.Methods C6 cells were cultured under three different growth conditions, i.e., adherent growth in conventional 10% serum medium, non-adherent spheres growth in serum-free medium, as well as adherent growth on laminin-coated flask in serum-free medium. Growth characteristics were detected contrastively through neurosphere formation assay and cell cycle analysis. Markers were determined by immunofluorescence, relative-quantitative reverse transcription (RT)-PCR,Western blotting and flow cytometry. Side population cells were analyzed via flow cytometry. Tumor models were detected by magnetic resonance imaging and hematoxylin & eosin staining. Data analyses were performed with SPSS software (17.0).Results C6 cells (C6-Adh, C6-SC-Sph and C6-SC-Adh) showed distinctive growth patterns and proliferation capacity.Compared to suspending C6-SC-Sph, adherent C6-Adh and C6-SC-Adh displayed higher growth ratio. C6-SC-Sph and C6-SC-Adh showed enhanced capability of neurosphere formation and self-renewal. High side population ratio was detected in C6-SC-Sph and C6-SC-Adh. CD133 was not detected in all three kinds of cells. Conversely, Nestin and β-Ⅲ-tubulin were demonstrated positive, nonetheless with no statistical significance (P >0.05). Interestingly, lower expression of glial fibrillary acidic protein was demonstrated in C6-SC-Sph and C6-SC-Adh. C6-Adh, C6-SC-Sph and C6-SC-Adh were all displayed in situ oncogenicity, while statistical difference of survival time was not confirmed.Conclusions C6 glioma cell line is endowed with some GSC

  13. Design and Performance of an Automated Bioreactor for Cell Culture Experiments in a Microgravity Environment

    Science.gov (United States)

    Kim, Youn-Kyu; Park, Seul-Hyun; Lee, Joo-Hee; Choi, Gi-Hyuk

    2015-03-01

    In this paper, we describe the development of a bioreactor for a cell-culture experiment on the International Space Station (ISS). The bioreactor is an experimental device for culturing mouse muscle cells in a microgravity environment. The purpose of the experiment was to assess the impact of microgravity on the muscles to address the possibility of longterm human residence in space. After investigation of previously developed bioreactors, and analysis of the requirements for microgravity cell culture experiments, a bioreactor design is herein proposed that is able to automatically culture 32 samples simultaneously. This reactor design is capable of automatic control of temperature, humidity, and culture-medium injection rate; and satisfies the interface requirements of the ISS. Since bioreactors are vulnerable to cell contamination, the medium-circulation modules were designed to be a completely replaceable, in order to reuse the bioreactor after each experiment. The bioreactor control system is designed to circulate culture media to 32 culture chambers at a maximum speed of 1 ml/min, to maintain the temperature of the reactor at 36°C, and to keep the relative humidity of the reactor above 70%. Because bubbles in the culture media negatively affect cell culture, a de-bubbler unit was provided to eliminate such bubbles. A working model of the reactor was built according to the new design, to verify its performance, and was used to perform a cell culture experiment that confirmed the feasibility of this device.

  14. Evaluation of Lo"wenstein-Jensen Medium Culture,MGIT 960 Culture and Different Specimen Types inDiagnosis of Bone and Joint Tuberculosis

    Institute of Scientific and Technical Information of China (English)

    Guirong Wang[1; Weijie Dong[2; Liping Zhao[1; Xia Yu[1; Suting Chen[1; Yuhong Fu[1; Shibing Qin[1; Hairong Huang[1

    2015-01-01

    Objective: The aim of this study was to evaluate L-J (Lo"wenstein-Jensen) medium culture, MGIT 960 culture anddifferent specimen types in diagnosis of BJTB (bone and joint tuberculosis). Methods:: Specimens of pus, caseous necrosis,tuberculous granuloma and sequestrum were collected from 52 BJTB patients. All specimens were cultured using both MGIT 960system and L-J medium; and all pus were amplified using real-time PCR to detect the presence of M. tuberculosis DNA. KeyFindings: A total of 191 specimens were collected. Granuloma had better chance to produce positive outcomes by L-J mediumculture, while for sequestrum MGIT 960 culture had higher yield, but there was no significant difference in the recovery rates amongdifferent types of specimen either by L-J culture (Z2 = 0.638, P = 0.888) or by MGIT960 culture (Z2 = 1.399, P = 0.706). MGIT960culture had significantly higher recovery rate than L-J culture, With a combined culture and PCR-based test, the recovery rate of pusspecimen was significantly higher than that of either method alone (P 〈 0.05). Conclusion: MGIT 960 culture is superior to L-Jculture in BJTB diagnosis; pus, sequestrum, granuloma and caseous necrosis are usable specimen for mycobacterial culture;combination of culture and molecular techniques can provide a better diagnostic significance.

  15. Enhanced cell attachment using a novel cell culture surface presenting functional domains from extracellular matrix proteins

    OpenAIRE

    Cooke, M. J.; Phillips, S R; Shah, D. S. H.; Athey, D.; Lakey, J H; Przyborski, S A

    2008-01-01

    Many factors contribute to the creation and maintenance of a realistic environment for cell growth in vitro, e.g. the consistency of the growth medium, the addition of supplements, and the surface on which the cells grow. The nature of the surface on which cells are cultured plays an important role in their ability to attach, proliferate, migrate and function. Components of the extracellular matrix (ECM) are often used to coat glass or plastic surfaces to enhance cell attachment in vitro. Fra...

  16. 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.

  17. Culture and transfection of axolotl cells.

    Science.gov (United States)

    Denis, Jean-François; Sader, Fadi; Ferretti, Patrizia; Roy, Stéphane

    2015-01-01

    The use of cells grown in vitro has been instrumental for multiple aspects of biomedical research and especially molecular and cellular biology. The ability to grow cells from multicellular organisms like humans, squids, or salamanders is important to simplify the analyses and experimental designs to help understand the biology of these organisms. The advent of the first cell culture has allowed scientists to tease apart the cellular functions, and in many situations these experiments help understand what is happening in the whole organism. In this chapter, we describe techniques for the culture and genetic manipulation of an established cell line from axolotl, a species widely used for studying epimorphic regeneration.

  18. Synergistic effect of cyclodextrins and methyl jasmonate on taxane production in Taxus x media cell cultures.

    Science.gov (United States)

    Sabater-Jara, Ana-Belén; Onrubia, Miriam; Moyano, Elisabeth; Bonfill, Mercedes; Palazón, Javier; Pedreño, María A; Cusidó, Rosa M

    2014-10-01

    Methyl jasmonate and cyclodextrins are proven effective inducers of secondary metabolism in plant cell cultures. Cyclodextrins, which are cyclic oligosaccharides, can form inclusion complexes with nonhydrophilic secondary products, thus increasing their excretion from the producer cells to the culture medium. In the present work, using a selected Taxus x media cell line cultured in a two-stage system, the relationship between taxane production and the transcript profiles of several genes involved in taxol metabolism was studied to gain more insight into the mechanism by which these two elicitors regulate the biosynthesis and excretion of taxol and related taxanes. Gene expression was not clearly enhanced by the presence of cyclodextrins in the culture medium and variably induced by methyl jasmonate, but when the culture was supplemented with both elicitors, a synergistic effect on transcript accumulation was observed. The BAPT and DBTNBT genes, which encode the last two transferases involved in the taxol pathway, appeared to control limiting biosynthetic steps. In the cell cultures treated with both elicitors, the produced taxanes were found mainly in the culture medium, which limited retroinhibition processes and taxane toxicity for the producer cells. The expression level of a putative ABC gene was found to have increased, suggesting it played a role in the taxane excretion. Taxol biosynthesis was clearly increased by the joint action of methyl jasmonate and cyclodextrins, reaching production levels 55 times higher than in nonelicited cultures.

  19. Mechanism of initial attachment of cells derived from human bone to commonly used prosthetic materials during cell culture.

    Science.gov (United States)

    Howlett, C R; Evans, M D; Walsh, W R; Johnson, G; Steele, J G

    1994-02-01

    The suitability of polymeric biomaterials as surfaces for the attachment and growth of cells has often been investigated in cell culture. In this study the contribution that serum fibronectin (Fn) or vitronectin (Vn) make to the attachment and spreading of cells cultured from explanted human bone (bone-derived cells) during the first 90 min of culture was determined for metallic and ceramic surfaces. The requirement for Fn or Vn for attachment and spreading of bone-derived cells onto stainless steel 316 (SS), titanium (Ti) and alumina (Al2O3) and to polyethyleneterephthalate (PET) was directly tested by selective removal of Fn or Vn from the serum prior to addition to the culture medium. Attachment and spreading of bone-derived cells onto SS, Ti and Al2O3 surfaces were reduced by 73-83% when the cells were seeded in medium containing serum from which the Vn had been removed. Cell attachment and spreading on these surfaces when seeded in medium containing Fn-depleted serum (which contained Vn) were not reduced to the same extent as in the medium containing Vn-depleted serum. The bone-derived cells failed to attach to the surfaces to the same extent when seeded in medium containing serum depleted of both Vn and Fn. Our results show that for human bone-derived cells, the attachment and spreading of cells onto SS, Ti and Al2O3 as well as PET during the first 90 min of a cell culture attachment assay are a function of adsorption of serum Vn onto the surface. PMID:7515290

  20. Metabolome analysis of Saccharomyces cerevisiae and optimization of culture medium for S-adenosyl-L-methionine production.

    Science.gov (United States)

    Hayakawa, Kenshi; Matsuda, Fumio; Shimizu, Hiroshi

    2016-12-01

    S-Adenosyl-L-methionine (SAM) is a fine chemical used as a nutritional supplement and a prescription drug. It is industrially produced using Saccharomyces cerevisiae owing to its high SAM content. To investigate the optimization of culture medium components for higher SAM production, metabolome analysis was conducted to compare the intracellular metabolite concentrations between Kyokai no. 6 (high SAM-producing) and laboratory yeast S288C (control) under different SAM production conditions. Metabolome analysis and the result of principal component analysis showed that the rate-limiting step for SAM production was ATP supply and the levels of degradation products of adenosine nucleotides were higher in Kyokai 6 strain than in the S288C strain under the L-methionine supplemented condition. Analysis of ATP accumulation showed that the levels of intracellular ATP in the Kyokai 6 strain were also higher compared to those in the S288C strain. Furthermore, as expected from metabolome analysis, the SAM content of Kyokai 6 strain cultivated in the medium without yeast extract increased by 2.5-fold compared to that in the additional condition, by increasing intracellular ATP level with inhibited cell growth. These results suggest that high SAM production is attributed to the enhanced ATP supply with L-methionine condition and high efficiency of intracellular ATP consumption. PMID:27277079

  1. Metabolome analysis of Saccharomyces cerevisiae and optimization of culture medium for S-adenosyl-L-methionine production.

    Science.gov (United States)

    Hayakawa, Kenshi; Matsuda, Fumio; Shimizu, Hiroshi

    2016-12-01

    S-Adenosyl-L-methionine (SAM) is a fine chemical used as a nutritional supplement and a prescription drug. It is industrially produced using Saccharomyces cerevisiae owing to its high SAM content. To investigate the optimization of culture medium components for higher SAM production, metabolome analysis was conducted to compare the intracellular metabolite concentrations between Kyokai no. 6 (high SAM-producing) and laboratory yeast S288C (control) under different SAM production conditions. Metabolome analysis and the result of principal component analysis showed that the rate-limiting step for SAM production was ATP supply and the levels of degradation products of adenosine nucleotides were higher in Kyokai 6 strain than in the S288C strain under the L-methionine supplemented condition. Analysis of ATP accumulation showed that the levels of intracellular ATP in the Kyokai 6 strain were also higher compared to those in the S288C strain. Furthermore, as expected from metabolome analysis, the SAM content of Kyokai 6 strain cultivated in the medium without yeast extract increased by 2.5-fold compared to that in the additional condition, by increasing intracellular ATP level with inhibited cell growth. These results suggest that high SAM production is attributed to the enhanced ATP supply with L-methionine condition and high efficiency of intracellular ATP consumption.

  2. 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.

  3. 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

  4. Oxidative Stress Induces Senescence in Cultured RPE Cells.

    Science.gov (United States)

    Aryan, Nona; Betts-Obregon, Brandi S; Perry, George; Tsin, Andrew T

    2016-01-01

    The aim of this research is to determine whether oxidative stress induces cellular senescence in human retinal pigment epithelial cells. Cultured ARPE19 cells were subjected to different concentrations of hydrogen peroxide to induce oxidative stress. Cells were seeded into 24-well plates with hydrogen peroxide added to cell medium and incubated at 37°C + 5% CO2 for a 90-minute period [at 0, 300, 400 and 800 micromolar (MCM) hydrogen peroxide]. The number of viable ARPE19 cells were recorded using the Trypan Blue Dye Exclusion Method and cell senescence was measured by positive staining for senescence-associated beta-galactosidase (SA-beta-Gal) protein. Without hydrogen peroxide treatment, the number of viable ARPE19 cells increased significantly from 50,000 cells/well to 197,000 within 72 hours. Treatment with hydrogen peroxide reduced this level of cell proliferation significantly (to 52,167 cells at 400 MCM; to 49,263 cells at 800 MCM). Meanwhile, cells with a high level of positive senescence-indicator SA-Beta-Gal-positive staining was induced by hydrogen peroxide treatment (from a baseline level of 12% to 80% at 400 MCM and at 800 MCM). Our data suggests that oxidative stress from hydrogen peroxide treatment inhibited ARPE19 cell proliferation and induced cellular senescence. PMID:27651846

  5. Endothelial Cells from Embryonic Stem Cells in a Chemically Defined Medium

    OpenAIRE

    Blancas, Alicia A.; Albert J. Shih; Lauer, Nicholas E.; McCloskey, Kara E.

    2011-01-01

    Endothelial cells (ECs) are desired for their therapeutic potential in a variety of areas including gene therapy, cardiac regeneration, development of tissue-engineered vascular grafts, and prevascularized tissue transplants. Pluripotent embryonic stem cells (ESCs) can be induced to differentiate into ECs in vitro using embryoid bodies, monolayer cultures, or by genetic manipulation and immortalization. However, obtaining homogeneous cultures of proliferating ESC-derived ECs without genetic m...

  6. Use of a stationary bed reactor and serum-free medium for the production of recombinant proteins in insect cells.

    Science.gov (United States)

    Kompier, R; Kislev, N; Segal, I; Kadouri, A

    1991-10-01

    Insect cells (Spodoptera frugiperda) have been cultured in a stationary bed reactor, packed with a fibrous polyester carrier. When the bioreactor was perfused with serum-supplemented medium, a cell density of 6 x 10(6) cells ml-1 packed carrier was reached. Scanning electron microscopy investigations have shown that the insect cells grew along the three-dimensionally oriented fibers of the Fibra-cel carrier. After infection of the logarithmically growing cells with a recombinant baculovirus (Autographa californica) containing the gene coding for beta-galactosidase, the medium in the bioreactor was changed to serum-free medium. At day 13 postinfection (p.i.), a beta-galactosidase level of 320 microgram ml-1 and, at day 17 p.i., a virus titer of 2.1 x 10(8) TCID50 units ml-1 (day 17 p.i.) were reached. In another bioreactor, operated in a similar way but with serum-containing medium, a beta-galactosidase concentration of 360 microgram ml-1 and a virus titer of 2.3 x 10(8) TCID50 units ml-1 were obtained. These results indicate the potential use of this production system for the production of recombinant protein and baculovirus in insect cells. PMID:1367637

  7. [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

  8. Optimization of the Liquid Culture Medium Composition to Obtain the Mycelium of Agaricus bisporus Rich in Essential Minerals.

    Science.gov (United States)

    Krakowska, Agata; Reczyński, Witold; Muszyńska, Bożena

    2016-09-01

    Agaricus bisporus species (J.E. Lange) Imbach one of the most popular Basidiomycota species was chosen for the research because of its dietary and medicinal value. The presented herein studies included determination of essential mineral accumulation level in the mycelium of A. bisporus, cultivated on liquid cultures in the medium supplemented with addition of the chosen metals' salts. Quantitative analyses of Zn, Cu, Mg, and Fe in liquid cultures made it possible to determine the relationship between accumulation of the selected mineral in A. bisporus mycelium and the culture conditions. Monitoring of the liquid cultures and determination of the elements' concentrations in mycelium of A. bisporus were performed using the flame technique of AAS method. Concentration of Zn in the mycelium, maintained in the medium with the addition of its salt, was in a very wide range from 95.9 to 4462.0 mg/g DW. In the analyzed A. bisporus mycelium, cultured in the medium enriched with copper salt, this metal concentration changed from 89.79 to 7491.50 mg/g DW; considering Mg in liquid cultured mycelium (medium with Mg addition), its concentration has changed from 0.32 to 10.55 mg/g DW. The medium enriched with iron salts has led to bioaccumulation of Fe in mycelia of A. bisporus. Determined Fe concentration was in the range from 0.62 to 161.28 mg/g DW. The proposed method of liquid A. bisporus culturing on medium enriched with the selected macro- and microelements in proper concentrations ratio have led to obtaining maximal growth of biomass, characterized by high efficiency of the mineral accumulation. As a result, a dietary component of increased nutritive value was obtained. PMID:26857993

  9. Induction of growth in kidney epithelial cells in culture by Na+.

    OpenAIRE

    Toback, F G

    1980-01-01

    The role of Na+ in the regulation of cell growth was examined in quiescent, high-density cultures of kidney epithelial cells of the BSC-1 line. The addition of NaCl to the medium increased the number of cells initiating DNA synthesis in a concentration-dependent manner after serum stimulation. In the presence of added NaCl, cells in confluent cultures grew to high density at an increased rate, whereas growth in sparse cultures was retarded. These results suggest that, in the presence of serum...

  10. Screening and optimization of low-cost medium for Pseudomonas putida Rs-198 culture using RSM

    Directory of Open Access Journals (Sweden)

    Yanjie Peng

    2014-12-01

    Full Text Available The plant growth-promoting rhizobacterial strain Pseudomonas putida Rs-198 was isolated from salinized soils from Xinjiang Province. We optimized the composition of the low-cost medium of P. putida Rs-198 based on its bacterial concentration, as well as its phosphate-dissolving and indole acetic acid (IAA-producing capabilities using the response surface methodology (RSM, and a mathematical model was developed to show the effect of each medium component and its interactions on phosphate dissolution and IAA production. The model predicted a maximum phosphate concentration in medium containing 63.23 mg/L inorganic phosphate with 49.22 g/L corn flour, 14.63 g/L soybean meal, 2.03 g/L K2HPO4, 0.19 g/L MnSO4 and 5.00 g/L NaCl. The maximum IAA concentration (18.73 mg/L was predicted in medium containing 52.41 g/L corn flour, 15.82 g/L soybean meal, 2.40 g/L K2HPO4, 0.17 g/L MnSO4 and 5.00 g/L NaCl. These predicted values were also verified through experiments, with a cell density of 10(13 cfu/mL, phosphate dissolution of 64.33 mg/L, and IAA concentration of 18.08 mg/L. The excellent correlation between predicted and measured values of each model justifies the validity of both the response models. The study aims to provide a basis for industrialized fermentation using P. putida Rs-198.

  11. Influence of co-culture on osteogenesis and angiogenesis of bone marrow mesenchymal stem cells and aortic endothelial cells.

    Science.gov (United States)

    Gurel Pekozer, Gorke; Torun Kose, Gamze; Hasirci, Vasif

    2016-11-01

    Co-culture of bone forming cells and endothelial cells to induce pre-vascularization is one of the strategies used to solve the insufficient vascularization problem in bone tissue engineering attempts. In the study, primary cells isolated from 2 different tissues of the same animal, rat bone marrow stem cells (RBMSCs) and rat aortic endothelial cells (RAECs) were co-cultured to study the effects of co-culturing on both osteogenesis and angiogenesis. The formation of tube like structure in 2D culture was observed for the first time in the literature by the co-culture of primary cells from the same animal and also osteogenesis and angiogenesis were investigated at the same time by using this co-culture system. Co-cultured cells mineralized and formed microvasculature beginning from 14days of incubation. After 28days of incubation in the osteogenic medium, expression of osteogenic genes in co-cultures was significantly upregulated compared to RBMSCs cultured alone. These results suggest that the co-culture of endothelial cells with mesenchymal stem cells induces both osteogenesis and angiogenesis.

  12. Sponge cell culture? A molecular identification method for sponge cells

    NARCIS (Netherlands)

    Sipkema, D.; Heilig, G.H.J.; Akkermans, A.D.L.; Osinga, R.; Tramper, J.; Wijffels, R.H.

    2003-01-01

    Dissociated sponge cells are easily confused with unicellular organisms. This has been an obstacle in the development of sponge-cell lines. We developed a molecular detection method to identify cells of the sponge Dysidea avara in dissociated cell cultures. The 18S ribosomal RNA gene from a Dysidea

  13. 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.

  14. PREPARATION AND THE CULTURE OF LO2 CELLS ON PVA-BASED MICROCARRIERS

    Institute of Scientific and Technical Information of China (English)

    ZHANG Hong; YU Yaoting; PAN Jilun; WANG lianyong

    2001-01-01

    Using polyvinyl alcohol (PVA) as raw material and vacuum pump oil as oil phase medium,PVA-based microcarriers were prepared by suspension method The diameters of the beads were 100-180 μ m. LO2 cells were cultured on PVA-based microcarriers and cytodexⅢ microcarriers.Morphology, attachment and growth rate of LO2 cells were studied.

  15. PREPARATION AND THE CULTURE OF LO2 CELLS ON PVA—BASED MICROCARRIERS

    Institute of Scientific and Technical Information of China (English)

    ZHANGHong; YUYaoting; 等

    2001-01-01

    Using polyvinyl alcohol(PVA) as raw material and vacuum pump oil as oil phase medium,PVA-based microcarriers were prepared by suspension method,The diameters of the beads were 100-180um,LO2 cells were cultured on PVA-based microcarriers and cytodexⅢ microcarriers.Morphology,attachment and growth rate of LO2 cells were studied.

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

    DEFF Research Database (Denmark)

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

    2011-01-01

    of active membrane bound rFVIII to the culture medium. Moreover, the attachment of rFVIII to cell membranes of un-transfected HEK293 cells was studied in the presence of compounds that competes for interactions between rFVIII and PS. Competitive assays between iodinated rFVIII (¹²5I-rFVIII) and annexin V...

  17. Keeping the intracellular vitamin C at a physiologically relevant level in endothelial cell culture

    DEFF Research Database (Denmark)

    Frikke-Schmidt, Henriette Rønne; Lykkesfeldt, Jens

    2010-01-01

    It is generally accepted that the addition of vitamin C to cell culture medium improves cell growth. However, once added, the vitamin C concentration declines rapidly. This situation differs from the in vivo environment where the endothelium is constantly supplied with ascorbate from the blood...

  18. Development and application of a high-throughput platform for perfusion-based cell culture processes.

    Science.gov (United States)

    Villiger-Oberbek, Agata; Yang, Yang; Zhou, Weichang; Yang, Jianguo

    2015-10-20

    A high-throughput (HT) cell culture model has been established for the support of perfusion-based cell culture processes operating at high cell densities. To mimic perfusion, the developed platform takes advantage of shake tubes and operates them in a batch-refeed mode with daily medium exchange to supply the cultures with nutrients and remove toxic byproducts. By adjusting the shaking parameters, such as the speed and setting angle, we have adapted the shake tubes to a semi-continuous production of a recombinant enzyme in a perfusion-like mode. We have demonstrated that the developed model can be used to select clones and cell culture media ahead of process optimization studies in bioreactors and confirmed the applicability of shake tubes to a perfusion-like cell culture reaching ∼50E6 viable cells/mL. Furthermore, through regular cell mass removal and periodic medium exchange we have successfully maintained satellite cultures of bench-top perfusion bioreactors, achieving a sustainable cell culture performance at ≥30E6 viable cells/mL and viabilities >80% for over 58 days. The established HT model is a unique and powerful tool that can be used for the development and screening of media formulations, or for testing selected process parameters during both process optimization and manufacturing support campaigns.

  19. Development and application of a high-throughput platform for perfusion-based cell culture processes.

    Science.gov (United States)

    Villiger-Oberbek, Agata; Yang, Yang; Zhou, Weichang; Yang, Jianguo

    2015-10-20

    A high-throughput (HT) cell culture model has been established for the support of perfusion-based cell culture processes operating at high cell densities. To mimic perfusion, the developed platform takes advantage of shake tubes and operates them in a batch-refeed mode with daily medium exchange to supply the cultures with nutrients and remove toxic byproducts. By adjusting the shaking parameters, such as the speed and setting angle, we have adapted the shake tubes to a semi-continuous production of a recombinant enzyme in a perfusion-like mode. We have demonstrated that the developed model can be used to select clones and cell culture media ahead of process optimization studies in bioreactors and confirmed the applicability of shake tubes to a perfusion-like cell culture reaching ∼50E6 viable cells/mL. Furthermore, through regular cell mass removal and periodic medium exchange we have successfully maintained satellite cultures of bench-top perfusion bioreactors, achieving a sustainable cell culture performance at ≥30E6 viable cells/mL and viabilities >80% for over 58 days. The established HT model is a unique and powerful tool that can be used for the development and screening of media formulations, or for testing selected process parameters during both process optimization and manufacturing support campaigns. PMID:26197419

  20. Histology of embryoid development in oil palm (Elaeis guineensis Jacq. cell suspension culture

    Directory of Open Access Journals (Sweden)

    Songrat Tinnongjig

    2001-11-01

    Full Text Available Embryos of oil palm (Elaeis guineensis Jacq. variety tenera were cultured on Eeuwens or Y3 (1976; 1978 medium supplemented with 2 mg/l 2,4-D. Calluses were initiated from these embryos. The eight-weekold calluses derived from embryos were transferred to modified Y3 liquid medium devoid of 2,4-D and supplemented with NAA, BA and coconut water to establish cell suspension culture. After a period of culture,these cells were then subcultured to the same medium without plant growth regulators to induce embryoid formation. The calluses and embryoids were harvested at various times, fixed, sectioned, stained and examined microscopically. Histological study revealed that embryoid occurred from meristematic cells with dense cytoplasm along the callus clumps.

  1. 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

  2. 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)

  3. Maturation and fertilisation of sheep oocytes cultured in serum-free medium containing silk protein sericin.

    Science.gov (United States)

    Yasmin, Cut; Otoi, Takeshige; Setiadi, Mohamad Agus; Karja, Ni Wayan Kurniani

    2015-03-01

    Sericin is a water-soluble component of silk and has been used as a biomaterial due to its antibacterial and ultraviolet radiation-resistant properties. This study was designed to evaluate the effect of sericin supplementation in a maturation medium on the meiotic competence and fertilisability of sheep oocytes. Cumulus-oocyte complexes (COCs) were cultured in TCM199 supplemented with sericin at various concentrations of 0 (control), 0.1, 0.25 and 0.5%, either with or without bovine serum albumin (BSA). When the COCs were matured without BSA, the supplementation of 0.1% sericin significantly increased the rates of maturation to metaphase II and the total fertilisation of oocytes compared with the other concentrations of sericin. When the COCs were matured with BSA, the beneficial effects of 0.1% sericin supplementation on the maturation and fertilisation of oocytes were not observed. Our findings indicate that supplementation with 0.1% sericin during maturation culture may improve the nuclear maturation and fertilisability of sheep oocytes. Moreover, it may be possible to replace BSA with sericin in chemically defined media without the risk of disease transmission. PMID:25655418

  4. Study on recycling of waste rubbers as medium components for hydroponic culture of rose

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jin-Kuk; Lee, Hyung-Gyu; Jeong, Byoung-Ryong; Hwang, Seung-Jae [Gyeongsang National Univ., Kumi(Korea)

    2000-06-30

    Recently, the efficient disposal of the waste rubber is necessary due to increasing amount of the waste rubbers. In this paper, method of recycling waste rubbers as components of medium for hydroponic rose culture was suggested. We investigated growth of rose, and macro- and micro-elements, pH and EC of the media amended with waste rubber. In the beginning of culture, stress symptoms such as thin brittle stem and incipient wilting were observed, but they disappeared in a few weeks. Concentration of Zn{sup 2+} in media at flowering increased in proportion to contents of waste tire in the media. pH of media at flowering were in the range of 5.70 to 6.35. Rose growth in all media, except in waste rock wool mixed with EPDM powder at 9:3 ratio, was normal and equivalent to the control in terms of stem length, number of stems harvested and fresh weight. (author). 10 refs., 5 tabs., 4 figs.

  5. Effects of taurine depletion on cell migration and NCAM expression in cultures of dissociated mouse cerebellum and N2A cells

    DEFF Research Database (Denmark)

    Maar, T E; Lund, Trine Meldgaard; Gegelashvili, G;

    1998-01-01

    Cultures of dissociated cerebellum from 5- to 6-day-old mice as well as of the N2A neuronal cell line were exposed to guanidino ethane sulfonate (GES, 2-5 mM) to reduce the cellular taurine content. Control cultures were kept in culture medium or medium containing 2-5 mM GES plus 2-5 mM taurine...

  6. In vitro plant regeneration of two cucumber (Cucumis sativum L. genotypes: Effects of explant types and culture medium

    Directory of Open Access Journals (Sweden)

    Grozeva Stanislava

    2014-01-01

    Full Text Available The effect of different phytohormone concentrations on callusogenesis and organogenesis in two cucumber genotypes were studied. It was established that the rate of plant regeneration depends on genotype, explant type and culture medium. Hypocotyls were found to be more responsive than cotyledons in morphogenesis. In vitro planlet-regenerants have been obtained in hypocotyls explants on culture medium with 1.0 and 2.0 mgL-1 BA for cultivar Gergana and in 1.0 and 3.0 mgL-1K-line 15B. Induction of regeneration in cotyledons were established only in cultivar Gergana on culture medium supplemented with 3.0 mgL-1 BA and in combination of 0.5 mgL-1IAA.

  7. Flux analysis of mammalian cell culture

    NARCIS (Netherlands)

    Martens, D.E.; Tramper, J.

    2010-01-01

    Animal cells are used for the production of vaccines and pharmaceutical proteins. The increase in demand for these products requires an increase in volumetric productivity of animal cell culture processes, which can be attained through an increase in biomass concentration and/or specific productivit

  8. 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)

  9. Immunologically active polysaccharides of Arnica montana cell cultures.

    Science.gov (United States)

    Puhlmann, J; Zenk, M H; Wagner, H

    1991-01-01

    From the nutrition medium of Arnica montana cell cultures two homogeneous polysaccharides, an acidic arabino-3,6-galactan-protein with mean Mr of 100,000 and a neutral fucogalactoxyloglucan with mean Mr of 22,500 have been isolated by DEAE-Sepharose CL-6B and Sephacryl S-400 column chromatography. Their structures were elucidated mainly by methylation analysis, partial acidic and enzymatic hydrolysis and 13C NMR spectroscopy. The fucogalactoxyloglucan shows a pronounced enhancement of phagocytosis in vivo. The arabino-3,6-galactan-protein displays a strong anticomplementary effect and stimulates macrophages to excrete the tumour necrosis factor (TNF alpha).

  10. [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

  11. 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.

  12. 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

    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.

  13. Differentiation of P19 embryonal carcinoma stem cells into insulin-producing cells promoted by pancreas-conditioned medium.

    Science.gov (United States)

    Mansouri, Akram; Esmaeili, Fariba; Nejatpour, Azadeh; Houshmand, Fariba; Shabani, Leila; Ebrahimie, Esmaeil

    2016-07-01

    The ability of embryonal carcinoma )EC (stem cells to generate insulin-producing cells (IPCs) is still unknown. We examined the trophic effects of pancreas-conditioned medium (PCM) on in vitro production of IPCs. Initially, P19 EC cells were characterized by the expression of stem cell markers, Oct3/4, Sox-2 and Nanog. To direct differentiation, P19-derived embryoid bodies (EBs) were induced by selection of nestin-positive cells and treatment with different concentrations of PCM. Morphological studies documented the presence of islet-like cell IPCs clusters. The differentiated cells were immunoreactive for β cell-specific proteins, including insulin, proinsulin, C-peptide and insulin receptor-β. The expression of genes related to pancreatic β cell development and function (PDX-1, INS1, INS2, EP300 and CREB1) was confirmed by qPCR. During differentiation, the expression of EP300 and CREB1 increased by 2.5 and 3.1 times, respectively. In contrast, a sharp decrease in the expression of Oct3/4, Sox-2 and Nanog by 4, 1.5 and 1.5 times, respectively, was observed. The differentiated cells were functionally active, synthesizing and secreting insulin in a glucose-regulated manner. Network prediction highlighted crosstalk between PDX-1 transcription factor and INS2 ligand in IPC generation and revealed positive regulatory effects of EP300, CREB1, PPARA, EGR, KIT, GLP1R, and PKT2 on activation of PDX-1 and INS2. This is the first report of the induction of IPC differentiation from EC cells by using neonate mouse PCM. Since P19 EC cells are widely available, easily cultured without feeders and do not require special growth conditions, they would provide a valuable tool for studying pancreatic β cell differentiation and development. Copyright © 2016 John Wiley & Sons, Ltd. PMID:25044225

  14. The Role of Glucose, Serum, and Three-Dimensional Cell Culture on the Metabolism of Bone Marrow-Derived Mesenchymal Stem Cells

    OpenAIRE

    Byron Deorosan; Nauman, Eric A.

    2011-01-01

    Mesenchymal stem cells (MSCs) have become a critical addition to all facets of tissue engineering. While most in vitro research has focused on their behavior in two-dimensional culture, relatively little is known about the cells' behavior in three-dimensional culture, especially with regard to their metabolic state. To evaluate MSC metabolism during twodimensional culture, murine bone marrow-derived MSCs were cultured for one week using twelve different medium compositions, varying in both gl...

  15. Transcription Factors and Medium Suitable for Initiating the Differentiation of Human-Induced Pluripotent Stem Cells to the Hepatocyte Lineage.

    Science.gov (United States)

    Tomizawa, Minoru; Shinozaki, Fuminobu; Motoyoshi, Yasufumi; Sugiyama, Takao; Yamamoto, Shigenori; Ishige, Naoki

    2016-09-01

    Transcription factors and culture media were investigated to determine the condition to initiate the differentiation of human-induced pluripotent stem (iPS) cells most efficiently. The expression of genes in human adult liver was compared with that in 201B7 cells (iPS cells) using cDNA microarray analysis. Episomal plasmids expressing transcription factors were constructed. 201B7 cells were transfected with the episomal plasmids and cultured in ReproFF (feeder-free media maintaining pluripotency), Leibovitz-15 (L15), William's E (WE), or Dulbecco's modified Eagle medium/Nutrient F-12 Ham (DF12) for 7 days. RNA was isolated and subjected to real-time quantitative PCR to analyze the expression of alpha-feto protein (AFP) and albumin. cDNA microarray analysis revealed 16 transcription factors that were upregulated in human adult liver relative to that in 201B7 cells. Episomal plasmids expressing these 16 genes were transfected into 201B7 cells. CCAAT/enhancer-binding protein alpha (CEBPA), CCAAT/enhancer-binding protein beta (CEBPB), forkhead box A1 (FOXA1), and forkhead box A3 (FOXA3) up-regulated AFP and down-regulated Nanog. These four genes were further analyzed. The expression of AFP and albumin was the highest in 201B7 cells transfected with the combination of CEBPA, CEBPB, FOXA1, and FOXA3 and cultured in WE. The combination of CEBPA, CEBPB, FOXA1, and FOXA3 was suitable for 201B7 cells to initiate differentiation to the hepatocyte lineage and WE was the most suitable medium for culture after transfection. J. Cell. Biochem. 117: 2001-2009, 2016. © 2016 Wiley Periodicals, Inc. PMID:26773721

  16. The effects of direct irradiation and bystander medium on EPC and CHSE cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Olwell, P.; Mothersill, C.; Seymour, C.; Cottell, D.C.; Lyng, F.M. [Dublin Institute of Technology, Radiation and Environmental Science Centre, Dublin (Ireland)

    2004-07-01

    The majority of studies involving the effects of direct radiation on cell lines use mammalian cells and the effects of bystander medium have all exclusively dealt with mammalian cells. There is increasing evidence that the effects of radiation differ in severity between different species. Two fish cell lines were irradiated in order to establish the radiosensitivity of fish cells. These cell lines, fibroblast-like CHSE 214 and epithelial-like EPC, were irradiated and compared to non-irradiated controls using three investigative parameters; lactate dehydrogenase (LDH) release, surface morphology and reproductive integrity. The same cell lines were also incubated with medium from irradiated cells (bystander medium) and compared to controls using the same methods as were used for directly irradiated cell lines. LDH is released when the plasma membrane of a cell is ruptured indicating non-lethal damage. Both cell lines were shown to exhibit less LDH release following direct radiation and exposure to bystander medium than mammalian cell lines of the same cell type. The surface of CHSE 214 cells showed an increase in surface features following direct radiation and exposure to bystander medium. The surface of EPC cells showed no significant surface differences following irradiation. Clonogenic studies of both cell lines, which detect effects in the cloning ability of cells, showed results similar to those seen in mammalian studies. The results are discussed and compared to studies using mammalian cells. (author)

  17. Production of inactivated influenza H5N1 vaccines from MDCK cells in serum-free medium.

    Directory of Open Access Journals (Sweden)

    Alan Yung-Chih Hu

    Full Text Available BACKGROUND: Highly pathogenic influenza viruses pose a constant threat which could lead to a global pandemic. Vaccination remains the principal measure to reduce morbidity and mortality from such pandemics. The availability and surging demand for pandemic vaccines needs to be addressed in the preparedness plans. This study presents an improved high-yield manufacturing process for the inactivated influenza H5N1 vaccines using Madin-Darby canine kidney (MDCK cells grown in a serum-free (SF medium microcarrier cell culture system. PRINCIPAL FINDING: The current study has evaluated the performance of cell adaptation switched from serum-containing (SC medium to several commercial SF media. The selected SF medium was further evaluated in various bioreactor culture systems for process scale-up evaluation. No significant difference was found in the cell growth in different sizes of bioreactors studied. In the 7.5 L bioreactor runs, the cell concentration reached to 2.3 × 10(6 cells/mL after 5 days. The maximum virus titers of 1024 Hemagglutinin (HA units/50 µL and 7.1 ± 0.3 × 10(8 pfu/mL were obtained after 3 days infection. The concentration of HA antigen as determined by SRID was found to be 14.1 µg/mL which was higher than those obtained from the SC medium. A mouse immunogenicity study showed that the formalin-inactivated purified SF vaccine candidate formulated with alum adjuvant could induce protective level of virus neutralization titers similar to those obtained from the SC medium. In addition, the H5N1 viruses produced from either SC or SF media showed the same antigenic reactivity with the NIBRG14 standard antisera. CONCLUSIONS: The advantages of this SF cell-based manufacturing process could reduce the animal serum contamination, the cost and lot-to-lot variation of SC medium production. This study provides useful information to manufacturers that are planning to use SF medium for cell-based influenza vaccine production.

  18. Human Umbilical Cord Blood-Derived Serum for Culturing the Supportive Feeder Cells of Human Pluripotent Stem Cell Lines

    Directory of Open Access Journals (Sweden)

    Ruttachuk Rungsiwiwut

    2016-01-01

    Full Text Available Although human pluripotent stem cells (hPSCs can proliferate robustly on the feeder-free culture system, genetic instability of hPSCs has been reported in such environment. Alternatively, feeder cells enable hPSCs to maintain their pluripotency. The feeder cells are usually grown in a culture medium containing fetal bovine serum (FBS prior to coculture with hPSCs. The use of FBS might limit the clinical application of hPSCs. Recently, human cord blood-derived serum (hUCS showed a positive effect on culture of mesenchymal stem cells. It is interesting to test whether hUCS can be used for culture of feeder cells of hPSCs. This study was aimed to replace FBS with hUCS for culturing the human foreskin fibroblasts (HFFs prior to feeder cell preparation. The results showed that HFFs cultured in hUCS-containing medium (HFF-hUCS displayed fibroblastic features, high proliferation rates, short population doubling times, and normal karyotypes after prolonged culture. Inactivated HFF-hUCS expressed important genes, including Activin A, FGF2, and TGFβ1, which have been implicated in the maintenance of hPSC pluripotency. Moreover, hPSC lines maintained pluripotency, differentiation capacities, and karyotypic stability after being cocultured for extended period with inactivated HFF-hUCS. Therefore, the results demonstrated the benefit of hUCS for hPSCs culture system.

  19. Synthesis of calcium oxalate crystals in culture medium irradiated with non-equilibrium atmospheric-pressure plasma

    Science.gov (United States)

    Kurake, Naoyuki; Tanaka, Hiromasa; Ishikawa, Kenji; Nakamura, Kae; Kajiyama, Hiroaki; Kikkawa, Fumitaka; Mizuno, Masaaki; Yamanishi, Yoko; Hori, Masaru

    2016-09-01

    Octahedral particulates several tens of microns in size were synthesized in a culture medium irradiated through contact with a plume of non-equilibrium atmospheric-pressure plasma (NEAPP). The particulates were identified in the crystalline phase as calcium oxalate dihydrate (COD). The original medium contained constituents such as NaCl, d-glucose, CaCl2, and NaHCO3 but not oxalate or oxalic acid. The oxalate was clearly synthesized and crystallized in the medium as thermodynamically unstable COD crystals after the NEAPP irradiation.

  20. Approaches to Optimizing Animal Cell Culture Process: Substrate Metabolism Regulation and Protein Expression Improvement

    Science.gov (United States)

    Zhang, Yuanxing

    Some high value proteins and vaccines for medical and veterinary applications by animal cell culture have an increasing market in China. In order to meet the demands of large-scale productions of proteins and vaccines, animal cell culture technology has been widely developed. In general, an animal cell culture process can be divided into two stages in a batch culture. In cell growth stage a high specific growth rate is expected to achieve a high cell density. In production stage a high specific production rate is stressed for the expression and secretion of qualified protein or replication of virus. It is always critical to maintain high cell viability in fed-batch and perfusion cultures. More concern has been focused on two points by the researchers in China. First, the cell metabolism of substrates is analyzed and the accumulation of toxic by-products is decreased through regulating cell metabolism in the culture process. Second, some important factors effecting protein expression are understood at the molecular level and the production ability of protein is improved. In pace with the rapid development of large-scale cell culture for the production of vaccines, antibodies and other recombinant proteins in China, the medium design and process optimization based on cell metabolism regulation and protein expression improvement will play an important role. The chapter outlines the main advances in metabolic regulation of cell and expression improvement of protein in animal cell culture in recent years.

  1. 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... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products § 864.2280 Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in...

  2. Fatty Acid Profiles for Differentiating Growth Medium Formulations Used to Culture Bacillus cereus T-strain Spores.

    Science.gov (United States)

    Ehrhardt, Christopher J; Murphy, Devonie L; Robertson, James M; Bannan, Jason D

    2015-07-01

    Microbial biomarkers that indicate aspects of an organism's growth conditions are important targets of forensic research. In this study, we examined fatty acid composition as a signature for the types of complex nutrients in the culturing medium. Bacillus cereus T-strain spores were grown in medium formulations supplemented with one of the following: peptone (meat protein), tryptone (casein protein), soy protein, and brain-heart infusion. Cellular biomass was profiled with fatty acid methyl ester (FAME) analysis. Results showed peptone cultures produced spores enriched in straight-chained lipids. Tryptone cultures produced spores enriched in branched-odd lipids when compared with peptone, soy, and brain-heart formulations. The observed FAME variation was used to construct a set of discriminant functions that could help identify the nutrients in a culturing recipe for an unknown spore sample. Blinded classification tests were most successful for spores grown on media containing peptone and tryptone, showing 88% and 100% correct identification, respectively. PMID:25854710

  3. Pinoresinol from Ipomoea cairica cell cultures.

    Science.gov (United States)

    Páska, Csilla; Innocenti, Gabbriella; Ferlin, Mariagrazia; Kunvári, Mónika; László, Miklós

    2002-10-01

    Ipomoea cairica cell cultures produced a tetrahydrofuran lignan, (+)-pinoresinol, identified by UV, IR, MS and NMR methods, not yet found in the intact plant, and new in the Convolvulaceae family. Pinoresinol was found to have antioxidant and Ca2+ antagonist properties. As it could be requested for its biological activity, we examined the possibility to raise the pinoresinol yield of I. cairica cultures, as well as we continued investigations on lignans' response to optimization.

  4. 补肾活血中药血清对高糖状态下纯化培养的视网膜神经节细胞活力的影响%Effect of Serum Contained Chinese Drugs for Nourishing Shen and Activating Blood on Activity of Purified Retinal Ganglion Cells Cultured in High Glucose Medium

    Institute of Scientific and Technical Information of China (English)

    马荣; 谢学军; 万李; 马殿伟

    2009-01-01

    目的 探讨补肾活血中药血清对体外高糖状态下纯化培养的视网膜神经节细胞活力的影响.方法 体外纯化SD大鼠视网膜神经节细胞(retinal ganglion cells,RGCs),模拟稳定高糖(50 mmol/L)及糖波动环境进行培养,以补肾活血中药血清进行干预,检测RGCs乳酸脱氢酶(lactate dehydrogenase,LDH)漏出量,以推测RGCs活力.结果 RGCs的LDH漏出量(U/L)稳定高糖组24 h(1 349.17±215.50)、48 h(1 220.24±124.53)及72 h(1 982.14±219.03)均较正常对照组(1 628.10±122.10、1 484.13±127.55及2 155.75±140.44)降低(P<0.05);而糖波动组的LDH漏出量在72 h(2 299.60±88.35)较正常对照组增加(P<0.05),且糖波动组各时段的LDH漏出量均较稳定高糖组增加(P<0.05);稳定高糖中药干预组的LDH漏出量在72 h(1 797.62±146.40)时较稳定高糖组减少(P<0.05);糖波动中药干预组的LDH漏出量在48 h(1 259.92±87.74)和72 h(1 940.40±155.47)时均较糖波动组减少(P<0.05).结论 糖波动能明显降低RGCs细胞膜的稳定性,增加细胞膜通透性,降低细胞活力;补肾活血中药血清能降低本实验中稳定高糖及糖波动条件下RGCs的细胞膜通透性,提高细胞膜稳定性,增强其细胞活力,这可能是补肾活血中药复方防治糖尿病视网膜病变(diabetic retinopathy,DRP)的药物干预途径之一.%Objective To investigate the effect of serum contained Chinese drugs for nourishing Shen and activating blood(S-NSAB)on activity of purified retinal ganglion cells(RGCs)cultured in high glucose medium.Methods Purified RGCs of SD rats were cultured in simulative stable high glucose(50 mmol/L)condition(SHG)and fluctuated glucose condition(FGC)separately,they were intervened with S-NSAB,and the lactate dehydrogenase(LDH)leakage was detected by spectrophotometer for estimating the activity of RGCs.Results LDH leakage(U/L)in SHG culture was 1 349.17±215.50 at 24 h,1 220.24±124.53 at 48 h and 1 982.14±219.03 at 72 h,all significantly lower

  5. Urban secondary sewage: an alternative medium for the culture of Tetraselmis chuii (Prasinophyceae) and Dunaliella viridis (Chlorophyceae)

    OpenAIRE

    Rauquírio André Albuquerque Marinho da Costa; Maria Luise Koening; Sílvio José de Macedo

    2004-01-01

    The effect of different concentrations (10, 20, 30 and 40%) of urban secondary sewage on the growth of Tetraselmis chuii (Prasinophyceae) and Dunaliella viridis (Chlorophyceae) was examined to verify the possibility of its use as an alternative culture medium for these species. Stocks and 700mL duplicate batch cultures were maintained under controlled laboratory conditions. 30% and 40% additions were the most efficient for the growth of both species. Statistical analysis of the adjusted growt...

  6. Optimization of an effective growth medium for culturing probiotic bacteria for applications in strict vegetarian food products

    OpenAIRE

    Manju Pathak; Danik Martirosyan

    2012-01-01

    Background: This study aimed to modify de Man Rogosa Sharpe culture medium (termed MRS) for selective cultivation of probiotics strain for the consumption by the strictly vegetarian human population. Vegetarian probiotic foods by definition must be free from all animal-derived ingredients. This not only includes the product ingredients but the probiotic inoculum as well. Probiotic starter cultures are traditionally grown and stored in media containing milk or meatderived ingredients. The pre...

  7. In-vitro Knock Down of VEGFR2 Expression Using Specific siRNA in the Culture Medium

    Directory of Open Access Journals (Sweden)

    Moslem Jafarisani

    2015-09-01

    Full Text Available Introduction:The use of siRNAforsilencinggene expressionis expandingtoday. Knock down ofvascular endothelial growth factor is one of theobjectives ofthistechnology. In this study we aimed to inhibit the expression of receptor type II of VEGF (VEGFR-2 using specific siRNA in the culture medium.Methods: First, according to the target gene sequences, sequence of the specific siRNA were designed; blasted and built. Using RT-PCR, cDNA of HUVEC was synthesized and PCR with specific primers was reproduced. PEFGP-N1 expression vector was cloned with target gene and confirmed. Then Hela cells which has not any expression of the target genes were transfected whit cloned pEGFP-N1 vector using lipofectamin. GFP expression rate in the Hela cellsContaining initial vector and cloned vector in presence and absence of specific siRNA was assessed. Through evaluation of gene inhibition, decreasing of green fluorescence from GFP, RT-PCR was investigated.Results: The results of the two used siRNA, indicate reduced gene expression 56% and 63% in comparison with the control group (P<0.05.Conclusion: Transfection of specific VEGFR-2 siRNA using lipofectamin significantly inhibits expression of receptor. In fact,it cancut thesignal transduction pathwayand preventsthe creation ofnewblood vessels. Therefore, probably it can act as a new treatment factor for prevention or reduction of neovascularization.

  8. Transcriptomic comparisons between cultured human adipose tissue-derived pericytes and mesenchymal stromal cells

    Directory of Open Access Journals (Sweden)

    Lindolfo da Silva Meirelles

    2016-03-01

    Full Text Available Mesenchymal stromal cells (MSCs, sometimes called mesenchymal stem cells, are cultured cells able to give rise to mature mesenchymal cells such as adipocytes, osteoblasts, and chondrocytes, and to secrete a wide range of trophic and immunomodulatory molecules. Evidence indicates that pericytes, cells that surround and maintain physical connections with endothelial cells in blood vessels, can give rise to MSCs (da Silva Meirelles et al., 2008 [1]; Caplan and Correa, 2011 [2]. We have compared the transcriptomes of highly purified, human adipose tissue pericytes subjected to culture-expansion in pericyte medium or MSC medium, with that of human adipose tissue MSCs isolated with traditional methods to test the hypothesis that their transcriptomes are similar (da Silva Meirelles et al., 2015 [3]. Here, we provide further information and analyses of microarray data from three pericyte populations cultured in pericyte medium, three pericyte populations cultured in MSC medium, and three adipose tissue MSC populations deposited in the Gene Expression Omnibus under accession number GSE67747.

  9. Rheological characteristics of cell suspension and cell culture of Perilla frutescens.

    Science.gov (United States)

    Zhong, J J; Seki, T; Kinoshita, S; Yoshida, T

    1992-12-01

    Physical properties such as viscosity, fluid dynamic behavior of cell suspension, and size distribution of cell aggregates of a plant, Perilla frustescens, cultured in a liquid medium were studied. As a result of investigations using cells harvester after 12 days of cultivation in a flask, it was found that the apparent viscosity of the cell suspension did not change with any variation of cell concentration below 5 g dry cell/L but markedly increased when the cell concentration increased over 12.8 g dry cell/L. The cell suspension exhibited the characteristics of a Bingham plastic fluid with a small yield stress. The size of cell aggregates in the range 74 to 500 mum did not influence the rheological characteristics of the cell suspension. The rheological characteristics of cultivation mixtures of P. frutescens cultivated in a flask and in a bioreactor were also investigated. The results showed that the flow characteristics of the cell culture could be described by a Bingham plastic model. At the later stage of cultivation, the apparent viscosity increased steadily, even though the biomass concentration (by dry weight) decreased, due to the increase of individual cell size.

  10. Epithelial Cell Culture from Human Adenoids: A Functional Study Model for Ciliated and Secretory Cells

    Directory of Open Access Journals (Sweden)

    Claudia González

    2013-01-01

    Full Text Available Background. Mucociliary transport (MCT is a defense mechanism of the airway. To study the underlying mechanisms of MCT, we have both developed an experimental model of cultures, from human adenoid tissue of ciliated and secretory cells, and characterized the response to local chemical signals that control ciliary activity and the secretion of respiratory mucins in vitro. Materials and Methods. In ciliated cell cultures, ciliary beat frequency (CBF and intracellular Ca2+ levels were measured in response to ATP, UTP, and adenosine. In secretory cultures, mucin synthesis and secretion were identified by using immunodetection. Mucin content was taken from conditioned medium and analyzed in the presence or absence of UTP. Results. Enriched ciliated cell monolayers and secretory cells were obtained. Ciliated cells showed a basal CBF of 10.7 Hz that increased significantly after exposure to ATP, UTP, or adenosine. Mature secretory cells showed active secretion of granules containing different glycoproteins, including MUC5AC. Conclusion. Culture of ciliated and secretory cells grown from adenoid epithelium is a reproducible and feasible experimental model, in which it is possible to observe ciliary and secretory activities, with a potential use as a model to understand mucociliary transport control mechanisms.

  11. General overview of neuronal cell culture.

    Science.gov (United States)

    Gordon, Jennifer; Amini, Shohreh; White, Martyn K

    2013-01-01

    In this introductory chapter, we provide a general overview of neuronal cell culture. This is a rapidly evolving area of research and we provide an outline and contextual framework for the different chapters of this book. These chapters were all contributed by scientists actively working in the field who are currently using state-of-the-art techniques to advance our understanding of the molecular and cellular biology of the central nervous system. Each chapter provides detailed descriptions and experimental protocols for a variety of techniques ranging in scope from basic neuronal cell line culturing to advanced and specialized methods.

  12. Effect of Culture Medium Supplementation with b-mercaptoethanol and Amino Acid on Canine Intergeneric Embryo Development with Porcine Oocyte Cytoplasm Recipient

    Directory of Open Access Journals (Sweden)

    Yuda Heru Fibrianto

    2015-11-01

    Full Text Available The present study investigated the effect of culture medium supplementation with mercaptoethanol ( MEand amino acid (AA on canine intergeneric embryo development with porcine oocyte cytoplasm. Porcine cumulusoocyte complexes (COCs were collected from slaughterhouse and matured in TCM-199 supplemented with 26.2mM NaHCO, 3.05 mM glucose, 0.91 mM sodium pyruvate, 0.57 mM L-cysteine, 75 mg/l kanamycin, 10 ng/ml 3epidermal growth factor, equine chorionic gonadotropin (eCG, 10 IU/ml human chorionic gonadotropin (hCG,and 10% (v/v porcine follicular fluid (pFF at 39 °C in a humidified atmosphere of 5% CO for 42-44 h and donor cell 2collected from ear skin afghanhound male dog. After somatic cell nuclear transfer (SCNT, embryo developmentwere examined for cleavage rate and 144 hr for final development after cultured in media. The result shows that,amino acid and mercapoethanol addition in culture medium (NCSU-23 have no effect on embryo development.The development rate of embryo until 16 cell stage in NCSU and NCSU supplement are 4.67% and morula stage are3.73% and 4.67%.Key words : intergeneric clone embryo, canine, ( amino acid (AA

  13. Mesenchymal stem cell-conditioned medium accelerates skin wound healing: An in vitro study of fibroblast and keratinocyte scratch assays

    International Nuclear Information System (INIS)

    We have used in vitro scratch assays to examine the relative contribution of dermal fibroblasts and keratinocytes in the wound repair process and to test the influence of mesenchymal stem cell (MSC) secreted factors on both skin cell types. Scratch assays were established using single cell and co-cultures of L929 fibroblasts and HaCaT keratinocytes, with wound closure monitored via time-lapse microscopy. Both in serum supplemented and serum free conditions, wound closure was faster in L929 fibroblast than HaCaT keratinocyte scratch assays, and in co-culture the L929 fibroblasts lead the way in closing the scratches. MSC-CM generated under serum free conditions significantly enhanced the wound closure rate of both skin cell types separately and in co-culture, whereas conditioned medium from L929 or HaCaT cultures had no significant effect. This enhancement of wound closure in the presence of MSC-CM was due to accelerated cell migration rather than increased cell proliferation. A number of wound healing mediators were identified in MSC-CM, including TGF-β1, the chemokines IL-6, IL-8, MCP-1 and RANTES, and collagen type I, fibronectin, SPARC and IGFBP-7. This study suggests that the trophic activity of MSC may play a role in skin wound closure by affecting both dermal fibroblast and keratinocyte migration, along with a contribution to the formation of extracellular matrix.

  14. Mesenchymal stem cell-conditioned medium accelerates skin wound healing: An in vitro study of fibroblast and keratinocyte scratch assays

    Energy Technology Data Exchange (ETDEWEB)

    Walter, M.N.M. [Institute for Science and Technology in Medicine, Keele University RJAH Orthopaedic Hospital, Oswestry, SY10 7AG (United Kingdom); School of Life and Health Science, Aston University, Aston Triangle, Birmingham, B4 7EJ (United Kingdom); Wright, K.T.; Fuller, H.R. [Institute for Science and Technology in Medicine, Keele University RJAH Orthopaedic Hospital, Oswestry, SY10 7AG (United Kingdom); MacNeil, S. [Kroto Research Institute and Centre for Nanoscience and Technology, Sheffield University, Sheffield, S1 2UE (United Kingdom); Johnson, W.E.B., E-mail: w.e.johnson@aston.ac.uk [School of Life and Health Science, Aston University, Aston Triangle, Birmingham, B4 7EJ (United Kingdom)

    2010-04-15

    We have used in vitro scratch assays to examine the relative contribution of dermal fibroblasts and keratinocytes in the wound repair process and to test the influence of mesenchymal stem cell (MSC) secreted factors on both skin cell types. Scratch assays were established using single cell and co-cultures of L929 fibroblasts and HaCaT keratinocytes, with wound closure monitored via time-lapse microscopy. Both in serum supplemented and serum free conditions, wound closure was faster in L929 fibroblast than HaCaT keratinocyte scratch assays, and in co-culture the L929 fibroblasts lead the way in closing the scratches. MSC-CM generated under serum free conditions significantly enhanced the wound closure rate of both skin cell types separately and in co-culture, whereas conditioned medium from L929 or HaCaT cultures had no significant effect. This enhancement of wound closure in the presence of MSC-CM was due to accelerated cell migration rather than increased cell proliferation. A number of wound healing mediators were identified in MSC-CM, including TGF-{beta}1, the chemokines IL-6, IL-8, MCP-1 and RANTES, and collagen type I, fibronectin, SPARC and IGFBP-7. This study suggests that the trophic activity of MSC may play a role in skin wound closure by affecting both dermal fibroblast and keratinocyte migration, along with a contribution to the formation of extracellular matrix.

  15. Biocompatibility Studies on Fibrin Glue Cultured with Bone Marrow Mesenchymal Stem Cells in Vitro

    Institute of Scientific and Technical Information of China (English)

    方煌; 彭松林; 陈安民; 黎逢峰; 任凯; 胡宁

    2004-01-01

    Summary: By culturing bone marrow mesenchymal stem cells of rabbits with fibrin glue in vitro,the biocompatibility of fibrin glue was investigated to study whether this material can be used as scaffolds in bone tissue engineering. After 2-months old New Zealand rabbits had been anesthetized, about 4-6 ml of bone marrow were aspirated from rabbit femoral trochanter. The monocytes suspension was aspirated after bone marrow was centrifuged with lymphocyte separating medium and cultured primarily. Then the cells were divided into two groups: one was cultured with complete medium and the other with induced medium. The cells of the two groups were collected and inoculated to the culture plate containing fibrin glue. In the control group, cells were inoculated without fibrin glue. The implanted cells and materials were observed at different stages under a phase-contrast microscope and scanning electron microscope. MTT and alkaline phosphatase (ALP) were measured. Bone marrow mesenchymal stem cells grew on the surface of fibrin glue and adhered to it gradually. Cells light absorption value (A value) and the ALP content showed no significant difference. Fibrin glue had no inhibitory effect on cell morphology, growth, proliferation and differentiation. It has good biocompatibility and can be used as scaffold materials for bone marrow mesenchymal stem cells in bone tissue engineering.

  16. Organisational culture as a part in the development of open innovation - the perspective of small and medium-sized enterprises

    Directory of Open Access Journals (Sweden)

    Szymańska Katarzyna

    2016-05-01

    Full Text Available The ability to introduce various concepts and business models is nowadays a prerequisite of creating a competitive advantage. This is to a large extent closely linked to the ability of enterprises to create, implement and disseminate a variety of innovative solutions. Today the use of open innovation is a necessity. This applies not only to large organisations, but also to small and medium-sized enterprises. In order to implement open innovation, small and medium-sized enterprises need to effectively manage their own growth through the preparation of appropriate strategies and the development of a model that encompasses all changes, taking into account a number of factors related to the growth dynamics of this sector. It is understood that an appropriate organisational culture plays an important role in the implementation of innovation in the sector of small and medium-sized enterprises. There are many indications that a cultural mismatch and misunderstanding are the main reasons for major problems related to the low level of implementation of innovation by small and medium-sized enterprises. The aim of the paper is to outline the issue of the impact of organisational culture on the development of the concept of open innovation in the sector of small and medium-sized enterprises.

  17. Testicular cell-conditioned medium supports embryonic stem cell differentiation toward germ lineage and to spermatocyte- and oocyte-like cells.

    Science.gov (United States)

    Shah, Syed M; Saini, Neha; Singh, Manoj K; Manik, Radheysham; Singla, Suresh K; Palta, Prabhat; Chauhan, Manmohan S

    2016-08-01

    Testicular cells are believed to secrete various growth factors that activate signaling pathways finally leading to gametogenesis. In vitro gametogenesis is an obscure but paramountly important task primarily because of paucity of the precursor cells and first trimester gonadal tissues. To overcome these limitations for development of in vitro gametes, the present study was designed to induce differentiation of buffalo embryonic stem (ES) cells into germ lineage cells on stimulation by testicular cell-conditioned medium (TCM), on the basis of the assumption that ES cells have the intrinsic property to differentiate into any cell type and TCM would provide the necessary growth factors for differentiation toward germ cell lineage. For this purpose, buffalo ES cells were differentiated as embryoid bodies (EB) in floating cultures and as monolayer adherent cultures in different doses (10%, 20%, and 40%) of TCM for different culture intervals (4, 8, and 14 days), to identify the optimum dose-and-time period. We observed that 40% TCM dose induces highest expression of primordial germ cell-specific (DAZL, VASA, and PLZF), meiotic (SYCP3, MLH1, TNP1/2, and PRM2), spermatocyte-specific (BOULE and TEKT1), and oocyte-specific genes (GDF9 and ZP2/3) for a culture period of 14 days under both floating and adherent differentiation. Immunocytochemical analysis of EBs and adherent cultures revealed presence of primordial germ cell markers (c-KIT, DAZL, and VASA), meiotic markers (SYCP3, MLH1 and PROTAMINE1), spermatocyte markers (ACROSIN and HAPRIN), and oocyte markers (GDF9 and ZP4), indicating progression into post-meiotic gametogenesis. The detection of germ cell-specific proteins in Day 14 EBs like VASA, GDF9, and ZP4 by Western blotting further confirmed germ lineage differentiation. The significantly lower (P embryonic development and progressed through two-cell, four-cell, eight-cell, morula, and blastocyst-like structures, indicative of their developmental competence

  18. Comparative Analysis of KnockOut™ Serum with Fetal Bovine Serum for the In Vitro Long-Term Culture of Human Limbal Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Shaokun Zhang

    2016-01-01

    Full Text Available The limbal epithelial cells can be maintained on 3T3 feeder layer with fetal bovine serum supplemented culture medium, and these cells have been used to successfully treat limbal stem cell deficiency. However, fetal bovine serum contains unknown components and displays quantitative and qualitative lot-to-lot variations. To improve the culture condition, the defined KnockOut serum replacement was investigated to replace fetal bovine serum for culturing human limbal epithelial cell. Human primary limbal epithelial cells were cultured in KnockOut serum and fetal bovine serum supplemented medium, respectively. The cell growth rate, gene expression, and maintenance of limbal epithelial stem cells were studied and compared between these two groups. Human primary limbal epithelial cells were isolated and successfully serially cultivated in this novel KnockOut serum supplemented medium; the cell proliferation and stem cell maintenance were similar to those of cells grown in fetal bovine serum supplemented medium. These data suggests that this KnockOut serum supplemented medium is an efficient replacement to traditional fetal bovine serum supplemented medium for limbal epithelial cell culture, and this medium has great potential for long term maintenance of limbal epithelial cells, limbal epithelial stem cells transplantation, and tissue regeneration.

  19. 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

  20. Children’s picturebook on sexual educationas a cultural and political medium

    Directory of Open Access Journals (Sweden)

    Małgorzata Cackowska

    2011-11-01

    Full Text Available In my article I deal with a social construction of meanings of picturebooks’ content and form in Poland and abroad, so also with what kinds of discourses and ideologies determine the conditions of picturebooks’ production in societies under analysis. For the analysis I have chosen picturebooks which deal with sexual education. The methodology applied in the research consists mostly of content analysis and critical discourse analysis. The research is a part of a bigger collaborative project called “Discursive construction of subjectivity” financed by Ministry of Science and Higher Education in Poland, grant no. N 10702632/3637, and conducted at the University of Gdansk. I present, basing on the empirical material, the critique of the dominant discourse in Poland which is powerful in the production of picturebooks, which is based on the conservative ideology and social and sexual roles defined in stereotypical, hierarchical and heterosexual terms. In this aura discourses based on liberal or radical ideologies are marginalized.The results show the knowledge/power relations, symptoms of symbolic violence in exemplified discourses and explain to what practices of ideological and political control the subject is exposed. In this context a picturebook is seen as a meaningful cultural and political medium, within the content and form of which various (possible ideologies and conceptions of the child are included to or excluded from social environment, what can occur as a real issue for educational theory and practice.

  1. Structure of polysaccharides from mycelium and culture medium of Phellinus nigricans using submerged fermentation

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Two water-soluble polysaccharides, PNW1 and PNM1, were respectively isolated from the mycelium and its culture medium of Phellinus nigricans using submerged fermentation before determining their effects on inhibiting the growth of transplantable tumors in mice. The results of the pharmaceutical experiments showed that oral administration of PNW1 and PNM1 (at a dose of 400 mg/kg) inhibited the growth of tumor of mice-transplanted Sarcoma 180 in vivo. Moreover, a higher inhibition ratio of PNW1 (74.70%) was obtained as compared with PNM1 (55.84%). The averaged molecular weight of PNW1 and PNM1 was determined to be 33 and 29 kD, respectively. Both PNW1 and PNM1 were consisted of glu- cose, galactose, mannose, arabinose and fucose. The major structural features of PNW1 and PNM1 were elucidated using partial acid hydrolysis, periodate oxidation, Smith degradation, 13C-NMR, me- thylation and GC-MS. On the basis of these results, the repeating units of PNW1 and PNM1 were estab- lished.

  2. Structure of polysaccharides from mycelium and culture medium of Phellinus nigricans using submerged fermentation

    Institute of Scientific and Technical Information of China (English)

    LI Xia; JIAO LiLi; ZHANG Xu; TIAN WenMin; CHEN Shan; ZHANG LiPing

    2008-01-01

    Two water-soluble polysaccharides, PNW1 and PNM1, were respectively isolated from the mycelium and its culture medium of Phellinus nigricans using submerged fermentation before determining their effects on inhibiting the growth of transplantable tumors in mice. The results of the pharmaceutical experiments showed that oral administration of PNW1 and PNM1 (at a dose of 400 mg/kg) inhibited the growth of tumor of mice-transplanted Sarcoma 180 in vivo. Moreover, a higher inhibition ratio of PNW1 (74.70%) was obtained as compared with PNM1 (55.84%). The averaged molecular weight of PNW1 and PNM1 was determined to be 33 and 29 kD, respectively. Both PNW1 and PNM1 were consisted of glu-cose, galactose, mannose, arabinose and fucose. The major structural features of PNW1 and PNM1 were elucidated using partial acid hydrolysis, periodate oxidation, Smith degradation,13C-NMR, me-thylation and GC-MS. On the basis of these results, the repeating units of PNW1 and PNM1 were estab-lished.

  3. 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.

  4. Statistical optimization of medium composition and culture condition for the production of recombinant antilipopolysaccharide factor of Eriocheir sinensis in Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    JIANG Shan; LIU Mei; WANG Baojie; JIANG Keyong; WANG Lei

    2011-01-01

    Anti-lipopolysaccharide factors (ALFs) are important antimicrobial peptides that are isolated from some aquatic species.In a previous study,we isolated ALF genes from Chinese mitten crab,Eriocheir sinensis.In this study,we optimized the production of a recombinant ALF by expressing E.sinensis ALF genes in Escherichia coli maintained in shake-flasks.In particular,we focused on optimization of both the medium composition and the culture condition.Various medium components were analyzed by the Plackett-Burman design,and two significant screened factors,(NH4)2SO4 and KH2PO4,were further optimized via the central composite design (CCD).Based on the CCD analysis,we investigated the induction start-up time,the isopropylthio-D-galactoside (IPTG) concentration,the post-induction time,and the temperature by response surface methodology.We found that the highest level of ALF fusion protein was achieved in the medium containing 1.89g/L (NH4)2SO4 and 3.18 g/L KH2PO4,with a cell optical density of 0.8 at 600 nm before induction,an IPTG concentration of 0.5 mmol/L,a post-induction temperature of 32.7℃,and a post-induction time of 4 h.Applying the whole optimization strategy using all optimal factors improved the target protein content from 6.1% (without optimization) to 13.2%.We further applied the optimized medium and conditions in high cell density cultivation,and determined that the soluble target protein constituted 10.5% of the total protein.Our identification of the economic medium composition,optimal culture conditions,and details of the fermentation process should facilitate the potential application of ALF for further research.

  5. Statistical optimization of medium composition and culture condition for the production of recombinant anti-lipopolysaccharide factor of Eriocheir sinensis in Escherichia coli

    Science.gov (United States)

    Jiang, Shan; Liu, Mei; Wang, Baojie; Jiang, Keyong; Wang, Lei

    2011-11-01

    Anti-lipopolysaccharide factors (ALFs) are important antimicrobial peptides that are isolated from some aquatic species. In a previous study, we isolated ALF genes from Chinese mitten crab, Eriocheir sinensis. In this study, we optimized the production of a recombinant ALF by expressing E. sinensis ALF genes in Escherichia coli maintained in shake-flasks. In particular, we focused on optimization of both the medium composition and the culture condition. Various medium components were analyzed by the Plackett-Burman design, and two significant screened factors, (NH4)2SO4 and KH2PO4, were further optimized via the central composite design (CCD). Based on the CCD analysis, we investigated the induction start-up time, the isopropylthio-D-galactoside (IPTG) concentration, the post-induction time, and the temperature by response surface methodology. We found that the highest level of ALF fusion protein was achieved in the medium containing 1.89 g/L (NH4)2SO4 and 3.18 g/L KH2PO4, with a cell optical density of 0.8 at 600 nm before induction, an IPTG concentration of 0.5 mmol/L, a post-induction temperature of 32.7°C, and a post-induction time of 4 h. Applying the whole optimization strategy using all optimal factors improved the target protein content from 6.1% (without optimization) to 13.2%. We further applied the optimized medium and conditions in high cell density cultivation, and determined that the soluble target protein constituted 10.5% of the total protein. Our identification of the economic medium composition, optimal culture conditions, and details of the fermentation process should facilitate the potential application of ALF for further research.

  6. In vitro differentiation of cultured human CD34+ cells into astrocytes

    Directory of Open Access Journals (Sweden)

    Katari Venkatesh

    2013-01-01

    Full Text Available Background: Astrocytes are abundantly present as glial cells in the brain and play an important role in the regenerative processes. The possible role of stem cell derived astrocytes in the spinal cord injuries is possible related to their influence at the synaptic junctions. Aim: The present study is focused on in vitro differentiation of cultured human CD34+ cells into astrocytes. Materials and Methods: Granulocyte-colony stimulating factor mobilized human CD34+ cells were isolated from peripheral blood using apheresis method from a donor. These cells were further purified by fluorescence-activated cell sorting and cultured in Dulbecco′s modified eagle′s medium. Thus, cultured cells were induced with astrocyte defined medium (ADM and in the differentiated astrocytes serine/threonine protein kinases (STPK and glutamine synthetase (GLUL activities were estimated. The expression of glial fibrillary acidic protein (GFAP and GLUL were confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR. Results: The cultured human CD34+ cells differentiated into astrocytes after 11 h of incubation in ADM. The RT-PCR experiment showed the expression of GLUL (1.5 kb and GFAP (2.9 kb in differentiated astrocytes. The high enzyme activities of GLUL and STPK in differentiated astrocytes compared with cultured human CD34+ cells confirmed astrocyte formation. Conclusion: In the present study, in vitro differentiation of stem cells with retinoic acid induction may result in the formation of astrocytes.

  7. Simple suspension culture system of human iPS cells maintaining their pluripotency for cardiac cell sheet engineering.

    Science.gov (United States)

    Haraguchi, Yuji; Matsuura, Katsuhisa; Shimizu, Tatsuya; Yamato, Masayuki; Okano, Teruo

    2015-12-01

    In this study, a simple three-dimensional (3D) suspension culture method for the expansion and cardiac differentiation of human induced pluripotent stem cells (hiPSCs) is reported. The culture methods were easily adapted from two-dimensional (2D) to 3D culture without any additional manipulations. When hiPSCs were directly applied to 3D culture from 2D in a single-cell suspension, only a few aggregated cells were observed. However, after 3 days, culture of the small hiPSC aggregates in a spinner flask at the optimal agitation rate created aggregates which were capable of cell passages from the single-cell suspension. Cell numbers increased to approximately 10-fold after 12 days of culture. The undifferentiated state of expanded hiPSCs was confirmed by flow cytometry, immunocytochemistry and quantitative RT-PCR, and the hiPSCs differentiated into three germ layers. When the hiPSCs were subsequently cultured in a flask using cardiac differentiation medium, expression of cardiac cell-specific genes and beating cardiomyocytes were observed. Furthermore, the culture of hiPSCs on Matrigel-coated dishes with serum-free medium containing activin A, BMP4 and FGF-2 enabled it to generate robust spontaneous beating cardiomyocytes and these cells expressed several cardiac cell-related genes, including HCN4, MLC-2a and MLC-2v. This suggests that the expanded hiPSCs might maintain the potential to differentiate into several types of cardiomyocytes, including pacemakers. Moreover, when cardiac cell sheets were fabricated using differentiated cardiomyocytes, they beat spontaneously and synchronously, indicating electrically communicative tissue. This simple culture system might enable the generation of sufficient amounts of beating cardiomyocytes for use in cardiac regenerative medicine and tissue engineering.

  8. Method and Apparatus for a Miniature Bioreactor System for Long-Term Cell Culture

    Science.gov (United States)

    Kleis, Stanley J. (Inventor); Geffert, Sandra K. (Inventor); Gonda, Steve R. (Inventor)

    2015-01-01

    A bioreactor and method that permits continuous and simultaneous short, moderate, or long term cell culturing of one or more cell types or tissue in a laminar flow configuration is disclosed, where the bioreactor supports at least two laminar flow zones, which are isolated by laminar flow without the need for physical barriers between the zones. The bioreactors of this invention are ideally suited for studying short, moderate and long term studies of cell cultures and the response of cell cultures to one or more stressors such as pharmaceuticals, hypoxia, pathogens, or any other stressor. The bioreactors of this invention are also ideally suited for short, moderate or long term cell culturing with periodic cell harvesting and/or medium processing for secreted cellular components.

  9. 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...

  10. 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

  11. 3D culture for cardiac cells.

    Science.gov (United States)

    Zuppinger, Christian

    2016-07-01

    This review discusses historical milestones, recent developments and challenges in the area of 3D culture models with cardiovascular cell types. Expectations in this area have been raised in recent years, but more relevant in vitro research, more accurate drug testing results, reliable disease models and insights leading to bioartificial organs are expected from the transition to 3D cell culture. However, the construction of organ-like cardiac 3D models currently remains a difficult challenge. The heart consists of highly differentiated cells in an intricate arrangement.Furthermore, electrical “wiring”, a vascular system and multiple cell types act in concert to respond to the rapidly changing demands of the body. Although cardiovascular 3D culture models have been predominantly developed for regenerative medicine in the past, their use in drug screening and for disease models has become more popular recently. Many sophisticated 3D culture models are currently being developed in this dynamic area of life science. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.

  12. 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

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

    Science.gov (United States)

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

    2015-12-01

    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-unit culture medium perfusion. Experimental results revealed that the flow of culture medium could be pneumatically driven in a flow-rate uniform manner. We used the system to successfully perform a perfusion 3-D cell culture of mesenchymal stem cells (MSCs) for up to 16 days. Moreover, we investigated the effects of various cell culture models on the physiology of MSCs. The physiological nature of MSCs can vary with respect to the cell culture model used. Using the perfusion 3-D cell culture format might affect the proliferation and osteogenic differentiation of MSCs. Overall, we have developed a cell culture system that can achieve multi-well microplate-based perfusion 3-D cell culture in an efficient, cost-effective, and user-friendly manner. These features could facilitate the widespread application of perfusion cell culture models for cell-based assays.

  14. Ameloblasts serum-free conditioned medium: bone morphogenic protein 4-induced odontogenic differentiation of mouse induced pluripotent stem cells.

    Science.gov (United States)

    Liu, Li; Liu, Ying-Feng; Zhang, Jing; Duan, Yin-Zhong; Jin, Yan

    2016-06-01

    Induced pluripotent stem (iPS) cells possess the ability of self-renewal and can differentiate into cells of the three germ layers, both in vitro and in vivo. Here we report a new method to efficiently induce differentiation of mouse iPS cells into the odontogenic lineage. Using ameloblasts serum-free conditioned medium (ASF-CM), we successfully generated ameloblast-like cells from mouse iPS cells. Importantly, culturing mouse iPS cells in ASF-CM supplemented with BMP4 (ASF-BMP4) promoted odontogenic differentiation, which was evident by the upregulation of ameloblast-specific as well as odontoblast-specific genes. On the other hand, culturing mouse iPS cells in ASF-CM supplemented with noggin (ASF-noggin), an inhibitor of BMP4, abrogated this effect. These results suggest that mouse iPS cells can be induced by ASF-BMP4 to differentiate into ameloblast-like and odontoblast-like cells. The results of our study raise the possibility of using patient-specific iPS cells for tooth regeneration in the future. Copyright © 2016 John Wiley & Sons, Ltd. PMID:23606575

  15. Differentiation of rat embryonic neural stem cells promoted by co-cultured Schwann cells

    Institute of Scientific and Technical Information of China (English)

    万虹; 安沂华; 张泽舜; 张亚卓; 王忠诚

    2003-01-01

    Objective To explore the factors which induce differentiation of embryonic neural stem cells. Methods Rat embryonic neural stem cells were co-cultured with newborn rat Schwann cells in serum-free medium. The phenotype and specific-markers including tubulin-β, glial fibrillary acidic protein (GFAP) and galactorcerebroside (GalC), were domonstrated by phase contrast microscopy and double immunofluorescence staining. Results Overall, 80%±5% of neural stem cells protruded several elongated processes and expressed tubulin-β antigen at high levels, while 20±3% of them protruded several short processes and were GalC or GFAP positive. Conclusion The factors secreted by Schwann cells could induce rat embryonic neural stem cell to differentiate.

  16. 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

  17. 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

    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...

  18. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.

    2007-07-01

    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  19. Signalling pathways induced in cells exposed to medium from irradiated cells

    Energy Technology Data Exchange (ETDEWEB)

    Lyng, F.M.; Maguire, P. (Radiation and Environmental Science Centre, Focas Institute, Dublin Institute of Technology, Dublin (Ireland)); McClean, B.; Seymour, C.; Mothersill, C. (St Luke' s Hospital, Dublin (Ireland))

    2008-12-15

    In recent years, radiation induced bystander effects have been reported in cells which were not themselves irradiated but were either in the vicinity of irradiated cells or exposed to medium from irradiated cells. The effects have been clearly shown to occur both in vivo and in vitro. This work has led to a paradigm shift in radiobiology over the last 5 - 10 years. The target theory of radiation induced effects is now being challenged because of an increasing number of studies which demonstrate non(DNA)-targeted effects. These effects appear to be particularly important at low doses. Considerable evidence now exists relating to radiation-induced bystander effects but the mechanisms involved in the transduction of the signal are still unclear. Cell - cell communication through gap junctions and / or secretion of a cytotoxic factor into the medium are thought to be involved in the transduction of the bystander signal. Oxidative metabolism has been shown to be important in both mechanisms. Signalling pathways leading to apoptosis, such as calcium, MAP kinase, mitochondrial and reactive oxygen species (ROS) signalling are discussed. The importance of oxidative metabolism and calcium signalling in bystander responses are demonstrated. Further investigations of these signalling pathways may aid in the identification of novel therapeutic targets. (orig.)

  20. Conditioned medium from activated spleen cells supports the survival of rat retinal cells in vitro

    Directory of Open Access Journals (Sweden)

    A. Sholl-Franco

    1997-11-01

    Full Text Available Cytokines are a heterogeneous group of molecules that have been associated with several functions in the nervous system, such as survival and differentiation of neuronal and glial cells. In the present study, we demonstrated that conditioned medium from spleen cells activated with concanavalin A increased neuritogenesis and survival of retinal cells, as measured by biochemical and morphological criteria. Our data showed that conditioned medium induced a five-fold increase in the amount of protein after 120 h in vitro. This effect was not inhibited by the blockade of voltage-dependent L-type calcium channels with 5.0 µM nifedipine. However, the use of an intracellular calcium chelator (15.0 µM BAPTA-AM inhibited this effect. Our results support the idea that factors secreted by activated lymphocytes, such as cytokines, can modulate the maintenance and the differentiation of rat retinal cells in vitro, indicating a possible role of these molecules in the development of retinal cells, as well as in its protection against pathological conditions

  1. Electrochemical cells for medium- and large-scale energy storage

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Wei; Wei, Xiaoliang; Choi, Daiwon; Lu, Xiaochuan; Yang, G.; Sun, C.

    2014-12-12

    This is one of the chapters in the book titled “Advances in batteries for large- and medium-scale energy storage: Applications in power systems and electric vehicles” that will be published by the Woodhead Publishing Limited. The chapter discusses the basic electrochemical fundamentals of electrochemical energy storage devices with a focus on the rechargeable batteries. Several practical secondary battery systems are also discussed as examples

  2. 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

  3. Dopamine facilitates dendritic spine formation by cultured striatal medium spiny neurons through both D1 and D2 dopamine receptors.

    Science.gov (United States)

    Fasano, Caroline; Bourque, Marie-Josée; Lapointe, Gabriel; Leo, Damiana; Thibault, Dominic; Haber, Michael; Kortleven, Christian; Desgroseillers, Luc; Murai, Keith K; Trudeau, Louis-Éric

    2013-04-01

    Variations of dopamine (DA) levels induced by drugs of abuse or in the context of Parkinson's disease modulate the number of dendritic spines in medium spiny neurons (MSNs) of the striatum, showing that DA plays a major role in the structural plasticity of MSNs. However, little is presently known regarding early spine development in MSNs occurring before the arrival of cortical inputs and in particular about the role of DA and D1 (D1R) and D2 (D2R) DA receptors. A cell culture model reconstituting early cellular interactions between MSNs, intrinsic cholinergic interneurons and DA neurons was used to study the role of DA in spine formation. After 5 or 10 days in vitro, the presence of DA neurons increased the number of immature spine-like protrusions. In MSN monocultures, chronic activation of D1R or D2R also increased the number of spines and spinophilin expression in MSNs, suggesting a direct role for these receptors. In DA-MSN cocultures, chronic blockade of D1R or D2R reduced the number of dendritic spines. Interestingly, the combined activation or blockade of both D1R and D2R failed to elicit more extensive spine formation, suggesting that both receptors act through a mechanism that is not additive. Finally, we found increased ionotropic glutamate receptor responsiveness and miniature excitatory postsynaptic current (EPSC) frequency in DA-MSN co-cultures, in parallel with a higher number of spines containing PSD-95, suggesting that the newly formed spines present functional post-synaptic machinery preparing the MSNs to receive additional glutamatergic contacts. These results represent a first step in the understanding of how dopamine neurons promote the structural plasticity of MSNs during the development of basal ganglia circuits.

  4. 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 foc...... metaheuristic for experimental design generation for the cell culture microfluidic biochips, and we have evaluated our approach using multiple experimental setups....... in this paper is on the optimization of how a biochemical application is performed on a biochip. In this paper, we consider cell culture biochips, where several cell colonies are exposed to soluble compounds and monitored in real-time to determine the right combination of factors that leads to the desired...

  5. Isolation and mycelial growth of Diehliomyces microsporus: effect of culture medium and incubation temperature

    Directory of Open Access Journals (Sweden)

    José Soares do Nascimento

    2007-07-01

    Full Text Available The false truffle is one of the main problems in the production of the Agaricus brasiliensis in Brazil and the control of this fungal competitor has been rather difficult due to difficulties in the isolation and cultivation of this pathogen. This experiment was conducted in three stages, the first consisting of the isolation of Diehliomyces microsporus starting from portions of the fruiting body and through the ascospores suspension; second, D. microsporus cultivated in vitro at 15, 20, 25, 30 and 35ºC in six different culture media (CSDA, OCDA, PCDA, ODA, PDA, CDA; third, D. microsporus was inoculated on sterilized compost for formation of the fruiting body. The colony formation from tissue of D. microsporus starting from portions of fruiting body was more efficient than germination of the ascospores. Compost medium (CDA allowed a larger diameter of the D. microsporus colony, followed by the medium made up of compost and potato mixture, favoring a denser composition. The largest mycelial growth speed of D. microsporus occurred when the culture was incubated at 28 and 30ºC. Incubation temperatures lower than 15ºC or above 35ºC inhibited the mycelial growth of D. microsporus completely. The fruiting bodies were obtained easily in sterilized compost and later inoculated along with mycelial competitor.A falsa trufa está sendo um dos principais problemas na produção do Agaricus brasiliensis cultivado no Brasil e o controle deste fungo competidor tem sido difícil, devido às dificuldades encontradas no isolamento e cultivo do patógeno. Este experimento foi conduzido em três etapas, sendo a primeira constituída pelo isolamento de Diehliomyces microsporus a partir de porções do ascostroma e através da suspensão de ascósporos; a segunda, o cultivo in vitro de D. microsporus nas temperaturas de 15, 20, 25, 30 e 35ºC e em seis meios de cultura (CTDA, ACDA, BCDA, ADA, BDA e CDA e a terceira pela inoculação de D. microsporus no composto

  6. Dual polarization of microglia isolated from mixed glial cell cultures.

    Science.gov (United States)

    Ju, Lili; Zeng, Hui; Chen, Yun; Wu, Yanhong; Wang, Beibei; Xu, Qunyuan

    2015-09-01

    Microglia are versatile immune effector cells of the CNS and are sensitive to various stimuli. The different methods used to isolate microglia may affect some of their characteristics, such as their polarization state. The influence of cell sorting methods on the polarization state of microglia has never been studied. Mixed glial culture system (MGCS) and magnetic activated cell sorting (MACS) are two methods that are commonly used to purify microglia. This study compares the immunological states between microglia isolated by MGCS and microglia isolated by MACS. We show that microglia isolated by MGCS exhibit a stronger immune-activated state than microglia isolated by MACS. They present an elevated phagocytic ability and high levels of markers associated with classical activation (M1) and alternative activation (M2). In addition, high levels of M1-type and M2-type chemokine (C-C motif) ligand 2 and transforming growth factor-β1 were detected in the culture medium of mixed glial cells. Our results show that microglia isolated by MGCS are in an immune-activated state, whereas microglia isolated by MACS appear to be closer to their primary in vivo state. Therefore, the immune status of microglia, depending on the protocol used to purify them, should be carefully considered in neuropathology research.

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

    Science.gov (United States)

    Cao, Ting-Ting; Zhang, Yu-Qing

    2015-09-01

    Cell cultures often require the addition of animal serum and other supplements. In this study, silk sericin, a bioactive protein, recovered from the waste of silk floss production was hydrolysed into three pepsin-degraded sericin peptides with different ranges of molecular mass. Normal animal cells, tumour cells and hybridoma cells were cultured systematically in FBS culture media containing sericin as a supplement or serum substitute. The culture test and microscopic observation of L929 cells showed that the smaller molecular weight of the degraded sericin is most suitable for cell culture. The cell culture results showed that with the degradation of sericin, for normal mouse fibroblast L929 cells, addition of 0.75 % sericin into FBS culture medium yields cell viability that is superior to FBS culture medium alone. When all serum was replaced by sericin, cell viability in the sericin medium could reach about one half of that in FBS medium. When in a medium containing a mixture of FBS: sericin (6:4, v/v), the cell culture effect is about 80 %. For the cultures of four tumour and one hybridoma cells, regardless of the molecular weight range, these degraded sericin peptides could substitute all serum in FBS media. The cell viability and proliferation of these tumour and hybridoma cells are equivalent or superior to that in FBS medium. In other words, cell viability and proliferation of these tumour and hybridoma cells in sericin media are more preferable to serum media. The mechanism of the sericin protein to promote cell growth and proliferation will be further investigated later. PMID:25895088

  8. Metabolic labeling with (14C)-glucose of bloodstream and cell culture trypanosoma cruzi trypomastigotes:

    International Nuclear Information System (INIS)

    Trypomastigote forms of Trypanosoma cruzi from infected mouse blood and from cell culture were metabolically labeled by incubation with D-(14C)-glucose. Analysis by polyacrylamide gel electrophoresis of lysates from parasites of two strains (RA and CA1) showed a significantly different pattern. The difference was mainly quantitative when the blood and cell culture trypomastigotes of the RA strain were compared. Analysis of the culture medium by paper electrophoresis showed an anionic exometabolite only in the blood forms of both strains. (Author)

  9. Medium-size droplets of methyl ricinoleate are reduced by cell-surface activity in the gamma-decalactone production by Yarrowia lipolytica.

    Science.gov (United States)

    Waché, Y; Bergmark, K; Courthaudon, J L; Aguedo, M; Nicaud, J M; Belin, J M

    2000-03-01

    Size of methyl ricinoleate droplets during biotransformation into gamma-decalactone by Yarrowia lipolytica was measured in both homogenized and non-homogenized media. In non-homogenized but shaken medium, droplets had an average volume surface diameter d32 of 2.5 microm whereas it was 0.7 microm in homogenized and shaken medium. But as soon as yeast cells were inoculated, both diameters became similar at about 0.7 microm and did not vary significantly until the end of the culture. The growth of Y. lipolytica in both media was very similar except for the lag phase which was lowered in homogenized medium conditions. PMID:10747247

  10. Regulation of Schwann cell proliferation in cultured segments of the adult rat sciatic nerve

    DEFF Research Database (Denmark)

    Svenningsen, Åsa Fex; Kanje, M

    1998-01-01

    of Schwann cells. Removal of extracellular Ca2+ by addition of EGTA to the culture medium suppressed [3H] thymidine incorporation as did the calmodulin inhibitor 48/80. The Ca2+ ionophore A23187 increased incorporation. Staurosporin, an inhibitor of protein kinase C (PKC), suppressed [3H] thymidine...

  11. In vitro isolation, culture and identification of adipose-derived stem cells*

    Institute of Scientific and Technical Information of China (English)

    Du Guo-jia; Chen Xiao-hong; Zhu Guo-hua; Fan Yan-dong; Wang Yun; Dang Mu-ren

    2013-01-01

    BACKGROUND:Adipose-derived stem cells are easily col ected and abundantly cultured, which can proliferate rapidly when being cultured in vitro. With multi-directional differentiation potential, adipose-derived stem cells are expected as seed cells for tissue engineering. OBJECTIVE:To isolate, culture and identify of adipose-derived stem cells from Sprague-Dawley rats in vitro. METHODS:The subcutaneous adipose tissue was obtained from the iliac region of rats under the aseptic condition, and then was digested with 0.075%type Ⅰ col agenase and cultured in vitro. The morphology and proliferation characteristics of the cells were observed under an inverted phase contrast microscope. The third passage was put into gauge for growth curve by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, and the cells were also identified by CD44, a stem cellmarker, with immunofluorescence staining. Adipose-derived stem cells were induced and differentiated into adipocytes in Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 containing 10%fetal bovine serum, dexamethasone and insulin, and then the cells were identified with oil red“O”staining. Adipose-derived stem cells were induced and differentiated into neural cells, and then the cells were identified with immunohistochemical staining. RESULTS AND CONCLUSION:The growth curve of adipose-derived stem cells was opposite-like“S”shape, and it strongly expressed CD44 that can designate a stem cell. The passage cells were exposed to a defined medium for adipocyte differentiation, and then could be stained with oil red. After being induced and differentiated into nerve cells, the cells expressed neuron-specific enolase. The adipose-derived stem cells of Sprague-Dawley rats are characterized by easy isolation, culture and proliferation in vitro, expressing related phenotypes of mesenchymal stem cells, as wel as induction and differentiation under certain conditions.

  12. Elicitation Phenolic Compounds in Cell Culture of Vitis vinifera L. by Phaeomoniella chlamydospora

    Directory of Open Access Journals (Sweden)

    Sák Martin

    2014-12-01

    Full Text Available The in vitro cell cultures of Vitis vinifera L. cv. St. Laurent were treated with two elicitors - synthetic methyl jasmonate and natural, prepared from grapevine plant infected with the Phaeomoniella chlamydospora, the agent causing the Esca disease of grapevine. Efficiency of phenolic compounds production after elicitation of cell culture was analysed immediately after treatment (15 min, 30 min, 60 min and later (after 24, 48, and 72 hours. The cell growth and content of phenolic compounds (+-catechin, (--epicatechin, p-coumaric acid, syringaldehyde, rutin, vanillic acid, and trans-resveratrol were analysed in cultivated cells as well as in cultivation medium. Pch-treatment increased production of total polyphenols the most significantly 15 min after the elicitation and in optimal time was 2.86 times higher than in nonelicited culture and 1.44 times higher than in MeJa induced cell culture.

  13. Using CR1aa versus KSOM as the culture medium for in vitro embryo production of cattle

    Directory of Open Access Journals (Sweden)

    Endang Triwulaninngsih

    2002-03-01

    Full Text Available This research has been conducted at the laboratory of in vitro fertilization in the Department of Animal Science University of Wisconsin, USA. These embryos can be used for improving genetic value of Indonesian cattle. Oocytes were matured in TCM- 199 medium (in 5% CO2 incubator and at 390C enriched with follicle stimulating hormone (FSH 10 μl/ml, oestradiol 17 β 1μl/ml and 10% Fetal Calf Serum (FCS. The oocytes were fertilized in vitro with motile sperm and incubation between sperm and oocytes in fertilization medium Tyroide Albumin Lactate Pyruvate (TALP for 20 hours. All zygotes were cultured in CR1aa (n=1549 medium versus modification of protein-free pottasium simplex optimized medium (KSOM (n=675 up to blastocyst stage and were fed FCS 5 μl/50 μl medium on day 6, as treatment A and B respectively. Data were analyzed by completely randomized design with SAS program. Percentages of cleavage, morula, blastocyst, expanded blastocyst, unfertilized and degenerated ova in this study were 91.4% vs 75.6 %; 75.6% vs 58.9%; 61.5% vs 38.5%; 31.2% vs 5.1%, 8.6% vs 24.4%, 15.7% vs 8% which were significantly different (P<0.01 for treatment CR1aa and KSOM respectively. Based on this study, CR1aa medium is better culture medium than KSOM for efficient in vitro production (IVP of bovine embryos.

  14. Cd-induced phytochelatin synthesis in Dittrichia viscosa (L.) Greuter is determined by the dilution of the culture medium.

    Science.gov (United States)

    Fernández, R; Fernández-Fuego, D; Rodríguez-González, P; Alonso, J I García; Bertrand, A; González, A

    2014-01-01

    In this paper, we examined Cd accumulation and PC synthesis in two clones of Dittrichia viscosa, one with a metallicolous (DV-A) and the other with a non-metallicolous origin (DV-W). The clones were cultured in vitro with 0 and 10 mg Cd L(-1) in both short-term treatments (up to 72 h) and over 10 days. We also examined the influence of the culture medium dilution and the PC-synthesis inhibitor, L-buthionine-sulfoximine (BSO), on these parameters. Similar Cd accumulation values were found in the two clones. No synthesis of new thiolic compounds was observed in Cd-treated plants cultured in vitro in Murashige and Skoog medium up to 72 h when compared to controls. Dilution of the culture medium affected PC production, increasing it in 1/2 MS and especially in 1/4 MS. Cd uptake did not increase in the same way, but still hyperaccumulation levels were exceeded in all Cd treatments. BSO addition increased the sensitivity of D. viscosa to Cd and diminished Cd accumulation. Nevertheless, a poor correlation between PCs and Cd accumulation capacity was observed since the highest Cd content did not correspond to the highest PC levels. All these results obtained suggest that PCs are important in Cd accumulation and detoxification in D. viscosa and also that other mechanisms might be involved in these traits.

  15. A randomized clinical trial to evaluate the effect of granulocyte- macrophage colony-stimulating factor (GM-CSF) in embryo culture medium for in vitro fertilization

    DEFF Research Database (Denmark)

    Ziebe, Søren; Loft, Anne; Povlsen, Betina B.;

    2013-01-01

    To evaluate the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) in embryo culture medium on ongoing implantation rate (OIR).......To evaluate the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) in embryo culture medium on ongoing implantation rate (OIR)....

  16. Platelet-rich concentrate in serum free medium enhances osteogenic differentiation of bone marrow-derived human mesenchymal stromal cells

    Science.gov (United States)

    Ramasamy, Thamil Selvee; Karunanithi, Puvanan; Naveen, Sangeetha Vasudevaraj; Murali, Malliga Raman; Abbas, Azlina A.; Kamarul, Tunku

    2016-01-01

    Previous studies have shown that platelet concentrates used in conjunction with appropriate growth media enhance osteogenic differentiation of human mesenchymal stromal cells (hMSCs). However, their potential in inducing osteogenesis of hMSCs when cultured in serum free medium has not been explored. Furthermore, the resulting osteogenic molecular signatures of the hMSCs have not been compared to standard osteogenic medium. We studied the effect of infrequent supplementation (8-day interval) of 15% non-activated platelet-rich concentrate (PRC) in serum free medium on hMSCs proliferation and differentiation throughout a course of 24 days, and compared the effect with those cultured in a standard osteogenic medium (OM). Cell proliferation was analyzed by alamar blue assay. Gene expression of osteogenic markers (Runx2, Collagen1, Alkaline Phosphatase, Bone morphogenetic protein 2, Osteopontin, Osteocalcin, Osteonectin) were analyzed using Q-PCR. Immunocytochemical staining for osteocalcin, osteopontin and transcription factor Runx2 were done at 8, 16 and 24 days. Biochemical assays for the expression of ALP and osteocalcin were also performed at these time-points. Osteogenic differentiation was further confirmed qualitatively by Alizarin Red S staining that was quantified using cetylpyridinium chloride. Results showed that PRC supplemented in serum free medium enhanced hMSC proliferation, which peaked at day 16. The temporal pattern of gene expression of hMSCs under the influence of PRC was comparable to that of the osteogenic media, but at a greater extent at specific time points. Immunocytochemical staining revealed stronger staining for Runx2 in the PRC-treated group compared to OM, while the staining for Osteocalcin and Osteopontin were comparable in both groups. ALP activity and Osteocalcin/DNA level were higher in the PRC group. Cells in the PRC group had similar level of bone mineralization as those cultured in OM, as reflected by the intensity of Alizarin red

  17. Rational design of medium supplementation strategy for improved influenza viruses production based on analyzing nutritional requirements of MDCK Cells.

    Science.gov (United States)

    Huang, Ding; Xia-Hou, Kang; Liu, Xu-Ping; Zhao, Liang; Fan, Li; Ye, Zhaoyang; Tan, Wen-Song; Luo, Jian; Chen, Ze

    2014-12-12

    Influenza vaccine production using cell culture technology has become popular nowadays. However, to meet the ever increasing demand of influenza vaccine, it is prerequisite to improve the yield of influenza virus in cells. To achieve this, in the present study, the nutritional requirements of MDCK cells in the virus production process were analyzed and a nutrient-feeding strategy was developed accordingly. Based on the consumption rates and corresponding concentration optimization, glucose and fast metabolized amino acids were supplemented into the maintaining medium at the time of infection. Compared with the non-supplemented culture, the average cell specific death rate during 0-48 h post-infection was 0.013 h(-1), which was 40.91% lower in the nutrient-supplemented culture. Total virus titer, HA antigen protein concentration and cell-specific virus yield were (1.88±0.23)×10(3) HA units/50μL, 11.70±0.22 μg/mL and (10.06±1.16)×10(3) virions/cell, respectively, which were 84.04±22.50%, 31.46±2.87% and 86.64±25.81% higher than those in the control, respectively. These data showed that the appropriate supplementation of nutrients during virus production process could reduce cell death, and improve cell-specific virus yield and total influenza virus output. This study laid foundation for the development of cell culture technology for influenza vaccine production.

  18. Chemical properties and antioxidant activity of exopolysaccharides fractions from mycelial culture of Inonotus obliquus in a ground corn stover medium.

    Science.gov (United States)

    Xiang, Yuling; Xu, Xiangqun; Li, Juan

    2012-10-15

    The medicinal mushroom Inonotus obliquus has been a folk remedy for a long time in East-European and Asian countries. We first reported the enhancement in production and antioxidant activity of exopolysaccharides by I. obliquus culture under lignocellulose decomposition. In this study, the two different sources of exopolysaccharides from the control medium and the lignocellulose (corn stover) containing medium by I. obliquus in submerged fermentation were fractionated and purified by chromatography. The exopolysaccharides from the corn stover-containing medium presented significantly stronger hydroxyl and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging activity than the control. Three fractions from the control medium and the corn stover-containing medium were isolated respectively. The fraction of DEPL3 from the corn stover-containing medium with the highest protein content (38.3%), mannose content (49.6%), and the lowest molecular weight (29 kDa) had the highest antioxidant activity with the lowest IC50 values. In conclusion, lignocellulose decomposition changed the chemical characterisation and significantly enhanced the antioxidant activity of exopolysaccharide fractions.

  19. Chemical properties and antioxidant activity of exopolysaccharides fractions from mycelial culture of Inonotus obliquus in a ground corn stover medium.

    Science.gov (United States)

    Xiang, Yuling; Xu, Xiangqun; Li, Juan

    2012-10-15

    The medicinal mushroom Inonotus obliquus has been a folk remedy for a long time in East-European and Asian countries. We first reported the enhancement in production and antioxidant activity of exopolysaccharides by I. obliquus culture under lignocellulose decomposition. In this study, the two different sources of exopolysaccharides from the control medium and the lignocellulose (corn stover) containing medium by I. obliquus in submerged fermentation were fractionated and purified by chromatography. The exopolysaccharides from the corn stover-containing medium presented significantly stronger hydroxyl and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging activity than the control. Three fractions from the control medium and the corn stover-containing medium were isolated respectively. The fraction of DEPL3 from the corn stover-containing medium with the highest protein content (38.3%), mannose content (49.6%), and the lowest molecular weight (29 kDa) had the highest antioxidant activity with the lowest IC50 values. In conclusion, lignocellulose decomposition changed the chemical characterisation and significantly enhanced the antioxidant activity of exopolysaccharide fractions. PMID:23442636

  20. 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.

  1. Simple and sensitive method for monitoring drug-induced cell injury in cultured cells

    Energy Technology Data Exchange (ETDEWEB)

    Shirhatti, V.; Krishna, G.

    1985-06-01

    A simple, sensitive method has been developed for evaluating cell injury noninvasively in monolayer cells in culture. The cell ATP pool was radiolabeled by incubating the cells with (/sup 14/C)adenine. The uptake and incorporation of (/sup 14/C)adenine was shown to proportional to the number of cells. As determined by HPLC, about 65-70% of the incorporated /sup 14/C label was in the ATP pool, 15-20% was in the ADP pool, and the rest was in the 5'-AMP pool. When prelabeled cells were exposed to toxic drugs (acetaminophen, calcium ionophore A-23187, or daunomycin) there was a marked decrease in cell ATP with a concomitant increase in leakage of labeled nucleotides, mainly 5'-AMP and 5'IMP. The authors have shown that leakage of /sup 14/C label into the medium from the prelabeled cells may be employed for quantitation of cell injury. This new measure of toxicity was shown to correlate very well with LDH leakage from the cells, which is a well accepted measure of cell injury. The leakage of 5'-(/sup 14/C)AMP also correlated very well with the reduction of cell ATP in cardiac myocytes. This method has been used for monitoring drug-induced toxicity in liver cells, cardiac myocytes, and LB cells.

  2. Bovine colostrum supports the serum-free proliferation of epithelial cells but not of fibroblasts in long-term culture

    OpenAIRE

    Klagsbrun, M

    1980-01-01

    Medium lacking serum but supplemented with milk will support the growth of sparse cells in culture. Milk obtained within 8 h after the birth of a calf (day 1 colostrum) is the most effective in supporting proliferation. In mixed cultures of early-passage bovine embryonic kidney (BEK) or early-passage calf kidney (CK) cells, both epithelial cells and fibroblasts grow in Dulbecco’s modified eagle’s medium (DMEM) supplemented with serum. However, only cells that appear to be epithelial-like grow...

  3. Human autologous serum as a substitute for fetal bovine serum in human Schwann cell culture.

    Directory of Open Access Journals (Sweden)

    Parisa Goodarzi

    2014-04-01

    Full Text Available Nowadays, cell -based and tissue engineered products have opened new horizons in treatment of incurable nervous system disorders. The number of studies on the role of Schwann cells (SC in treating nervous disorders is higher than other cell types. Different protocols have been suggested for isolation and expansion of SC which most of them have used multiple growth factors, mitogens and fetal bovine sera (FBS in culture medium. Because of potential hazards of animal-derived reagents, this study was designed to evaluate the effect of replacing FBS with human autologous serum (HAS on SC's yield and culture parameters. Samples from 10 peripheral nerve biopsies were retrieved and processed under aseptic condition. The isolated cells cultured in FBS (1st group or autologous serum (2nd group. After primary culture the cells were seeded at 10000 cell/cm2 in a 12 wells cell culture plate for each group. At 100% confluency, the cell culture parameters (count, viability, purity and culture duration of 2 groups were compared using paired t-test. The average donors' age was 35.80 (SD=13.35 and except for 1 sample the others cultured successfully. In first group, the averages of cell purity, viability and culture duration were 97% (SD=1.32, 97/33% (SD=1.22 and 11.77 (SD=2.58 days respectively. This parameters were 97.33% (SD=1.00, 97.55% (SD=1.33 and 10.33 days (SD=1.65 in second group. The difference of cell count, purity and viability were not significant between 2 groups (P>0.05. The cells of second group reached to 100% confluency in shorter period of time (P=0.03. The results of this study showed that autologous serum can be a good substitute for FBS in human SC culture. This can reduce the costs and improve the safety of cell product for clinical application.

  4. Human Dermal Stem/Progenitor Cell-Derived Conditioned Medium Improves Senescent Human Dermal Fibroblasts

    Directory of Open Access Journals (Sweden)

    Ji-Yong Jung

    2015-08-01

    Full Text Available Adult skin stem cells are recognized as potential therapeutics to rejuvenate aged skin. We previously demonstrated that human dermal stem/progenitor cells (hDSPCs with multipotent capacity could be enriched from human dermal fibroblasts using collagen type IV. However, the effects of hDSPCs on cellular senescence remain to be elucidated. In the present study, we investigated whether conditioned medium (CM collected from hDSPC cultures (hDSPC-CM exhibits beneficial effects on senescent fibroblasts. We found that hDSPC-CM promoted proliferation and decreased the expression level of senescence-associated β-galactosidase in senescent fibroblasts. In addition, p53 phosphorylation and p21 expression were significantly reduced in senescent fibroblasts treated with hDSPC-CM. hDSPC-CM restored the expression levels of collagen type I, collagen type III, and tissue inhibitor of metalloproteinase, and antagonized the increase of matrix metalloproteinase 1 expression. Finally, we demonstrated that hDSPC-CM significantly reduced reactive oxygen species levels by specifically up-regulating the expression level of superoxide dismutase 2. Taken together, these data suggest that hDSPC-CM can be applied as a potential therapeutic agent for improving human aged skin.

  5. Bioleaching of sphalerite by Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans cultured in 9K medium modified with pyrrhotite

    Institute of Scientific and Technical Information of China (English)

    CHEN Song; QIU Guan-zhou; QIN Wen-qing; LAN Zhuo-yue

    2008-01-01

    Elective culture of Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans in 9K medium modified with pyrrhotite was studied. Bioleaching of flotation concentrate of sphalerite by the selected bacteria was carried out. The results show that the microorganisms cultured by pyrrhotite are a mixture of Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans, of which the capability to oxidize ferrous to ferric irons is enhanced by the high mass ratio of Fe to S in pyrrhotite. Three pyrrhotite samples were separated into various parts with corresponding S/Fe ratios by magnetic separation and were used to culture the elective bacteria as the substrate. The association of the cultures could provide a more rapid and complete oxidation of sphalerite than that of bacteria cultivated by conventional methods.

  6. 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........

  7. Sphingomonas sp. is a novel cell culture contaminant.

    Science.gov (United States)

    Asghar, Muhammad Tahir; Al-Ghanim, K; Mahboob, Shahid; Sharif, Muhammad; Nazir, Jawad; Shakoori, Abdul Rauf

    2015-06-01

    A novel contaminant was isolated from Madin Darby Bovine Kidney (MDBK) cells. The organism was unable to grow on standard microbiological media by conventional techniques, but grew well in Dulbecco's Modified Eagle's Medium (DMEM) containing high glucose concentration. The organism formed a white biofilm on the bottom without any signs of turbidity. Upon genome sequence analysis of 16 S rDNA, the contaminant was identified as Sphingomonas sp. Shah, a member of the group α-Proteobacteria. Neutral red dye uptake method confirmed clear cytotoxic potential of the bacterium on A-549 cells. The organism was capable of invading and infecting different mammalian cell lines: MDBK, ZZ-R, 293-T, A549, and HeLa cells. Infected cells showed a variety of cytopathic effects including vacuolation at perinuclear area, cytoplasmic granulation and membrane blebbing. Microscopic analysis of the infected cells revealed the presence of cytoplasmic vacuoles harboring motile organisms. Apparently local serum preparations seem to be the source of this contamination, which is imperceptibly passed on from one culture passage to the other and ultimately leading to serious cytopathic manifestations.

  8. Growth of hybridoma cells in serum-free medium: ethanolamine is an essential component.

    OpenAIRE

    Murakami, H.; Masui, H; Sato, G H; Sueoka, N; Chow, T P; Kano-Sueoka, T

    1982-01-01

    A serum-free medium supplemented with a few growth factors was devised to grow lymphocyte hybridomas. The medium was developed with the hybridoma line MPC11-BL, a fusion product between a mouse plasmacytoma cell line (MPC11TG70na3) and mouse (BALB/c) spleen cells. In the process of developing the medium, ethanolamine was found to be an essential growth factor for the hybridoma. Phosphoethanolamine at 10-fold higher concentration could substitute for ethanolamine. Long-term cultivation of the ...

  9. Medium-intensity acute exhaustive exercise induces neural cell apoptosis in the rat hippocampus★

    OpenAIRE

    Li, Shanni; Jin LIU; Yan, Hengmei

    2013-01-01

    The present study assessed the influence of medium-intensity (treadmill at a speed of 19.3 m/min until exhaustion) and high-intensity (treadmill at a speed of 26.8 m/min until exhaustion) acute exhaustive exercise on rat hippocampal neural cell apoptosis. TUNEL staining showed significantly increased neural cell apoptosis in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise, particularly the medium-intensity acute exhaustive exercise, when compared ...

  10. Influences of Plant Growth Regulators,Basal Media and Carbohydrate Levels on Cell Suspension Culture of Panax ginseng

    Institute of Scientific and Technical Information of China (English)

    TangWei; WuJiongyuan; 等

    1995-01-01

    A cell suspension culture of Panax ginseng which may be continuously subcultured has been established.Embryogenic callus derived from clutured young leaves was used to initiate the culture,Plant growth regulators,basal medium formula and carbohydrate levels were examined to determine their various effects on suspension culture cell growth and development ,The best selection of plant growth regulator,basal medium and carbohydrate level is 2mg/L 2,4-D:0.5mg/L KT,MS and 3% sucrose respectively.

  11. 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, ...

  12. Influence of the Addition of Riboflavin in Culture Medium on Delivering Biomass Using Yeast Strains of Saccharomyces Carlsbengensis

    Directory of Open Access Journals (Sweden)

    Cornelia Nicoară

    2010-05-01

    Full Text Available Yeasts requirements for growth factors should be considered both in terms of ability to summarize the simpleaverage and the dependence on external supplies. Vitamins are components of coenzymes or enzymes prostheticgroups and thus they are growth factors for yeast. The study concerns about the influence of the addition ofriboflavin in culture medium in different quantities, the accumulation of yeast biomass under the action of yeaststrains of beer. The process of cultivation has been made for 24 hours at a temperature of 220C. The addition ofriboflavin in culture medium of yeast biomass increased in each strain of yeast compared with the witness - thesample without added riboflavin. Biomass obtained by follow this procedure could be used to create new foodproducts with high ration nutritional value.

  13. Efficient flotation of yeast cells grown in batch culture.

    Science.gov (United States)

    Palmieri, M C; Greenhalf, W; Laluce, C

    1996-05-01

    A fast flotation assay was used to select new floating yeast strains. The flotation ability did not seem to be directly correlated to total extracellular protein concentration of the culture. However, the hydrophobicity of the cell was definitely correlated to the flotation capacity. The Saccharomyces strains (FLT strains) were highly hydrophobic and showed an excellent flotation performance in batch cultures without additives (flotation agents) and with no need for a special flotation chamber or flotation column. A stable and well-organized structure was evident in the dried foam as shown by scanning electron microscopy which revealed its unique structure showing mummified cells (dehydrated) attached to each other. The attachment among the cells and the high protein concentration of the foams indicated that proteins might be involved in the foam formation. The floating strains (strains FLT) which were not flocculent and showed no tendency to aggregate, were capable of growing and producing ethanol in a synthetic medium containing high glucose concentration as a carbon source. The phenomenon responsible for flotation seems to be quite different from the flocculation phenomenon. PMID:18626952

  14. Nonlinear Dielectric Properties of Yeast Cells Cultured in Different Environmental Conditions

    Science.gov (United States)

    Kawanishi, Gomon; Fukuda, Naoki; Muraji, Masafumi

    The harmonics of the electric current through yeast suspensions, the nonlinear dielectric properties of yeast cells, have particular patterns according to the biological activity of the cells and the measurement of these patterns is a technique for determining the activity of living cells. The concentration of glucose and oxygen in yeast culture medium influences the manifestation of fermentation or respiration of yeast cells. Measurements were made with yeast cells (Saccharomyces cerevisiae) cultured aerobically and anaerobically in sufficient glucose concentration, aerobic fermentation and anaerobic fermentation, and aerobically in limited glucose concentration, respiration. The results showed that the harmonics were barely apparent for yeast cells in aerobic fermentation and respiratory; however, cells in the anaerobic fermentation displayed substantial third and fifth harmonics. We can say that environmental condition affects the yeast cells' nonlinear properties, from another viewpoint, the measurements of the nonlinear properties are available to determine the activity of yeast cells adjusted to the conditions of their cultivation.

  15. 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...

  16. A comprehensive characterization of cell cultures and xenografts derived from a human verrucous penile carcinoma

    DEFF Research Database (Denmark)

    Muñoz, Juan J; Drigo, Sandra A; Kuasne, Hellen;

    2016-01-01

    , and cultivated in KSFM/DF12 medium. Cell cultures were evaluated at passage 5 (P5) using migration and invasion assays and were serially propagated, in vivo, in BALB/c nude mice until passage 3 (X1-X3). Immunophenotypic characterization of cultures and xenografts was performed. Genomic (CytoScan HD, Affymetrix...... xenograft origin. Cell cultures and xenografts retained the genomic alterations present in the parental tumor. Compared to VSCC, differentially expressed transcripts detected in all experimental conditions were associated with cellular morphology, movement, and metabolism and organization pathways......This study aimed to establish and characterize primary cell cultures and xenografts derived from penile carcinoma (PeCa) in order to provide experimental models for cellular processes and efficacy of new treatments. A verrucous squamous cell carcinoma (VSCC) was macrodissected, dissociated...

  17. Using tobacco waste extract in pre-culture medium to improve xylose utilization for l-lactic acid production from cellulosic waste by Rhizopus oryzae.

    Science.gov (United States)

    Zheng, Yuxi; Wang, Yuanliang; Zhang, Jianrong; Pan, Jun

    2016-10-01

    The aim of this work was to study the high-titer l-lactic acid production from cellulosic waste using Rhizopus oryzae. The tobacco waste water-extract (TWE) added with 5g/L glucose and 0.1g/L vitamin C was optimized as pre-culture medium for R. oryzae. Results found that compared to traditional pre-culture medium, it improved xylose consumption rate up to 2.12-fold and enhanced l-lactic acid yield up to 1.73-fold. The highest l-lactic acid concentration achieved was 173.5g/L, corresponding to volumetric productivity of 1.45g/Lh and yield of 0.860g/g total reducing sugar in fed-batch fermentation. This process achieves efficient production of polymer-grade l-lactic acid from cellulosic feedstocks, lowers the cost of fungal cell pre-culture and provides a novel way for re-utilization of tobacco waste. PMID:27376833

  18. A simple and efficient method for the long-term preservation of plant cell suspension cultures

    Directory of Open Access Journals (Sweden)

    Boisson Anne-Marie

    2012-01-01

    Full Text Available Abstract Background The repeated weekly subculture of plant cell suspension is labour intensive and increases the risk of variation from parental cells lines. Most of the procedures to preserve cultures are based on controlled freezing/thawing and storage in liquid nitrogen. However, cells viability after unfreezing is uncertain. The long-term storage and regeneration of plant cell cultures remains a priority. Results Sycamore (Acer pseudoplatanus and Arabidopsis cell were preserved over six months as suspensions cultures in a phosphate-free nutrient medium at 5°C. The cell recovery monitored via gas exchange measurements and metabolic profiling using in vitro and in vivo 13C- and 31P-NMR took a couple of hours, and cell growth restarted without appreciable delay. No measurable cell death was observed. Conclusion We provide a simple method to preserve physiologically homogenous plant cell cultures without subculture over several months. The protocol based on the blockage of cell growth and low culture temperature is robust for heterotrophic and semi-autotrophic cells and should be adjustable to cell lines other than those utilised in this study. It requires no specialized equipment and is suitable for routine laboratory use.

  19. 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

  20. Accurate and noninvasive embryos screening during in vitro fertilization (IVF) assisted by Raman analysis of embryos culture medium

    International Nuclear Information System (INIS)

    In combination with morphological evaluation tests, we employ Raman spectroscopy to select higher potential reproductive embryos during in vitro fertilization (IVF) based on chemical composition of embryos culture medium. In this study, 57 Raman spectra are acquired from both higher and lower quality embryos culture medium (ECM) from 10 patients which have been preliminarily confirmed by clinical assay. Data are fit by using a linear combination model of least squares method in which 12 basis spectra represent the chemical features of ECM. The final fitting coefficients provide insight into the chemical compositions of culture medium samples and are subsequently used as criterion to evaluate the quality of embryos. The relative fitting coefficients ratios of sodium pyruvate/albumin and phenylalanine/albumin seem act as key roles in the embryo screening, attaining 85.7% accuracy in comparison with clinical pregnancy. The good results demonstrate that Raman spectroscopy therefore is an important candidate for an accurate and noninvasive screening of higher quality embryos, which potentially decrease the time-consuming clinical trials during IVF

  1. Pre-culturing of nodal explants in thidiazuron supplemented liquid medium improves in vitro shoot multiplication of Cassia angustifolia.

    Science.gov (United States)

    Siddique, I; Abdullwahab Bukhari, N; Perveen, K; Siddiqui, I; Anis, M

    2013-09-01

    An in vitro propagation system for Cassia angustifolia Vahl. has been developed. Due to the presence of sennosides, the demand of this plant has increased manyfold in global market. Multiple shoots were induced by culturing nodal explants excised from mature plants on a liquid Murashige and Skoog [8] medium supplemented with 5-100 μM of thidiazuron (TDZ) for different treatment duration (4, 8, 12 and 16 d). The optimal level of TDZ supplemented to the culture medium was 75 μM for 12 d induction period followed by subculturing in MS medium devoid of TDZ as it produced maximum regeneration frequency (87%), mean number of shoots (9.6 ± 0.33) and shoot length (4.4 ± 0.46 cm) per explant. A culture period longer than 12 d with TDZ resulted in the formation of fasciated or distorted shoots. Ex vitro rooting was achieved when the basal cut end of regenerated shoots was dipped in 200 μM indole-3-butyric acid (IBA) for half an hour followed by their transplantation in plastic pots filled with sterile soilrite where 85% plantlets grew well and all exhibited normal development. The present findings describe an efficient and rapid plant regeneration protocol that can further be used for genetic transformation studies.

  2. Influence of auxins and sucrose in monoterpenoid oxindole alkaloid production by Uncaria tomentosa cell suspension cultures.

    Science.gov (United States)

    Luna-Palencia, Gabriela R; Cerda-García-Rojas, Carlos M; Rodríguez-Monroy, Mario; Ramos-Valdivia, Ana C

    2005-01-01

    Growth and alkaloid production in Uncaria tomentosa cell suspension cultures were studied in Murashige and Skoog medium supplemented with 10 microM 2,4-dichlorophenoxyacetic acid, 10 microM kinetin, and 58 mM sucrose for maintenance and with 10 microM indole-3-acetic acid, 10 microM kinetin, and 58 mM sucrose for production. A U. tomentosa pale Uth-3 cell line, cultured in the production medium, showed a reduced lag phase and a specific growth rate (mu) of 0.27 day(-1), while cells growing in the maintenance medium showed mu = 0.20 day(-1). U. tomentosa cells growing in the production medium produced monoterpenoid oxindole alkaloids (MOA) in amounts of 10.2 +/- 1.6 microg g(-1) dry weight (DW). The chemical profile of MOA produced by in vitro cell cultures was similar to that found in the plant. After 10 subcultures, maximum MOA production decreased to 2.0 +/- 0.7 microg g(-1) DW, while tryptamine alkaloids (TA) were produced with a maximum of 6.2 +/- 0.4 microg g(-1) DW. The increase of initial sucrose concentration up to 145 mM in the production medium enhanced the cell biomass by 3.2-fold (from 10.2 +/- 0.1 to 32.8 +/- 1.1 g DW L(-1)), reduced mu from 0.27 to 0.23 day(-1), and provoked a substantial accumulation of TA (23.1 +/- 4.7 microg g(-1) DW). A high sucrose concentration stimulated MOA production in the maintenance medium (2.7 +/- 0.5 microg g(-1) DW), even in the presence of 2,4-dichlorophenoxyacetic acid.

  3. The presence of cortical neurons in striatal-cortical co-cultures alters the effects of dopamine and BDNF on Medium Spiny Neuron dendritic development

    Directory of Open Access Journals (Sweden)

    Rachel D Penrod

    2015-07-01

    Full Text Available Medium spiny neurons (MSNs are the major striatal neuron and receive synaptic input from both glutamatergic and dopaminergic afferents. These synapses are made on MSN dendritic spines, which undergo density and morphology changes in association with numerous disease and experience-dependent states. Despite wide interest in the structure and function of mature MSNs, relatively little is known about MSN development. Furthermore, most in vitro studies of MSN development have been done in simple striatal cultures that lack any type of non-autologous synaptic input, leaving open the question of how MSN development is affected by a complex environment that includes other types of neurons, glia, and accompanying secreted and cell-associated cues. Here we characterize the development of MSNs in striatal-cortical co-culture, including quantitative morphological analysis of dendritic arborization and spine development, describing progressive changes in density and morphology of developing spines. Overall, MSN growth is much more robust in the striatal-cortical co-culture compared to striatal mono-culture. Inclusion of dopamine in the co-culture further enhances MSN dendritic arborization and spine density, but the effects of dopamine on dendritic branching are only significant at later times in development. In contrast, exogenous Brain Derived Neurotrophic Factor (BDNF has only a minimal effect on MSN development in the co-culture, but significantly enhances MSN dendritic arborization in striatal mono-culture. Importantly, inhibition of NMDA receptors in the co-culture significantly enhances the effect of exogenous BDNF, suggesting that the efficacy of BDNF depends on the cellular environment. Combined, these studies identify specific periods of MSN development that may be particularly sensitive to perturbation by external factors and demonstrate the importance of studying MSN development in a complex signaling environment.

  4. Solutions to academic failure: The cognitive and cultural realities ofEnglish as the medium of instruction among black learners

    Directory of Open Access Journals (Sweden)

    R. Gamaroff

    2013-02-01

    Full Text Available In South Africa, black learners who are speakers of Bantu languages have to use a second language, namely English, as the medium of instruction from Std 3 onwards. The differences between English language-culture and Bantu languages-culture(s have generated a host of problems (and pseudo-problems?, where the main problem is academic failure. Three solutions to academic failure are discussed in the light of cultural and cognitive factors in multicultural education: 1. The use of the mother tongue as the exclusive medium of instruction 2. Critical Language Study (CLS and People's English 3. The separation of high ability learners from limited ability learners in the teaching situation. It is emphasised that culture is closely connected to a symbolic system, and thus an understanding of cognitive processes in academic learning requires an understanding of culture, and vice versa. Ultimately, of primary importance in academic study are the cognitive underpinnings of Cognitive Academic Language Proficiency (CALP developed in the first language. In Suid-Afrika word swart leerders wie se moedertaal een van die Afrika tale is, tans vanaf st. 3 in 'n tweede taal, naamlik Engels, onderrig. As gevolg van die verskille tussen die Engelse taalkultuur en die taalkulture van die A.frika tale het daar 'n groot aantal probleme (en pseudoprobleme? ontstaan, waarvan akademiese mislukking die belangrikste is. Drie oplossings vir hierdie akademiese mislukking word bespreek aan die hand van kulturele en kognitiewe faktore in multikulturele onderwys: 1. Die gebruik van die moedertaal as eksklusiewe medium van onderrig 2. "Critical Language Study" (CLS en "People's English" 3. Die afsonderlike hantering van hoogsbegaafde en minder begaafde leerlinge. Dit moet beklemtoon word dat kultuur nouverwant is aan 'n simbolesisteem. Gevolglik is 'n be grip van die kognitiewe prosesse betrokke by akademiese leer 'n voorvereiste vir 'n be grip van kultuur, en omgekeerd. Vera

  5. The Influence of the Intangible Cultural Heritage About propagation medium in the Big data Era

    Directory of Open Access Journals (Sweden)

    Jin Zhuo

    2016-01-01

    Full Text Available Film and television works have been attracting the attention of the modern people with their own characteristics. Image, vivid, intuitive, popular, the media uses its own advantage to spread out the information quickly, the transmission efficiency and the promotion is unmatched by other any medias, it also plays an important role for the dissemination of intangible cultural heritage. With our country pay more attention on protecting the intangible cultural heritage, digital media can use its own advantages to protect, promote, promote the intangible cultural heritage, to visual intangible cultural heritage, and present it to the public, in order to let the public understand what is intangible cultural heritage, take the initiative to protect the intangible cultural heritage. This paper discusses the influence of digital media on protecting intangible cultural heritage, with the power of digital media, we expect to protect it better, leave a valuable legacy for future generations.

  6. 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.

  7. Effect of nitrogen concentration in culture mediums on growth and enzyme production of Phanerochaete chrysosporium

    Institute of Scientific and Technical Information of China (English)

    GAO Da-wen; WEN Xiang-hua; QIAN Yi

    2005-01-01

    Effect of different nitrogen concentration in the mediums on growth and enzyme production of Phanerochaete chrysosporium was studied when glucose concentration was 10 g/L. The results showed that the medium contained 0.8 g/L ammonium tartrate is the best. It not only supply abundant nutrients for the growth of Phanerochaete chrysosporium, which make mycelia the best grow compared with the other medium, but also produce higher manganese-dependent peroxidase(Mnp) and laccase(Lac) activity. In addition, it is observed that the variation of mycelia surface is related to ligninolytic enzyme secreted by Phanerochaete chrysosporium. When the surface of mycelium pellets appeared burs, it predicts secondary metabolism begin. This experimentation demonstrated that when the ratio of carbon and nitrogen in nitrogen limited medium is equal to 100:8, growth and enzyme production of Phanerochaete chrysosporium is the best, it could achieve the maximum Mnp and Lac activity.

  8. Insect cell culture in research: Indian scenario.

    Science.gov (United States)

    Sudeep, A B; Mourya, D T; Mishra, A C

    2005-06-01

    Insect cell cultures are widely used in viral diagnosis and biotechnology, for the production of recombinant proteins, viral pesticides and vaccines as well as in basic research in genetics, molecular biology, biochemistry, endocrinology and virology. Following KRP Singh's pioneering research in 1967, a large number of cell lines from diptera, hemiptera, and lepidopteran insects were established and characterized in India. With the availability of the modern tools in molecular biology and the advancements made in biotechnology, the indigenous cell lines may prove useful in creating a future without biohazardous chemical pesticides as well as producing life saving pharmaceuticals and vaccines for many diseases. This review summarizes information gathered regarding the insect cell lines established so far in India. It also covers the familiarization of the well characterized continuous cell lines and their potential applications. Special attention is given to virus susceptibility of the cell lines, the yield of virus with a comparative analysis with other conventional systems. The potential applications of dipteran and lepidopteran cell lines in agriculture and biotechnology are also briefly discussed for prospective studies.

  9. Cytotoxicity effects of amiodarone on cultured cells.

    Science.gov (United States)

    Golli-Bennour, Emna El; Bouslimi, Amel; Zouaoui, Olfa; Nouira, Safa; Achour, Abdellatif; Bacha, Hassen

    2012-07-01

    Amiodarone is a potent anti-arrhythmic drug used for the treatment of cardiac arrhythmias. Although, the effects of amiodarone are well characterized on post-ischemic heart and cardiomyocytes, its toxicity on extra-cardiac tissues is still poorly understood. To this aim, we have monitored the cytotoxicity effects of this drug on three cultured cell lines including hepatocytes (HepG2), epithelial cells (EAhy 926) and renal cells (Vero). We have investigated the effects of amiodarone on (i) cell viabilities, (ii) heat shock protein expressions (Hsp 70) as a parameter of protective and adaptive response and (iii) oxidative damage.Our results clearly showed that amiodarone inhibits cell proliferation, induces an over-expression of Hsp 70 and generates significant amount of reactive oxygen species as measured by lipid peroxidation occurrence. However, toxicity of amiodarone was significantly higher in renal and epithelial cells than in hepatocytes. Vitamin E supplement restores the major part of cell mortalities induced by amiodarone showing that oxidative damage is the predominant toxic effect of the drug.Except its toxicity for the cardiac system, our findings demonstrated that amiodarone can target other tissues. Therefore, kidneys present a high sensibility to this drug which may limit its use with subjects suffering from renal disorders.

  10. In vitro Culture of Bone Marrow Mesenchymal Stem Cells in Rats and Differentiation into Retinal Neural-like Cells

    Institute of Scientific and Technical Information of China (English)

    SUN Xufang; JIANG Huanrong; YANG Hong

    2007-01-01

    In order to study the in vitro culture and expansion of bone marrow mesenchymal stem cells in rats (rMSCs) and the possibility of rMSCs differentiation into retinal neural cells, the bone marrow-derived cells in SD rats were isolated and cultured in vitro. The retinal neural cells in SD rats were cultured and the supernatants were collected to prepare conditioned medium. The cultured rMSCs were induced to differentiate by two steps. Imrnunofluorescence method and anti-nestin, anti-NeuN, anti-GFAP and anti-Thy1.1 antibodies were used to identify the cells derived from the rMSCs. The results showed that the in vitro cultured rMSCs grew well and expanded quickly. After induction with two conditioned media, rMSCs was induced to differentiate into neural progenitor cells, then into retinal neural-like cells which were positive for nestin, NeuN, GFAP and Thy1.1 de-tected by fluorescence method. The findings suggested that rMSCs could be culture and expanded in vitro, and induced to differentiate into retinal neural-like cells.

  11. Neutrophil-induced transmigration of tumour cells treated with tumour-conditioned medium is facilitated by granulocyte-macrophage colony-stimulating factor.

    LENUS (Irish Health Repository)

    Wu, Q D

    2012-02-03

    OBJECTIVE: To investigate the effect of different cytokines that are present in tumour-conditioned medium on human neutrophil (PMN)-induced tumour cell transmigration. DESIGN: Laboratory study. SETTING: University hospital, Ireland. MATERIAL: Isolated human PMN and cultured human breast tumour cell line, MDA-MB-231. Interventions: Human PMN treated with either tumour-conditioned medium or different media neutralised with monoclonal antibodies (MoAb), and MDA-MB-231 cells were plated on macrovascular and microvascular endothelial monolayers in collagen-coated transwells to assess migration of tumour cells. MAIN OUTCOME MEASURES: Cytokines present in tumour-conditioned medium, PMN cytocidal function and receptor expression, and tumour cell transmigration. RESULTS: tumour-conditioned medium contained high concentrations of granulocyte-macrophage colony-stimulating factor (GM-CSF), vascular endothelial growth factor (VEGF), and interleukin 8 (IL-8), but not granulocyte colony-stimulating factor (G-CSF) and interleukin 3 (IL-3). Anti-GM-CSF MoAb significantly reduced PMN-induced transmigration of tumour cells treated with tumour-conditioned medium (p < 0.05), whereas anti-VEGF and anti-IL-8 MoAbs did not affect their migration. In addition, anti-GM-CSF MoAb, but not anti-VEGF or anti-IL-8 MoAb, reduced PMN CD11b and CD18 overexpression induced by tumour-conditioned medium (p < 0.05). CONCLUSION: These results indicate that the GM-CSF that is present in tumour-conditioned medium may be involved, at least in part, in alterations in PMN function mediated by the medium and subsequently PMN-induced transmigration of tumour cells.

  12. Tick cell culture isolation and growth of Rickettsia raoultii from Dutch Dermacentor reticulatus ticks.

    Science.gov (United States)

    Alberdi, M Pilar; Nijhof, Ard M; Jongejan, Frans; Bell-Sakyi, Lesley

    2012-12-01

    Tick cell lines play an important role in research on ticks and tick-borne pathogenic and symbiotic microorganisms. In an attempt to derive continuous Dermacentor reticulatus cell lines, embryo-derived primary cell cultures were set up from eggs laid by field ticks originally collected as unfed adults in The Netherlands and maintained for up to 16 months. After several months, it became evident that cells in the primary cultures were infected with a Rickettsia-like intracellular organism. Supernatant medium containing some D. reticulatus cells was inoculated into cultures of 2 Rhipicephalus (Boophilus) microplus cell lines, BME/CTVM2 and BME/CTVM23, where abundant growth of the bacteria occurred intracellularly on transfer to both cell lines. Bacterial growth was monitored by light (live, inverted microscope, Giemsa-stained cytocentrifuge smears) and transmission electron microscopy revealing heavy infection with typical intracytoplasmic Rickettsia-like bacteria, not present in uninfected cultures. DNA was extracted from bacteria-infected and uninfected control cultures, and primers specific for Rickettsia 16S rRNA, ompB, and sca4 genes were used to generate PCR products that were subsequently sequenced. D. reticulatus primary cultures and both infected tick cell lines were positive for all 3 Rickettsia genes. Sequencing of PCR products revealed 99-100% identity with published Rickettsia raoultii sequences. The R. raoultii also grew abundantly in the D. nitens cell line ANE58, poorly in the D. albipictus cell line DALBE3, and not at all in the D. andersoni cell line DAE15. In conclusion, primary tick cell cultures and cell lines are useful systems for isolation and propagation of fastidious tick-borne microorganisms. In vitro isolation of R. raoultii from Dutch D. reticulatus confirms previous PCR-based detection in field ticks, and presence of the bacteria in the tick eggs used to initiate the primary cultures confirms that transovarial transmission of this

  13. Comparative Analysis of KnockOut™ Serum with Fetal Bovine Serum for the In Vitro Long-Term Culture of Human Limbal Epithelial Cells

    OpenAIRE

    Shaokun Zhang; Zaoxia Liu; Guanfang Su; Hong Wu

    2016-01-01

    The limbal epithelial cells can be maintained on 3T3 feeder layer with fetal bovine serum supplemented culture medium, and these cells have been used to successfully treat limbal stem cell deficiency. However, fetal bovine serum contains unknown components and displays quantitative and qualitative lot-to-lot variations. To improve the culture condition, the defined KnockOut serum replacement was investigated to replace fetal bovine serum for culturing human limbal epithelial cell. Human prima...

  14. 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...

  15. Optimization of flask culture medium and conditions for hyaluronic acid production by a streptococcus equisimilis mutant nc2168

    Directory of Open Access Journals (Sweden)

    Yong-Hao Chen

    2012-12-01

    Full Text Available A mutant designated NC2168, which was selected from wild-type Streptococcus equisimilis CVCC55116by ultraviolet ray combined with60Co-γ ray treatment and does not produce streptolysin, was employed to produce hyaluronic acid (HA. In order to increase the output of HA in a flask, the culture medium and conditions for NC2168 were optimized in this study. The influence of culture medium ingredients including carbon sources, nitrogen sources and metal ions on HA production was evaluated using factional factorial design. The mathematical model, which represented the effect of each medium component and their interaction on the yield of HA, was established by the quadratic rotary combination design and response surface method. The model estimated that, a maximal yield of HA could be obtained when the concentrations of yeast extract, peptone, glucose, and MgSO4 were set at 3 g/100 mL, 2 g/100 mL, 0.5 g/100 mL and 0.15 g/100 mL, respectively. Compared with the values obtained by other runs in the experimental design, the optimized medium resulted in a remarkable increase in the output of HA and the maximum of the predicted HA production was 174.76 mg/L. The model developed was accurate and reliable for predicting the production of HA by NC2168.Cultivation conditions were optimized by an orthogonal experimental design and the optimal conditions were as follows: temperature 33ºC, pH 7.8, agitation speed 200 rpm, medium volume 20 mL.

  16. Nanoscale imaging of untreated mammalian cells in a medium with low radiation damage using scanning electron-assisted dielectric microscopy

    Science.gov (United States)

    Okada, Tomoko; Ogura, Toshihiko

    2016-07-01

    Imaging of untreated living cells in a medium at a nanometre-scale resolution under physiological conditions is a significant challenge. Scanning electron microscopy (SEM) is widely used to observe cells in various atmospheric holders or special equipment. However, untreated biological specimens in aqueous solution generally incur heavy radiation damage from the direct electron beam (EB); and these images exhibit very poor contrast. Therefore, a new method for generating high-contrast images of living cells under physiological conditions without radiation damage has been strongly desired. Here, we demonstrate the first nanoscale observation of living cultured mammalian cells using our newly developed scanning-electron assisted dielectric microscopy (SE-ADM) method with a culture dish holder. Using the difference in relative permittivity between water and specimens, our SE-ADM system aids in the visualisation of untreated biological samples in aqueous solution. In addition, specimens incurred only a low level of radiation damage because the tungsten (W)-coated silicon nitride (SiN) film absorbs irradiated electrons. Untreated cells and organelles are clearly visible in high-contrast and high-resolution images without staining and fixation. Furthermore, our method enables the detection of changes in organelle structures within cells via time-lapse imaging with minimal radiation damage.

  17. Nanoscale imaging of untreated mammalian cells in a medium with low radiation damage using scanning electron-assisted dielectric microscopy

    Science.gov (United States)

    Okada, Tomoko; Ogura, Toshihiko

    2016-01-01

    Imaging of untreated living cells in a medium at a nanometre-scale resolution under physiological conditions is a significant challenge. Scanning electron microscopy (SEM) is widely used to observe cells in various atmospheric holders or special equipment. However, untreated biological specimens in aqueous solution generally incur heavy radiation damage from the direct electron beam (EB); and these images exhibit very poor contrast. Therefore, a new method for generating high-contrast images of living cells under physiological conditions without radiation damage has been strongly desired. Here, we demonstrate the first nanoscale observation of living cultured mammalian cells using our newly developed scanning-electron assisted dielectric microscopy (SE-ADM) method with a culture dish holder. Using the difference in relative permittivity between water and specimens, our SE-ADM system aids in the visualisation of untreated biological samples in aqueous solution. In addition, specimens incurred only a low level of radiation damage because the tungsten (W)-coated silicon nitride (SiN) film absorbs irradiated electrons. Untreated cells and organelles are clearly visible in high-contrast and high-resolution images without staining and fixation. Furthermore, our method enables the detection of changes in organelle structures within cells via time-lapse imaging with minimal radiation damage. PMID:27375121

  18. 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.

  19. Fabrication of mouse embryonic stem cell-derived layered cardiac cell sheets using a bioreactor culture system.

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    Katsuhisa Matsuura

    Full Text Available Bioengineered functional cardiac tissue is expected to contribute to the repair of injured heart tissue. We previously developed cardiac cell sheets using mouse embryonic stem (mES cell-derived cardiomyocytes, a system to generate an appropriate number of cardiomyocytes derived from ES cells and the underlying mechanisms remain elusive. In the present study, we established a cultivation system with suitable conditions for expansion and cardiac differentiation of mES cells by embryoid body formation using a three-dimensional bioreactor. Daily conventional medium exchanges failed to prevent lactate accumulation and pH decreases in the medium, which led to insufficient cell expansion and cardiac differentiation. Conversely, a continuous perfusion system maintained the lactate concentration and pH stability as well as increased the cell number by up to 300-fold of the seeding cell number and promoted cardiac differentiation after 10 days of differentiation. After a further 8 days of cultivation together with a purification step, around 1 × 10(8 cardiomyocytes were collected in a 1-L bioreactor culture, and additional treatment with noggin and granulocyte colony stimulating factor increased the number of cardiomyocytes to around 5.5 × 10(8. Co-culture of mES cell-derived cardiomyocytes with an appropriate number of primary cultured fibroblasts on temperature-responsive culture dishes enabled the formation of cardiac cell sheets and created layered-dense cardiac tissue. These findings suggest that this bioreactor system with appropriate medium might be capable of preparing cardiomyocytes for cell sheet-based cardiac tissue.

  20. Uptake and metabolism of sugars by suspension-cultured catharanthus roseus cells

    Energy Technology Data Exchange (ETDEWEB)

    Ashihara, Hiroshi; Sagishima, Kyoko; Kubota, Kaoru (Ochanomizu Univ., Tokyo (Japan))

    1989-04-01

    The Uptake and metabolism of sugars by suspension-cultured Catharanthus roseus cells were investigated. Substantially all the sucrose in the culture medium was hydrolyzed to glucose and fructose before being taken up by the cells. The activity of invertase bound to cell walls, determined in situ, was high at the early stage of culture. Glucose was more easily taken up by the cells than was fructose. Tracer experiments using (U-{sup 14}C)glucose and (U-{sup 14}C)fructose indicated that glucose is a better precursor for respiration than fructose, while fructose is preferentially utilized for the synthesis of sucrose, especially in the early phase of cell growth. These results suggest that fructose is utilized for the synthesis of sucrose via the reaction catalyzed by sucrose synthase, prior to the phosphorylation by hexokinase or fructokinase.

  1. Bystander effects induced by medium from carbon-ion irradiated human cancer cells

    Institute of Scientific and Technical Information of China (English)

    WANG Ju-Fang; LI Wen-Jian; ZHOU Guang-Ming; DANG Bing-Rong; MA Qiu-Feng; FENG Yan

    2005-01-01

    The bystander effects induced by medium from human hepatoma SMMC-7721 and adenocarcinoma F56 cells irradiated with carbon ions were investigated. It was found that the survival fraction (SF) of the irradiated cells decreased exponentially along with the increased dose. SMMC-7721 cells were more radiosensitive than F56 cells.The plating efficiency (PE) of the non-irradiated cells treated with irradiated conditioned medium (ICM) was obviously lower than the PE of control cells for SMMC-7721 cells but not for F56 cells. Moreover, the reduced PE and SF by ICM treatment were more significant for 1Gy irradiation than for 6Gy irradiation on SMMC-7721 cells. These results suggest that the irradiated cells can secrete factor(s) into medium that is cytotoxic to bystander non-irradiated cells. The bystander effects are dependent on cell genotype presented at the time of irradiation and radiation dose.This makes impact on the precise estimation of the effects of radiation and tumor radiotherapy.

  2. 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)

  3. The Kinetic of Growth Cell of Neurospora Sitophila at Phythohormone Production Medium

    International Nuclear Information System (INIS)

    The study of growth cell kinetic of Neurospora Sitophila at Phythohormone production medium has been done. The growth of this mould at Mendel broth medium is affected by pH and glucose concentration. In the enriched medium by 2 % glucose content and pH 4.5 shows the optimal growth of cell with specific growth rate by 0.0785 per hour. The changing of pH connected with the growth cell curve, so this moment as an indicator of cell harvest time. At the stationary phase to dead phase is explained by pH changed between 2.80-2.85. This phase may be estimated as a time of Phythohormone synthesize by mold. (author)

  4. Urban secondary sewage: an alternative medium for the culture of Tetraselmis chuii (Prasinophyceae and Dunaliella viridis (Chlorophyceae

    Directory of Open Access Journals (Sweden)

    Rauquírio André Albuquerque Marinho da Costa

    2004-07-01

    Full Text Available The effect of different concentrations (10, 20, 30 and 40% of urban secondary sewage on the growth of Tetraselmis chuii (Prasinophyceae and Dunaliella viridis (Chlorophyceae was examined to verify the possibility of its use as an alternative culture medium for these species. Stocks and 700mL duplicate batch cultures were maintained under controlled laboratory conditions. 30% and 40% additions were the most efficient for the growth of both species. Statistical analysis of the adjusted growth (cell.mL-1 and biomass (µg chlorophyll-a.mL-1 curves performed using the Chi-square test (pFoi estudado o efeito das diferentes concentrações de esgoto urbano secundário (10%, 20%, 30% e 40% sobre o crescimento de Tetraselmis chuii (Prasinophyceae e Dunaliellla viridis (Chlorophyceae para verificar a possibilidade do seu uso como meio de cultura altenativo para estas espécies. Culturas stocks duplicatas de 700mL foram mantidas sob condições de laboratório controladas. As adições de 30% e 40% foram as mais eficientes para o crescimento de ambas as espécies. Análises estatísticas para o ajuste do crescimento (cel.mL-1 e curvas de biomassa (µg clorofila-a.mL-1 em relação ao tempo de cultura desenvolvidas usando Chi-square test (p<0.05, demonstraram diferenças significativas entre o meio nutritivo f/2 e a adição de 40% usada para estas duas espécies. Os resultados sugerem que algumas das concentrações testadas, rendem densidades celulares e taxas de crescimento satisfatórias quando comparados com outros meios de cultura (macroalga, estratos de esterco bovino e de galinha sendo um meio altenativo que pode ser usado para sistemas de aquicultura semi intensivos devido aos baixos custos e fácil aquisição.

  5. Semi-selective culture medium for Exserohilum turcicum isolation from corn seeds

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    Roberto Luis De Rossi

    2014-06-01

    Full Text Available Northern corn leaf blight, caused by Exserohilum turcicum (Et, is a disease of widespread occurrence in regions where corn, sweetcorn and popcorn are grown. This disease has great potential to cause damage and has been studied for years, but the association of its causal agent with seeds remains unconfirmed. Thus, the availability of a sensitive method to detect and quantify the inoculum in seeds, even at low incidence, is essential. The aim of this study was to develop a method to detect and quantify the presence of the fungus infecting and infesting corn and popcorn seeds. Artificially and naturally infected seeds were employed to develop the medium. The semi-selective medium was composed of carbendazim (active ingredient (60 mg/L, captan (30 mg/L, streptomycin sulfate (500 mg/L and neomycin sulfate (600 mg/L aggregated to the medium lactose casein hydrolysate agar medium. By using this, Et was detected in naturally infected corn seeds, showing 0.124% incidence, in four out of ten analyzed samples. In addition, 1.04 conidia were detected per infested seed. By means of isolation, pathogenicity test, morphological characterization and comparison with descriptions of the species in the literature, the fungus isolated from the seeds was confirmed to be Et. Both infection and infestation were considered low; thus, for studies of Et detection in corn seeds, the use of semi-selective medium and more than 1,200 seeds/sample is suggested.

  6. α1-antitrypsin and its C-terminal fragment attenuate effects of degranulated neutrophil-conditioned medium on lung cancer HCC cells, in vitro

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    Westin Ulla

    2004-11-01

    Full Text Available Abstract Background Tumor microenvironment, which is largely affected by inflammatory cells, is a crucial participant in the neoplastic process through promotion of cell proliferation, survival and migration. We measured the effects of polymorphonuclear neutrophil (PMN conditioned medium alone, and supplemented with serine proteinase inhibitor α-1 antitrypsin (AAT or its C-terminal fragment (C-36 peptide, on cultured lung cancer cells. Methods Lung cancer HCC cells were grown in a regular medium or in a PMN-conditioned medium in the presence or absence of AAT (0.5 mg/ml or its C-36 peptide (0.06 mg/ml for 24 h. Cell proliferation, invasiveness and release of IL-8 and VEGF were analyzed by [3H]-thymidine incorporation, Matrigel invasion and ELISA methods, respectively. Results Cells exposed to PMN-conditioned medium show decreased proliferation and IL-8 release by 3.9-fold, p Conclusions Our data provide evidence that neutrophil derived factors decrease lung cancer HCC cell proliferation and IL-8 release, but increase cell invasiveness. These effects were found to be modulated by exogenously present serine proteinase inhibitor, AAT, and its C-terminal fragment, which points to a complexity of the relationships between tumor cell biological activities and local microenvironment.

  7. Production of Fusaric Acid by Fusarium spp. in Pure Culture and in Solid Medium Co-Cultures

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    Nadine Bohni

    2016-03-01

    Full Text Available The ability of fungi isolated from nails of patients suffering from onychomycosis to induce de novo production of bioactive compounds in co-culture was examined. Comparison between the metabolite profiles produced by Sarocladium strictum, by Fusarium oxysporum, and by these two species in co-culture revealed de novo induction of fusaric acid based on HRMS. Structure confirmation of this toxin, using sensitive microflow NMR, required only three 9-cm Petri dishes of fungal culture. A targeted metabolomics study based on UHPLC-HRMS confirmed that the production of fusaric acid was strain-dependent. Furthermore, the detected toxin levels suggested that onychomycosis-associated fungal strains of the F. oxysporum and F. fujikuroi species complexes are much more frequently producing fusaric acid, and in higher amount, than strains of the F. solani species complex. Fusarium strains producing no significant amounts of this compound in pure culture, were shown to de novo produce that compound when grown in co-culture. The role of fusaric acid in fungal virulence and defense is discussed.

  8. Production of Fusaric Acid by Fusarium spp. in Pure Culture and in Solid Medium Co-Cultures.

    Science.gov (United States)

    Bohni, Nadine; Hofstetter, Valérie; Gindro, Katia; Buyck, Bart; Schumpp, Olivier; Bertrand, Samuel; Monod, Michel; Wolfender, Jean-Luc

    2016-03-18

    The ability of fungi isolated from nails of patients suffering from onychomycosis to induce de novo production of bioactive compounds in co-culture was examined. Comparison between the metabolite profiles produced by Sarocladium strictum, by Fusarium oxysporum, and by these two species in co-culture revealed de novo induction of fusaric acid based on HRMS. Structure confirmation of this toxin, using sensitive microflow NMR, required only three 9-cm Petri dishes of fungal culture. A targeted metabolomics study based on UHPLC-HRMS confirmed that the production of fusaric acid was strain-dependent. Furthermore, the detected toxin levels suggested that onychomycosis-associated fungal strains of the F. oxysporum and F. fujikuroi species complexes are much more frequently producing fusaric acid, and in higher amount, than strains of the F. solani species complex. Fusarium strains producing no significant amounts of this compound in pure culture, were shown to de novo produce that compound when grown in co-culture. The role of fusaric acid in fungal virulence and defense is discussed.

  9. Isolation, culture and characterization of porcine primordial germ cells%猪原始生殖细胞的分离、培养与鉴定

    Institute of Scientific and Technical Information of China (English)

    李世杰; 郑瑞珍; 岳奎忠; 陈瑛; 杨增明

    2003-01-01

    Embryonic germ cells (EG cells) are pluripotential undifferentiated stem cells isolated from cultured primordial germ cells (PGCs). Like ES cells, EG cells are of importance for gene targeting, therapeutical cloning and organ trans-plantation. The aim of this study was to isolate and characterize EG cells from porcine PGCs. The genital ridges from 24- 26 days old porcine embryos were treated in 0.02% EDTA for 20 min and pricked with a needle to release PGCs. The isolated PGCs were cultured on a SNL feeder layer in an EG cell medium. The EG cell medium consisted of Dulbecco''s modified Eagle'' s medium (DMEM) supplemented with 20 % Buffalo rat lever (BRL) cell-conditioned medium, 15 % fe-tal bovine serum, 1 mmol L-glutamine, 0.1 mol nonessential amino acids, 10 μmol β-mercaptoethanol and antibiotics.The freshly isolated PGCs were positive for alkaline phosphatase activity and Periodic acid-Schiff'' s staining. Under this culture regime, PGCs could be maintained in an undifferentiated state and used for further cultures. One strain of the cul-tured PGCs was cultured 8 times, and alkaline phosphatase activity was detected in the colony formed from this strain.These cultured PGCs could spontaneously differentiate into fibroblast-like cells. These data suggested that we had success-fully isolated EG-like cells from oorcine PGCs.

  10. Culture conditions for yellow pigment formation byMonascus sp. KB 10 grown on cassava medium.

    Science.gov (United States)

    Yongsmith, B; Tabloka, W; Yongmanitchai, W; Bavavoda, R

    1993-01-01

    An isolate ofMonascus, from a commercial, fermented soybean curd (sufu) was grown on a cassava medium. With medium at an initial pH of 7.0 an orange-red pigmentation was produced but with an initial pH below 4, a light golden pigment was obtained. A medium containing, w/v, 3% cassava starch, 0.4% peptone and 0.1% glutamic acid, with an initial pH of 2.5 was optimal for the production of this yellow pigment, which had a single maximum absorption spectrum at 330 nm. The spectroscopic characterization of the purified yellow pigments demonstrated a monascin-ankaflavin-monascorubrin skeleton.

  11. Application of Different Culture Medium in Onion Tissue Culture%不同培养基在洋葱组培中的应用

    Institute of Scientific and Technical Information of China (English)

    潘美红; 陈振泰; 薛萍; 杨海峰

    2012-01-01

    为完善洋葱组培技术,针对不同的洋葱品种,使用不同的基础培养基,添加不同的生长调节剂进行培养,结果发现:洋葱品种早春黄在培养基MS+6 mg,L-16-BA+0.1 mg,L-1NAA上增殖效果最好,连葱6号在培养基3g·L1花宝+6 mg,L-16-BA+0.1 mg,L-1NAA上增殖效果最好;在继代培养过程中添加0.25 mg,L-1多效唑可以提高增殖率.%In order to improve onion (Allium cepa L. ) tissue culture technology, this study used different basal medium, added different growth regulators on different onion varieties. The results showed that the onion variety ' Zaochunhuang' in culture medium of MS+6 mg · L-1 6-BA+0.1 mg · L-1 NAA had the best proliferation effect. The onion variety ' Liancong No. 6' in culture medium of 3 g · L-1 Hyponex +6 mg · L-1 6-BA +0.1 mg · L NAA had the best proliferation effect. In successive transfer culture, adding 0.25 mg · L paclobutrazol could raise the proliferation rate.

  12. Evaluation of dry sheet medium culture plate (Compactdry TC) method for determining numbers of bacteria in food samples.

    Science.gov (United States)

    Mizuochi, S; Kodaka, H

    2000-05-01

    The Compactdry, a ready-to-use and self-diffusible dry medium sheet culture system, has been developed by the Nissui Pharmaceutical Co. Ltd. for enumerating bacteria in food. The Compactdry consists of special spread sheet with culture medium that is the same as standard method nutrients, a cold water-soluble gelling agent, and a unique plastic dish. The procedure for bacterial examination in a sample solution (1 ml) is to just inoculate a test solution into the center of the self-diffusible medium and incubate at 35 degrees C for 48 h. The Compactdry TC (CTC) for the enumeration of total aerobic bacteria from 97 food samples was compared with the standard plate count (SPC) method and 3M Petrifilm aerobic count plates (PAC). The correlation coefficients between the CTC and SPC method, the CTC and PAC, and the PAC and SPC method were 0.97, 0.99, and 0.97, respectively. The Compactdry system is useful for the enumeration of total aerobic bacteria in food and may be a possible suitable alternative to the conventional pour-plate or the Petrifilm plate methods.

  13. Influence of culture medium on the production of eif antigen from Leishmania chagasi in recombinant Escherichia coli

    Directory of Open Access Journals (Sweden)

    Michelle Rossana Ferreira Vaz

    2011-12-01

    Full Text Available With the advent of recombinant DNA technology, recombinant protein expression has become an important tool in the study of the structure, function and identification of new proteins, especially those with therapeutic functions. Escherichia coli has been the predominant prokaryote used in genetic engineering studies due to the abundance of information about its metabolism. Despite significant advances in molecular biology and immunology of infections, there are as yet no prophylactic drugs capable of preventing visceral leishmaniasis. It is therefore important to identify specific antigens in order to develop vaccines and diagnostic kits against this disease. The objective of this study was to evaluate the influence of culture medium on the production of eIF antigen from Leishmania chagasi in recombinant Escherichia coli. An induction procedure using IPTG was carried out in a series of trials, to observe the influence of culture medium (2xTY, TB under expression of the recombinant eIF protein. Results showed that recombinant protein expression was associated to growth and that the highest eIF antigen expression was obtained in the 2xTY medium.

  14. Influence of the Culture Medium in Dose-Response Effect of the Chlorhexidine on Streptococcus mutans Biofilms.

    Science.gov (United States)

    de Queiroz, Vanessa Salvadego; Ccahuana-Vásquez, Renzo Alberto; Tedesco, Alcides Fabiano; Lyra, Luzia; Cury, Jaime Aparecido; Schreiber, Angélica Zaninelli

    2016-01-01

    The aim of this study was to evaluate the influence of culture medium on dose-response effect of chlorhexidine (CHX) on Streptococcus mutans UA159 biofilm and validate the use of the cation-adjusted-Müller-Hinton broth (MH) for the evaluation of antibacterial activity. Ultrafiltered Tryptone-Yeast Extract Broth (UTYEB) was compared against MH and MH with blood supplementation (MHS). For each medium, six groups (n = 4) were assessed: two negative control groups (baseline 48 and 120 h) and four experimental groups (0.0001, 0.001, 0.012, and 0.12% CHX). S. mutans biofilm grew on glass slides of each media containing 1% sucrose. After 48 h of growth, biofilms of baseline 48 h were collected and the other groups were treated for 1 min, twice a day, for 3 days, with their respective treatments. The media were changed daily and pH was measured. After 120 h, biofilms were collected and dry weight and viable microorganisms were determined. Results showed CHX dose-response effect being observed in all media for all the variables. However, MH and MHS showed higher sensitivity than UTYEB (p culture medium does influence dose-response effect of CHX on Streptococcus mutans biofilm and that MH can be used for antibacterial activity.

  15. Influence of the Culture Medium in Dose-Response Effect of the Chlorhexidine on Streptococcus mutans Biofilms

    Science.gov (United States)

    de Queiroz, Vanessa Salvadego; Ccahuana-Vásquez, Renzo Alberto; Tedesco, Alcides Fabiano; Lyra, Luzia; Cury, Jaime Aparecido; Schreiber, Angélica Zaninelli

    2016-01-01

    The aim of this study was to evaluate the influence of culture medium on dose-response effect of chlorhexidine (CHX) on Streptococcus mutans UA159 biofilm and validate the use of the cation-adjusted-Müller-Hinton broth (MH) for the evaluation of antibacterial activity. Ultrafiltered Tryptone-Yeast Extract Broth (UTYEB) was compared against MH and MH with blood supplementation (MHS). For each medium, six groups (n = 4) were assessed: two negative control groups (baseline 48 and 120 h) and four experimental groups (0.0001, 0.001, 0.012, and 0.12% CHX). S. mutans biofilm grew on glass slides of each media containing 1% sucrose. After 48 h of growth, biofilms of baseline 48 h were collected and the other groups were treated for 1 min, twice a day, for 3 days, with their respective treatments. The media were changed daily and pH was measured. After 120 h, biofilms were collected and dry weight and viable microorganisms were determined. Results showed CHX dose-response effect being observed in all media for all the variables. However, MH and MHS showed higher sensitivity than UTYEB (p < 0.05). We can conclude that the culture medium does influence dose-response effect of CHX on Streptococcus mutans biofilm and that MH can be used for antibacterial activity. PMID:27293967

  16. DNA MUTAGENESIS IN PANAX GINSENG CELL CULTURES

    Directory of Open Access Journals (Sweden)

    Kiselev K.V.

    2012-08-01

    Full Text Available At the present time, it is well documented that plant tissue culture induces a number of mutations and chromosome rearrangements termed “somaclonal variations”. However, little is known about the nature and the molecular mechanisms of the tissue culture-induced mutagenesis and the effects of long-term subculturing on the rate and specific features of the mutagenesis. The aim of the present study was to investigate and compare DNA mutagenesis in different genes of Panax ginseng callus cultures of different age. It has previously been shown that the nucleotide sequences of the Agrobacterium rhizogenes rolC locus and the selective marker nptII developed mutations during long-term cultivation of transgenic cell cultures of P. ginseng. In the present work, we analyzed nucleotide sequences of selected plant gene families in a 2-year-old and 20-year-old P. ginseng 1c cell culture and in leaves of cultivated P. ginseng plants. We analysed sequence variability between the Actin genes, which are a family of house-keeping genes; the phenylalanine ammonia-lyase (PAL and dammarenediol synthase (DDS genes, which actively participate in the biosynthesis of ginsenosides; and the somatic embryogenesis receptor kinase (SERK genes, which control plant development. The frequency of point mutations in the Actin, PAL, DDS, and SERK genes in the 2-year-old callus culture was markedly higher than that in cultivated plants but lower than that in the 20-year-old callus culture of P. ginseng. Most of the mutations in the 2- and 20-year-old P. ginseng calli were A↔G and T↔C transitions. The number of nonsynonymous mutations was higher in the 2- and 20-year-old callus cultures than the number of nonsynonymous mutations in the cultivated plants of P. ginseng. Interestingly, the total number of N→G or N→C substitutions in the analyzed genes was 1.6 times higher than the total number of N→A or N→T substitutions. Using methylation-sensitive DNA fragmentation

  17. Production Pattern of Ajmalicine in Catharanthus roseus (L.) G. Don. Cell Aggregates Culture in the Airlift Bioreactor

    OpenAIRE

    RIZKITA RACHMI ESYANTI; AIDA MUSPIAH

    2006-01-01

    A research has been conducted to optimize the rate of aeration and initial weight of cell aggregates in the production of ajmalicine in Catharanthus roseus cell culture in airlift bioreactor. Catharanthus roseus culture were grown in Zenk medium with the addition of 2.50 x 10-6 M naphthalene acetic acid (NAA) and 10-5 M benzyl amino purine (BAP). Cell aggregates were sub-cultured two times before transferring 20 and 30 g/fw of cell aggregates into bioreactor, respectively, and aerated with th...

  18. Innovative culture, management control systems and performance in small and medium-sized Spanish family firms

    Directory of Open Access Journals (Sweden)

    Antonio Duréndez

    2011-06-01

    Full Text Available The aim of this paper is to research on organizational culture and management of family firms. We identify family-firms innovative culture and assess the relationship between organizational culture, management control systems (MCS use and their effects on performance of SME family-firms. With this purpose, we carry out an empirical analysis on a sample of Spanish SMEs (285 family and 151 non-family firms. Results show that (1 family-firms have a more hierarchical culture and a lesser extent of MCS use than non-family firms have, and (2 an innovative culture and the use of MCS have positive influences on family-firm performance.

  19. Teaching Sociolinguistics: A Medium for Cultural Awareness of Indonesian University Foreign Language Learners

    Directory of Open Access Journals (Sweden)

    Agnes Herawati

    2014-05-01

    Full Text Available This paper tries to show the evidences that indicate how teaching Sociolinguistics can result in a number of valuable outcomes, including helping students understand and appreciate other cultures different from theirs. Sociolinguistics provides useful examples of language usage in different genres, including how culture influences people in using a language. The opportunities of learning other cultures through language will take the students to the higher level of appreciation of the culture of the target language. To determine how this outcome can be achieved in the language classrooms, this paper provides a review of closely connected literature about how to bridge the gap between cultures in particular. However, to increase its completeness and relevance, this paper also provides some research results that reveal how teaching Sociolinguistics has taken its new applicability and importance, and furthermore adds the effects on how students become more proficient and enthusiastic about their learning.

  20. Medium-intensity acute exhaustive exercise induces neural cell apoptosis in the rat hippocampus

    Institute of Scientific and Technical Information of China (English)

    Shanni Li; Jin Liu; Hengmei Yan

    2013-01-01

    The present study assessed the influence of medium-intensity (treadmill at a speed of 19.3 m/min until exhaustion) and high-intensity (treadmill at a speed of 26.8 m/min until exhaustion) acute exhaustive exercise on rat hippocampal neural cell apoptosis. TUNEL staining showed significantly increased neural cell apoptosis in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise, particularly the medium-intensity acute exhaustive exercise, when compared with the control. Immunohistochemistry showed significantly increased expression of the antiapoptotic protein Bcl-2 and the proapoptotic protein Bax in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise. Additionally, the ratio of Bax to Bcl-2 increased in both exercise groups. In particular, the medium-intensity acute exhaustive exercise group had significantly higher Bax and Bcl-2 protein expression and a higher Bax/Bcl-2 ratio. These findings indicate that acute exhaustive exercise of different intensities can induce neural cell apoptosis in the hippocampus, and that medium-intensity acute exhaustive exercise results in greater damage when compared with high-intensity exercise.

  1. Medium-intensity acute exhaustive exercise induces neural cell apoptosis in the rat hippocampus.

    Science.gov (United States)

    Li, Shanni; Liu, Jin; Yan, Hengmei

    2013-01-15

    The present study assessed the influence of medium-intensity (treadmill at a speed of 19.3 m/min until exhaustion) and high-intensity (treadmill at a speed of 26.8 m/min until exhaustion) acute exhaustive exercise on rat hippocampal neural cell apoptosis. TUNEL staining showed significantly increased neural cell apoptosis in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise, particularly the medium-intensity acute exhaustive exercise, when compared with the control. Immunohistochemistry showed significantly increased expression of the antiapoptotic protein Bcl-2 and the proapoptotic protein Bax in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise. Additionally, the ratio of Bax to Bcl-2 increased in both exercise groups. In particular, the medium-intensity acute exhaustive exercise group had significantly higher Bax and Bcl-2 protein expression and a higher Bax/Bcl-2 ratio. These findings indicate that acute exhaustive exercise of different intensities can induce neural cell apoptosis in the hippocampus, and that medium-intensity acute exhaustive exercise results in greater damage when compared with high-intensity exercise. PMID:25206482

  2. 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…

  3. 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....

  4. Human cultured cells are capable to incorporate isolated plant mitochondria loaded with exogenous DNA

    Directory of Open Access Journals (Sweden)

    Laktionov P. P.

    2012-07-01

    Full Text Available Aim. To investigate the possibility of human cultured cells to incorporate isolated mitochondria together with exogenous DNA introduced into organelles. Methods. Two approaches were used for this purpose, fluorescent labelling of mitochondria and/or DNA with subsequent analysis of the cells subjected to incubation by microscopy or by quantitative PCR. Results. We have shown that human cultured cells lines, HeLa and HUVEC, are capable to uptake isolated plant mitochondria and that this process depends on the incubation time and concentration of organelles present in medium. The incorporated mitochondria can serve as vehicles to deliver exogenous DNA into human cells, this DNA is then distributed in different cell compartments. Conclusions. These results are preliminary and need further investigations, including testing the possibility of human cells to incorporate the mitochondria of human or animal origin and creating genetic construction which could provide certain selectivity or stability of the transferred exogenous DNA upon cell uptake of the mitochondria as vectors.

  5. Effects of calcitriol, seocalcitol, and medium-chain triglyceride on a canine transitional cell carcinoma cell line

    DEFF Research Database (Denmark)

    Kaewsakhorn, T.; Kisseberth, W.C.; Capen, C.C.;

    2005-01-01

    Background: Transitional cell carcinoma (TCC) in dogs is associated with high morbidity and mortality. Calcitriol and its analog seocalcitol, combined with medium-chain triglyceride (MCT), have potential for the treatment of this disease. Materials and Methods: TCC cells were treated with calcitr......Background: Transitional cell carcinoma (TCC) in dogs is associated with high morbidity and mortality. Calcitriol and its analog seocalcitol, combined with medium-chain triglyceride (MCT), have potential for the treatment of this disease. Materials and Methods: TCC cells were treated...

  6. Transdifferentiation of mouse adipose-derived stromal cells into acinar cells of the submandibular gland using a co-culture system

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jingu; Park, Sangkyu; Roh, Sangho, E-mail: sangho@snu.ac.kr

    2015-05-15

    A loss of salivary gland function often occurs after radiation therapy in head and neck tumors, though secretion of saliva by the salivary glands is essential for the health and maintenance of the oral environment. Transplantation of salivary acinar cells (ACs), in part, may overcome the side effects of therapy. Here we directly differentiated mouse adipose-derived stromal cells (ADSCs) into ACs using a co-culture system. Multipotent ADSCs can be easily collected from stromal vascular fractions of adipose tissues. The isolated ADSCs showed positive expression of markers such as integrin beta-1 (CD29), cell surface glycoprotein (CD44), endoglin (CD105), and Nanog. The cells were able to differentiate into adipocytes, osteoblasts, and neural-like cells after 14 days in culture. ADSCs at passage 2 were co-cultured with mouse ACs in AC culture medium using the double-chamber (co-culture system) to avoid mixing the cell types. The ADSCs in this co-culture system expressed markers of ACs, such as α-amylases and aquaporin5, in both mRNA and protein. ADSCs cultured in AC-conditioned medium also expressed AC markers. Cellular proliferation and senescence analyses demonstrated that cells in the co-culture group showed lower senescence and a higher proliferation rate than the AC-conditioned medium group at Days 14 and 21. The results above imply direct conversion of ADSCs into ACs under the co-culture system; therefore, ADSCs may be a stem cell source for the therapy for salivary gland damage. - Highlights: • ADSCs could transdifferentiate into acinar cells (ACs) using ACs co-culture (CCA). • Transdifferentiated ADSCs expressed ACs markers such as α-amylase and aquaporin5. • High proliferation and low senescence were presented in CCA at Day 14. • Transdifferentiation of ADSCs into ACs using CCA may be an appropriate method for cell-based therapy.

  7. Transdifferentiation of mouse adipose-derived stromal cells into acinar cells of the submandibular gland using a co-culture system

    International Nuclear Information System (INIS)

    A loss of salivary gland function often occurs after radiation therapy in head and neck tumors, though secretion of saliva by the salivary glands is essential for the health and maintenance of the oral environment. Transplantation of salivary acinar cells (ACs), in part, may overcome the side effects of therapy. Here we directly differentiated mouse adipose-derived stromal cells (ADSCs) into ACs using a co-culture system. Multipotent ADSCs can be easily collected from stromal vascular fractions of adipose tissues. The isolated ADSCs showed positive expression of markers such as integrin beta-1 (CD29), cell surface glycoprotein (CD44), endoglin (CD105), and Nanog. The cells were able to differentiate into adipocytes, osteoblasts, and neural-like cells after 14 days in culture. ADSCs at passage 2 were co-cultured with mouse ACs in AC culture medium using the double-chamber (co-culture system) to avoid mixing the cell types. The ADSCs in this co-culture system expressed markers of ACs, such as α-amylases and aquaporin5, in both mRNA and protein. ADSCs cultured in AC-conditioned medium also expressed AC markers. Cellular proliferation and senescence analyses demonstrated that cells in the co-culture group showed lower senescence and a higher proliferation rate than the AC-conditioned medium group at Days 14 and 21. The results above imply direct conversion of ADSCs into ACs under the co-culture system; therefore, ADSCs may be a stem cell source for the therapy for salivary gland damage. - Highlights: • ADSCs could transdifferentiate into acinar cells (ACs) using ACs co-culture (CCA). • Transdifferentiated ADSCs expressed ACs markers such as α-amylase and aquaporin5. • High proliferation and low senescence were presented in CCA at Day 14. • Transdifferentiation of ADSCs into ACs using CCA may be an appropriate method for cell-based therapy

  8. Phenomenological modelling and simulation of cell clusters in 3D cultures.

    Science.gov (United States)

    González-Valverde, I; Semino, C; García-Aznar, J M

    2016-10-01

    Cell clustering and aggregation are fundamental processes in the development of several tissues and the progression of many diseases. The formation of these aggregates also has a direct impact on the oxygen concentration in their surroundings due to cellular respiration and poor oxygen diffusion through clusters. In this work, we propose a mathematical model that is capable of simulating cell cluster formation in 3D cultures through combining a particle-based and a finite element approach to recreate complex experimental conditions. Cells are modelled considering cell proliferation, cell death and cell-cell mechanical interactions. Additionally, the oxygen concentration profile is calculated through finite element analysis using a reaction-diffusion model that considers cell oxygen consumption and diffusion through the extracellular matrix and the cell clusters. In our model, the local oxygen concentration in the medium determines both cell proliferation and cell death. Numerical predictions are also compared with experimental data from the literature. The simulation results indicate that our model can predict cell clustering, cluster growth and oxygen distribution in 3D cultures. We conclude that the initial cell distribution, cell death and cell proliferation dynamics determine the size and density of clusters. Moreover, these phenomena are directly affected by the oxygen transport in the 3D culture. PMID:27615191