WorldWideScience

Sample records for cell culture harvest

  1. Discovering and differentiating new and emerging clonal populations of Chlamydia trachomatis with a novel shotgun cell culture harvest assay.

    Science.gov (United States)

    Somboonna, Naraporn; Mead, Sally; Liu, Jessica; Dean, Deborah

    2008-03-01

    Chlamydia trachomatis is the leading cause of preventable blindness and bacterial sexually transmitted diseases worldwide. Plaque assays have been used to clonally segregate laboratory-adapted C. trachomatis strains from mixed infections, but no assays have been reported to segregate clones from recent clinical samples. We developed a novel shotgun cell culture harvest assay for this purpose because we found that recent clinical samples do not form plaques. Clones were strain-typed by using outer membrane protein A and 16S rRNA sequences. Surprisingly, ocular trachoma reference strain A/SA-1 contained clones of Chlamydophila abortus. C. abortus primarily infects ruminants and pigs and has never been identified in populations where trachoma is endemic. Three clonal variants of reference strain Ba/Apache-2 were also identified. Our findings reflect the importance of clonal isolation in identifying constituents of mixed infections containing new or emerging strains and of viable clones for research to more fully understand the dynamics of in vivo strain-mixing, evolution, and disease pathogenesis.

  2. Treatment and electricity harvesting from sulfate/sulfide-containing wastewaters using microbial fuel cell with enriched sulfate-reducing mixed culture

    International Nuclear Information System (INIS)

    Lee, Duu-Jong; Lee, Chin-Yu; Chang, Jo-Shu

    2012-01-01

    Highlights: ► We started up microbial fuel cell (MFC) using enriched sulfate-reducing mixed culture. ► Sulfate-reducing bacteria and anode-respiring bacteria were enriched in anodic biofilms. ► The MFC effectively remove sulfate to elementary sulfur in the presence of lactate. ► The present device can treat sulfate laden wastewaters with electricity harvesting. - Abstract: Anaerobic treatment of sulfate-laden wastewaters can produce excess sulfide, which is corrosive to pipelines and is toxic to incorporated microorganisms. This work started up microbial fuel cell (MFC) using enriched sulfate-reducing mixed culture as anodic biofilms and applied the so yielded MFC for treating sulfate or sulfide-laden wastewaters. The sulfate-reducing bacteria in anodic biofilm effectively reduced sulfate to sulfide, which was then used by neighboring anode respiring bacteria (ARB) as electron donor for electricity production. The presence of organic carbons enhanced MFC performance since the biofilm ARB were mixotrophs that need organic carbon to grow. The present device introduces a route for treating sulfate laden wastewaters with electricity harvesting.

  3. Treatment and electricity harvesting from sulfate/sulfide-containing wastewaters using microbial fuel cell with enriched sulfate-reducing mixed culture

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Duu-Jong, E-mail: cedean@mail.ntust.edu.tw [Department of Chemical Engineering, National Taiwan University, Taipei, Taiwan (China); Department of Chemical Engineering, National Taiwan University of Science and Technology, Taipei, Taiwan (China); Lee, Chin-Yu [Department of Chemical Engineering, National Taiwan University, Taipei, Taiwan (China); Chang, Jo-Shu [Department of Chemical Engineering, National Cheng Kung University, Tainan, Taiwan (China); Center for Bioscience and Biotechnology, National Cheng Kung University, Tainan, Taiwan (China); Research Center for Energy Technology and Strategy, National Cheng Kung University, Tainan, Taiwan (China)

    2012-12-15

    Highlights: Black-Right-Pointing-Pointer We started up microbial fuel cell (MFC) using enriched sulfate-reducing mixed culture. Black-Right-Pointing-Pointer Sulfate-reducing bacteria and anode-respiring bacteria were enriched in anodic biofilms. Black-Right-Pointing-Pointer The MFC effectively remove sulfate to elementary sulfur in the presence of lactate. Black-Right-Pointing-Pointer The present device can treat sulfate laden wastewaters with electricity harvesting. - Abstract: Anaerobic treatment of sulfate-laden wastewaters can produce excess sulfide, which is corrosive to pipelines and is toxic to incorporated microorganisms. This work started up microbial fuel cell (MFC) using enriched sulfate-reducing mixed culture as anodic biofilms and applied the so yielded MFC for treating sulfate or sulfide-laden wastewaters. The sulfate-reducing bacteria in anodic biofilm effectively reduced sulfate to sulfide, which was then used by neighboring anode respiring bacteria (ARB) as electron donor for electricity production. The presence of organic carbons enhanced MFC performance since the biofilm ARB were mixotrophs that need organic carbon to grow. The present device introduces a route for treating sulfate laden wastewaters with electricity harvesting.

  4. Harvest of Plasmodium falciparum merozoites from continuous culture.

    Science.gov (United States)

    Mrema, J E; Campbell, G H; Jaramillo, A L; Miranda, R; Rieckmann, K H

    1979-01-01

    Spontaneously released merozoites were harvested from cultures in which 42-90% of the erythrocytes had been infected with mature forms of Plasmodium falciparum at the start of incubation. The mature forms had been extracted from asynchronous cultures by the use of Ficoll and Plasmagel gradients. As the mature forms consisted of both trophozoites and schizonts, merozoites were released into the culture medium over a long period of time. The synchrony of merozoite release did not appear to be improved by prior exposure of parasites to sorbitol. Over this prolonged period of incubation, the yield of merozoites was disappointingly low in cultures containing 2.5% of erythrocytes. At erythrocyte concentrations of 0.01-0.25%, 3-10 times more merozoites were released into the medium; 0.4-2.3 merozoites per initial mature form were harvested over a 15-19-hour period. In addition to merozoites, contents of the culture medium included intact erythrocytes, ghost cells, and other cellular fragments. Only intact erythrocytes were effectively removed from the medium by simple or Ficoll gradient centrifugation. Merozoite preparations that are free from host cellular material are important in the development of a human malaria vaccine.

  5. Allogeneic Peripheral Blood Stem Cell Harvest

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. Allogeneic Peripheral Blood Stem Cell Harvest. Mobilization protocol. G-CSF 10 mcg/Kg / day for 5 days. Pheresis. Cobe Spectra; Haemonetics mcs+. Enumeration. CD34 counts; Cfu-GM assays.

  6. Microalgal Mass Culture Room Harvest Records

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The microalgal mass culture room, housed at the NOAA Fisheries' Milford CT laboratory, provides research grade microalgae (phytoplankton) to in-house and...

  7. Applications of chitosan-based thermo-sensitive copolymers for harvesting living cell sheet

    International Nuclear Information System (INIS)

    Chen, J.-P.; Yang, T.-F.

    2008-01-01

    A thermo-sensitive chitosan-based copolymer hydrogel was used for harvesting living cell sheets. The hydrogel was tested for harvesting 3T3 cells after carrying out cell culture at 37 deg. C and incubating the confluent cells at 20 deg. C for spontaneous detachment of cell sheets from hydrogel surface without enzyme treatment. Results from cell viability assay and microscopy observations demonstrated that cells could attach to the hydrogel surface and maintain high viability and proliferation ability. Cell detachment efficiency from the hydrogel was about 80%. The detached cell sheet retained high viability and could proliferate again after transferred to a new culture surface

  8. Microalgae Harvest through Fungal Pelletization—Co-Culture of Chlorella vulgaris and Aspergillus niger

    Directory of Open Access Journals (Sweden)

    Sarman Oktovianus Gultom

    2014-07-01

    Full Text Available Microalgae harvesting is a labor- and energy-intensive process and new approaches to harvesting microalgae need to be developed in order to decrease the costs. In this study; co-cultivatation of filamentous fungus (Aspergillus niger and microalgae (Chlorella vulgaris to form cell pellets was evaluated under different conditions, including organic carbon source (glucose; glycerol; and sodium acetate concentration; initial concentration of fungal spores and microalgal cells and light. Results showed that 2 g/L of glucose with a 1:300 ratio of fungi to microalgae provided the best culturing conditions for the process to reach >90% of cell harvest efficiency. The results also showed that an organic carbon source was required to sustain the growth of fungi and form the cell pellets. The microalgae/fungi co-cultures at mixotrophic conditions obtained much higher total biomass than pure cultures of each individual strains; indicating the symbiotic relationship between two strains. This can benefit the microbial biofuel production in terms of cell harvest and biomass production.

  9. Insect Cell Culture

    NARCIS (Netherlands)

    Oers, van M.M.; Lynn, D.E.

    2010-01-01

    Insect cell cultures are widely used in studies on insect cell physiology, developmental biology and microbial pathology. In particular, insect cell culture is an indispensable tool for the study of insect viruses. The first continuously growing insect cell cultures were established from

  10. Periodic harvesting of embryonic stem cells from a hollow-fiber membrane based four-compartment bioreactor.

    Science.gov (United States)

    Knöspel, Fanny; Freyer, Nora; Stecklum, Maria; Gerlach, Jörg C; Zeilinger, Katrin

    2016-01-01

    Different types of stem cells have been investigated for applications in drug screening and toxicity testing. In order to provide sufficient numbers of cells for such in vitro applications a scale-up of stem cell culture is necessary. Bioreactors for dynamic three-dimensional (3D) culture of growing cells offer the option for culturing large amounts of stem cells at high densities in a closed system. We describe a method for periodic harvesting of pluripotent stem cells (PSC) during expansion in a perfused 3D hollow-fiber membrane bioreactor, using mouse embryonic stem cells (mESC) as a model cell line. A number of 100 × 10(6) mESC were seeded in bioreactors in the presence of mouse embryonic fibroblasts (MEF) as feeder cells. Over a cultivation interval of nine days cells were harvested by trypsin perfusion and mechanical agitation every second to third culture day. A mean of 380 × 10(6) mESC could be removed with every harvest. Subsequent to harvesting, cells continued growing in the bioreactor, as determined by increasing glucose consumption and lactate production. Immunocytochemical staining and mRNA expression analysis of markers for pluripotency and the three germ layers showed a similar expression of most markers in the harvested cells and in mESC control cultures. In conclusion, successful expansion and harvesting of viable mESC from bioreactor cultures with preservation of sterility was shown. The present study is the first one showing the feasibility of periodic harvesting of adherent cells from a continuously perfused four-compartment bioreactor including further cultivation of remaining cells. © 2015 American Institute of Chemical Engineers.

  11. Cell Culture Made Easy.

    Science.gov (United States)

    Dye, Frank J.

    1985-01-01

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

  12. Mammalian Cell Culture Simplified.

    Science.gov (United States)

    Moss, Robert; Solomon, Sondra

    1991-01-01

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

  13. Energy harvesting influences electrochemical performance of microbial fuel cells

    Science.gov (United States)

    Lobo, Fernanda Leite; Wang, Xin; Ren, Zhiyong Jason

    2017-07-01

    Microbial fuel cells (MFCs) can be effective power sources for remote sensing, wastewater treatment and environmental remediation, but their performance needs significant improvement. This study systematically analyzes how active harvesting using electrical circuits increased MFC system outputs as compared to passive resistors not only in the traditional maximal power point (MPP) but also in other desired operating points such as the maximum current point (MCP) and the maximum voltage point (MVP). Results show that active harvesting in MPP increased power output by 81-375% and active harvesting in MCP increased Coulombic efficiency by 207-805% compared with resisters operated at the same points. The cyclic voltammograms revealed redox potential shifts and supported the performance data. The findings demonstrate that active harvesting is a very effective approach to improve MFC performance across different operating points.

  14. 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...

  15. Impulsive control of a continuous-culture and flocculation harvest chemostat model

    Science.gov (United States)

    Zhang, Tongqian; Ma, Wanbiao; Meng, Xinzhu

    2017-12-01

    In this paper, a new mathematical model describing the process of continuous culture and harvest of microalgaes is proposed. By inputting medium and flocculant at two different fixed moments periodically, continuous culture and harvest of microalgaes is implemented. The mathematical analysis is conducted and the whole dynamics of model is investigated by using theory of impulsive differential equations. We find that the model has a microalgaes-extinction periodic solution and it is globally asymptotically stable when some certain threshold value is less than the unit. And the model is permanent when some certain threshold value is larger than the unit. Then, according to the threshold, the control strategies of continuous culture and harvest of microalgaes are discussed. The results show that continuous culture and harvest of microalgaes can be archived by adjusting suitable input time, input amount of medium or flocculant. Finally, some numerical simulations are carried out to verify the control strategy.

  16. Liver Cell Culture Devices

    NARCIS (Netherlands)

    Andria, B.; Bracco, A.; Cirino, G.; Chamuleau, R. A. F. M.

    2010-01-01

    In the last 15 years many different liver cell culture devices, consisting of functional liver cells and artificial materials, have been developed. They have been devised for numerous different applications, such as temporary organ replacement (a bridge to liver transplantation or native liver

  17. PDADMAC flocculation of Chinese hamster ovary cells: enabling a centrifuge-less harvest process for monoclonal antibodies.

    Science.gov (United States)

    McNerney, Thomas; Thomas, Anne; Senczuk, Anna; Petty, Krista; Zhao, Xiaoyang; Piper, Rob; Carvalho, Juliane; Hammond, Matthew; Sawant, Satin; Bussiere, Jeanine

    2015-01-01

    High titer (>10 g/L) monoclonal antibody (mAb) cell culture processes are typically achieved by maintaining high viable cell densities over longer culture durations. A corresponding increase in the solids and sub-micron cellular debris particle levels are also observed. This higher burden of solids (≥15%) and sub-micron particles typically exceeds the capabilities of a continuous centrifuge to effectively remove the solids without a substantial loss of product and/or the capacity of the harvest filtration train (depth filter followed by membrane filter) used to clarify the centrate. We discuss here the use of a novel and simple two-polymer flocculation method used to harvest mAb from high cell mass cell culture processes. The addition of the polycationic polymer, poly diallyldimethylammonium chloride (PDADMAC) to the cell culture broth flocculates negatively-charged cells and cellular debris via an ionic interaction mechanism. Incorporation of a non-ionic polymer such as polyethylene glycol (PEG) into the PDADMAC flocculation results in larger flocculated particles with faster settling rate compared to PDADMAC-only flocculation. PDADMAC also flocculates the negatively-charged sub-micron particles to produce a feed stream with a significantly higher harvest filter train throughput compared to a typical centrifuged harvest feed stream. Cell culture process variability such as lactate production, cellular debris and cellular densities were investigated to determine the effect on flocculation. Since PDADMAC is cytotoxic, purification process clearance and toxicity assessment were performed.

  18. Thermo-responsive poly(N-isopropylacrylamide)-grafted hollow fiber membranes for osteoblasts culture and non-invasive harvest

    Energy Technology Data Exchange (ETDEWEB)

    Zhuang, Meiling, E-mail: zhuangmeiling2006@126.com; Liu, Tianqing, E-mail: liutq@dlut.edu.cn; Song, Kedong, E-mail: kedongsong@dlut.edu.cn; Ge, Dan, E-mail: gedan@dlut.edu.cn; Li, Xiangqin, E-mail: xiangqinli@163.com

    2015-10-01

    Hollow fiber membrane (HFM) culture system is one of the most important bioreactors for the large-scale culture and expansion of therapeutic cells. However, enzymatic and mechanical treatments are traditionally applied to harvest the expanded cells from HFMs, which inevitably causes harm to the cells. In this study, thermo-responsive cellulose acetate HFMs for cell culture and non-invasive harvest were prepared for the first time via free radical polymerization in the presence of cerium (IV). ATR-FTIR and elemental analysis results indicated that the poly(N-isopropylacrylamide) (PNIPAAm) was covalently grafted on HFMs successfully. Dynamic contact angle measurements at different temperatures revealed that the magnitude of volume phase transition was decreased with increasing grafted amount of PNIPAAm. And the amount of serum protein adsorbed on HFMs surface also displayed the same pattern. Meanwhile osteoblasts adhered and spread well on the surface of PNIPAAm-grafted HFMs at 37 °C. And Calcein-AM/PI staining, AB assay, ALP activity and OCN protein expression level all showed that PNIPAAm-grafted HFMs had good cell compatibility. After incubation at 20 °C for 120 min, the adhering cells on PNIPAAm-grafted HFMs turned to be round and detached after being gently pipetted. These results suggest that thermo-responsive HFMs are attractive cell culture substrates which enable cell culture, expansion and the recovery without proteolytic enzyme treatment for the application in tissue engineering and regenerative medicine. - Highlights: • PNIPAAm-grafted HFMs exhibited thermoresponsive characteristic. • The OB cells could adhere and spread well on the surface of PNIPAAm-grafted HFMs. • PNIPAAm-grafted HFMs do not significantly impact ALP activity and OCN protein expression level of OB cells. • Cell could be detached from PNIPAAm-grafted HFMs when temperature decreased from 37 °C to 20 °C.

  19. 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.

  20. 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

  1. Plant cell culture initiation

    NARCIS (Netherlands)

    Hall, R.D.

    2000-01-01

    The use of cultured plant cells in either organized or unorganized form has increased vey considerably in the last 10-15 yr. Many new technologies have been developed and applications in both fundamental and applied research have led to the development of some powerful tools for improving our

  2. An Investigation of Equine Mesenchymal Stem Cell Characteristics from Different Harvest Sites: More Similar Than Not

    Directory of Open Access Journals (Sweden)

    Karla eLombana

    2015-12-01

    Full Text Available Diseases of the musculoskeletal system are a major cause of loss of use and retirement in sport horses. The use of bone marrow derived mesenchymal stem cells (BMDMSCs for healing of traumatized tissue has gained substantial favor in clinical settings and can assist healing and tissue regeneration in orthopaedic injuries. There are two common sites of harvest of BMDMSCs, the sternum and ilium. Our objective was to determine if any differences exist in BMDMSCs acquired from the sternum and the ilium. We compared the two harvest sites in their propensity to undergo multilineage differentiation, differences in cell surface markers or gene transduction efficiencies.BMDMSCs were isolated and culture-expanded from five mL aspirates of bone marrow from sternum and ilium. The cells were then plated and cultured with appropriate differentiation medium to result in multi-lineage differentiation and cell characteristics were compared between sternal and ilial samples. Cell surface antibody expression of CD11a/18, CD34, CD44 and CD90 were evaluated using flow cytometry and gene transduction efficiencies were evaluated using GFP scAAV. There were no statistically significant differences in cell characteristics between MSCs cultured from sternum and ilium under any circumstances.

  3. Semiconductor Nanocrystals as Light Harvesters in Solar Cells

    Directory of Open Access Journals (Sweden)

    Lioz Etgar

    2013-02-01

    Full Text Available Photovoltaic cells use semiconductors to convert sunlight into electrical current and are regarded as a key technology for a sustainable energy supply. Quantum dot-based solar cells have shown great potential as next generation, high performance, low-cost photovoltaics due to the outstanding optoelectronic properties of quantum dots and their multiple exciton generation (MEG capability. This review focuses on QDs as light harvesters in solar cells, including different structures of QD-based solar cells, such as QD heterojunction solar cells, QD-Schottky solar cells, QD-sensitized solar cells and the recent development in organic-inorganic perovskite heterojunction solar cells. Mechanisms, procedures, advantages, disadvantages and the latest results obtained in the field are described. To summarize, a future perspective is offered.

  4. Semiconductor Nanocrystals as Light Harvesters in Solar Cells.

    Science.gov (United States)

    Etgar, Lioz

    2013-02-04

    Photovoltaic cells use semiconductors to convert sunlight into electrical current and are regarded as a key technology for a sustainable energy supply. Quantum dot-based solar cells have shown great potential as next generation, high performance, low-cost photovoltaics due to the outstanding optoelectronic properties of quantum dots and their multiple exciton generation (MEG) capability. This review focuses on QDs as light harvesters in solar cells, including different structures of QD-based solar cells, such as QD heterojunction solar cells, QD-Schottky solar cells, QD-sensitized solar cells and the recent development in organic-inorganic perovskite heterojunction solar cells. Mechanisms, procedures, advantages, disadvantages and the latest results obtained in the field are described. To summarize, a future perspective is offered.

  5. Semiconductor Nanocrystals as Light Harvesters in Solar Cells

    Science.gov (United States)

    Etgar, Lioz

    2013-01-01

    Photovoltaic cells use semiconductors to convert sunlight into electrical current and are regarded as a key technology for a sustainable energy supply. Quantum dot-based solar cells have shown great potential as next generation, high performance, low-cost photovoltaics due to the outstanding optoelectronic properties of quantum dots and their multiple exciton generation (MEG) capability. This review focuses on QDs as light harvesters in solar cells, including different structures of QD-based solar cells, such as QD heterojunction solar cells, QD-Schottky solar cells, QD-sensitized solar cells and the recent development in organic-inorganic perovskite heterojunction solar cells. Mechanisms, procedures, advantages, disadvantages and the latest results obtained in the field are described. To summarize, a future perspective is offered. PMID:28809318

  6. Epithelial Cell Cultures

    Directory of Open Access Journals (Sweden)

    Imran S. Chaudhry

    2011-01-01

    Full Text Available The biological effects of only a finite number of tobacco toxins have been studied. Here, we describe exposure of cultures of human bronchial epithelial cells to low concentrations of tobacco carcinogens: nickel sulphate, benzo(bfluoranthene, N-nitrosodiethylamine, and 4-(methylnitrosamino-1-(3-pyridyl-1-butanone (NNK. After a 24-hour exposure, EGFR was expressed in cell membrane and cytoplasm, BCL-2 was expressed only in the irregular nuclei of large atypical cells, MKI67 was expressed in nuclei with no staining in larger cells, cytoplasmic BIRC5 with stronger nuclear staining was seen in large atypical cells, and nuclear TP53 was strongly expressed in all cells. After only a 24-hour exposure, cells exhibited atypical nuclear and cytoplasmic features. After a 48-hour exposure, EGFR staining was localized to the nucleus, BCL-2 was slightly decreased in intensity, BIRC5 was localized to the cytoplasm, and TP53 staining was increased in small and large cells. BCL2L1 was expressed in both the cytoplasm and nuclei of cells at 24- and 48-hour exposures. We illustrate that short-termexposure of a bronchial epithelial cell line to smoking-equivalent concentrations of tobacco carcinogens alters the expression of key proliferation regulatory genes, EGFR, BCL-2, BCL2L1, BIRC5, TP53, and MKI67, similar to that reported in biopsy specimens of pulmonary epithelium described to be preneoplastic lesions.

  7. 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....

  8. 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.

  9. Iatrogenic giant cell tumor at bone graft harvesting site

    Directory of Open Access Journals (Sweden)

    Zile S Kundu

    2013-01-01

    Full Text Available 30 year old female patient with giant cell tumor of the distal tibia initially treated at a peripheral nononcological center by curettage and autologous bone grafting from the ipsilateral iliac crest reported to us with local recurrence and an implantation giant cell tumor at the graft harvesting site which required extensive surgeries at both sites. The risk of iatrogenic direct implantation of tumor, often attributable to inadequate surgical planning or poor surgical techniques, and the steps to prevent such complication is reported here.

  10. Recent developments in production and purification of malaria antigens: Harvest of Plasmodium falciparum merozoites from continuous culture*

    Science.gov (United States)

    Mrema, J. E. K.; Campbell, G. H.; Jaramillo, A. L.; Miranda, R.; Rieckmann, K. H.

    1979-01-01

    Spontaneously released merozoites were harvested from cultures in which 42-90% of the erythrocytes had been infected with mature forms of Plasmodium falciparum at the start of incubation. The mature forms had been extracted from asynchronous cultures by the use of Ficoll and Plasmagel gradients. As the mature forms consisted of both trophozoites and schizonts, merozoites were released into the culture medium over a long period of time. The synchrony of merozoite release did not appear to be improved by prior exposure of parasites to sorbitol. Over this prolonged period of incubation, the yield of merozoites was disappointingly low in cultures containing 2.5% of erythrocytes. At erythrocyte concentrations of 0.01-0.25%, 3-10 times more merozoites were released into the medium; 0.4-2.3 merozoites per initial mature form were harvested over a 15-19-hour period. In addition to merozoites, contents of the culture medium included intact erythrocytes, ghost cells, and other cellular fragments. Only intact erythrocytes were effectively removed from the medium by simple or Ficoll gradient centrifugation. Merozoite preparations that are free from host cellular material are important in the development of a human malaria vaccine. ImagesFig. 3 PMID:397009

  11. Human mixed lymphocyte cultures. Evaluation of microculture technique utilizing the multiple automated sample harvester (MASH)

    Science.gov (United States)

    Thurman, G. B.; Strong, D. M.; Ahmed, A.; Green, S. S.; Sell, K. W.; Hartzman, R. J.; Bach, F. H.

    1973-01-01

    Use of lymphocyte cultures for in vitro studies such as pretransplant histocompatibility testing has established the need for standardization of this technique. A microculture technique has been developed that has facilitated the culturing of lymphocytes and increased the quantity of cultures feasible, while lowering the variation between replicate samples. Cultures were prepared for determination of tritiated thymidine incorporation using a Multiple Automated Sample Harvester (MASH). Using this system, the parameters that influence the in vitro responsiveness of human lymphocytes to allogeneic lymphocytes have been investigated. PMID:4271568

  12. Harvesting, processing and inventory management of peripheral blood stem cells

    Directory of Open Access Journals (Sweden)

    Mijovic Aleksandar

    2007-01-01

    Full Text Available By 2003, 97% autologous transplants and 65% of allogeneic transplants in Europe used mobilised peripheral blood stem cells (PBSC. Soon after their introduction in the early 1990′s, PBSC were associated with faster haemopoietic recovery, fewer transfusions and antibiotic usage, and a shorter hospital stay. Furthermore, ease and convenience of PBSC collection made them more appealing than BM harvests. Improved survival has hitherto been demonstrated in patients with high risk AML and CML. However, the advantages of PBSC come at a price of a higher incidence of extensive chronic GVHD. In order to be present in the blood, stem cells undergo the process of "mobilisation" from their bone marrow habitat. Mobilisation, and its reciprocal process - homing - are regulated by a complex network of molecules on the surface of stem cells and stromal cells, and enzymes and cytokines released from granulocytes and osteoclasts. Knowledge of these mechanisms is beginning to be exploited for clinical purposes. In current practice, stem cell are mobilised by use of chemotherapy in conjunction with haemopoietic growth factors (HGF, or with HGF alone. Granulocyte colony stimulating factor has emerged as the single most important mobilising agent, due to its efficacy and a relative paucity of serious side effects. Over a decade of use in healthy donors has resulted in vast experience of optimal dosing and administration, and safety matters. PBSC harvesting can be performed on a variety of cell separators. Apheresis procedures are nowadays routine, but it is important to be well versed in the possible complications in order to avoid harm to the patient or donor. To ensure efficient collection, harvesting must begin when sufficient stem cells have been mobilised. A rapid, reliable, standardized blood test is essential to decide when to begin harvesting; currently, blood CD34+ cell counting by flow cytometry fulfils these criteria. Blood CD34+ cell counts strongly

  13. Arthroscopic Harvest of Adipose-Derived Mesenchymal Stem Cells From the Infrapatellar Fat Pad.

    Science.gov (United States)

    Dragoo, Jason L; Chang, Wenteh

    2017-11-01

    The successful isolation of adipose-derived mesenchymal stem cells (ADSCs) from the arthroscopically harvested infrapatellar fat pad (IFP) would provide orthopaedic surgeons with an autologous solution for regenerative procedures. To demonstrate the quantity and viability of the mesenchymal stem cell population arthroscopically harvested from the IFP as well as the surrounding synovium. Descriptive laboratory study. The posterior border of the IFP, including the surrounding synovial tissue, was harvested arthroscopically from patients undergoing anterior cruciate ligament reconstruction. Tissue was then collected in an AquaVage adipose canister, followed by fat fractionization using syringe emulsification and concentration with an AdiPrep device. In the laboratory, the layers of tissue were separated and then digested with 0.3% type I collagenase. The pelleted stromal vascular fraction (SVF) cells were then immediately analyzed for viability, mesenchymal cell surface markers by fluorescence-activated cell sorting, and clonogenic capacity. After culture expansion, the metabolic activity of the ADSCs was assessed by an AlamarBlue assay, and the multilineage differentiation capability was tested. The transition of surface antigens from the SVF toward expanded ADSCs at passage 2 was further evaluated. SVF cells were successfully harvested with a mean yield of 4.86 ± 2.64 × 10 5 cells/g of tissue and a mean viability of 69.03% ± 10.75%, with ages ranging from 17 to 52 years (mean, 35.14 ± 13.70 years; n = 7). The cultured ADSCs composed a mean 5.85% ± 5.89% of SVF cells with a mean yield of 0.33 ± 0.42 × 10 5 cells/g of tissue. The nonhematopoietic cells (CD45 - ) displayed the following surface antigens as a percentage of the viable population: CD44 + (52.21% ± 4.50%), CD73 + CD90 + CD105 + (19.20% ± 17.04%), and CD44 + CD73 + CD90 + CD105 + (15.32% ± 15.23%). There was also a significant increase in the expression of ADSC markers CD73 (96.97% ± 1.72%; P

  14. Green grasses as light harvesters in dye sensitized solar cells

    Science.gov (United States)

    Shanmugam, Vinoth; Manoharan, Subbaiah; Sharafali, A.; Anandan, Sambandam; Murugan, Ramaswamy

    2015-01-01

    Chlorophylls, the major pigments presented in plants are responsible for the process of photosynthesis. The working principle of dye sensitized solar cell (DSSC) is analogous to natural photosynthesis in light-harvesting and charge separation. In a similar way, natural dyes extracted from three types of grasses viz. Hierochloe Odorata (HO), Torulinium Odoratum (TO) and Dactyloctenium Aegyptium (DA) were used as light harvesters in dye sensitized solar cells (DSSCs). The UV-Vis absorption spectroscopy, Fourier transform infrared (FT-IR), and liquid chromatography-mass spectrometry (LC-MS) were used to characterize the dyes. The electron transport mechanism and internal resistance of the DSSCs were investigated by the electrochemical impedance spectroscopy (EIS). The performance of the cells fabricated with the grass extract shows comparable efficiencies with the reported natural dyes. Among the three types of grasses, the DSSC fabricated with the dye extracted from Hierochloe Odorata (HO) exhibited the maximum efficiency. LC-MS investigations indicated that the dominant pigment present in HO dye was pheophytin a (Pheo a).

  15. Green grasses as light harvesters in dye sensitized solar cells.

    Science.gov (United States)

    Shanmugam, Vinoth; Manoharan, Subbaiah; Sharafali, A; Anandan, Sambandam; Murugan, Ramaswamy

    2015-01-25

    Chlorophylls, the major pigments presented in plants are responsible for the process of photosynthesis. The working principle of dye sensitized solar cell (DSSC) is analogous to natural photosynthesis in light-harvesting and charge separation. In a similar way, natural dyes extracted from three types of grasses viz. Hierochloe Odorata (HO), Torulinium Odoratum (TO) and Dactyloctenium Aegyptium (DA) were used as light harvesters in dye sensitized solar cells (DSSCs). The UV-Vis absorption spectroscopy, Fourier transform infrared (FT-IR), and liquid chromatography-mass spectrometry (LC-MS) were used to characterize the dyes. The electron transport mechanism and internal resistance of the DSSCs were investigated by the electrochemical impedance spectroscopy (EIS). The performance of the cells fabricated with the grass extract shows comparable efficiencies with the reported natural dyes. Among the three types of grasses, the DSSC fabricated with the dye extracted from Hierochloe Odorata (HO) exhibited the maximum efficiency. LC-MS investigations indicated that the dominant pigment present in HO dye was pheophytin a (Pheo a). Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Cell culture density affects the stemness gene expression of adipose tissue-derived mesenchymal stem cells.

    Science.gov (United States)

    Kim, Dae Seong; Lee, Myoung Woo; Lee, Tae-Hee; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee

    2017-03-01

    The results of clinical trials using mesenchymal stem cells (MSCs) are controversial due to the heterogeneity of human MSCs and differences in culture conditions. In this regard, it is important to identify gene expression patterns according to culture conditions, and to determine how the cells are expanded and when they should be clinically used. In the current study, stemness gene expression was investigated in adipose tissue-derived MSCs (AT-MSCs) harvested following culture at different densities. AT-MSCs were plated at a density of 200 or 5,000 cells/cm 2 . After 7 days of culture, stemness gene expression was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. The proliferation rate of AT-MSCs harvested at a low density (~50% confluent) was higher than that of AT-MSCs harvested at a high density (~90% confluent). Although there were differences in the expression levels of stemness gene, such as octamer-binding transcription factor 4, nanog homeobox ( Nanog ), SRY-box 2, Kruppel like factor 4, v-myc avian myelocytomatosis viral oncogene homolog ( c-Myc ), and lin-28 homolog A, in the AT-MSCs obtained from different donors, RT-qPCR analysis demonstrated differential gene expression patterns according to the cell culture density. Expression levels of stemness genes, particularly Nanog and c-Myc , were upregulated in AT-MSCs harvested at a low density (~50% confluent) in comparison to AT-MSCs from the same donor harvested at a high density (~90% confluent). These results imply that culture conditions, such as the cell density at harvesting, modulate the stemness gene expression and proliferation of MSCs.

  17. Photovoltaic characteristics of natural light harvesting dye sensitized solar cells

    Science.gov (United States)

    Hafez, H. S.; Shenouda, S. S.; Fadel, M.

    2018-03-01

    In this work of research, anthocyanin as a natural dye obtained from raspberry fruits, was used and tested as a photon harvesting/electron donating dye in titanium dioxide nanoparticle-based DSSCs. A working photoelectrode made from TiO2 nanoparticles with an average particle size (10-40 nm) that is coated on Florine doped tin-oxide substrate, was prepared via a simple and low cost hydrothermal method. A detailed structural and morphological analysis of the TiO2 photoactive electrode was investigated by X-ray diffraction (XRD), diffuse reflectance spectrometer, transmission electron microscope (TEM) and scanning electron microscope (SEM). Complete photovoltaic characteristics including (current, voltage, outpower, and responsivity) of the natural anthocyanin based dye sensitized solar cell have been investigated under different illumination intensity ranging from 10 to 100 mW.cm- 2. The cell responsivity and efficiency of the fabricated solar cell under different illumination intensity were found to be in the range (R = 15.6-23.8 mA.W- 1 and η = 0.13-0.25) at AM = 1.5 conditions. This study is important for enhancing the future applications of the promising DSSC technology.

  18. Photovoltaic characteristics of natural light harvesting dye sensitized solar cells.

    Science.gov (United States)

    Hafez, H S; Shenouda, S S; Fadel, M

    2018-03-05

    In this work of research, anthocyanin as a natural dye obtained from raspberry fruits, was used and tested as a photon harvesting/electron donating dye in titanium dioxide nanoparticle-based DSSCs. A working photoelectrode made from TiO 2 nanoparticles with an average particle size (10-40nm) that is coated on Florine doped tin-oxide substrate, was prepared via a simple and low cost hydrothermal method. A detailed structural and morphological analysis of the TiO 2 photoactive electrode was investigated by X-ray diffraction (XRD), diffuse reflectance spectrometer, transmission electron microscope (TEM) and scanning electron microscope (SEM). Complete photovoltaic characteristics including (current, voltage, outpower, and responsivity) of the natural anthocyanin based dye sensitized solar cell have been investigated under different illumination intensity ranging from 10 to 100mW.cm -2 . The cell responsivity and efficiency of the fabricated solar cell under different illumination intensity were found to be in the range (R=15.6-23.8mA.W -1 and η=0.13-0.25) at AM=1.5 conditions. This study is important for enhancing the future applications of the promising DSSC technology. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Principles of cancer cell culture.

    Science.gov (United States)

    Cree, Ian A

    2011-01-01

    The basics of cell culture are now relatively common, though it was not always so. The pioneers of cell culture would envy our simple access to manufactured plastics, media and equipment for such studies. The prerequisites for cell culture are a well lit and suitably ventilated laboratory with a laminar flow hood (Class II), CO(2) incubator, benchtop centrifuge, microscope, plasticware (flasks and plates) and a supply of media with or without serum supplements. Not only can all of this be ordered easily over the internet, but large numbers of well-characterised cell lines are available from libraries maintained to a very high standard allowing the researcher to commence experiments rapidly and economically. Attention to safety and disposal is important, and maintenance of equipment remains essential. This chapter should enable researchers with little prior knowledge to set up a suitable laboratory to do basic cell culture, but there is still no substitute for experience within an existing well-run laboratory.

  20. Adipogenic differentiation by adipose-derived stem cells harvested from GFP transgenic mice - including relationship of sex differences

    International Nuclear Information System (INIS)

    Ogawa, Rei; Mizuno, Hiroshi; Watanabe, Atsushi; Migita, Makoto; Hyakusoku, Hiko; Shimada, Takashi

    2004-01-01

    We have previously demonstrated that adipose-derived stromal cells (ASCs) as well as bone marrow-derived stromal cells (BSCs) differentiate into a variety of cell lineages both in vitro and in vivo. Both types are considered to include mesenchymal stem cells. Taking advantage of homogeneously marked cells from green fluorescent protein (GFP) transgenic mice, we have also previously reported the plasticity of BSCs and ASCs. In this study, we focused on adipogenic differentiation in vitro by ASCs harvested from GFP transgenic mice. Moreover, preadipocytes and mature adipocytes were harvested at the same time, and the cells were cultured to compare them with ASCs. Inguinal fat pads from GFP transgenic mice were used for the isolation of ASCs, preadipocytes, and mature adipocytes. After expansion to three passages of ASCs, the cells were incubated in an adipogenic medium for two weeks. Adipogenic differentiation of ASCs was assessed by Oil Red O staining and the expression of the adipocyte specific peroxisome proliferative activated receptor γ2 (PPAR-γ2) gene. These ASCs stained positively, and expression of PPAR-γ2 was detected. Moreover, we also tried to characterize the influence of sex differences on the adipogenic differentiation of ASCs harvested from both male and female mice. This was assessed by the expression levels of the PPAR-γ2 gene using real-time PCR. The results showed that the expression levels of ASCs harvested from female mice were a maximum of 2.89 times greater than those harvested from male mice. This suggests that the adipogenic differentiation of ASCs is closely related to sex differences

  1. Flexible hybrid energy cell for simultaneously harvesting thermal, mechanical, and solar energies.

    Science.gov (United States)

    Yang, Ya; Zhang, Hulin; Zhu, Guang; Lee, Sangmin; Lin, Zong-Hong; Wang, Zhong Lin

    2013-01-22

    We report the first flexible hybrid energy cell that is capable of simultaneously or individually harvesting thermal, mechanical, and solar energies to power some electronic devices. For having both the pyroelectric and piezoelectric properties, a polarized poly(vinylidene fluoride) (PVDF) film-based nanogenerator (NG) was used to harvest thermal and mechanical energies. Using aligned ZnO nanowire arrays grown on the flexible polyester (PET) substrate, a ZnO-poly(3-hexylthiophene) (P3HT) heterojunction solar cell was designed for harvesting solar energy. By integrating the NGs and the solar cells, a hybrid energy cell was fabricated to simultaneously harvest three different types of energies. With the use of a Li-ion battery as the energy storage, the harvested energy can drive four red light-emitting diodes (LEDs).

  2. Development of model for analysing respective collections of intended hematopoietic stem cells and harvests of unintended mature cells in apheresis for autologous hematopoietic stem cell collection.

    Science.gov (United States)

    Hequet, O; Le, Q H; Rodriguez, J; Dubost, P; Revesz, D; Clerc, A; Rigal, D; Salles, G; Coiffier, B

    2014-04-01

    Hematopoietic stem cells (HSCs) required to perform peripheral hematopoietic autologous stem cell transplantation (APBSCT) can be collected by processing several blood volumes (BVs) in leukapheresis sessions. However, this may cause granulocyte harvest in graft and decrease in patient's platelet blood level. Both consequences may induce disturbances in patient. One apheresis team's current purpose is to improve HSC collection by increasing HSC collection and prevent increase in granulocyte and platelet harvests. Before improving HSC collection it seemed important to know more about the way to harvest these types of cells. The purpose of our study was to develop a simple model for analysing respective collections of intended CD34+ cells among HSC (designated here as HSC) and harvests of unintended platelets or granulocytes among mature cells (designated here as mature cells) considering the number of BVs processed and factors likely to influence cell collection or harvest. For this, we processed 1, 2 and 3 BVs in 59 leukapheresis sessions and analysed corresponding collections and harvests with a referent device (COBE Spectra). First we analysed the amounts of HSC collected and mature cells harvested and second the evolution of the respective shares of HSC and mature cells collected or harvested throughout the BV processes. HSC collections and mature cell harvests increased globally (pcollections and harvests, which showed that only pre-leukapheresis blood levels (CD34+cells and platelets) influenced both cell collections and harvests (CD34+cells and platelets) (pcollections and mature unintended cells harvests (pcollections or unintended mature cell harvests were pre-leukapheresis blood cell levels. Our model was meant to assist apheresis teams in analysing shares of HSC collected and mature cells harvested with new devices or with new types of HSC mobilization. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Improved endothelial cell seeding with cultured cells and fibronectin-coated grafts

    International Nuclear Information System (INIS)

    Seeger, J.M.; Klingman, N.

    1985-01-01

    A possible approach to the low seeding efficiency of endothelial cells into prosthetic grafts is to increase the number of cells to be seeded in cell culture and improve seeding efficiency by graft precoating with fibronectin. The effect of cell culture on cell adhesion is unknown, however, and fibronectin also binds fibrin, which may increase the thrombogenicity of the graft luminal surface. To investigate these questions, freshly harvested canine jugular vein endothelial cells from six animals and similar cells harvested from six primary and eight secondary cell cultures were labeled with 111 Indium and seeded into 5 cm, 4 mm PTFE grafts coated with fibronectin, using similar uncoated PTFE grafts as controls. Platelet accumulation and distribution on six similar coated and uncoated grafts placed in canine carotid, external jugular arterial venous shunts for 2 hr were also determined using autogenous 111 Indium-labeled platelets. Significant differences between group means were determined using the paired Student's t test. Results reveal that seeding efficiency is significantly better in all groups of coated grafts compared to uncoated grafts (P less than 0.01). Cells derived from cell culture also had significantly higher seeding efficiencies than freshly harvested cells when seeded into coated grafts (P less than 0.05) and tended to have higher seeding efficiencies than harvested cells when seeded into uncoated grafts (P = 0.53). Fibronectin coating increased mean platelet accumulation on the entire graft luminal surface, but not to a statistically significant degree (P greater than 0.1). Whether this increased seeding efficiency will improve graft endothelialization remains to be investigated

  4. Mutation in cultured mammalian cells

    International Nuclear Information System (INIS)

    Nakamura, N.; Okada, S.

    1982-01-01

    Mammalian cell cultures were exposed to gamma-rays at various dose rates. Dose-rate effects were observed in cultured somatic cells of the mouse for cell killing and mutations resistant to 6-thioguanine (TGsup(r)) and to methotrexate (MTXsup(r)). Linear quadratic model may be applied to cell killing and TGsup(r) mutations in some cases but can not explain the whole data. Results at low doses with far low dose-rate were not predictable from data at high doses with acute or chronic irradiation. Radioprotective effects of dimethyl sulfoxide were seen only after acute exposure but not after chronic one, suggesting that damages by indirect action of radiations may be potentially reparable by cells. TGsup(r) mutations seem to contain gross structural changes whereas MTXsup(r) ones may have smaller alterations. (Namekawa, K.)

  5. Energy harvesting from organic liquids in micro-sized microbial fuel cells

    KAUST Repository

    Mink, J.E.; Qaisi, R.M.; Logan, B.E.; Hussain, Muhammad Mustafa

    2014-01-01

    Micro-sized microbial fuel cells (MFCs) are miniature energy harvesters that use bacteria to convert biomass from liquids into usable power. The key challenge is transitioning laboratory test beds into devices capable of producing high power using

  6. Energy transfer in nanowire solar cells with photon-harvesting shells

    KAUST Repository

    Peters, C. H.; Guichard, A. R.; Hryciw, A. C.; Brongersma, M. L.; McGehee, M. D.

    2009-01-01

    The concept of a nanowire solar cell with photon-harvesting shells is presented. In this architecture, organic molecules which absorb strongly in the near infrared where silicon absorbs weakly are coupled to silicon nanowires (SiNWs). This enables

  7. Iron Administration before Stem Cell Harvest Enables MR Imaging Tracking after Transplantation

    OpenAIRE

    Khurana, Aman; Chapelin, Fanny; Beck, Graham; Lenkov, Olga D.; Donig, Jessica; Nejadnik, Hossein; Messing, Solomon; Derugin, Nikita; Chan, Ray Chun-Fai; Gaur, Amitabh; Sennino, Barbara; McDonald, Donald M.; Kempen, Paul J.; Tikhomirov, Grigory A.; Rao, Jianghong

    2013-01-01

    Transplanted mesenchymal stem cells (MSCs) could be detected and tracked with MR imaging, if the donor is treated with an intravenous injection of the Food and Drug Administration–approved iron supplement ferumoxytol prior to MSC harvesting.

  8. 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.

  9. Light harvesting via energy transfer in the dye solar cell

    Energy Technology Data Exchange (ETDEWEB)

    Siegers, Conrad

    2007-11-09

    The PhD-thesis ''Light Harvesting via Energy Transfer in the Dye Solar Cell'' (University of Freiburg, July 2007) describes the conceptual design, synthesis and testing of energy donor acceptor sensitizers for the dye solar cell (DSC). Under monochromatic illumination solar cells sensitized with the novel donor acceptor systems revealed a higher power conversion efficiency than cells containing exclusively the acceptor component. The following approach led to this conclusion: (i) the choice of suitable chromophores as energy donor and acceptor moieties according to the Foerster-theory, (ii) the synthesis of different donor acceptor systems, (iii) the development of a methodology allowing the quantification of energy transfer within dye solar cells, and (iv) the evaluation of characteristics of DSCs that were sensitized with the different donor acceptor systems. The acceptor chromophores used in this work were derived from [Ru(dcbpy)2acac]Cl (dcbpy = 4,4'-dicarboxy-2,2'-bipyridin, acac = acetylacetonato). This complex offered the opportunity to introduce substituents at the acac-ligand's terminal CH3 groups without significantly affecting its excellent photoelectrochemical properties. Alkylated 4-amino-1,8-naphthalimides (termed Fluorols in the following) were used as energy donor chromophores. This class of compounds fulfils the requirements for efficient energy transfer to [Ru(dcbpy)2acac]Cl. Covalently linking donor and acceptor chromophores to one another was achieved by two different concepts. A dyad comprising one donor and one acceptor chromophore was synthesized by subsequent hydrosilylation steps of an olefin-bearing donor and an acceptor precursor to the dihydrosilane HSiMe2-CH2CH2-SiMe2H. A series of polymers comprising multiple donor and acceptor units was made by the addition of alkyne-bearing chromophores to hyperbranched polyglycerol azide (''Click-chemistry''). In this series the donor acceptor

  10. Cultured skin substitutes reduce donor skin harvesting for closure of excised, full-thickness burns.

    Science.gov (United States)

    Boyce, Steven T; Kagan, Richard J; Yakuboff, Kevin P; Meyer, Nicholas A; Rieman, Mary T; Greenhalgh, David G; Warden, Glenn D

    2002-02-01

    Comparison of cultured skin substitutes (CSS) and split-thickness skin autograft (AG) was performed to assess whether donor-site harvesting can be reduced quantitatively and whether functional and cosmetic outcome is similar qualitatively in the treatment of patients with massive cutaneous burns. Cultured skin substitutes consisting of collagen-glycosaminoglycan substrates populated with autologous fibroblasts and keratinocytes have been shown to close full-thickness skin wounds in preclinical and clinical studies with acceptable functional and cosmetic results. Qualitative outcome was compared between CSS and AG in 45 patients on an ordinal scale (0, worst; 10, best) with primary analyses at postoperative day 28 and after about 1 year for erythema, pigmentation, pliability, raised scar, epithelial blistering, and surface texture. In the latest 12 of the 45 patients, tracings were performed of donor skin biopsies and wounds treated with CSS at postoperative days 14 and 28 to calculate percentage engraftment, the ratio of closed wound:donor skin areas, and the percentage of total body surface area closed with CSS. Measures of qualitative outcome of CSS or AG were not different statistically at 1 year after grafting. Engraftment at postoperative day 14 exceeded 75% in the 12 patients evaluated. The ratio of closed wound:donor skin areas for CSS at postoperative day 28 was significantly greater than for conventional 4:1 meshed autografts. The percentage of total body surface area closed with CSS at postoperative day 28 was significantly less than with AG. The requirement for harvesting of donor skin for CSS was less than for conventional skin autografts. These results suggest that acute-phase recovery of patients with extensive burns is facilitated and that complications are reduced by the use of CSS together with conventional skin grafting.

  11. 9 CFR 101.6 - Cell cultures.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Cell cultures. 101.6 Section 101.6..., SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS DEFINITIONS § 101.6 Cell cultures. When used in conjunction with or in reference to cell cultures, which may be referred to as tissue cultures...

  12. Youth Culture and Cell Phone

    Directory of Open Access Journals (Sweden)

    mohammad saeed zokaei

    2009-11-01

    Full Text Available Iranian youth’s leisure culture has been immediately affected by the digital media culture. As a communicative media, cell phone has crossed borders of youth norms and identity; and in addition to facilitating their communication, has changed its patterns. Applying Bourdieu’s concepts of habitus and field, and relied on the qualitative and quantitative data gathered from the mobile youth users, the present study argues that mobile has produced a new field in which youth’s opportunities for leisure, entertainment, communication, and independence have extended. In addition, cell phone has facilitated and compensated for some defects in public sphere, and therefore empowered youth agency, individuality, and power. Despite this strengthening, cell phone does not cross borders of gender and class differences, or the levels of social capital.

  13. Comparison of the effect of three autogenous bone harvesting methods on cell viability in rabbits

    Science.gov (United States)

    Moradi Haghgoo, Janet; Arabi, Seyed Reza; Hosseinipanah, Seyyed Mohammad; Solgi, Ghasem; Rastegarfard, Neda; Farhadian, Maryam

    2017-01-01

    Background. This study was designed to compare the viability of autogenous bone grafts, harvested using different methods, in order to determine the best harvesting technique with respect to more viable cells. Methods. In this animal experimental study, three harvesting methods, including manual instrument (chisel), rotary device and piezosurgery, were used for harvesting bone grafts from the lateral body of the mandible on the left and right sides of 10 rabbits. In each group, 20 bone samples were collected and their viability was assessed using MTS kit. Statistical analyses, including ANOVA and post hoc Tukey tests, were used for evaluating significant differences between the groups. Results. One-way ANOVA showed significant differences between all the groups (P=0.000). Data analysis using post hoc Tukey tests indicated that manual instrument and piezosurgery had no significant differences with regard to cell viability (P=0.749) and the cell viability in both groups was higher than that with the use of a rotary instrument (P=0.000). Conclusion. Autogenous bone grafts harvested with a manual instrument and piezosurgery had more viable cells in comparison to the bone chips harvested with a rotary device. PMID:28748046

  14. A study of chromosomal aberrations in amniotic fluid cell cultures.

    Science.gov (United States)

    Wolstenholme, J; Crocker, M; Jonasson, J

    1988-06-01

    This paper represents the analysis of 1916 routine amniotic fluid specimens harvested by an in situ fixation technique in a prospective study with regard to cultural chromosome anomalies. Excluding constitutional abnormalities, 2.9 per cent of 19,432 cells analysed showed some form of chromosome anomaly, terminal deletions (57 per cent) and chromatid/chromosome breaks and gaps (18 per cent) being the most frequent, followed by interchange aberrations (13 per cent) and trisomy (5 per cent). No case was found of more than one colony from the same culture showing the same anomaly without it being present in other cultures from the same fluid. The wholly abnormal colonies had a surplus of trisomies and from the mathematical considerations presented one may infer that these are likely to reflect the presence of abnormal cells in the amniotic fluid. Partly abnormal colonies appeared at a frequency that would correspond to virtual absence of selection against chromosomally abnormal cells when cultured in vitro. The aberrations found were similar to those seen as single cell anomalies, except for chromatid breaks and exchanges. The data suggest a basic preferential induction of trisomy for chromosomes 2, 18, 21, and the Y-chromosome. Structural aberrations showed a marked clustering of breakpoints around the centromeres. The frequency of mutant cells was low (1.4 X 10(-3)) before culture was initiated. At harvest, the frequency of abnormal cells was much higher (3 X 10(-2)) corresponding to 3 X 10(-3) mutations per cell per generation accumulating over approximately ten generations in vitro.

  15. Compact hybrid cell based on a convoluted nanowire structure for harvesting solar and mechanical energy

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Chen; Wang, Zhong Lin [School of Materials Science and Engineering, Georgia Institute of Technology, Atlanta, GA 30332 (United States)

    2011-02-15

    A fully integrated, solid-state, compact hybrid cell (CHC) that comprises ''convoluted'' ZnO nanowire structures for concurrent harvesting of both solar and mechanical energy is demonstrated. The compact hybrid cell is based on a conjunction design of an organic solid-state dye-sensitized solar cell (DSSC) and piezoelectric nanogenerator in one compact structure. The CHC shows a significant increase in output power, clearly demonstrating its potential for simultaneously harvesting multiple types of energy for powering small electronic devices for independent, sustainable, and mobile operation. (Copyright copyright 2011 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  16. The impact of donor characteristics on the immune cell composition of mixture allografts of granulocyte-colony-stimulating factor-mobilized marrow harvests and peripheral blood harvests.

    Science.gov (United States)

    Wang, Yu-Tong; Zhao, Xiang-Yu; Zhao, Xiao-Su; Xu, Lan-Ping; Zhang, Xiao-Hui; Wang, Yu; Liu, Kai-Yan; Chang, Ying-Jun; Huang, Xiao-Jun

    2015-12-01

    The association of donor characteristics with immune cell composition in allografts remains poorly understood. In this retrospective study, the effects of donor characteristics on immune cell composition in allografts were investigated. The correlations of donor characteristics with the immune cell composition in mixture allografts of granulocyte-colony-stimulating factor-mobilized marrow harvests and peripheral blood harvests of 390 healthy donors (male, 240; female, 150; median age, 40 years old) were analyzed. The median doses of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD3+CD4-CD8- T cells, and monocytes in mixture allografts were 160.57 × 10(6), 89.29 × 10(6), 56.16 × 10(6), 10.87 × 10(6), and 137.94 × 10(6)/kg, respectively. Multivariate analysis showed that younger donor age was associated with a higher dose of CD3+ T cells (p = 0.006), CD3+CD8+ T cells (p donor weight with CD3+ T cells (p blood lymphocyte pre-peripheral blood apheresis was correlated with the yield of CD3+ T cells (p blood monocyte count before marrow harvest predicted the monocyte dose (p = 0.002). The results suggested that older and overweight donors should not be chosen. The monocyte and lymphocyte counts before harvest could predict the yield of immune cells in allografts. © 2015 AABB.

  17. Efficient light harvesting from flexible perovskite solar cells under indoor white light-emitting diode illumination

    NARCIS (Netherlands)

    Lucarelli, G.; Di Giacomo, F.; Zardetto, V.; Creatore, M.; Brown, T.M.

    2017-01-01

    This is the first report of an investigation on flexible perovskite solar cells for artificial light harvesting by using a white light-emitting diode (LED) lamp as a light source at 200 and 400 lx, values typically found in indoor environments. Flexible cells were developed using either

  18. Peripheral blood stem cell harvest in patients with limited stage small-cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Katakami, Nobuyuki; Takakura, Shunji; Fujii, Hiroshi; Nishimura, Takashi; Umeda, Bunichi [Kobe City General Hospital (Japan)

    2000-06-01

    Chemotherapy plus granulocyte colony-stimulating factor (G-CSF) induced mobilization of peripheral blood stem cells (PBSC) was performed in patients with limited stage small-cell lung cancer. Chemotherapy consisted of cisplatin/etoposide or cisplatin/adriamycin/etoposide. The amounts of CD34 positive cells and granulocyte-macrophage colony forming units (CFU-GM) collected during 2-3 courses of apheresis were 3.1{+-}2.9 x 10{sup 6}/kg (n=10) and 3.1{+-}1.5 x 10{sup 5}/kg (n=8) , respectively. Adequate amounts of PBSC were also harvested even in patients treated with concurrent chemoradiotherapy. Eight patients were successfully treated with high-dose chemotherapy consisting of ifosfamide, carboplatin and etoposide with PBSC transfusion. The patients'-bone marrow reconstruction was rapid and no treatment-related death was observed. (author)

  19. Multistage carcinogenesis in cell culture.

    Science.gov (United States)

    Rubin, H

    2001-01-01

    Rodent fibroblasts explanted from embryos to culture undergo a period of declining growth rate in serial passages leading to crisis, followed by the appearance of variants which can multiply indefinitely. If the "immortal" cell line was established by low density passage, i.e., 3T3 cells, it has a low saturation density and is non-tumorigenic. If it was established by high density passage, it has a high saturation density and is tumorigenic. The establishment of cells goes through successive stages, including increased capacity to multiply in low serum concentration, growth to high saturation density, growth in suspension, assisted tumour formation in susceptible hosts and unassisted tumour formation. Chromosome aberrations and aneuploidy occur long before the capacity to produce tumours appears. Contrary to conventional belief, human fibroblast populations also undergo a continuous loss of capacity to multiply from the time of explantation, with only the longest surviving clone reaching the Hayflick limit. Neoplastic transformation of rodent cells is strongly favoured by maintaining them in a quiescent state at confluence for prolonged periods, which results in genetic damage to the cells. It also produces a large variety of chromosomal aberrations in human cells and extends their replicative lifespan. Individual clones are more susceptible to spontaneous transformation than their heterogeneous parental cultures. The implications of these results for tumour development in vivo are that oncogenic genetic changes may be common under stressful conditions which restrict replication, and that such changes are maximized when a rogue clone reaches a critical size that reduces stabilizing interactions with neighbouring clones. An alternative explanation, described in the Addendum, which we retrospectively favor is that the easily transformed clones are a minority in the uncloned parental population. The reason they transform before the parental population is that when

  20. Pros and cons of fish skin cells in culture: long-term full skin and short-term scale cell culture from rainbow trout, Oncorhynchus mykiss.

    Science.gov (United States)

    Rakers, Sebastian; Klinger, Matthias; Kruse, Charli; Gebert, Marina

    2011-12-01

    Here, we report the establishment of a permanent skin cell culture from rainbow trout (Oncorhynchus mykiss). The cells of the fish skin cell culture could be propagated over 60 passages so far. Furthermore, we show for the first time that it is possible to integrate freshly harvested rainbow trout scales into this new fish skin cell culture. We further demonstrated that epithelial cells derived from the scales survived in the artificial micro-environment of surrounding fibroblast-like cells. Also, antibody staining indicated that both cell types proliferated and started to build connections with the other cell type. It seems that it is possible to generate an 'artificial skin' with two different cell types. This could lead to the development of a three-dimensional test system, which might be a better in vitro representative of fish skin in vivo than individual skin cell lines. Copyright © 2011 Elsevier GmbH. All rights reserved.

  1. Plant cell tissue culture: A potential source of chemicals

    Energy Technology Data Exchange (ETDEWEB)

    Scott, C.D.; Dougall, D.K.

    1987-08-01

    Higher plants produce many industrially important products. Among these are drugs and medicinal chemicals, essential oils and flavors, vegetable oils and fats, fine and specialty chemicals, and even some commodity chemicals. Although, currently, whole-plant extraction is the primary means of harvesting these materials, the advent of plant cell tissue culture could be a much more effective method of producing many types of phytochemicals. The use of immobilized plant cells in an advanced bioreactor configuration with excretion of the product into the reactor medium may represent the most straightforward way of commercializing such techniques for lower-value chemicals. Important research and development opportunities in this area include screening for plant cultures for nonmedical, lower-value chemicals; understanding and controlling plant cell physiology and biochemistry; optimizing effective immobilization methods; developing more efficient bioreactor concepts; and perfecting product extraction and purification techniques. 62 refs., 2 figs.

  2. 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.

  3. Cultural plant harvests on federal lands: perspectives from members of the Northwest Native American Basketweavers Association

    Science.gov (United States)

    Rebecca Dobkins; Ceara Lewis; Susan Hummel; Emily. Dickey

    2016-01-01

    Native Americans who wish to harvest forest plants for traditional uses report difficulties gaining access to federal lands in the northwestern United States. To learn more about this issue, we reviewed the published literature on site access and resource harvests by tribal members and discussed it with Native American traditional users of plant resources. Specifically...

  4. Response of Soil Bulk Density and Mineral Nitrogen to Harvesting and Cultural Treatments

    Science.gov (United States)

    Minyi Zhou; Mason C. Carter; Thomas J. Dean

    1998-01-01

    The interactive effects of harvest intensity, site preparation, and fertilization on soil compaction and nitrogen mineralization were examined in a loblolly pine (Pinus taeda L.) stand growing on a sandy, well-drained soil in eastern Texas. The experimental design was 2 by 2 by 2 factorial, consisting of two harvesting treatments (mechanical whole-...

  5. Cell culture media impact on drug product solution stability.

    Science.gov (United States)

    Purdie, Jennifer L; Kowle, Ronald L; Langland, Amie L; Patel, Chetan N; Ouyang, Anli; Olson, Donald J

    2016-07-08

    To enable subcutaneous administration of monoclonal antibodies, drug product solutions are often needed at high concentrations. A significant risk associated with high drug product concentrations is an increase in aggregate level over the shelf-life dating period. While much work has been done to understand the impact of drug product formulation on aggregation, there is limited understanding of the link between cell culture process conditions and soluble aggregate growth in drug product. During cell culture process development, soluble aggregates are often measured at harvest using cell-free material purified by Protein A chromatography. In the work reported here, cell culture media components were evaluated with respect to their impact on aggregate levels in high concentration solution drug product during accelerated stability studies. Two components, cysteine and ferric ammonium citrate, were found to impact aggregate growth rates in our current media (version 1) leading to the development of new chemically defined media and concentrated feed formulations. The new version of media and associated concentrated feeds (version 2) were evaluated across four cell lines producing recombinant IgG4 monoclonal antibodies and a bispecific antibody. In all four cell lines, the version 2 media reduced aggregate growth over the course of a 12 week accelerated stability study compared with the version 1 media, although the degree to which aggregate growth decreased was cell line dependent. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:998-1008, 2016. © 2016 American Institute of Chemical Engineers.

  6. Design of Transparent Anodes for Resonant Cavity Enhanced Light Harvesting in Organic Solar Cells

    KAUST Repository

    Sergeant, Nicholas P.; Hadipour, Afshin; Niesen, Bjoern; Cheyns, David; Heremans, Paul; Peumans, Peter; Rand, Barry P.

    2012-01-01

    The use of an ITO-free MoO 3/Ag/MoO 3 anode to control the photon harvesting in PCDTBT:PC 70BM solar cells is proposed. At first sight, the fact that these anodes possess reduced far-field transmission compared to ITO may seem to be a disadvantage

  7. Microfluidic cell culture systems for drug research.

    Science.gov (United States)

    Wu, Min-Hsien; Huang, Song-Bin; Lee, Gwo-Bin

    2010-04-21

    In pharmaceutical research, an adequate cell-based assay scheme to efficiently screen and to validate potential drug candidates in the initial stage of drug discovery is crucial. In order to better predict the clinical response to drug compounds, a cell culture model that is faithful to in vivo behavior is required. With the recent advances in microfluidic technology, the utilization of a microfluidic-based cell culture has several advantages, making it a promising alternative to the conventional cell culture methods. This review starts with a comprehensive discussion on the general process for drug discovery and development, the role of cell culture in drug research, and the characteristics of the cell culture formats commonly used in current microfluidic-based, cell-culture practices. Due to the significant differences in several physical phenomena between microscale and macroscale devices, microfluidic technology provides unique functionality, which is not previously possible by using traditional techniques. In a subsequent section, the niches for using microfluidic-based cell culture systems for drug research are discussed. Moreover, some critical issues such as cell immobilization, medium pumping or gradient generation in microfluidic-based, cell-culture systems are also reviewed. Finally, some practical applications of microfluidic-based, cell-culture systems in drug research particularly those pertaining to drug toxicity testing and those with a high-throughput capability are highlighted.

  8. A flexible thermoresponsive cell culture substrate for direct transfer of keratinocyte cell sheets.

    Science.gov (United States)

    Praveen, Wulligundam; Madathil, Bernadette K; Sajin Raj, R S; Kumary, T V; Anil Kumar, P R

    2017-10-25

    Most cell sheet engineering systems require a support or carrier to handle the harvested cell sheets. In this study, polyethylene terephthalate-based overhead projection transparency sheets (OHPS) were subjected to surface hydrolysis by alkali treatment to increase pliability and hydrophilicity and enable poly(N-isopropylacrylamide-co-glycidylmethacrylate) copolymer (NGMA) coating to impart thermoresponsiveness. NGMA was applied on the modified OHPS by the technique of spin coating using an indigenously designed spin coater. The spin coating had the advantage of using low volumes of the polymer and a reduced coating time. The surface chemistry and thermoresponsive coating was analyzed by Fourier transform infrared spectroscopy and water contact angle. Human keratinocyte cells were cultured on the spin coated surface and scaffold-free cell sheets were successfully harvested by simple variation of temperature. These cell sheets were found to be viable, exhibited epithelial characteristic and cell-cell contact as confirmed by positive immunostaining for ZO-1. The integrity and morphology of the cell sheet was confirmed by stereomicroscopy and E-SEM. These results highlight the potential of the NGMA spin coated modified OHPS to serve as a thermoresponsive culture surface-cum-flexible transfer tool.

  9. Enhanced Light Harvesting in Dye-Sensitized Solar Cell Using External Lightguide

    Directory of Open Access Journals (Sweden)

    Chi-Hui Chien

    2011-01-01

    Full Text Available An external lightguide (EL for enhancing the light-harvesting efficiency of dye-sensitized solar cells (DSSCs was designed and developed. The EL attached to the exterior of a DSSC photoelectrode directed light on a dye-covered nanoporous TiO2 film (D-NTF of the photoelectrode. Experimental tests confirmed that the EL increased the light-harvesting efficiency of a DSSC with an active area of 0.25 cm2 by 30.69%. Photocurrent density and the power conversion efficiency were also increased by 38.12% and 25.09%, respectively.

  10. Self-assembled photosynthesis-inspired light harvesting material and solar cells containing the same

    Science.gov (United States)

    Lindsey, Jonathan S [Raleigh, NC; Chinnasamy, Muthiah [Raleigh, NC; Fan, Dazhong [Raleigh, NC

    2009-12-15

    A solar cell is described that comprises: (a) a semiconductor charge separation material; (b) at least one electrode connected to the charge separation material; and (c) a light-harvesting film on the charge separation material, the light-harvesting film comprising non-covalently coupled, self-assembled units of porphyrinic macrocycles. The porphyrinic macrocycles preferably comprise: (i) an intramolecularly coordinated metal; (ii) a first coordinating substituent; and (iii) a second coordinating substituent opposite the first coordinating substituent. The porphyrinic macrocycles can be assembled by repeating intermolecular coordination complexes of the metal, the first coordinating substituent and the second coordinating substituent.

  11. 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)

  12. Cell culture techniques in honey bee research

    Science.gov (United States)

    Cell culture techniques are indispensable in most if not all life science disciplines to date. Wherever cell culture models are lacking scientific development is hampered. Unfortunately this has been and still is the case in honey bee research because permanent honey bee cell lines have not yet been...

  13. Enteric neurospheres are not specific to neural crest cultures : Implications for neural stem cell therapies

    NARCIS (Netherlands)

    Binder, E. (Ellen); D. Natarajan (Dipa); J.E. Cooper (Julie E.); Kronfli, R. (Rania); Cananzi, M. (Mara); J.-M. Delalande (Jean-Marie); C. Mccann; A.J. Burns (Alan); N. Thapar (Nikhil)

    2015-01-01

    textabstractObjectives Enteric neural stem cells provide hope of curative treatment for enteric neuropathies. Current protocols for their harvesting from humans focus on the generation of 'neurospheres' from cultures of dissociated gut tissue. The study aims to better understand the derivation,

  14. Photothermally Activated Pyroelectric Polymer Films for Harvesting of Solar Heat with a Hybrid Energy Cell Structure.

    Science.gov (United States)

    Park, Teahoon; Na, Jongbeom; Kim, Byeonggwan; Kim, Younghoon; Shin, Haijin; Kim, Eunkyoung

    2015-12-22

    Photothermal effects in poly(3,4-ethylenedioxythiophene)s (PEDOTs) were explored for pyroelectric conversion. A poled ferroelectric film was coated on both sides with PEDOT via solution casting polymerization of EDOT, to give highly conductive and effective photothermal thin films of PEDOT. The PEDOT films not only provided heat source upon light exposure but worked as electrodes for the output energy from the pyroelectric layer in an energy harvester hybridized with a thermoelectric layer. Compared to a bare thermoelectric system under NIR irradiation, the photothermal-pyro-thermoelectric device showed more than 6 times higher thermoelectric output with the additional pyroelectric output. The photothermally driven pyroelectric harvesting film provided a very fast electric output with a high voltage output (Vout) of 15 V. The pyroelectric effect was significant due to the transparent and high photothermal PEDOT film, which could also work as an electrode. A hybrid energy harvester was assembled to enhance photoconversion efficiency (PCE) of a solar cell with a thermoelectric device operated by the photothermally generated heat. The PCE was increased more than 20% under sunlight irradiation (AM 1.5G) utilizing the transmitted light through the photovoltaic cell as a heat source that was converted into pyroelectric and thermoelectric output simultaneously from the high photothermal PEDOT electrodes. Overall, this work provides a dynamic and static hybrid energy cell to harvest solar energy in full spectral range and thermal energy, to allow solar powered switching of an electrochromic display.

  15. Energy harvesting using TEG and PV cell for low power application

    Science.gov (United States)

    Tawil, Siti Nooraya Mohd; Zainal, Mohd Zulkarnain

    2018-02-01

    A thermoelectric generator (TEG) module and photovoltaic cell (PV) were utilized to harvest energy from temperature gradients of heat sources from ambient heat and light of sun. The output of TEG and PV were connected to a power management circuit consist of step-up dc-dc converter in order to increase the output voltage to supply a low power application such as wireless communication module and the photovoltaic cell for charging an energy storage element in order to switch on a fan for cooling system of the thermoelectric generator. A switch is used as a selector to choose the input of source either from photovoltaic cell or thermoelectric generator to switch on DC-DC step-up converter. In order to turn on the DC-DC step-up converter, the input must be greater than 3V. The energy harvesting was designed so that it can be used continuously and portable anywhere. Multiple sources used in this energy harvesting system is to ensure the system can work in whatever condition either in good weather or not good condition of weather. This energy harvesting system has the potential to be used in military operation and environment that require sustainability of energy resources.

  16. Incomplete Exciton Harvesting from Fullerenes in Bulk Heterojunction Solar Cells

    KAUST Repository

    Burkhard, George F.

    2009-12-09

    We investigate the internal quantum efficiencies (IQEs) of high efficiency poly-3-hexylthiophene:[6,6]-phenyl-C61-butyric acid methyl ester (P3HT:PCBM) solar cells and find them to be lower at wavelengths where the PCBM absorbs. Because the exciton diffusion length in PCBM is too small, excitons generated in PCBM decay before reaching the donor-acceptor interface. This result has implications for most state of the art organic solar cells, since all of the most efficient devices use fullerenes as electron acceptors. © 2009 American Chemical Society.

  17. Incomplete Exciton Harvesting from Fullerenes in Bulk Heterojunction Solar Cells

    KAUST Repository

    Burkhard, George F.; Hoke, Eric T.; Scully, Shawn R.; McGehee, Michael D.

    2009-01-01

    We investigate the internal quantum efficiencies (IQEs) of high efficiency poly-3-hexylthiophene:[6,6]-phenyl-C61-butyric acid methyl ester (P3HT:PCBM) solar cells and find them to be lower at wavelengths where the PCBM absorbs. Because the exciton diffusion length in PCBM is too small, excitons generated in PCBM decay before reaching the donor-acceptor interface. This result has implications for most state of the art organic solar cells, since all of the most efficient devices use fullerenes as electron acceptors. © 2009 American Chemical Society.

  18. Energy harvesting by implantable abiotically catalyzed glucose fuel cells

    Science.gov (United States)

    Kerzenmacher, S.; Ducrée, J.; Zengerle, R.; von Stetten, F.

    Implantable glucose fuel cells are a promising approach to realize an autonomous energy supply for medical implants that solely relies on the electrochemical reaction of oxygen and glucose. Key advantage over conventional batteries is the abundant availability of both reactants in body fluids, rendering the need for regular replacement or external recharging mechanisms obsolete. Implantable glucose fuel cells, based on abiotic catalysts such as noble metals and activated carbon, have already been developed as power supply for cardiac pacemakers in the late-1960s. Whereas, in vitro and preliminary in vivo studies demonstrated their long-term stability, the performance of these fuel cells is limited to the μW-range. Consequently, no further developments have been reported since high-capacity lithium iodine batteries for cardiac pacemakers became available in the mid-1970s. In recent years research has been focused on enzymatically catalyzed glucose fuel cells. They offer higher power densities than their abiotically catalyzed counterparts, but the limited enzyme stability impedes long-term application. In this context, the trend towards increasingly energy-efficient low power MEMS (micro-electro-mechanical systems) implants has revived the interest in abiotic catalysts as a long-term stable alternative. This review covers the state-of-the-art in implantable abiotically catalyzed glucose fuel cells and their development since the 1960s. Different embodiment concepts are presented and the historical achievements of academic and industrial research groups are critically reviewed. Special regard is given to the applicability of the concept as sustainable micro-power generator for implantable devices.

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

    Science.gov (United States)

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

    1992-12-05

    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. (c) 1992 John Wiley & Sons, Inc.

  20. Realizing Efficient Energy Harvesting from Organic Photovoltaic Cells

    Science.gov (United States)

    Zou, Yunlong

    Organic photovoltaic cells (OPVs) are emerging field of research in renewable energy. The development of OPVs in recent years has made this technology viable for many niche applications. In order to realize widespread application however, the power conversion efficiency requires further improvement. The efficiency of an OPV depends on the short-circuit current density (JSC), open-circuit voltage (VOC) and fill factor (FF). For state-of-the-art devices, JSC is mostly optimized with the application of novel low-bandgap materials and a bulk heterojunction device architecture (internal quantum efficiency approaching 100%). The remaining limiting factors are the low VOC and FF. This work focuses on overcoming these bottlenecks for improved efficiency. Temperature dependent measurements of device performance are used to examine both charge transfer and exciton ionization process in OPVs. The results permit an improved understanding of the intrinsic limit for VOC in various device architectures and provide insight on device operation. Efforts have also been directed at engineering device architecture for optimized FF, realizing a very high efficiency of 8% for vapor deposited small molecule OPVs. With collaborators, new molecules with tailored desired energy levels are being designed for further improvements in efficiency. A new type of hybrid organic-inorganic perovskite material is also included in this study. By addressing processing issues and anomalous hysteresis effects, a very high efficiency of 19.1% is achieved. Moving forward, topics including engineering film crystallinity, exploring tandem architectures and understanding degradation mechanisms will further push OPVs toward broad commercialization.

  1. 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.

  2. Replication of cultured lung epithelial cells

    International Nuclear Information System (INIS)

    Guzowski, D.; Bienkowski, R.

    1986-01-01

    The authors have investigated the conditions necessary to support replication of lung type 2 epithelial cells in culture. Cells were isolated from mature fetal rabbit lungs (29d gestation) and cultured on feeder layers of mitotically inactivated 3T3 fibroblasts. The epithelial nature of the cells was demonstrated by indirect immunofluorescent staining for keratin and by polyacid dichrome stain. Ultrastructural examination during the first week showed that the cells contained myofilaments, microvilli and lamellar bodies (markers for type 2 cells). The following changes were observed after the first week: increase in cell size; loss of lamellar bodies and appearance of multivesicular bodies; increase in rough endoplasmic reticulum and golgi; increase in tonafilaments and well-defined junctions. General cell morphology was good for up to 10 wk. Cells cultured on plastic surface degenerated after 1 wk. Cell replication was assayed by autoradiography of cultures exposed to ( 3 H)-thymidine and by direct cell counts. The cells did not replicate during the first week; however, between 2-10 wk the cells incorporated the label and went through approximately 6 population doublings. They have demonstrated that lung alveolar epithelial cells can replicate in culture if they are maintained on an appropriate substrate. The coincidence of ability to replicate and loss of markers for differentiation may reflect the dichotomy between growth and differentiation commonly observed in developing systems

  3. 3D Cell Culture in Alginate Hydrogels

    Directory of Open Access Journals (Sweden)

    Therese Andersen

    2015-03-01

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

  4. Energy Harvesting by Subcutaneous Solar Cells: A Long-Term Study on Achievable Energy Output.

    Science.gov (United States)

    Bereuter, L; Williner, S; Pianezzi, F; Bissig, B; Buecheler, S; Burger, J; Vogel, R; Zurbuchen, A; Haeberlin, A

    2017-05-01

    Active electronic implants are powered by primary batteries, which induces the necessity of implant replacement after battery depletion. This causes repeated interventions in a patients' life, which bears the risk of complications and is costly. By using energy harvesting devices to power the implant, device replacements may be avoided and the device size may be reduced dramatically. Recently, several groups presented prototypes of implants powered by subcutaneous solar cells. However, data about the expected real-life power output of subcutaneously implanted solar cells was lacking so far. In this study, we report the first real-life validation data of energy harvesting by subcutaneous solar cells. Portable light measurement devices that feature solar cells (cell area = 3.6 cm 2 ) and continuously measure a subcutaneous solar cell's output power were built. The measurement devices were worn by volunteers in their daily routine in summer, autumn and winter. In addition to the measured output power, influences such as season, weather and human activity were analyzed. The obtained mean power over the whole study period was 67 µW (=19 µW cm -2 ), which is sufficient to power e.g. a cardiac pacemaker.

  5. Subsequent donation requests among 2472 unrelated hematopoietic progenitor cell donors are associated with bone marrow harvest

    Science.gov (United States)

    Lown, Robert N.; Tulpule, Sameer; Russell, Nigel H.; Craddock, Charles F.; Roest, Rochelle; Madrigal, J. Alejandro; Shaw, Bronwen E.

    2013-01-01

    Approximately 1 in 20 unrelated donors are asked to make a second donation of hematopoietic progenitor cells, the majority for the same patient. Anthony Nolan undertook a study of subsequent hematopoietic progenitor cell donations made by its donors from 2005 to 2011, with the aims of predicting those donors more likely to be called for a second donation, assessing rates of serious adverse reactions and examining harvest yields. This was not a study of factors predictive of second allografts. During the study period 2591 donations were made, of which 120 (4.6%) were subsequent donations. The median time between donations was 179 days (range, 21–4016). Indications for a second allogeneic transplant included primary graft failure (11.7%), secondary graft failure (53.2%), relapse (30.6%) and others (1.8%). On multivariate analysis, bone marrow harvest at first donation was associated with subsequent donation requests (odds ratio 2.00, P=0.001). The rate of serious adverse reactions in donors making a subsequent donation appeared greater than the rate in those making a first donation (relative risk=3.29, P=0.005). Harvest yields per kilogram recipient body weight were equivalent between donations, although females appeared to have a lower yield at the subsequent donation. Knowledge of these factors will help unrelated donor registries to counsel their donors. PMID:23812935

  6. Recent trends in mesoscopic solar cells based on molecular and nanopigment light harvesters

    KAUST Repository

    Grä tzel, Carole; Zakeeruddin, Shaik M.

    2013-01-01

    Mesoscopic solar cells are one of the most promising photovoltaic technologies among third generation photovoltaics due to their low cost and high efficiency. The morphology of wide-band semiconductors, sensitized with molecular or nanosized light harvesters, used as electron collectors contribute substantially to the device performance. Recent developments in the use of organic-inorganic layer structured perovskites as light absorbers and as electron or hole transport materials allows reduction in the thickness of photoanodes to the submicron level and have raised the power conversion efficiency of solid state mesoscopic solar cells above the 10% level.

  7. Quantum dot sensitized solar cells: Light harvesting versus charge recombination, a film thickness consideration

    Science.gov (United States)

    Wang, Xiu Wei; Wang, Ye Feng; Zeng, Jing Hui; Shi, Feng; Chen, Yu; Jiang, Jiaxing

    2017-08-01

    Sensitizer loading level is one of the key factors determined the performance of sensitized solar cells. In this work, we systemically studied the influence of photo-anode thicknesses on the performance of the quantum-dot sensitized solar cells. It is found that the photo-to-current conversion efficiency enhances with increased film thickness and peaks at around 20 μm. The optimal value is about twice as large as the dye counterparts. Here, we also uncover the underlying mechanism about the influence of film thickness over the photovoltaic performance of QDSSCs from the light harvesting and charge recombination viewpoint.

  8. Advances in cell culture: anchorage dependence

    Science.gov (United States)

    Merten, Otto-Wilhelm

    2015-01-01

    Anchorage-dependent cells are of great interest for various biotechnological applications. (i) They represent a formidable production means of viruses for vaccination purposes at very large scales (in 1000–6000 l reactors) using microcarriers, and in the last decade many more novel viral vaccines have been developed using this production technology. (ii) With the advent of stem cells and their use/potential use in clinics for cell therapy and regenerative medicine purposes, the development of novel culture devices and technologies for adherent cells has accelerated greatly with a view to the large-scale expansion of these cells. Presently, the really scalable systems—microcarrier/microcarrier-clump cultures using stirred-tank reactors—for the expansion of stem cells are still in their infancy. Only laboratory scale reactors of maximally 2.5 l working volume have been evaluated because thorough knowledge and basic understanding of critical issues with respect to cell expansion while retaining pluripotency and differentiation potential, and the impact of the culture environment on stem cell fate, etc., are still lacking and require further studies. This article gives an overview on critical issues common to all cell culture systems for adherent cells as well as specifics for different types of stem cells in view of small- and large-scale cell expansion and production processes. PMID:25533097

  9. Bio-Photoelectrochemical Solar Cells Incorporating Reaction Center and Reaction Center Plus Light Harvesting Complexes

    Science.gov (United States)

    Yaghoubi, Houman

    Harvesting solar energy can potentially be a promising solution to the energy crisis now and in the future. However, material and processing costs continue to be the most important limitations for the commercial devices. A key solution to these problems might lie within the development of bio-hybrid solar cells that seeks to mimic photosynthesis to harvest solar energy and to take advantage of the low material costs, negative carbon footprint, and material abundance. The bio-photoelectrochemical cell technologies exploit biomimetic means of energy conversion by utilizing plant-derived photosystems which can be inexpensive and ultimately the most sustainable alternative. Plants and photosynthetic bacteria harvest light, through special proteins called reaction centers (RCs), with high efficiency and convert it into electrochemical energy. In theory, photosynthetic RCs can be used in a device to harvest solar energy and generate 1.1 V open circuit voltage and ~1 mA cm-2 short circuit photocurrent. Considering the nearly perfect quantum yield of photo-induced charge separation, efficiency of a protein-based solar cell might exceed 20%. In practice, the efficiency of fabricated devices has been limited mainly due to the challenges in the electron transfer between the protein complex and the device electrodes as well as limited light absorption. The overarching goal of this work is to increase the power conversion efficiency in protein-based solar cells by addressing those issues (i.e. electron transfer and light absorption). This work presents several approaches to increase the charge transfer rate between the photosynthetic RC and underlying electrode as well as increasing the light absorption to eventually enhance the external quantum efficiency (EQE) of bio-hybrid solar cells. The first approach is to decrease the electron transfer distance between one of the redox active sites in the RC and the underlying electrode by direct attachment of the of protein complex

  10. A rapid microassay for detecting antibodies against poliovirus based on [14C]thymidine uptake of treated cell cultures

    International Nuclear Information System (INIS)

    Hilfenhaus, J.; Damm, H.; Ziegelmaier, R.; Gruschkau, H.

    1977-01-01

    DNA synthesis of mammalian cells propagated in microplates can easily be measured if cell cultures incubated with [ 14 C]thymidine are harvested on to glass fibre filters by a semiautomatic harvesting technique. Soon after infection with poliovirus, [ 14 C]thymidine uptake of U cells (established, human amniotic cell line) is inhibited. This inhibition can be prevented by previous virus neutralization with antibody. Based on this effect a rapid, precise assay method was set up to determine neutralizing antibody titres against poliovirus. There was a good correlation between titres obtained by this assay and those obtained by 50% endpoint titrations in cytopathogenic effect inhibition assays

  11. Advances in 3D neuronal cell culture

    NARCIS (Netherlands)

    Frimat, Jean Philippe; Xie, Sijia; Bastiaens, Alex; Schurink, Bart; Wolbers, Floor; Den Toonder, Jaap; Luttge, Regina

    2015-01-01

    In this contribution, the authors present our advances in three-dimensional (3D) neuronal cell culture platform technology contributing to controlled environments for microtissue engineering and analysis of cellular physiological and pathological responses. First, a micromachined silicon sieving

  12. Identification and prevention of antibody disulfide bond reduction during cell culture manufacturing.

    Science.gov (United States)

    Trexler-Schmidt, Melody; Sargis, Sandy; Chiu, Jason; Sze-Khoo, Stefanie; Mun, Melissa; Kao, Yung-Hsiang; Laird, Michael W

    2010-06-15

    In the biopharmaceutical industry, therapeutic monoclonal antibodies are primarily produced in mammalian cell culture systems. During the scale-up of a monoclonal antibody production process, we observed excessive mechanical cell shear as well as significant reduction of the antibody's interchain disulfide bonds during harvest operations. This antibody reduction event was catastrophic as the product failed to meet the drug substance specifications and the bulk product was lost. Subsequent laboratory studies have demonstrated that cells subjected to mechanical shear release cellular enzymes that contribute to this antibody reduction phenomenon (manuscript submitted; Kao et al., 2009). Several methods to prevent this antibody reduction event were developed using a lab-scale model to reproduce the lysis and reduction events. These methods included modifications to the cell culture media with chemicals (e.g., cupric sulfate (CuSO(4))), pre- and post-harvest chemical additions to the cell culture fluid (CCF) (e.g., CuSO(4), EDTA, L-cystine), as well as lowering the pH and air sparging of the harvested CCF (HCCF). These methods were evaluated for their effectiveness in preventing disulfide bond reduction and their impact to product quality. Effective prevention methods, which yielded acceptable product quality were evaluated for their potential to be implemented at manufacturing-scale. The work described here identifies numerous effective reduction prevention measures from lab-scale studies; several of these methods were then successfully translated into manufacturing processes. 2010 Wiley Periodicals, Inc.

  13. Morphology and steroidogenesis of cultured granulosa cells obtained from ovaries of women treated with ionizing radiation

    International Nuclear Information System (INIS)

    Skrzypczak, J.

    1997-01-01

    The object of the study was the morphology and steroidogenesis of cultured granulosa cells obtained from 6 women aged 28-39 years who, because of Ib cervix carcinoma, were treated with ionizing radiation and later underwent surgery. It was observed that the granulosa cells were viable, had strong proliferative ability, and formed a monolayer on day 2 of culture. Contrary to our expectations, these cells produced larger amounts of steroids in culture than the control cells harvested from normal ovaries in late follicular phase. It was also found that the cells treated with ionizing radiation responded to exogenous gonadotropins with higher production of progesterone and estradiol than the controls. It is concluded that the increase in metabolic activity by granulosa cells from ovaries which had been indirectly affected by ionizing radiation is manifested by the stimulating influence of radiation on steroidogenesis. (author)

  14. Culture of Mouse Neural Stem Cell Precursors

    OpenAIRE

    Currle, D. Spencer; Hu, Jia Sheng; Kolski-Andreaco, Aaron; Monuki, Edwin S.

    2007-01-01

    Primary neural stem cell cultures are useful for studying the mechanisms underlying central nervous system development. Stem cell research will increase our understanding of the nervous system and may allow us to develop treatments for currently incurable brain diseases and injuries. In addition, stem cells should be used for stem cell research aimed at the detailed study of mechanisms of neural differentiation and transdifferentiation and the genetic and environmental signals that direct the...

  15. 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

  16. Validation of cell-free culture using scanning electron microscopy (SEM) and gene expression studies.

    Science.gov (United States)

    Yang, R; Elankumaran, Y; Hijjawi, N; Ryan, U

    2015-06-01

    A cell-free culture system for Cryptosporidium parvum was analysed using scanning electron microscopy (SEM) to characterise life cycle stages and compare gene expression in cell-free culture and cell culture using HCT-8 cells. Cryptosporidium parvum samples were harvested at 2 h, 8 h, 14 h, 26 h, 50 h, 74 h, 98 h, 122 h and 170 h, chemically fixed and specimens were observed using a Zeiss 1555 scanning electron microscope. The presence of sporozoites, trophozoites and type I merozoites were identified by SEM. Gene expression in cell culture and cell-free culture was studied using reverse transcriptase quantitative PCR (RT-qPCR) of the sporozoite surface antigen protein (cp15), the glycoprotein 900 (gp900), the Cryptosporidium oocyst wall protein (COWP) and 18S ribosomal RNA (rRNA) genes in both cell free and conventional cell culture. In cell culture, cp15 expression peaked at 74 h, gp900 expression peaked at 74 h and 98 h and COWP expression peaked at 50 h. In cell-free culture, CP15 expression peaked at 98 h, gp900 expression peaked at 74 h and COWP expression peaked at 122 h. The present study is the first to compare gene expression of C. parvum in cell culture and cell-free culture and to characterise life cycle stages of C. parvum in cell-free culture using SEM. Findings from this study showed that gene expression patterns in cell culture and cell-free culture were similar but in cell-free culture, gene expression was delayed for CP15 and COWP in cell free culture compared with the cell culture system and was lower. Although three life cycle stageswere conclusively identified, improvements in SEM methodology should lead to the detection of more life cycle stages. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Melphalan metabolism in cultured cells

    International Nuclear Information System (INIS)

    Seagrave, J.C.; Valdez, J.G.; Tobey, R.A.; Gurley, L.R.

    1985-06-01

    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 ZnC1 2 , one being increased in amount while the other was reduced to an insignificant level. In ZnC1 2 -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 ZnC1 2 -treated cells. While ZnC1 2 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 ZnC1 2 -treated or untreated cells. Formation of derivatives of melphalan with glutathione catabolic products in ZnC1 2 -treated cells has not yet been eliminated, however. 17 refs., 5 figs., 1 tab

  18. Design of Transparent Anodes for Resonant Cavity Enhanced Light Harvesting in Organic Solar Cells

    KAUST Repository

    Sergeant, Nicholas P.

    2012-01-03

    The use of an ITO-free MoO 3/Ag/MoO 3 anode to control the photon harvesting in PCDTBT:PC 70BM solar cells is proposed. At first sight, the fact that these anodes possess reduced far-field transmission compared to ITO may seem to be a disadvantage. But, despite this, we show that by carefully tuning the resonant optical cavity we can enhance the external quantum efficiency close to the band edge of PCDTBT, resulting in high photocurrent and power conversion efficiency on par with ITO. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Data of continuous harvest of stem cells via partial detachment from thermoresponsive nanobrush surfaces

    Directory of Open Access Journals (Sweden)

    Chin-Chen Yeh

    2016-03-01

    Full Text Available This data article contains two figures and one table supporting the research article entitled: “Continuous harvest of stem cells via partial detachment from thermoresponsive nanobrush surface” [1]. The table shows coating conditions of three copolymers, poly(styrene-co-acrylic acid grafted with oligovitronectin, poly(styrene-co-N-isopropylacrylamide and poly(styrene-co-polyethylene glycol methacrylate to prepare thermoresponsive surface. XPS spectra show the nitrogen peak of the polystyrene surface coated with poly(styrene-co-acrylic acid grafted with oligovitronectin. The surface coating density analyzed from sorption of poly(styrene-co-acrylic acid grafted with oligovitronectin by UV–vis spectroscopy is also presented.

  20. Data of continuous harvest of stem cells via partial detachment from thermoresponsive nanobrush surfaces

    Science.gov (United States)

    Yeh, Chin-Chen; Muduli, Saradaprasan; Peng, I-Chia; Lu, Yi-Tung; Ling, Qing-Dong; Alarfaj, Abdullah A.; Munusamy, Murugan A.; Kumar, S. Suresh; Murugan, Kadarkarai; Chen, Da-Chung; Lee, Hsin-chung; Chang, Yung; Higuchi, Akon

    2016-01-01

    This data article contains two figures and one table supporting the research article entitled: “Continuous harvest of stem cells via partial detachment from thermoresponsive nanobrush surface” [1]. The table shows coating conditions of three copolymers, poly(styrene-co-acrylic acid) grafted with oligovitronectin, poly(styrene-co-N-isopropylacrylamide) and poly(styrene-co-polyethylene glycol methacrylate) to prepare thermoresponsive surface. XPS spectra show the nitrogen peak of the polystyrene surface coated with poly(styrene-co-acrylic acid) grafted with oligovitronectin. The surface coating density analyzed from sorption of poly(styrene-co-acrylic acid) grafted with oligovitronectin by UV–vis spectroscopy is also presented. PMID:26909373

  1. 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

  2. Quantitative volumetric Raman imaging of three dimensional cell cultures

    KAUST Repository

    Kallepitis, Charalambos; Bergholt, Mads S.; Mazo, Manuel M.; Leonardo, Vincent; Skaalure, Stacey C.; Maynard, Stephanie A.; Stevens, Molly M.

    2017-01-01

    in conventional cell culture systems and mesenchymal stem cells inside biomimetic hydrogels that supplied a 3D cell culture environment. We demonstrate visualization and quantification of fine details in cell shape, cytoplasm, nucleus, lipid bodies

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

    Science.gov (United States)

    Stampfer, Martha R; Garbe, James C

    2015-02-24

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

  4. Circulating hematopoietic progenitors and CD34(+) cells predicted successful hematopoietic stem cell harvest in myeloma and lymphoma patients: experiences from a single institution.

    Science.gov (United States)

    Yu, Jui-Ting; Cheng, Shao-Bin; Yang, Youngsen; Chang, Kuang-Hsi; Hwang, Wen-Li; Teng, Chieh-Lin Jerry

    2016-01-01

    Previous studies have shown that the numbers of both circulating hematopoietic progenitor cell (HPC) and CD34(+) cell are positively correlated with CD34(+) cell harvest yield. However, the minimal numbers of both circulating HPCs and CD34(+) cells required for performing an efficient hematopoietic stem cell (HSC) harvest in lymphoma and myeloma patients have not been defined in our institution. Medical records of 50 lymphoma and myeloma patients undergoing peripheral blood HSC harvest in our institution were retrospectively reviewed. The minimal and optimal HSC harvest yield required for the treatment was considered to be ≥2×10(6) CD34(+) cells/kg and ≥5×10(6) CD34(+) cells/kg, respectively. The minimally required or optimal HSC yield obtained was not influenced by age (≥60 years), sex, underlying malignancies, disease status, multiple rounds of chemotherapy, or history of radiotherapy. The numbers of both circulating HPC and CD34(+) cell were higher in patients with minimally required HSC yields (P=0.000 for HPC and P=0.000 for CD34(+) cell) and also in patients with optimal HSC yields (P=0.011 for HPC and P=0.006 for CD34(+) cell). The cell count cutoff for obtaining minimally required HSC harvest was determined to be 20/mm(3) for HPCs and 10/mm(3) for CD34(+) cells. Furthermore, the cell count cutoff for obtaining optimal HSC harvest was determined to be 60/mm(3) for HPCs and 35/mm(3) for CD34(+) cells. A total of 60/mm(3) of HPCs and 35/mm(3) of CD34(+) cells in peripheral blood predicted optimal HSC harvest in lymphoma and myeloma patients.

  5. Do institutions, inequality and cultural differences affect cadaveric versus live-kidney harvesting?

    OpenAIRE

    Nejat Anbarci; Mustafa Caglayan

    2010-01-01

    This paper empirically investigates the role of institutions, income inequality, cultural differences and health expenditures on cadaveric versus total kidney transplants scrutinizing information gathered from 63 countries over the period 1998-2002. We show that improvements in income equality and the rule of law encourage cadaveric kidney transplants in low-income countries. We find that cultural differences affect the number of cadaveric kidney transplants both in low- and high-income count...

  6. Substrate utilisation by plant-cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Fowler, M W

    1982-01-01

    Plant cell cultures have been grown on a wide range of carbon sources in addition to the traditional ones of sucrose and glucose. Biomass yields and growth rates vary greatly between the different carbon sources and there is a variation in response between different cell cultures to individual carbon sources. Some attempts have been made to grow cell cultures on 'waste' and related carbon sources, such as lactose, maltose, starch, molasses and milk whey. Only maltose was found to support growth to anything near the levels observed with glucose and sucrose. In the case of molasses carbon source cell growth was either non-existent or only just measurable. All the data point to glucose as being the most suitable carbon source, principally on the grounds of biomass yield and growth rate. It should be noted, however, that other carbon sources do appear to have a major (positive) influence on natural product synthesis. Uptake into the cell is an important aspect of carbohydrate utilisation. There is strong evidence that from disaccharides upwards, major degradation to smaller units occurs before uptake. In some cases the necessary enzymes appear to be excreted into the culture broth, in others they may be located within the cell wall; invertase that hydrolyses sucrose is a good example. Once the products of carbohydrate degradation and mobilisation enter the cell they may suffer one of two fates, oxidation or utilisation for biosynthesis. The precise split between these two varies depending on such factors as cell growth rate, cell size, nutrient broth composition and carbohydrate status of the cells. In general rapidly growing cells have a high rate of oxidation, whereas cells growing more slowly tend to be more directed towards biosynthesis. Carbohydrate utilisation is a key area of study, underpinning as it does both biomass yield and natural product synthesis. (Refs. 13).

  7. Floating microbial fuel cells as energy harvesters for signal transmission from natural water bodies

    Science.gov (United States)

    Schievano, Andrea; Colombo, Alessandra; Grattieri, Matteo; Trasatti, Stefano P.; Liberale, Alessandro; Tremolada, Paolo; Pino, Claudio; Cristiani, Pierangela

    2017-02-01

    A new type of floating microbial fuel cell (fMFC) was developed for power supply of remote environmental sensors and data transmission. Ten operating fMFCs generated a cell potential in the range 100-800 mV depending on the external resistance applied. Power production peaked around 3-3.5 mW (power density of 22-28 mW m-2 cathode) after about 20-30 days of start-up period. The average of daily electrical energy harvested ranged between 10 and 35 mWh/d. Long-term performances were ensured in the presence of dense rice plants (Oryza Sativa). A power management system, based on a step-up DC/DC converter and a low-power data transmission system via SIGFOX™ technology, have been set up for the fMFCs. The tested fMFCs systems allowed to: i) harvest produced energy, ii) supply electronic devices (intermittent LED-light and a buzzer); iii) transmit remote data at low speed (three message of 12 bites each, in 6 s). Several 'floating garden' MFCs were set in the context of demonstrative events at EXPO2015 world exposition held in Milan between May-October 2015. Some of the 'floating garden' MFCs were operating for more than one year.

  8. Metabolite extraction from adherently growing mammalian cells for metabolomics studies: optimization of harvesting and extraction protocols.

    Science.gov (United States)

    Dettmer, Katja; Nürnberger, Nadine; Kaspar, Hannelore; Gruber, Michael A; Almstetter, Martin F; Oefner, Peter J

    2011-01-01

    Trypsin/ethylenediaminetetraacetic acid (EDTA) treatment and cell scraping in a buffer solution were compared for harvesting adherently growing mammalian SW480 cells for metabolomics studies. In addition, direct scraping with a solvent was tested. Trypsinated and scraped cell pellets were extracted using seven different extraction protocols including pure methanol, methanol/water, pure acetone, acetone/water, methanol/chloroform/water, methanol/isopropanol/water, and acid-base methanol. The extracts were analyzed by GC-MS after methoximation/silylation and derivatization with propyl chloroformate, respectively. The metabolic fingerprints were compared and 25 selected metabolites including amino acids and intermediates of energy metabolism were quantitatively determined. Moreover, the influence of freeze/thaw cycles, ultrasonication and homogenization using ceramic beads on extraction yield was tested. Pure acetone yielded the lowest extraction efficiency while methanol, methanol/water, methanol/isopropanol/water, and acid-base methanol recovered similar metabolite amounts with good reproducibility. Based on overall performance, methanol/water was chosen as a suitable extraction solvent. Repeated freeze/thaw cycles, ultrasonication and homogenization did not improve overall metabolite yield of the methanol/water extraction. Trypsin/EDTA treatment caused substantial metabolite leakage proving it inadequate for metabolomics studies. Gentle scraping of the cells in a buffer solution and subsequent extraction with methanol/water resulted on average in a sevenfold lower recovery of quantified metabolites compared with direct scraping using methanol/water, making the latter one the method of choice to harvest and extract metabolites from adherently growing mammalian SW480 cells.

  9. Efficiency Enhancement of InGaN-Based Solar Cells via Stacking Layers of Light-Harvesting Nanospheres

    KAUST Repository

    Alamri, Amal M.; Fu, Po-Han; Lai, Kun-Yu; Wang, Hsin-Ping; Li, Lain-Jong; He, Jr-Hau

    2016-01-01

    An effective light-harvesting scheme for InGaN-based multiple quantum well solar cells is demonstrated using stacking layers of polystyrene nanospheres. Light-harvesting efficiencies on the solar cells covered with varied stacks of nanospheres are evaluated through numerical and experimental methods. The numerical simulation reveals that nanospheres with 3 stacking layers exhibit the most improved optical absorption and haze ratio as compared to those obtained by monolayer nanospheres. The experimental demonstration, agreeing with the theoretical analyses, shows that the application of 3-layer nanospheres improves the conversion efficiency of the solar cell by ~31%.

  10. Efficiency Enhancement of InGaN-Based Solar Cells via Stacking Layers of Light-Harvesting Nanospheres

    KAUST Repository

    Alamri, Amal M.

    2016-06-24

    An effective light-harvesting scheme for InGaN-based multiple quantum well solar cells is demonstrated using stacking layers of polystyrene nanospheres. Light-harvesting efficiencies on the solar cells covered with varied stacks of nanospheres are evaluated through numerical and experimental methods. The numerical simulation reveals that nanospheres with 3 stacking layers exhibit the most improved optical absorption and haze ratio as compared to those obtained by monolayer nanospheres. The experimental demonstration, agreeing with the theoretical analyses, shows that the application of 3-layer nanospheres improves the conversion efficiency of the solar cell by ~31%.

  11. Cell culture experiments planned for the space bioreactor

    Science.gov (United States)

    Morrison, Dennis R.; Cross, John H.

    1987-01-01

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

  12. Autologous peripheral blood stem cell harvest: Collection efficiency and factors affecting it

    Directory of Open Access Journals (Sweden)

    Aseem K Tiwari

    2016-01-01

    Full Text Available Background: Harvest of hematopoietic progenitor cells via leukapheresis is being used increasingly for transplants in India. Adequate yield of cells per kilogram body weight of recipient is required for successful engraftment. Collection efficiency (CE is an objective quality parameter used to assess the quality of leukapheresis program. In this study, we calculated the CE of the ComTec cell separator (Fresenius Kabi, Germany using two different formulae (CE1 and CE2 and analyzed various patient and procedural factors, which may affect it. Materials and Methods: One hundred and one consecutive procedures in 77 autologous donors carried out over 3 years period were retrospectively reviewed. Various characteristics like gender, age, weight, disease status, hematocrit, preprocedure total leukocyte count, preprocedure CD34 positive (CD34+ cells count, preprocedure absolute CD34+ cell count and processed apheresis volume effect on CE were compared. CE for each procedure was calculated using two different formulae, and results were compared using statistical correlation and regression analysis. Results: The mean CE1 and CE2 was 41.2 and 49.1, respectively. CE2 appeared to be more accurate indicator of overall CE as it considered the impact of continued mobilization of stem cells during apheresis procedure, itself. Of all the factors affecting CE, preprocedure absolute CD34+ was the only independent factor affecting CE. Conclusion: The only factor affecting CE was preprocedure absolute CD34+ cells. Though the mean CE2 was higher than CE1, it was not statistically significant.

  13. Integration of Aqueous Two-Phase Extraction as Cell Harvest and Capture Operation in the Manufacturing Process of Monoclonal Antibodies

    Directory of Open Access Journals (Sweden)

    Axel Schmidt

    2017-12-01

    Full Text Available Substantial improvements have been made to cell culturing processes (e.g., higher product titer in recent years by raising cell densities and optimizing cultivation time. However, this has been accompanied by an increase in product-related impurities and therefore greater challenges in subsequent clarification and capture operations. Considering the paradigm shift towards the design of continuously operating dedicated plants at smaller scales—with or without disposable technology—for treating smaller patient populations due to new indications or personalized medicine approaches, the rising need for new, innovative strategies for both clarification and capture technology becomes evident. Aqueous two-phase extraction (ATPE is now considered to be a feasible unit operation, e.g., for the capture of monoclonal antibodies or recombinant proteins. However, most of the published work so far investigates the applicability of ATPE in antibody-manufacturing processes at the lab-scale and for the most part, only during the capture step. This work shows the integration of ATPE as a combined harvest and capture step into a downstream process. Additionally, a model is applied that allows early prediction of settler dimensions with high prediction accuracy. Finally, a reliable process development concept, which guides through the necessary steps, starting from the definition of the separation task to the final stages of integration and scale-up, is presented.

  14. Significant efficiency enhancement of hybrid solar cells using core-shell nanowire geometry for energy harvesting.

    Science.gov (United States)

    Tsai, Shin-Hung; Chang, Hung-Chih; Wang, Hsin-Hua; Chen, Szu-Ying; Lin, Chin-An; Chen, Show-An; Chueh, Yu-Lun; He, Jr-Hau

    2011-12-27

    A novel strategy employing core-shell nanowire arrays (NWAs) consisting of Si/regioregular poly(3-hexylthiophene) (P3HT) was demonstrated to facilitate efficient light harvesting and exciton dissociation/charge collection for hybrid solar cells (HSCs). We experimentally demonstrate broadband and omnidirectional light-harvesting characteristics of core-shell NWA HSCs due to their subwavelength features, further supported by the simulation based on finite-difference time domain analysis. Meanwhile, core-shell geometry of NWA HSCs guarantees efficient charge separation since the thickness of the P3HT shells is comparable to the exciton diffusion length. Consequently, core-shell HSCs exhibit a 61% improvement of short-circuit current for a conversion efficiency (η) enhancement of 31.1% as compared to the P3HT-infiltrated Si NWA HSCs with layers forming a flat air/polymer cell interface. The improvement of crystal quality of P3HT shells due to the formation of ordering structure at Si interfaces after air mass 1.5 global (AM 1.5G) illumination was confirmed by transmission electron microscopy and Raman spectroscopy. The core-shell geometry with the interfacial improvement by AM 1.5G illumination promotes more efficient exciton dissociation and charge separation, leading to η improvement (∼140.6%) due to the considerable increase in V(oc) from 257 to 346 mV, J(sc) from 11.7 to 18.9 mA/cm(2), and FF from 32.2 to 35.2%, which is not observed in conventional P3HT-infiltrated Si NWA HSCs. The stability of the Si/P3HT core-shell NWA HSCs in air ambient was carefully examined. The core-shell geometry should be applicable to many other material systems of solar cells and thus holds high potential in third-generation solar cells.

  15. HEMATOPOIETIC PROGENITOR CELLS AS A PREDICTIVE OF CD34+ ENUMERATION PRIOR TO PERIPHERAL BLOOD STEM CELLS HARVESTING

    Directory of Open Access Journals (Sweden)

    Z. Zulkafli

    2014-09-01

    Full Text Available Background: To date, the CD34+ cell enumeration has relied predominantly on flow cytometry technique. However, flow cytometry is time consuming and operator dependent. The application of the hematopoietic progenitor cells (HPCs channel in Sysmex XE-2100, a fully automated hematology analyzer offers an alternative approach, which is with minimal sample manipulation and less operator dependent. This study evaluates the utility of HPC counts as a predictive of CD34+ enumeration prior to peripheral blood stem cells harvesting. Materials and methods: HPC, CD34+, white blood cell (WBC, reticulocytes (retic, immature platelet fraction (IPF and immature reticulocyte fraction (IRF were determined in 61 samples from 19 patients with hematological malignancies (15 lymphoma and 4 multiple myeloma patients at Hospital Universiti Sains Malaysia (Hospital USM who had received granulocyte-colony stimulating factor (G-CSF and planned for autologous transplantation. Results: CD34+ count showed strong and significant correlation with HPC. The receiver operating characteristics (ROC curve analysis revealed that HPC count > 21.5 x 106 / L can predicts a pre harvest CD34+ count of >20 x 106 / L with sensitivity of 77%, specificity of 64% and area under the curve (AUC of 0.802. Conclusion: We concluded that HPC count can be a useful potential parameter in optimizing timing for CD34+ enumeration prior to leukapheresis.

  16. Bystander responses in three-dimensional cultures containing radiolabelled and unlabelled human cells

    International Nuclear Information System (INIS)

    Pinto, M.; Azzam, E. I.; Howell, R. W.

    2006-01-01

    Research on the radiation-induced bystander effect has been carried out mainly in 2-D tissue culture systems. This study uses a 3-D model, wherein apparently normal human diploid fibroblasts (AG1522) are grown in a carbon scaffold, to investigate the induction of a G 1 checkpoint in bystander cells present alongside radiolabelled cells. Cultures were simultaneously pulse-labelled with 3 H-deoxycytidine ( 3 HdC) to selectively irradiate a minor fraction of cells, and bromodeoxyuridine (BrdU) to identify the radiolabelled cells. After thorough washing of cultures, iododeoxyuridine (IdU) was administered to detect proliferating bystander cells. The cultures were harvested at various times thereafter, and cells were reacted with two monoclonal antibodies specific to IdU/BrdU or BrdU, respectively, stained with propidium iodide, and subjected to multi-parameter flow cytometry. Cell-cycle progression was followed in radiolabelled cells (BrdU + ) that were chronically irradiated by low energy beta particles emitted by DNA-incorporated 3 H, and in unlabelled bystander cells (BrdU - ) by a flow cytometry based cumulative labelling index assay. As expected, radiolabelled cells were delayed, in a dose-dependent manner, in G 2 and subsequently G 1 . No delay occurred in progression of bystander cells through G 1 , when the labelled cells were irradiated at dose rates up to 0.32 Gy h -1 . (authors)

  17. The Study of the Viermicompost Influence on the Harvest and on the Quality of Forage Cultures

    Directory of Open Access Journals (Sweden)

    Larisa Cremeneac

    2011-10-01

    Full Text Available The purpose of the research was the appreciation of the viermicompost quality. This material was obtained as a result of bioconversion of organically wastes (obtained from cattle by worm cultivation, using wormculture (especially the Red Hybrid of California. The bioconversion process of organic wastes is implemented in the Experimental Section of the Scientific and Practical Institute of Biotechnologies in Animal Husbandry and Veterinary Medicine. As a result of the studies to effect in several states, including the Republic of Moldova, it has been stated that the veirmicompost has a positive influence on the productivity of cultures, diminishing the period of culture growing , increases their resistance to infavorable clamaterical conditions and to frequent phitotechnical maligns. In the agricultural production obtained as a result of cultivating on viermicompost. As a result of the research it has dean stated that the quantity of the C vitamin in some of the vegetables has increased by 1,5 – 9,4 times. What s more the viermicompost has influenced positively the quality of the alfalfa, maize and sugar fodder, in which the azoth compounds quantity has diminished by 1,32 – 2,66; 3,47 – 3,76 and 1,10 – 1,14 times, in comparison with cultures cultivars tied with the help of mineral fertilizer. So, the agricultural and ecological importance of the viermicompost consists in improving the quality of agricultural production.

  18. Manufacturing of Proteins and Antibodies: Chapter Downstream Processing Technologies : Harvest Operations.

    Science.gov (United States)

    Turner, Richard; Joseph, Adrian; Titchener-Hooker, Nigel; Bender, Jean

    2017-08-04

    Cell harvesting is the separation or retention of cells and cellular debris from the supernatant containing the target molecule Selection of harvest method strongly depends on the type of cells, mode of bioreactor operation, process scale, and characteristics of the product and cell culture fluid. Most traditional harvesting methods use some form of filtration, centrifugation, or a combination of both for cell separation and/or retention. Filtration methods include normal flow depth filtration and tangential flow microfiltration. The ability to scale down predictably the selected harvest method helps to ensure successful production and is critical for conducting small-scale characterization studies for confirming parameter targets and ranges. In this chapter we describe centrifugation and depth filtration harvesting methods, share strategies for harvest optimization, present recent developments in centrifugation scale-down models, and review alternative harvesting technologies.

  19. Transfection in Primary Cultured Neuronal Cells.

    Science.gov (United States)

    Marwick, Katie F M; Hardingham, Giles E

    2017-01-01

    Transfection allows the introduction of foreign nucleic acid into eukaryotic cells. It is an important tool in understanding the roles of NMDARs in neurons. Here, we describe using lipofection-mediated transfection to introduce cDNA encoding NMDAR subunits into postmitotic rodent primary cortical neurons maintained in culture.

  20. Cell Culture Microfluidic Biochips: Experimental Throughput Maximization

    DEFF Research Database (Denmark)

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

    2011-01-01

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

  1. Plant Cell Culture Initiation: practical tips

    NARCIS (Netherlands)

    Hall, R.D.

    2001-01-01

    The use of cultured plant cells in either organized or unorganized form has increased vey considerably in the last 10-15 yr. Many new technologies have been developed and applications in both fundamental and applied research have led to the development of some powerful tools for improving our

  2. 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

  3. 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

  4. Mild hypothermic culture conditions affect residual host cell protein composition post-Protein A chromatography.

    Science.gov (United States)

    Goey, Cher Hui; Bell, David; Kontoravdi, Cleo

    2018-04-01

    Host cell proteins (HCPs) are endogenous impurities, and their proteolytic and binding properties can compromise the integrity, and, hence, the stability and efficacy of recombinant therapeutic proteins such as monoclonal antibodies (mAbs). Nonetheless, purification of mAbs currently presents a challenge because they often co-elute with certain HCP species during the capture step of protein A affinity chromatography. A Quality-by-Design (QbD) strategy to overcome this challenge involves identifying residual HCPs and tracing their source to the harvested cell culture fluid (HCCF) and the corresponding cell culture operating parameters. Then, problematic HCPs in HCCF may be reduced by cell engineering or culture process optimization. Here, we present experimental results linking cell culture temperature and post-protein A residual HCP profile. We had previously reported that Chinese hamster ovary cell cultures conducted at standard physiological temperature and with a shift to mild hypothermia on day 5 produced HCCF of comparable product titer and HCP concentration, but with considerably different HCP composition. In this study, we show that differences in HCP variety at harvest cascaded to downstream purification where different residual HCPs were present in the two sets of samples post-protein A purification. To detect low-abundant residual HCPs, we designed a looping liquid chromatography-mass spectrometry method with continuous expansion of a preferred, exclude, and targeted peptide list. Mild hypothermic cultures produced 20% more residual HCP species, especially cell membrane proteins, distinct from the control. Critically, we identified that half of the potentially immunogenic residual HCP species were different between the two sets of samples.

  5. 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

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

    Science.gov (United States)

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

    1998-01-01

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

  7. 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

    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...... on cell survival. Low grade light exposure was however compatible with optical recordings as well as cell viability. These results strongly indicate that a cell culture chip could be constructed that allowed for on-line optical recording of cellular events without affecting the cell culturing condition...

  8. 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...... on cell survival. Low grade light exposure was however compatible with optical recordings as well as cell viability. These results strongly indicate that a cell culture chip could be constructed that allowed for on-line optical recording of cellular events without affecting the cell culturing condition...

  9. A cell culture technique for human epiretinal membranes to describe cell behavior and membrane contraction in vitro.

    Science.gov (United States)

    Wertheimer, Christian; Eibl-Lindner, Kirsten H; Compera, Denise; Kueres, Alexander; Wolf, Armin; Docheva, Denitsa; Priglinger, Siegfried G; Priglinger, Claudia; Schumann, Ricarda G

    2017-11-01

    To introduce a human cell culture technique for investigating in-vitro behavior of primary epiretinal cells and membrane contraction of fibrocellular tissue surgically removed from eyes with idiopathic macular pucker. Human epiretinal membranes were harvested from ten eyes with idiopathic macular pucker during standard vitrectomy. Specimens were fixed on cell culture plastic using small entomological pins to apply horizontal stress to the tissue, and then transferred to standard cell culture conditions. Cell behavior of 400 epiretinal cells from 10 epiretinal membranes was observed in time-lapse microscopy and analyzed in terms of cell migration, cell velocity, and membrane contraction. Immunocytochemistry was performed for cell type-specific antigens. Cell specific differences in migration behavior were observed comprising two phenotypes: (PT1) epiretinal cells moving fast, less directly, with small round phenotype and (PT2) epiretinal cells moving slowly, directly, with elongated large phenotype. No mitosis, no outgrowth and no migration onto the plastic were seen. Horizontal contraction measurements showed variation between specimens. Masses of epiretinal cells with a myofibroblast-like phenotype expressed cytoplasmatic α-SMA stress fibers and correlated with cell behavior characteristics (PT2). Fast moving epiretinal cells (PT1) were identified as microglia by immunostaining. This in-vitro technique using traction application allows for culturing surgically removed epiretinal membranes from eyes with idiopathic macular pucker, demonstrating cell behavior and membrane contraction of primary human epiretinal cells. Our findings emphasize the abundance of myofibroblasts, the presence of microglia and specific differences of cell behavior in these membranes. This technique has the potential to improve the understanding of pathologies at the vitreomacular interface and might be helpful in establishing anti-fibrotic treatment strategies.

  10. Lipoprotein receptors in cultured bovine endothelial cells

    International Nuclear Information System (INIS)

    Struempfer, A.E.M.

    1983-07-01

    In this study, receptors that may be involved in the uptake of low density lipoproteins (LDL) and low density lipoproteins which have been modified by acetylation (AcLDL), were characterized. Aortic epithelial cells were used and a cell culture system which closely resembled the in vivo monolayer was established. Endothelial cell and lipoprotein interactions were examined by incubating the cells with 125 l-labelled lipoproteins under various conditions. The receptor affinity of bovine aortic endothelial cells was higher for AcLDL than that for LDL. Competition studies demonstrated that there were two distinct receptors for LDL and AcLDL on the endothelial cells. AcLDL did not compete with LDL for the LDL receptor, and conversely LDL did not compete with AcLDL for the AcLDL receptor. The receptor activities for LDL and AcLDL were examined as a function of culture age. Whereas the LDL receptor could be regulated, the AcLDL receptor was not as susceptible to regulation. Upon exposing endothelial cells for 72 h to either LDL or AcLDL, it was found that the total amount of cellular cholesterol increased by about 50%. However, the increase of total cholesterol was largely in the form of free cholesterol. This is in contrast to macrophages, where the increase in total cholesterol upon exposure to AcLDL is largely in the form cholesteryl esters

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

    Science.gov (United States)

    Cogoli, Augusto

    1992-01-01

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

  12. 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... Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in vitro cultivated cell lines from the tissue of humans or other animals which are used in various diagnostic...

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

    Directory of Open Access Journals (Sweden)

    Rebecca C. Schugar

    2009-01-01

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

  14. Circulating hematopoietic progenitors and CD34+ cells predicted successful hematopoietic stem cell harvest in myeloma and lymphoma patients: experiences from a single institution

    Directory of Open Access Journals (Sweden)

    Yu JT

    2016-02-01

    Full Text Available Jui-Ting Yu,1,2,* Shao-Bin Cheng,3,* Youngsen Yang,1 Kuang-Hsi Chang,4 Wen-Li Hwang,1 Chieh-Lin Jerry Teng,1,5,6 1Division of Hematology/Medical Oncology, Department of Medicine, Taichung Veterans General Hospital, 2Division of Hematology/Medical Oncology, Tungs' Taichung MetroHarbor Hospital, 3Division of General Surgery, Department of Surgery, 4Department of Medical Research and Education, Taichung Veterans General Hospital, 5Department of Life Science, Tunghai University, 6School of Medicine, Chung Shan Medical University, Taichung, Taiwan, Republic of China *These authors contributed equally to this work Background: Previous studies have shown that the numbers of both circulating hematopoietic progenitor cell (HPC and CD34+ cell are positively correlated with CD34+ cell harvest yield. However, the minimal numbers of both circulating HPCs and CD34+ cells required for performing an efficient hematopoietic stem cell (HSC harvest in lymphoma and myeloma patients have not been defined in our institution. Patients and methods: Medical records of 50 lymphoma and myeloma patients undergoing peripheral blood HSC harvest in our institution were retrospectively reviewed. The minimal and optimal HSC harvest yield required for the treatment was considered to be ≥2×106 CD34+ cells/kg and ≥5×106 CD34+ cells/kg, respectively. Results: The minimally required or optimal HSC yield obtained was not influenced by age (≥60 years, sex, underlying malignancies, disease status, multiple rounds of chemotherapy, or history of radiotherapy. The numbers of both circulating HPC and CD34+ cell were higher in patients with minimally required HSC yields (P=0.000 for HPC and P=0.000 for CD34+ cell and also in patients with optimal HSC yields (P=0.011 for HPC and P=0.006 for CD34+ cell. The cell count cutoff for obtaining minimally required HSC harvest was determined to be 20/mm3 for HPCs and 10/mm3 for CD34+ cells. Furthermore, the cell count cutoff for obtaining

  15. 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........

  16. Thermo-responsive cell culture carrier: Effects on macrophage functionality and detachment efficiency.

    Science.gov (United States)

    Rennert, Knut; Nitschke, Mirko; Wallert, Maria; Keune, Natalie; Raasch, Martin; Lorkowski, Stefan; Mosig, Alexander S

    2017-01-01

    Harvesting cultivated macrophages for tissue engineering purposes by enzymatic digestion of cell adhesion molecules can potentially result in unintended activation, altered function, or behavior of these cells. Thermo-responsive polymer is a promising tool that allows for gentle macrophage detachment without artificial activation prior to subculture within engineered tissue constructs. We therefore characterized different species of thermo-responsive polymers for their suitability as cell substrate and to mediate gentle macrophage detachment by temperature shift. Primary human monocyte- and THP-1-derived macrophages were cultured on thermo-responsive polymers and characterized for phagocytosis and cytokine secretion in response to lipopolysaccharide stimulation. We found that both cell types differentially respond in dependence of culture and stimulation on thermo-responsive polymers. In contrast to THP-1 macrophages, primary monocyte-derived macrophages showed no signs of impaired viability, artificial activation, or altered functionality due to culture on thermo-responsive polymers compared to conventional cell culture. Our study demonstrates that along with commercially available UpCell carriers, two other thermo-responsive polymers based on poly(vinyl methyl ether) blends are attractive candidates for differentiation and gentle detachment of primary monocyte-derived macrophages. In summary, we observed similar functionality and viability of primary monocyte-derived macrophages cultured on thermo-responsive polymers compared to standard cell culture surfaces. While this first generation of custom-made thermo-responsive polymers does not yet outperform standard culture approaches, our results are very promising and provide the basis for exploiting the unique advantages offered by custom-made thermo-responsive polymers to further improve macrophage culture and recovery in the future, including the covalent binding of signaling molecules and the reduction of

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

    Science.gov (United States)

    Halldorsson, Skarphedinn; Lucumi, Edinson; Gómez-Sjöberg, Rafael; Fleming, Ronan M T

    2015-01-15

    Culture of cells using various microfluidic devices is becoming more common within experimental cell biology. At the same time, a technological radiation of microfluidic cell culture device designs is currently in progress. Ultimately, the utility of microfluidic cell culture will be determined by its capacity to permit new insights into cellular function. Especially insights that would otherwise be difficult or impossible to obtain with macroscopic cell culture in traditional polystyrene dishes, flasks or well-plates. Many decades of heuristic optimization have gone into perfecting conventional cell culture devices and protocols. In comparison, even for the most commonly used microfluidic cell culture devices, such as those fabricated from polydimethylsiloxane (PDMS), collective understanding of the differences in cellular behavior between microfluidic and macroscopic culture is still developing. Moving in vitro culture from macroscopic culture to PDMS based devices can come with unforeseen challenges. Changes in device material, surface coating, cell number per unit surface area or per unit media volume may all affect the outcome of otherwise standard protocols. In this review, we outline some of the advantages and challenges that may accompany a transition from macroscopic to microfluidic cell culture. We focus on decisive factors that distinguish macroscopic from microfluidic cell culture to encourage a reconsideration of how macroscopic cell culture principles might apply to microfluidic cell culture. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

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

    2000-01-01

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

  19. Comparison of different options for harvest of a therapeutic protein product from high cell density yeast fermentation broth.

    Science.gov (United States)

    Wang, Alice; Lewus, Rachael; Rathore, Anurag S

    2006-05-05

    Recovery of therapeutic protein from high cell density yeast fermentations at commercial scale is a challenging task. In this study, we investigate and compare three different harvest approaches, namely centrifugation followed by depth filtration, centrifugation followed by filter-aid enhanced depth filtration, and microfiltration. This is achieved by presenting a case study involving recovery of a therapeutic protein from Pichia pastoris fermentation broth. The focus of this study is on performance of the depth filtration and the microfiltration steps. The experimental data has been fitted to the conventional models for cake filtration to evaluate specific cake resistance and cake compressibility. In the case of microfiltration, the experimental data agrees well with flux predicted by shear induced diffusion model. It is shown that, under optimal conditions, all three options can deliver the desired product recovery ( >80%), harvest time ( making a final decision on a harvesting approach.

  20. Energy harvesting from organic liquids in micro-sized microbial fuel cells

    KAUST Repository

    Mink, J.E.

    2014-03-07

    Micro-sized microbial fuel cells (MFCs) are miniature energy harvesters that use bacteria to convert biomass from liquids into usable power. The key challenge is transitioning laboratory test beds into devices capable of producing high power using readily available fuel sources. Here, we present a pragmatic step toward advancing MFC applications through the fabrication of a uniquely mobile and inexpensive micro-sized device that can be fueled with human saliva. The 25-ll MFC was fabricated with graphene, a two-dimensional atomic crystal-structured material, as an anode for efficient current generation and with an air cathode for enabling the use of the oxygen present in air, making its operation completely mobile and free of the need for laboratory chemicals. With saliva as a fuel, the device produced higher current densities (1190 Am-3) than any previous aircathode micro-sized MFCs. The use of the graphene anode generated 40 times more power than that possible using a carbon cloth anode. Additional tests were performed using acetate, a conventional organic material, at high organic loadings that were comparable to those in saliva, and the results demonstrated a linear relationship between the organic loading and current. These findings open the door to saliva-powered applications of this fuel cell technology for Lab-on-a-Chip devices or portable point-of-care diagnostic devices. 2014 Nature Publishing Group All rights reserved 1884-4057/14.

  1. In silico characterization of cell-cell interactions using a cellular automata model of cell culture.

    Science.gov (United States)

    Kihara, Takanori; Kashitani, Kosuke; Miyake, Jun

    2017-07-14

    Cell proliferation is a key characteristic of eukaryotic cells. During cell proliferation, cells interact with each other. In this study, we developed a cellular automata model to estimate cell-cell interactions using experimentally obtained images of cultured cells. We used four types of cells; HeLa cells, human osteosarcoma (HOS) cells, rat mesenchymal stem cells (MSCs), and rat smooth muscle A7r5 cells. These cells were cultured and stained daily. The obtained cell images were binarized and clipped into squares containing about 10 4 cells. These cells showed characteristic cell proliferation patterns. The growth curves of these cells were generated from the cell proliferation images and we determined the doubling time of these cells from the growth curves. We developed a simple cellular automata system with an easily accessible graphical user interface. This system has five variable parameters, namely, initial cell number, doubling time, motility, cell-cell adhesion, and cell-cell contact inhibition (of proliferation). Within these parameters, we obtained initial cell numbers and doubling times experimentally. We set the motility at a constant value because the effect of the parameter for our simulation was restricted. Therefore, we simulated cell proliferation behavior with cell-cell adhesion and cell-cell contact inhibition as variables. By comparing growth curves and proliferation cell images, we succeeded in determining the cell-cell interaction properties of each cell. Simulated HeLa and HOS cells exhibited low cell-cell adhesion and weak cell-cell contact inhibition. Simulated MSCs exhibited high cell-cell adhesion and positive cell-cell contact inhibition. Simulated A7r5 cells exhibited low cell-cell adhesion and strong cell-cell contact inhibition. These simulated results correlated with the experimental growth curves and proliferation images. Our simulation approach is an easy method for evaluating the cell-cell interaction properties of cells.

  2. Three-dimensional culture of human mesenchymal stem cells in a polyethylene terephthalate matrix

    International Nuclear Information System (INIS)

    Cao Yanfen; Li Ding; Shang Chunhua; Wang Jufang; Wang Xiaoning; Yang Shangtian

    2010-01-01

    Polyethylene terephthalate (PET) was used as the scaffold material to support the proliferation of human mesenchymal stem cells (hMSCs). The cells were cultured either statically in multi-wells or in a spinner flask agitated at 80 rpm for up to 20 days. To optimize the cell expansion condition, effects of the initial cell density and basic fibroblast growth factor (bFGF) were examined. During culture, cell growth and metabolism were tested. After 20 days, cells were harvested and surface markers were identified and quantified with flow cytometry. The results showed that hMSCs seeded at the lowest density gave the highest expansion fold. hMSCs grown in porous three-dimensional (3D) matrices displayed significantly different characteristics in terms of their proliferation and metabolism. PET matrices with 3D space could sustain cell proliferation for a long time. In addition, a low concentration (5 ng mL -1 ) of bFGF significantly enhanced the expansion of hMSCs in PET. Cell attachment and distribution in PET matrices were studied with confocal laser microscopy and scanning electron microscopy, which also confirmed cell proliferation. Furthermore, most of the cells in PET matrices were CD29, CD44 and CD105 positive, and CD34, CD45 and CD14 negative, confirming that hMSCs cultured in 3D PET matrices can be expanded and maintained in their undifferentiated state for at least 20 days without subculturing.

  3. Comparison of Cellular Alterations in Fat Cells Harvested With Laser-Assisted Liposuction and Suction-Assisted Liposuction.

    Science.gov (United States)

    Yildiz, Kemalettin; Taşli, Pakize Neslihan; Şahin, Fikrettin; Güneren, Ethem

    2016-05-01

    The aim of the present study was to evaluate the viability and proliferative capacity of adipose-derived stem cells obtained by laser-assisted liposuction (LAL). Fat tissue was obtained from 7 male patients treated surgically for gynecomastia. On one side, harvesting was made before LAL, while it was implemented after LAL on the contralateral side. Viability, cell surface antigens, pluripotency, and apoptosis were assessed and compared in these samples. Cells harvested before and after LAL did not exhibit any significant difference in terms of surface cell markers. Number of viable stem cells was lower initially after exposure to laser, while this difference was reversed at the end of 72 hours. Genetic indicators of cellular differentiation were similar in both groups. Apoptosis indicators were increased remarkably after laser exposure in the first 24 hours, but this increase was absent 72 hours after LAL procedure. The authors' results have promising clinical relevance since mesenchymal stem cells harvested during LAL have maintained appropriate cellular features to be used for autologous fat transfer and fat grafting.

  4. Rapid and reliable identification of Gram-negative bacteria and Gram-positive cocci by deposition of bacteria harvested from blood cultures onto the MALDI-TOF plate.

    OpenAIRE

    Barnini, S; Ghelardi, Emilia; Brucculeri, V; Morici, Paola; Lupetti, Antonella

    2015-01-01

    Background Rapid identification of the causative agent(s) of bloodstream infections using the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) methodology can lead to increased empirical antimicrobial therapy appropriateness. Herein, we aimed at establishing an easier and simpler method, further referred to as the direct method, using bacteria harvested by serum separator tubes from positive blood cultures and placed onto the polished steel target plate for rapid identif...

  5. Comparison of Amicus and COBE Spectra for allogenic peripheral blood stem cell harvest: Study from tertiary care centre in India.

    Science.gov (United States)

    Setia, Rasika Dhawan; Arora, Satyam; Handoo, Anil; Dadu, Tina; Choudhary, Dharma; Sharma, Sajeev Kumar; Kharya, Gaurav; Khandelwal, Vipin; Sachdeva, Prerna; Doval, Divya; Bakliwal, Anamika; Kapoor, Meenu; Bajaj, Shalu; Bachchas, Virendra; Singh, Praveen

    2017-06-01

    Most common source of stem cell graft for both autologous and allogenic haematopoietic transplants are peripheral blood haematopoietic progenitor stem cells. Adequate collection of the CD34+ cells and safety of the allogenic donor during the leukapheresis are of prime importance to an apheresis physician. Our retrospective analysis is a comparison between of two platforms namely, COBE Spectra and Amicus, for CD34+ mononuclear cell collection. The study included the data of GSCF (Granulocyte-Colony-Stimulating Factor) mobilized allogenic PBSC collections at our centre from January 2015 to June 2016. The apheresis platforms used were COBE Spectra and Amicus. Blood cell counts were done using LH750 Beckman Coulter (Florida, Miami, USA). CD45+ & CD34+ cell counts were done using BD FACS Canto-II Flow-Cytometer by ISHAGE guidelines. A total of 170 PBSC (100 COBE Spectra & 70 Amicus) harvests were done on 143 donors, of which 116 completed the collection in a single session and 27 required a second session. Demographic details and pre harvest peripheral blood counts for both the groups did not show any statistical differences. Amicus processed higher blood volume with higher ACD exposure and procedure time compared to COBE Spectra. Higher platelets loss was with COBE Spectra harvests with higher product volumes collection. Collection efficiency (CE2), collection ratio, CD34+ cells dose was similar on both the platforms. RBC contamination, absolute lymphocyte and monocytes counts were significantly higher with Amicus harvest product compared with COBE Spectra. A total of 14 (8.2%; citrate toxicity) adverse reactions were reported out of 170 allogenic PBSC collections. Our study suggests that both Amicus and COBE Spectra platforms offer comparable results for allogenic PBSC collections. Amicus offers a concentrated PBSC product with lesser volume and platelets loss but higher RBC contamination. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. 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. © 2015 International Federation for Cell Biology.

  7. 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.

  8. Traditional and Modern Cell Culture in Virus Diagnosis.

    Science.gov (United States)

    Hematian, Ali; Sadeghifard, Nourkhoda; Mohebi, Reza; Taherikalani, Morovat; Nasrolahi, Abbas; Amraei, Mansour; Ghafourian, Sobhan

    2016-04-01

    Cell cultures are developed from tissue samples and then disaggregated by mechanical, chemical, and enzymatic methods to extract cells suitable for isolation of viruses. With the recent advances in technology, cell culture is considered a gold standard for virus isolation. This paper reviews the evolution of cell culture methods and demonstrates why cell culture is a preferred method for identification of viruses. In addition, the advantages and disadvantages of both traditional and modern cell culture methods for diagnosis of each type of virus are discussed. Detection of viruses by the novel cell culture methods is considered more accurate and sensitive. However, there is a need to include some more accurate methods such as molecular methods in cell culture for precise identification of viruses.

  9. 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.

  10. Energy transfer in nanowire solar cells with photon-harvesting shells

    KAUST Repository

    Peters, C. H.

    2009-01-01

    The concept of a nanowire solar cell with photon-harvesting shells is presented. In this architecture, organic molecules which absorb strongly in the near infrared where silicon absorbs weakly are coupled to silicon nanowires (SiNWs). This enables an array of 7-μm -long nanowires with a diameter of 50 nm to absorb over 85% of the photons above the bandgap of silicon. The organic molecules are bonded to the surface of the SiNWs forming a thin shell. They absorb the low-energy photons and subsequently transfer the energy to the SiNWs via Förster resonant energy transfer, creating free electrons and holes within the SiNWs. The carriers are then separated at a radial p-n junction in a nanowire and extracted at the respective electrodes. The shortness of the nanowires is expected to lower the dark current due to the decrease in p-n junction surface area, which scales linearly with wire length. The theoretical power conversion efficiency is 15%. To demonstrate this concept, we measure a 60% increase in photocurrent from a planar silicon-on-insulator diode when a 5 nm layer of poly[2-methoxy-5-(2′ -ethyl-hexyloxy)-1,4-phenylene vinylene is applied to the surface of the silicon. This increase is in excellent agreement with theoretical predictions. © 2009 American Institute of Physics.

  11. Good cell culture practices &in vitro toxicology.

    Science.gov (United States)

    Eskes, Chantra; Boström, Ann-Charlotte; Bowe, Gerhard; Coecke, Sandra; Hartung, Thomas; Hendriks, Giel; Pamies, David; Piton, Alain; Rovida, Costanza

    2017-12-01

    Good Cell Culture Practices (GCCP) is of high relevance to in vitro toxicology. The European Society of Toxicology In Vitro (ESTIV), the Center for Alternatives for Animal Testing (CAAT) and the In Vitro Toxicology Industrial Platform (IVTIP) joined forces to address by means of an ESTIV 2016 pre-congress session the different aspects and applications of GCCP. The covered aspects comprised the current status of the OECD guidance document on Good In Vitro Method Practices, the importance of quality assurance for new technological advances in in vitro toxicology including stem cells, and the optimized implementation of Good Manufacturing Practices and Good Laboratory Practices for regulatory testing purposes. General discussions raised the duality related to the difficulties in implementing GCCP in an academic innovative research framework on one hand, and on the other hand, the need for such GCCP principles in order to ensure reproducibility and robustness of in vitro test methods for toxicity testing. Indeed, if good cell culture principles are critical to take into consideration for all uses of in vitro test methods for toxicity testing, the level of application of such principles may depend on the stage of development of the test method as well as on the applications of the test methods, i.e., academic innovative research vs. regulatory standardized test method. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Harvesting Cultural Knowledge.

    Science.gov (United States)

    Keating, Joseph F.

    1997-01-01

    Describes a year-long course called Outdoor Science taught at an American Indian reservation high school that demonstrates to students the connection between traditional tribal knowledge and western science to spark student interest in science. Presents a list that contains references on the subject of ethnobotany. Includes specific references for…

  13. Enhanced infectivity of bluetongue virus in cell culture by centrifugation.

    OpenAIRE

    Sundin, D R; Mecham, J O

    1989-01-01

    The effects of centrifugation of the infection of cell culture with bluetongue virus (BTV) were investigated. Baby hamster kidney cells were infected with BTV with or without centrifugation. Viral antigen was detected by immunofluorescence at 24 h in both centrifuged and noncentrifuged cultures. However, after 24 h of infection, the production of PFU in centrifuged cell cultures was 10- to 20-fold greater than that seen in cultures not centrifuged. In addition, centrifugation enhanced the dir...

  14. 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…

  15. 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

  16. Turbulent Dynamics of Epithelial Cell Cultures

    Science.gov (United States)

    Blanch-Mercader, C.; Yashunsky, V.; Garcia, S.; Duclos, G.; Giomi, L.; Silberzan, P.

    2018-05-01

    We investigate the large length and long time scales collective flows and structural rearrangements within in vitro human bronchial epithelial cell (HBEC) cultures. Activity-driven collective flows result in ensembles of vortices randomly positioned in space. By analyzing a large population of vortices, we show that their area follows an exponential law with a constant mean value and their rotational frequency is size independent, both being characteristic features of the chaotic dynamics of active nematic suspensions. Indeed, we find that HBECs self-organize in nematic domains of several cell lengths. Nematic defects are found at the interface between domains with a total number that remains constant due to the dynamical balance of nucleation and annihilation events. The mean velocity fields in the vicinity of defects are well described by a hydrodynamic theory of extensile active nematics.

  17. Recombinant Protein Production and Insect Cell Culture and Process

    Science.gov (United States)

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

    1997-01-01

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

  18. Seamless Combination of Fluorescence-Activated Cell Sorting and Hanging-Drop Networks for Individual Handling and Culturing of Stem Cells and Microtissue Spheroids.

    Science.gov (United States)

    Birchler, Axel; Berger, Mischa; Jäggin, Verena; Lopes, Telma; Etzrodt, Martin; Misun, Patrick Mark; Pena-Francesch, Maria; Schroeder, Timm; Hierlemann, Andreas; Frey, Olivier

    2016-01-19

    Open microfluidic cell culturing devices offer new possibilities to simplify loading, culturing, and harvesting of individual cells or microtissues due to the fact that liquids and cells/microtissues are directly accessible. We present a complete workflow for microfluidic handling and culturing of individual cells and microtissue spheroids, which is based on the hanging-drop network concept: The open microfluidic devices are seamlessly combined with fluorescence-activated cell sorting (FACS), so that individual cells, including stem cells, can be directly sorted into specified culturing compartments in a fully automated way and at high accuracy. Moreover, already assembled microtissue spheroids can be loaded into the microfluidic structures by using a conventional pipet. Cell and microtissue culturing is then performed in hanging drops under controlled perfusion. On-chip drop size control measures were applied to stabilize the system. Cells and microtissue spheroids can be retrieved from the chip by using a parallelized transfer method. The presented methodology holds great promise for combinatorial screening of stem-cell and multicellular-spheroid cultures.

  19. Differential marker expression by cultures rich in mesenchymal stem cells

    Science.gov (United States)

    2013-01-01

    Background Mesenchymal stem cells have properties that make them amenable to therapeutic use. However, the acceptance of mesenchymal stem cells in clinical practice requires standardized techniques for their specific isolation. To date, there are no conclusive marker (s) for the exclusive isolation of mesenchymal stem cells. Our aim was to identify markers differentially expressed between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. We compared and contrasted the phenotype of tissue cultures in which mesenchymal stem cells are rich and rare. By initially assessing mesenchymal stem cell differentiation, we established that bone marrow and breast adipose cultures are rich in mesenchymal stem cells while, in our hands, foreskin fibroblast and olfactory tissue cultures contain rare mesenchymal stem cells. In particular, olfactory tissue cells represent non-stem cell mesenchymal cells. Subsequently, the phenotype of the tissue cultures were thoroughly assessed using immuno-fluorescence, flow-cytometry, proteomics, antibody arrays and qPCR. Results Our analysis revealed that all tissue cultures, regardless of differentiation potential, demonstrated remarkably similar phenotypes. Importantly, it was also observed that common mesenchymal stem cell markers, and fibroblast-associated markers, do not discriminate between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. Examination and comparison of the phenotypes of mesenchymal stem cell and non-stem cell mesenchymal cell cultures revealed three differentially expressed markers – CD24, CD108 and CD40. Conclusion We indicate the importance of establishing differential marker expression between mesenchymal stem cells and non-stem cell mesenchymal cells in order to determine stem cell specific markers. PMID:24304471

  20. 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)

  1. Protection of cultured mammalian cells by rebamipide

    International Nuclear Information System (INIS)

    Antoku, Shigetoshi; Aramaki, Ryoji; Tanaka, Hisashi; Kusumoto, Naotoshi.

    1997-01-01

    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 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 14 C increased with increasing contact time with rebamipide. These rebamipide mainly distributed in nucleus rather than cytoplasm. (author)

  2. Cultured circulating tumor cells and their derived xenografts for personalized oncology

    Directory of Open Access Journals (Sweden)

    Ruoxiang Wang

    2016-10-01

    Full Text Available Recent cancer research has demonstrated the existence of circulating tumor cells (CTCs in cancer patient's blood. Once identified, CTC biomarkers will be invaluable tools for clinical diagnosis, prognosis and treatment. In this review, we propose ex vivo culture as a rational strategy for large scale amplification of the limited numbers of CTCs from a patient sample, to derive enough CTCs for accurate and reproducible characterization of the biophysical, biochemical, gene expressional and behavioral properties of the harvested cells. Because of tumor cell heterogeneity, it is important to amplify all the CTCs in a blood sample for a comprehensive understanding of their role in cancer metastasis. By analyzing critical steps and technical issues in ex vivo CTC culture, we developed a cost-effective and reproducible protocol directly culturing whole peripheral blood mononuclear cells, relying on an assumed survival advantage in CTCs and CTC-like cells over the normal cells to amplify this specified cluster of cancer cells.

  3. The evolution of chicken stem cell culture methods.

    Science.gov (United States)

    Farzaneh, M; Attari, F; Mozdziak, P E; Khoshnam, S E

    2017-12-01

    1. The avian embryo is an excellent model for studying embryology and the production of pharmaceutical proteins in transgenic chickens. Furthermore, chicken stem cells have the potential for proliferation and differentiation and emerged as an attractive tool for various cell-based technologies. 2. The objective of these studies is the derivation and culture of these stem cells is the production of transgenic birds for recombinant biomaterials and vaccine manufacture, drug and cytotoxicity testing, as well as to gain insight into basic science, including cell tracking. 3. Despite similarities among the established chicken stem cell lines, fundamental differences have been reported between their culture conditions and applications. Recent conventional protocols used for expansion and culture of chicken stem cells mostly depend on feeder cells, serum-containing media and static culture. 4. Utilising chicken stem cells for generation of cell-based transgenic birds and a variety of vaccines requires large-scale cell production. However, scaling up the conventional adherent chicken stem cells is challenging and labour intensive. Development of a suspension cell culture process for chicken embryonic stem cells (cESCs), chicken primordial germ cells (PGCs) and chicken induced pluripotent stem cells (ciPSCs) will be an important advance for increasing the growth kinetics of these cells. 6. This review describes various approaches and suggestions to achieve optimal cell growth for defined chicken stem cells cultures and use in future manufacturing applications.

  4. In vitro expansion and differentiation of rat pancreatic duct-derived stem cells into insulin secreting cells using a dynamicthree-dimensional cell culture system.

    Science.gov (United States)

    Chen, X C; Liu, H; Li, H; Cheng, Y; Yang, L; Liu, Y F

    2016-06-27

    In this study, a dynamic three-dimensional cell culture technology was used to expand and differentiate rat pancreatic duct-derived stem cells (PDSCs) into islet-like cell clusters that can secrete insulin. PDSCs were isolated from rat pancreatic tissues by in situ collagenase digestion and density gradient centrifugation. Using a dynamic three-dimensional culture technique, the cells were expanded and differentiated into functional islet-like cell clusters, which were characterized by morphological and phenotype analyses. After maintaining 1 x 108 isolated rat PDSCs in a dynamic three-dimensional cell culture for 7 days, 1.5 x 109 cells could be harvested. Passaged PDSCs expressed markers of pancreatic endocrine progenitors, including CD29 (86.17%), CD73 (90.73%), CD90 (84.13%), CD105 (78.28%), and Pdx-1. Following 14 additional days of culture in serum-free medium with nicotinamide, keratinocyte growth factor (KGF), and b fibroblast growth factor (FGF), the cells were differentiated into islet-like cell clusters (ICCs). The ICC morphology reflected that of fused cell clusters. During the late stage of differentiation, representative clusters were non-adherent and expressed insulin indicated by dithizone (DTZ)-positive staining. Insulin was detected in the extracellular fluid and cytoplasm of ICCs after 14 days of differentiation. Additionally, insulin levels were significantly higher at this time compared with the levels exhibited by PDSCs before differentiation (P cell culture system, PDSCs can be expanded in vitro and can differentiate into functional islet-like cell clusters.

  5. Multiweek cell culture project for use in upper-level biology laboratories.

    Science.gov (United States)

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

    2012-06-01

    This article describes a laboratory protocol for a multiweek project piloted in a new upper-level biology laboratory (BIO 426) using cell culture techniques. Human embryonic kidney-293 cells were used, and several culture media and supplements were identified for students to design their own experiments. Treatments included amino acids, EGF, caffeine, epinephrine, heavy metals, and FBS. Students researched primary literature to determine their experimental variables, made their own solutions, and treated their cells over a period of 2 wk. Before this, a sterile technique laboratory was developed to teach students how to work with the cells and minimize contamination. Students designed their experiments, mixed their solutions, seeded their cells, and treated them with their control and experimental media. Students had the choice of manipulating a number of variables, including incubation times, exposure to treatment media, and temperature. At the end of the experiment, students observed the effects of their treatment, harvested and dyed their cells, counted relative cell numbers in control and treatment flasks, and determined the ratio of living to dead cells using a hemocytometer. At the conclusion of the experiment, students presented their findings in a poster presentation. This laboratory can be expanded or adapted to include additional cell lines and treatments. The ability to design and implement their own experiments has been shown to increase student engagement in the biology-related laboratory activities as well as develop the critical thinking skills needed for independent research.

  6. X-ray microanalysis of single and cultured cells

    International Nuclear Information System (INIS)

    Wroblewski, J.; Roomans, G.M.

    1984-01-01

    X-ray microanalysis of single or cultured cells is often a useful alternative or complement to the analysis of the corresponding tissue. It also allows the analysis of individual cells in a cell population. Preparation for X-ray microanalysis poses a number of typical problems. Suspensions of single cells can be prepared by either of two pathways: (1) washing - mounting - drying, or (2) centrifugation - freezing or fixation - sectioning. The washing step in the preparation of single or cultured cells presents the most severe problems. Cultured cells are generally grown on a substrate that is compatible with both the analysis and the culture, washed and dried. In some cases, sectioning of cultured cell monolayers has been performed. Special problems in quantitative analysis occur in those cases where the cells are analyzed on a thick substrate, since the substrate contributes to the spectral background

  7. Integrating a Silicon Solar Cell with a Triboelectric Nanogenerator via a Mutual Electrode for Harvesting Energy from Sunlight and Raindrops.

    Science.gov (United States)

    Liu, Yuqiang; Sun, Na; Liu, Jiawei; Wen, Zhen; Sun, Xuhui; Lee, Shuit-Tong; Sun, Baoquan

    2018-03-27

    Solar cells, as promising devices for converting light into electricity, have a dramatically reduced performance on rainy days. Here, an energy harvesting structure that integrates a solar cell and a triboelectric nanogenerator (TENG) device is built to realize power generation from both sunlight and raindrops. A heterojunction silicon (Si) solar cell is integrated with a TENG by a mutual electrode of a poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) film. Regarding the solar cell, imprinted PEDOT:PSS is used to reduce light reflection, which leads to an enhanced short-circuit current density. A single-electrode-mode water-drop TENG on the solar cell is built by combining imprinted polydimethylsiloxane (PDMS) as a triboelectric material combined with a PEDOT:PSS layer as an electrode. The increasing contact area between the imprinted PDMS and water drops greatly improves the output of the TENG with a peak short-circuit current of ∼33.0 nA and a peak open-circuit voltage of ∼2.14 V, respectively. The hybrid energy harvesting system integrated electrode configuration can combine the advantages of high current level of a solar cell and high voltage of a TENG device, promising an efficient approach to collect energy from the environment in different weather conditions.

  8. Establishment and characterization of American elm cell suspension cultures

    Science.gov (United States)

    Steven M. Eshita; Joseph C. Kamalay; Vicki M. Gingas; Daniel A. Yaussy

    2000-01-01

    Cell suspension cultures of Dutch elm disease (DED)-tolerant and DED-susceptible American elms clones have been established and characterized as prerequisites for contrasts of cellular responses to pathogen-derived elicitors. Characteristics of cultured elm cell growth were monitored by A700 and media conductivity. Combined cell growth data for all experiments within a...

  9. Electrospinning of microbial polyester for cell culture

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, Oh Hyeong [Department of Polymer Science and Engineering, Kumoh National Institute of Technology, 1 Yangho-dong, Gumi, Gyeongbuk 730-701 (Korea, Republic of); Lee, Ik Sang [Department of Polymer Science and Engineering, Kumoh National Institute of Technology, 1 Yangho-dong, Gumi, Gyeongbuk 730-701 (Korea, Republic of); Ko, Young-Gwang [Department of Polymer Science and Engineering, Kumoh National Institute of Technology, 1 Yangho-dong, Gumi, Gyeongbuk 730-701 (Korea, Republic of); Meng, Wan [Department of Polymer Science, Kyungpook National University, 1370 Sankyuk-dong, Buk-gu, Daegu 702-701 (Korea, Republic of); Jung, Kyung-Hye [Department of Polymer Science, Kyungpook National University, 1370 Sankyuk-dong, Buk-gu, Daegu 702-701 (Korea, Republic of); Kang, Inn-Kyu [Department of Polymer Science, Kyungpook National University, 1370 Sankyuk-dong, Buk-gu, Daegu 702-701 (Korea, Republic of); Ito, Yoshihiro [Kanagawa Academy of Science and Technology, KSP East 309, Sakado 3-2-1, Takatsu-ku, Kawasaki 213-0012 (Japan)

    2007-03-01

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

  10. Stimulation of the proliferation of hemopoietic stem cells in irradiated bone marrow cell culture

    International Nuclear Information System (INIS)

    Mori, K.J.; Izumi, H.; Seto, A.

    1981-01-01

    Long-term hemopoiesis was established in bone marrow cell culture in vitro. This culture was shown to support the recovery proliferation of hemopoietic stem cells completely in vitro after irradiation. Hemopoietic stem cells were stimulated into proliferation in culture when normal bone marrow cells were overlayed on top of the irradiated adherent cell colonies. These results indicate that proliferation and differentiation of hemopoietic stem cells in vitro are also supported by stromahemopoietic cell interactions

  11. Design of a hybrid power system based on solar cell and vibration energy harvester

    Science.gov (United States)

    Zhang, Bin; Li, Mingxue; Zhong, Shaoxuan; He, Zhichao; Zhang, Yufeng

    2018-03-01

    Power source has become a serious restriction of wireless sensor network. High efficiency, self-energized and long-life renewable source is the optimum solution for unmanned sensor network applications. However, single renewable power source can be easily affected by ambient environment, which influences stability of the system. In this work, a hybrid power system consists of a solar panel, a vibration energy harvester and a lithium battery is demonstrated. The system is able to harvest multiple types of ambient energy, which extends its applicability and feasibility. Experiments have been conducted to verify performance of the system.

  12. Wild Harvests from Scottish Woodlands Social, cultural and economic values of contemporary non-timber forest products

    Science.gov (United States)

    Marla Emery; Suzanne Martin; Alison Dyke; Alison Dyke

    2006-01-01

    More than 30 people were interviewed about the wild edibles, medicinals, and craft materials they collect and the part that collecting plays in their lives as part of the Wild Harvests from Scottish Woodlands project. Interviews were conducted in autumn 2004. Collecting non-timber forest products (NTFPs) is a source of joy and satisfaction for many of those interviewed...

  13. 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. © 2015 International Federation for Cell Biology.

  14. The effect of varying type and volume of sedimenting agents on leukocyte harvesting and labelling in sickle cell patients

    International Nuclear Information System (INIS)

    Webber, D.; Nunan, T.O.; O'Doherty, M.J.

    1994-01-01

    Leukocyte labelling in patients with sickle cell anaemia has been reported as difficult if not impossible due to the slow erythrocyte sedimentation rate (ESR) in these patients. This study investigated standard sedimentation methods in patients with sickle cell disease (n=16) and compared the results obtained with those following changes in the amount and type of sedimenting agent used. Labelling with either 111 In-oxine or 99 Tc m -exametazime was attempted in only five patients. Replacement of the commonly used 6% Hetastarch (Hespan) with Dextran or Haemaccel did not improve leukocyte harvesting, even when the proportions used of these agents were increased. In most cases where standard procedures for leukocyte collection did not lead to harvesting of viable samples, it was possible to collect reasonably pure samples by increasing the proportion of Hespan used. It is possible to obtain adequate leukocyte labelling in the majority of sickle cell patients using a minor modification of standard techniques. In this group of patients a ratio of 8 ml of Hespan to 16 ml of blood should be used for cell separation. If this fails then donor cells, anti-granulocyte antibody labelling or HIG should be considered. (author)

  15. Particle Trajectories in Rotating Wall Cell Culture Devices

    Science.gov (United States)

    Ramachandran N.; Downey, J. P.

    1999-01-01

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

  16. Microfluidic engineered high cell density three-dimensional neural cultures

    Science.gov (United States)

    Cullen, D. Kacy; Vukasinovic, Jelena; Glezer, Ari; La Placa, Michelle C.

    2007-06-01

    Three-dimensional (3D) neural cultures with cells distributed throughout a thick, bioactive protein scaffold may better represent neurobiological phenomena than planar correlates lacking matrix support. Neural cells in vivo interact within a complex, multicellular environment with tightly coupled 3D cell-cell/cell-matrix interactions; however, thick 3D neural cultures at cell densities approaching that of brain rapidly decay, presumably due to diffusion limited interstitial mass transport. To address this issue, we have developed a novel perfusion platform that utilizes forced intercellular convection to enhance mass transport. First, we demonstrated that in thick (>500 µm) 3D neural cultures supported by passive diffusion, cell densities =104 cells mm-3), continuous medium perfusion at 2.0-11.0 µL min-1 improved viability compared to non-perfused cultures (p death and matrix degradation. In perfused cultures, survival was dependent on proximity to the perfusion source at 2.00-6.25 µL min-1 (p 90% viability in both neuronal cultures and neuronal-astrocytic co-cultures. This work demonstrates the utility of forced interstitial convection in improving the survival of high cell density 3D engineered neural constructs and may aid in the development of novel tissue-engineered systems reconstituting 3D cell-cell/cell-matrix interactions.

  17. Quality of harvest and role of cell dose in unrelated bone marrow transplantation: an Italian Bone Marrow Donor Registry-Gruppo Italiano Trapianto di Midollo Osseo Study.

    Science.gov (United States)

    Fagioli, Franca; Quarello, Paola; Pollichieni, Simona; Lamparelli, Teresa; Berger, Massimo; Benedetti, Fabio; Barat, Veronica; Marciano, Renato; Rambaldi, Alessandro; Bacigalupo, Andrea; Sacchi, Nicoletta

    2014-01-01

    In this study, we investigated the factors affecting cell dose harvest and the role of cell dose on outcome. We analysed data from a cohort of 703 patients who underwent unrelated bone marrow transplantation facilitated by IBMDR in GITMO centers between 2002 and 2008. The median-infused cell doses is 3.7 × 10(8)/kg, the correlation between the nucleated cells requested from transplant centers and those harvested by collection centers was adequate. A harvested/requested cells ratio lower than 0.5 was observed only in 3% of harvests. A volume of harvested marrow higher than the median value of 1270 ml was related to a significant lower infused cell dose (χ(2): 44.4; P < 0.001). No patient- or donor-related variables significantly influenced the cell dose except for the recipient younger age (χ(2): 95.7; P < 0.001) and non-malignant diseases (χ(2): 33.8; P < 0.001). The cell dose resulted an independent predictor factor for a better outcome in patients affected by non-malignant disease (P = 0.05) while early disease malignant patients receiving a lower cell dose showed a higher risk of relapse (P = 0.05).

  18. Usability and Applicability of Microfluidic Cell Culture Systems

    DEFF Research Database (Denmark)

    Hemmingsen, Mette

    possibilities for, for example, precise control of the chemical environment, 3D cultures, controlled co-culture of different cell types or automated, individual control of up to 96 cell culture chambers in one integrated system. Despite the great new opportunities to perform novel experimental designs......Microfluidic cell culture has been a research area with great attention the last decade due to its potential to mimic the in vivo cellular environment more closely compared to what is possible by conventional cell culture methods. Many exciting and complex devices have been presented providing......, these devices still lack general implementation into biological research laboratories. In this project, the usability and applicability of microfluidic cell culture systems have been investigated. The tested systems display good properties regarding optics and compatibility with standard laboratory equipment...

  19. Residual Energy Harvesting from Light Transients Using Hematite as an Intrinsic Photocapacitor in a Symmetrical Cell

    NARCIS (Netherlands)

    Blom, Burgert; van Leeuwen, Nicole; Xie, Mengying; Adamaki, Vana; Bowen, Chris R.; de Araujo, Moises A.; Mascaro, Lucia H.; Cameron, Petra J.; Marken, Frank

    2017-01-01

    Hematite as a sustainable photoabsorber material offers a band gap close to 2 eV and photoanode characteristics, but usually requires additional catalysts to enhance surface redox chemistry during steady state light energy harvesting for water splitting. Here, for a highly doped hematite film,

  20. 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

  1. Evaluation of castor oil-based polyurethane membranes in rat bone-marrow cell culture.

    Science.gov (United States)

    Cerejo, Sofia de Amorim; Rahal, Sheila Canevese; Lima Neto, João Ferreira de; Voorwald, Fabiana Azevedo; Alvarenga, Fernanda da Cruz Landim e

    2011-10-01

    To evaluate three methods to isolate rats MSCs and to analyze the potential of a castor oil polyurethane base membrane as a scaffold for MSCs. Four male Wistar rats, aged 20-30 days were used. Bone marrow aspirates from femur and tibia were harvested using DMEM high glucose and heparin. The cell culture was performed in three different ways: direct culture and two types of density gradients. After 15 days, was made the 1st passage and analyzed cell viability with markers Hoerscht 33342 and propidium iodide. The MSCs were characterized by surface markers with the aid of flow cytometry. After this, three types of castor oil polyurethane membranes associated with the MSCs were kept on the 6-well plate for 5 days and were analyzed by optical microscopy to confirm cell aggregation and growth. Separation procedures 1 and 2 allowed adequate isolation of MSCs and favored cell growth with the passage being carried out at 70% confluence after 15 days in culture. The cells could not be isolated using procedure 3. When the 3 castor oil polyurethane membrane types were compared it was possible to observe that the growth of MSCs was around 80% in membrane type 3, 20% in type 2, and 10% in type 1. Both Ficoll-Hypaque densities allow isolation of rat MSCs, and especially castor oil-based membrane type 3 may be used as a scaffold for MSCs.

  2. Isolation and culture of larval cells from C. elegans.

    Directory of Open Access Journals (Sweden)

    Sihui Zhang

    Full Text Available Cell culture is an essential tool to study cell function. In C. elegans the ability to isolate and culture cells has been limited to embryonically derived cells. However, cells or blastomeres isolated from mixed stage embryos terminally differentiate within 24 hours of culture, thus precluding post-embryonic stage cell culture. We have developed an efficient and technically simple method for large-scale isolation and primary culture of larval-stage cells. We have optimized the treatment to maximize cell number and minimize cell death for each of the four larval stages. We obtained up to 7.8×10(4 cells per microliter of packed larvae, and up to 97% of adherent cells isolated by this method were viable for at least 16 hours. Cultured larval cells showed stage-specific increases in both cell size and multinuclearity and expressed lineage- and cell type-specific reporters. The majority (81% of larval cells isolated by our method were muscle cells that exhibited stage-specific phenotypes. L1 muscle cells developed 1 to 2 wide cytoplasmic processes, while L4 muscle cells developed 4 to 14 processes of various thicknesses. L4 muscle cells developed bands of myosin heavy chain A thick filaments at the cell center and spontaneously contracted ex vivo. Neurons constituted less than 10% of the isolated cells and the majority of neurons developed one or more long, microtubule-rich protrusions that terminated in actin-rich growth cones. In addition to cells such as muscle and neuron that are high abundance in vivo, we were also able to isolate M-lineage cells that constitute less than 0.2% of cells in vivo. Our novel method of cell isolation extends C. elegans cell culture to larval developmental stages, and allows use of the wealth of cell culture tools, such as cell sorting, electrophysiology, co-culture, and high-resolution imaging of subcellular dynamics, in investigation of post-embryonic development and physiology.

  3. Damage of Neuroblastoma Cell SH-SY5Y Mediated by MPP+ Inhibits Proliferation of T-Cell Leukemia Jurkat by Co-Culture System

    Directory of Open Access Journals (Sweden)

    Fuli Wang

    2014-06-01

    Full Text Available The adaptive immune system has implications in pathology of Parkinson’s disease (PD. Research data demonstrated that the peripheral CD4+ T-cell population decreased in pathogenesis of PD. The effect of damaged dopaminergic neurons on peripheral T cells of PD is still unknown. In this study, we constructed a neuronal and glial cells co-culture model by using human neuroblastoma cells SH-SY5Y and gliomas cells U87. After the co-culture cells were treated with neurotoxin 1-methyl-4-phenylpyridinium (MPP+ for 24 h, the conditioned media was harvested and used to cultivate T-cell leukemia Jurkat cells for another 24 h. We then analyzed the cell proliferation, cell cycle and necrosis effect of Jurkat cells. The results showed that co-culture medium of SH-SY5Y and U87 cells with MPP+ treatment inhibited the proliferation of Jurkat cells compared to control medium without MPP+, even though the same concentration of MPP+ had very little toxicity to the Jurkat cell. Furthermore, co-culture medium with low concentration of MPP+ (100 µM arrested Jurkat cells cycle in G2/M phase through increasing cell cycle division 2 (CDC2 and CyclinB1 expression level, whereas co-culture medium with high concentration of MPP+ (500 µM induced Jurkat cell necrosis through cellular swelling and membrane breakage. Our data implies that damaged dopamine neurons with glial cells can lead to the reduced number or inhibited proliferation activity of peripheral T cells.

  4. In vitro development of cloned bovine embryos produced by handmade cloning using somatic cells from distinct levels of cell culture confluence.

    Science.gov (United States)

    Gerger, R P C; Ribeiro, E S; Forell, F; Bertolini, L R; Rodrigues, J L; Ambrósio, C E; Miglino, M A; Mezzalira, A; Bertolini, M

    2010-02-18

    The relationship between the level of cell confluence near the plateau phase of growth and blastocyst yield following somatic cell cloning is not well understood. We examined the effect of distinct cell culture confluence levels on in vitro development of cloned bovine embryos. In vitro-matured bovine oocytes were manually bisected and selected by DNA staining. One or two enucleated hemi-cytoplasts were paired and fused with an adult skin somatic cell. Cultured skin cells from an adult Nellore cow harvested at three distinct culture confluence levels (70-80, 80-90, and >95%) were used for construction of embryos and hemi-embryos. After activation, structures were cultured in vitro as one embryo (1 x 100%) or as aggregates of two hemi-embryos (2 x 50%) per microwell. Fusion, cleavage and blastocyst rates were compared using the chi(2) test. The fusion rate for hemi-embryos (51.4%) was lower than for embryos (67.6%), with no influence of degree of cell confluence. However, blastocyst rates improved linearly (7.0, 17.5, and 29.4%) with increases in cell confluence. We conclude that degree of cell culture confluence significantly influences subsequent embryo development; use of a cell population in high confluence (>90%) for nuclear transfer significantly improved blastocyst yield after cloning.

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

    Science.gov (United States)

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

    2014-01-01

    Annually, more than 200,000 elective liposuction procedures are performed in the United States and over a million worldwide. The ease of harvest and abundance make human adipose-derived stromal/stem cells (hASCs) isolated from lipoaspirates an attractive, readily available source of adult stem cells that have become increasingly popular for use in many studies. Here, we describe common methods for hASC culture, preservation, and osteogenic differentiation. We introduce methods of ceramic, polymer, and composite scaffold synthesis with a description of morphological, chemical, and mechanical characterization techniques. Techniques for scaffold loading are compared, and methods for determining cell loading efficiency and proliferation are described. Finally, we provide both qualitative and quantitative techniques for in vitro assessment of hASC osteogenic differentiation. © 2014 Elsevier Inc. All rights reserved.

  6. Biogelx: Cell Culture on Self-Assembling Peptide Gels.

    Science.gov (United States)

    Harper, Mhairi M; Connolly, Michael L; Goldie, Laura; Irvine, Eleanore J; Shaw, Joshua E; Jayawarna, Vineetha; Richardson, Stephen M; Dalby, Matthew J; Lightbody, David; Ulijn, Rein V

    2018-01-01

    Aromatic peptide amphiphiles can form self-supporting nanostructured hydrogels with tunable mechanical properties and chemical compositions. These hydrogels are increasingly applied in two-dimensional (2D) and three-dimensional (3D) cell culture, where there is a rapidly growing need to store, grow, proliferate, and manipulate naturally derived cells within a hydrated, 3D matrix. Biogelx Limited is a biomaterials company, created to commercialize these bio-inspired hydrogels to cell biologists for a range of cell culture applications. This chapter describes methods of various characterization and cell culture techniques specifically optimized for compatibility with Biogelx products.

  7. Sustained release of stem cell factor in a double network hydrogel for ex vivo culture of cord blood-derived CD34+ cells.

    Science.gov (United States)

    Zhang, Yuanhao; Pan, Xiuwei; Shi, Zhen; Cai, Haibo; Gao, Yun; Zhang, Weian

    2018-04-01

    Stem cell factor (SCF) is considered as a commonly indispensable cytokine for proliferation of haematopoietic stem cells (HSCs), which is used in large dosages during ex vivo culture. The work presented here aimed to reduce the consumption of SCF by sustained release but still support cells proliferation and maintain the multipotency of HSCs. Stem cell factor was physically encapsulated within a hyaluronic acid/gelatin double network (HGDN) hydrogel to achieve a slow release rate. CD34 + cells were cultured within the SCF-loaded HGDN hydrogel for 14 days. The cell number, phenotype and functional capacity were investigated after culture. The HGDN hydrogels had desirable properties and encapsulated SCF kept being released for more than 6 days. SCF remained the native bioactivity, and the proliferation of HSCs within the SCF-loaded HGDN hydrogel was not affected, although the consumption of SCF was only a quarter in comparison with the conventional culture. Moreover, CD34 + cells harvested from the SCF-loaded HGDN hydrogels generated more multipotent colony-forming units (CFU-GEMM). The data suggested that the SCF-loaded HGDN hydrogel could support ex vivo culture of HSCs, thus providing a cost-effective culture protocol for HSCs. © 2017 John Wiley & Sons Ltd.

  8. Development of a microfluidic perfusion 3D cell culture system

    Science.gov (United States)

    Park, D. H.; Jeon, H. J.; Kim, M. J.; Nguyen, X. D.; Morten, K.; Go, J. S.

    2018-04-01

    Recently, 3-dimensional in vitro cell cultures have gained much attention in biomedical sciences because of the closer relevance between in vitro cell cultures and in vivo environments. This paper presents a microfluidic perfusion 3D cell culture system with consistent control of long-term culture conditions to mimic an in vivo microenvironment. It consists of two sudden expansion reservoirs to trap incoming air bubbles, gradient generators to provide a linear concentration, and microchannel mixers. Specifically, the air bubbles disturb a flow in the microfluidic channel resulting in the instability of the perfusion cell culture conditions. For long-term stable operation, the sudden expansion reservoir is designed to trap air bubbles by using buoyancy before they enter the culture system. The performance of the developed microfluidic perfusion 3D cell culture system was examined experimentally and compared with analytical results. Finally, it was applied to test the cytotoxicity of cells infected with Ewing’s sarcoma. Cell death was observed for different concentrations of H2O2. For future work, the developed microfluidic perfusion 3D cell culture system can be used to examine the behavior of cells treated with various drugs and concentrations for high-throughput drug screening.

  9. Systems Biology for Organotypic Cell Cultures

    Energy Technology Data Exchange (ETDEWEB)

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

    2016-08-04

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

  10. Embryonic stem cells in scaffold-free three-dimensional cell culture: osteogenic differentiation and bone generation.

    Science.gov (United States)

    Handschel, Jörg; Naujoks, Christian; Depprich, Rita; Lammers, Lydia; Kübler, Norbert; Meyer, Ulrich; Wiesmann, Hans-Peter

    2011-07-14

    Extracorporeal formation of mineralized bone-like tissue is still an unsolved challenge in tissue engineering. Embryonic stem cells may open up new therapeutic options for the future and should be an interesting model for the analysis of fetal organogenesis. Here we describe a technique for culturing embryonic stem cells (ESCs) in the absence of artificial scaffolds which generated mineralized miromasses. Embryonic stem cells were harvested and osteogenic differentiation was stimulated by the addition of dexamethasone, ascorbic acid, and ß-glycerolphosphate (DAG). After three days of cultivation microspheres were formed. These spherical three-dimensional cell units showed a peripheral zone consisting of densely packed cell layers surrounded by minerals that were embedded in the extracellular matrix. Alizarine red staining confirmed evidence of mineralization after 10 days of DAG stimulation in the stimulated but not in the control group. Transmission electron microscopy demonstrated scorching crystallites and collagenous fibrils as early indication of bone formation. These extracellular structures resembled hydroxyl apatite-like crystals as demonstrated by distinct diffraction patterns using electron diffraction analysis. The micromass culture technique is an appropriate model to form three-dimensional bone-like micro-units without the need for an underlying scaffold. Further studies will have to show whether the technique is applicable also to pluripotent stem cells of different origin. © 2011 Handschel et al; licensee BioMed Central Ltd.

  11. Embryonic stem cells in scaffold-free three-dimensional cell culture: osteogenic differentiation and bone generation

    Directory of Open Access Journals (Sweden)

    Meyer Ulrich

    2011-07-01

    Full Text Available Abstract Extracorporeal formation of mineralized bone-like tissue is still an unsolved challenge in tissue engineering. Embryonic stem cells may open up new therapeutic options for the future and should be an interesting model for the analysis of fetal organogenesis. Here we describe a technique for culturing embryonic stem cells (ESCs in the absence of artificial scaffolds which generated mineralized miromasses. Embryonic stem cells were harvested and osteogenic differentiation was stimulated by the addition of dexamethasone, ascorbic acid, and ß-glycerolphosphate (DAG. After three days of cultivation microspheres were formed. These spherical three-dimensional cell units showed a peripheral zone consisting of densely packed cell layers surrounded by minerals that were embedded in the extracellular matrix. Alizarine red staining confirmed evidence of mineralization after 10 days of DAG stimulation in the stimulated but not in the control group. Transmission electron microscopy demonstrated scorching crystallites and collagenous fibrils as early indication of bone formation. These extracellular structures resembled hydroxyl apatite-like crystals as demonstrated by distinct diffraction patterns using electron diffraction analysis. The micromass culture technique is an appropriate model to form three-dimensional bone-like micro-units without the need for an underlying scaffold. Further studies will have to show whether the technique is applicable also to pluripotent stem cells of different origin.

  12. A method for culturing human hair follicle cells.

    Science.gov (United States)

    Weterings, P J; Vermorken, A J; Bloemendal, H

    1981-01-01

    For the first time a method for culturing human hair follicle cells is described. The bovine eye lens capsule, a basement membrane-like structure, is used as the substrate for the cultures. In a culture medium supplemented with hydrocortisone and insulin about 70% of the original follicles will form growing colonies of diploid keratinocytes.

  13. Modelling in 3D the olive trees cultures in order to establish the forces (interval) needed for automatic harvesting

    Science.gov (United States)

    Babanatsas, T.; Glăvan, D. O.; Babanatis Merce, R. M.; Maris, S. A.

    2018-01-01

    The purpose of this study is to bring as much as possible, close to real situation the 3D modelling for the olive trees in order to establish the necessary forces for automatic harvesting (harvesting robots). To fulfil our goal we have at our disposal different ways to do modelling very close to the real situation. One way is to use reality capture software (its results being photos) that are converted into a real 3D model, the disadvantage of the method being a mesh model that is not accurate enough. The reasonable alternative is to develop an experiment by measuring a sample orchard of olive trees (experiment who took place in Halkidiki, Greece, measuring over 120 trees). After establishing the real dimensions, we adopted as model the media that we have measured (the height of the tree, the thickness of branches, number of branches, etc.), model which we consider closer to the reality and therefor more suitable for our simulation.

  14. Cell cycle regulation in human embryonic stem cells: links to adaptation to cell culture.

    Science.gov (United States)

    Barta, Tomas; Dolezalova, Dasa; Holubcova, Zuzana; Hampl, Ales

    2013-03-01

    Cell cycle represents not only a tightly orchestrated mechanism of cell replication and cell division but it also plays an important role in regulation of cell fate decision. Particularly in the context of pluripotent stem cells or multipotent progenitor cells, regulation of cell fate decision is of paramount importance. It has been shown that human embryonic stem cells (hESCs) show unique cell cycle characteristics, such as short doubling time due to abbreviated G1 phase; these properties change with the onset of differentiation. This review summarizes the current understanding of cell cycle regulation in hESCs. We discuss cell cycle properties as well as regulatory machinery governing cell cycle progression of undifferentiated hESCs. Additionally, we provide evidence that long-term culture of hESCs is accompanied by changes in cell cycle properties as well as configuration of several cell cycle regulatory molecules.

  15. Rabbit uterine epithelial cells: Co-culture with spermatozoa

    International Nuclear Information System (INIS)

    Boice, M.L.

    1988-01-01

    A primary culture of rabbit uterine epithelial cells was established and their effects on sperm function were examined in vitro. Epithelial cells were isolated from uteri of estrous rabbits and cultured on floating collagen gels in phenol red-free medium supplemented with 5% fetal bovine serum. Light microscopy and keratin staining showed that the epithelial cell population established in culture had morphological characteristics similar to that seen in the intact endometrium. Cells were cultured with 3 H-leucine and uptake of label by cells and its incorporation into cellular and secretory proteins determined. When compared to cells cultured for 24-48 h, incorporation of label into cellular protein was lower at 72-96 h, but secretion increased. Estradiol 17-β did not affect label uptake or incorporation, but did enhance proliferation of cells as judged by total DNA content of the cell population. Analysis of proteins in media by sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography suggested that epithelial and stromal cells synthesis proteins that may be secretory in nature during 72-96 h culture. Twenty-nine to thirty-one h after initiation of epithelial cultures, 1-2 x 10 6 sperm were co-incubated with cells and sperm viability, motility, loss of acrosome and fertilizing ability determined

  16. Long-term maintenance of human induced pluripotent stem cells by automated cell culture system.

    Science.gov (United States)

    Konagaya, Shuhei; Ando, Takeshi; Yamauchi, Toshiaki; Suemori, Hirofumi; Iwata, Hiroo

    2015-11-17

    Pluripotent stem cells, such as embryonic stem cells and induced pluripotent stem (iPS) cells, are regarded as new sources for cell replacement therapy. These cells can unlimitedly expand under undifferentiated conditions and be differentiated into multiple cell types. Automated culture systems enable the large-scale production of cells. In addition to reducing the time and effort of researchers, an automated culture system improves the reproducibility of cell cultures. In the present study, we newly designed a fully automated cell culture system for human iPS maintenance. Using an automated culture system, hiPS cells maintained their undifferentiated state for 60 days. Automatically prepared hiPS cells had a potency of differentiation into three germ layer cells including dopaminergic neurons and pancreatic cells.

  17. The impact of cell culture equipment on energy loss.

    Science.gov (United States)

    Davies, Lleucu B; Kiernan, Michael N; Bishop, Joanna C; Thornton, Catherine A; Morgan, Gareth

    2014-01-01

    Light energy of discrete wavelengths supplied via lasers and broadband intense pulsed light have been used therapeutically for many years. In vitro models complement clinical studies, especially for the elucidation of underlying mechanisms of action. Clarification that light energy reaches the cells is necessary when developing protocols for the treatment of cells using in vitro models. Few studies report on energy loss in cell culture equipment. The ability of energy from light with therapeutic potential to reach cells in culture needs to be determined; this includes determining the proportion of light energy lost within standard cell culture media and cell culture vessels. The energy absorption of cell culture media, with/without the pH indicator dye phenol red, and the loss of energy within different plastics and glassware used typically for in vitro cell culture were investigated using intense pulsed light and a yellow pulsed dye laser. Media containing phenol red have a distinctive absorption peak (560 nm) absent in phenol red-free media and restored by the addition of phenol red. For both light sources, energy loss was lowest in standard polystyrene tissue culture flasks or multi-well plates and highest in polypropylene vessels or glass tubes. The effects of phenol red-free media on the absorption of energy varied with the light source used. Phenol red-free media are the media of choice; polystyrene vessels with flat surfaces such as culture flasks or multi-well plates should be used in preference to polypropylene or glass vessels.

  18. Biona-C Cell Culture pH Monitoring System

    Science.gov (United States)

    Friedericks, C.

    1999-01-01

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

  19. Sequential cancer mutations in cultured human intestinal stem cells

    NARCIS (Netherlands)

    Drost, Jarno; van Jaarsveld, Richard H.; Ponsioen, Bas; Zimberlin, Cheryl; van Boxtel, Ruben; Buijs, Arjan; Sachs, Norman; Overmeer, René M.; Offerhaus, G. Johan; Begthel, Harry; Korving, Jeroen; van de Wetering, Marc; Schwank, Gerald; Logtenberg, Meike; Cuppen, Edwin; Snippert, Hugo J.; Medema, Jan Paul; Kops, Geert J. P. L.; Clevers, Hans

    2015-01-01

    Crypt stem cells represent the cells of origin for intestinal neoplasia. Both mouse and human intestinal stem cells can be cultured in medium containing the stem-cell-niche factors WNT, R-spondin, epidermal growth factor (EGF) and noggin over long time periods as epithelial organoids that remain

  20. PECULIARITIES OF SECONDARY METABOLITES BIOSYNTHESIS IN PLANT CELL CULTURES

    Directory of Open Access Journals (Sweden)

    A.M. NOSOV

    2014-06-01

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

  1. Comparison of ex vivo harvested and in vitro cultured materials from Echinococcus granulosus by measuring expression levels of five genes putatively involved in the development and maturation of adult worms.

    Science.gov (United States)

    Dezaki, Ebrahim Saedi; Yaghoubi, Mohammad Mehdi; Spiliotis, Markus; Boubaker, Ghalia; Taheri, Elham; Almani, Pooya Ghaseminejad; Tohidi, Farideh; Harandi, Majid Fasihi; Gottstein, Bruno

    2016-11-01

    Parts of the natural life cycle of Echinococcus granulosus can be retraced in vitro such as the development of protoscoleces into semiadult worms with three or more proglottids, or the redifferentiation of in vitro cultured protoscoleces into metacestode-like cystic structures. Most in vitro generated samples share-at the microscopical level-high similarities with those naturally grown, but developmental differences have also been documented, such as missing egg production in in vitro grown adults or unusual bladder/vesicle formation in protoscoleces cultured into the metacestode direction. The aim of the present study was to explore how far different in vitro generated stage-specific materials/structures match the natural situation on the transcriptome level, based on testing five exemplarily chosen different genes: the frizzled receptor eg-fz4 (posterior marker), the FGF receptor-like factor eg-fgfrl (anterior association), the cell differentiation protein eg-rcd1 (part of the CCR4-NOT complex, a key regulator of eukaryotic gene expression), the rapidly accelerated fibrosarcoma serin/threonin kinase eg-braf (part of the MAPK pathway involved, e.g., in EGF signaling) and the co-smad eg-smadD (downstream factor of TGFβ/BMP2/activin signaling). These genes-tested via qPCR-were selected such as to allow a discussion on their potential role in the development of E. granulosus into the adult stage. Thus, testing took place with three ex vivo isolated samples, namely (i) egg-containing adult worms, (ii) invaginated protoscoleces, and (iii) protoscolex-free germinal layer tissue. Respective data were compared (a) with in vitro generated metacestode-like microcysts developed from protoscolices, and (b) different development stages of protoscoleces in vitro cultured toward adult maturation. As a finding, only eg-smadD and partially eg-fz4 showed high expression similarities between ex vivo harvested and in vitro cultured E. granulosus, thus suggesting a putative role in

  2. Radiosensitivity of normal human epidermal cells in culture

    International Nuclear Information System (INIS)

    Dover, R.; Potten, C.S.

    1983-01-01

    Using an in vitro culture system the authors have derived #betta#-radiation survival curves over a dose range 0-8 Gy for the clonogenic cells of normal human epidermis. The culture system used allows the epidermal cells to stratify and form a multi-layered sheet of keratinizing cells. The cultures appear to be a very good model for epidermis in vivo. The survival curves show a population which is apparently more sensitive than murine epidermis in vivo. It remains unclear whether this is an intrinsic difference between the species or is a consequence of the in vitro cultivation of the human cells. (author)

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

    Science.gov (United States)

    Gebhard, C; Gabriel, C; Walter, I

    2016-06-01

    Spheroid cell culture emerges as powerful in vitro tool for experimental tumour research. In this study, we established a scaffold-free three-dimensional spheroid system built from canine osteosarcoma (OS) cells (D17). Spheroids (7, 14 and 19 days of cultivation) and monolayer cultures (2 and 7 days of cultivation) were evaluated and compared on light and electron microscopy. Monolayer and spheroid cultures were tested for vimentin, cytokeratin, alkaline phosphatase, osteocalcin and collagen I by means of immunohistochemistry. The spheroid cell culture exhibited a distinct network of collagen I in particular after 19-day cultivation, whereas in monolayer cultures, collagen I was arranged as a lamellar basal structure. Necrotic centres of large spheroids, as observed in 14- and 19-day cultures, were characterized by significant amounts of osteocalcin. Proliferative activity as determined by Ki-67 immunoreactivity showed an even distribution in two-dimensional cultures. In spheroids, proliferation was predominating in the peripheral areas. Metastasis-associated markers ezrin and S100A4 were shown to be continuously expressed in monolayer and spheroid cultures. We conclude that the scaffold-free spheroid system from canine OS cells has the ability to mimic the architecture of the in vivo tumour, in particular cell-cell and cell-matrix interactions. © 2015 The Authors. Anatomia, Histologia, Embryologia Published by Blackwell Verlag GmbH.

  4. 21st Century Cell Culture for 21st Century Toxicology.

    Science.gov (United States)

    Pamies, David; Hartung, Thomas

    2017-01-17

    There is no good science in bad models. Cell culture is especially prone to artifacts. A number of novel cell culture technologies have become more broadly available in the 21st century, which allow overcoming limitations of traditional culture and are more physiologically relevant. These include the use of stem-cell derived human cells, cocultures of different cell types, scaffolds and extracellular matrices, perfusion platforms (such as microfluidics), 3D culture, organ-on-chip technologies, tissue architecture, and organ functionality. The physiological relevance of such models is further enhanced by the measurement of biomarkers (e.g., key events of pathways), organ specific functionality, and more comprehensive assessment cell responses by high-content methods. These approaches are still rarely combined to create microphysiological systems. The complexity of the combination of these technologies can generate results closer to the in vivo situation but increases the number of parameters to control, bringing some new challenges. In fact, we do not argue that all cell culture needs to be that sophisticated. The efforts taken are determined by the purpose of our experiments and tests. If only a very specific molecular target to cell response is of interest, a very simple model, which reflects this, might be much more suited to allow standardization and high-throughput. However, the less defined the end point of interest and cellular response are, the better we should approximate organ- or tissue-like culture conditions to make physiological responses more probable. Besides these technologic advances, important progress in the quality assurance and reporting on cell cultures as well as the validation of cellular test systems brings the utility of cell cultures to a new level. The advancement and broader implementation of Good Cell Culture Practice (GCCP) is key here. In toxicology, this is a major prerequisite for meaningful and reliable results, ultimately

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

    Science.gov (United States)

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

    2015-01-01

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

  6. Radiosensitivity of primary cultured fish cells with different ploidy

    International Nuclear Information System (INIS)

    Mitani, Hiroshi; Egami, Nobuo; Kobayashi, Hiromu.

    1986-01-01

    The radiosensitivity of primary cultured goldfish cells (Carassius auratus) was investigated by colony formation assay. The radiosensitivity of cells from two varieties of goldfish, which show different sensitivity to lethal effect of ionizing radiation in vivo, was almost identical. Primary cultured cells from diploid, triploid and tetraploid fish retained their DNA content as measured by microfluorometry, and the nuclear size increases as ploidy increases. However, radiosensitivity was not related to ploidy. (author)

  7. Establishment of a Novel Lingual Organoid Culture System: Generation of Organoids Having Mature Keratinized Epithelium from Adult Epithelial Stem Cells

    Science.gov (United States)

    Hisha, Hiroko; Tanaka, Toshihiro; Kanno, Shohei; Tokuyama, Yoko; Komai, Yoshihiro; Ohe, Shuichi; Yanai, Hirotsugu; Omachi, Taichi; Ueno, Hiroo

    2013-11-01

    Despite the strong need for the establishment of a lingual epithelial cell culture system, a simple and convenient culture method has not yet been established. Here, we report the establishment of a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Histological analyses showed that the generated organoids had both a stratified squamous epithelial cell layer and a stratum corneum. Very recently, we showed via a multicolor lineage tracing method that Bmi1-positive stem cells exist at the base of the epithelial basal layer in the interpapillary pit. Using our new culture system, we found that organoids could be generated by single Bmi1-positive stem cells and that in the established organoids, multiple Bmi1-positive stem cells were generated at the outermost layer. Moreover, we observed that organoids harvested at an early point in culture could be engrafted and maturate in the tongue of recipient mice and that the organoids generated from carcinogen-treated mice had an abnormal morphology. Thus, this culture system presents valuable settings for studying not only the regulatory mechanisms of lingual epithelium but also lingual regeneration and carcinogenesis.

  8. Multizone Paper Platform for 3D Cell Cultures

    Science.gov (United States)

    Derda, Ratmir; Hong, Estrella; Mwangi, Martin; Mammoto, Akiko; Ingber, Donald E.; Whitesides, George M.

    2011-01-01

    In vitro 3D culture is an important model for tissues in vivo. Cells in different locations of 3D tissues are physiologically different, because they are exposed to different concentrations of oxygen, nutrients, and signaling molecules, and to other environmental factors (temperature, mechanical stress, etc). The majority of high-throughput assays based on 3D cultures, however, can only detect the average behavior of cells in the whole 3D construct. Isolation of cells from specific regions of 3D cultures is possible, but relies on low-throughput techniques such as tissue sectioning and micromanipulation. Based on a procedure reported previously (“cells-in-gels-in-paper” or CiGiP), this paper describes a simple method for culture of arrays of thin planar sections of tissues, either alone or stacked to create more complex 3D tissue structures. This procedure starts with sheets of paper patterned with hydrophobic regions that form 96 hydrophilic zones. Serial spotting of cells suspended in extracellular matrix (ECM) gel onto the patterned paper creates an array of 200 micron-thick slabs of ECM gel (supported mechanically by cellulose fibers) containing cells. Stacking the sheets with zones aligned on top of one another assembles 96 3D multilayer constructs. De-stacking the layers of the 3D culture, by peeling apart the sheets of paper, “sections” all 96 cultures at once. It is, thus, simple to isolate 200-micron-thick cell-containing slabs from each 3D culture in the 96-zone array. Because the 3D cultures are assembled from multiple layers, the number of cells plated initially in each layer determines the spatial distribution of cells in the stacked 3D cultures. This capability made it possible to compare the growth of 3D tumor models of different spatial composition, and to examine the migration of cells in these structures. PMID:21573103

  9. Influence of cell type and cell culture media on the propagation of foot-and-mouth disease virus with regard to vaccine quality.

    Science.gov (United States)

    Dill, Veronika; Hoffmann, Bernd; Zimmer, Aline; Beer, Martin; Eschbaumer, Michael

    2018-03-16

    Suspension culture of BHK cells allows large-scale virus propagation and cost-efficient vaccine production, while the shift to animal-component-free cell culture media without serum is beneficial for the quality and downstream processing of the product. Foot-and-mouth disease virus is still endemic in many parts of the world and high-quality vaccines are essential for the eradication of this highly contagious and economically devastating disease. Changes to the viral genome sequence during passaging in an adherent and a suspension cell culture system were compared and the impact of amino acid substitutions on receptor tropism, antigenicity and particle stability was examined. Virus production in suspension cells in animal-component-free media and in serum-containing media as well as in adherent cells in serum-containing media was compared. Infection kinetics were determined and the yield of intact viral particles was estimated in all systems using sucrose density gradient centrifugation. Capsid protein sequence alterations were serotype-specific, but varied between cell lines. But The A 24 -2P virus variant had expanded its receptor tropism, but virus neutralization tests found no changes in the antigenic profile in comparison to the original viruses. There were no differences in viral titer between a suspension and an adherent cell culture system, independent of the type of media used. Also, the usage of a serum-free suspension culture system promoted viral growth and allowed an earlier harvest. For serotype O isolates, no differences were seen in the yield of 146S particles. Serotype A preparations revealed a decreased yield of 146S particles in suspension cells independent of the culture media. The selective pressure of the available surface receptors in different cell culture systems may be responsible for alterations in the capsid coding sequence of culture-grown virus. Important vaccine potency characteristics such as viral titer and the neutralization

  10. Single-cell qPCR on dispersed primary pituitary cells -an optimized protocol

    Directory of Open Access Journals (Sweden)

    Haug Trude M

    2010-11-01

    Full Text Available Abstract Background The incidence of false positives is a potential problem in single-cell PCR experiments. This paper describes an optimized protocol for single-cell qPCR measurements in primary pituitary cell cultures following patch-clamp recordings. Two different cell harvesting methods were assessed using both the GH4 prolactin producing cell line from rat, and primary cell culture from fish pituitaries. Results Harvesting whole cells followed by cell lysis and qPCR performed satisfactory on the GH4 cell line. However, harvesting of whole cells from primary pituitary cultures regularly produced false positives, probably due to RNA leakage from cells ruptured during the dispersion of the pituitary cells. To reduce RNA contamination affecting the results, we optimized the conditions by harvesting only the cytosol through a patch pipette, subsequent to electrophysiological experiments. Two important factors proved crucial for reliable harvesting. First, silanizing the patch pipette glass prevented foreign extracellular RNA from attaching to charged residues on the glass surface. Second, substituting the commonly used perforating antibiotic amphotericin B with β-escin allowed efficient cytosol harvest without loosing the giga seal. Importantly, the two harvesting protocols revealed no difference in RNA isolation efficiency. Conclusion Depending on the cell type and preparation, validation of the harvesting technique is extremely important as contaminations may give false positives. Here we present an optimized protocol allowing secure harvesting of RNA from single cells in primary pituitary cell culture following perforated whole cell patch clamp experiments.

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

    Science.gov (United States)

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

    1996-01-01

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

  12. Post-thaw non-cultured and post-thaw cultured equine cord blood mesenchymal stromal cells equally suppress lymphocyte proliferation in vitro.

    Directory of Open Access Journals (Sweden)

    Lynn B Williams

    Full Text Available Multipotent mesenchymal stromal cells (MSC are receiving increased attention for their non-progenitor immunomodulatory potential. Cryopreservation is commonly used for long-term storage of MSC. Post-thaw MSC proliferation is associated with a lag-phase in vitro. How this lag-phase affect MSC immunomodulatory properties is unknown. We hypothesized that in vitro there is no difference in lymphocyte suppression potential between quick-thawed cryopreserved equine cord blood (CB MSC immediately included in mixed lymphocyte reaction (MLR and same MSC allowed post-thaw culture time prior to inclusion in MLR. Cryopreserved CB-MSC from five unrelated foals were compared using two-way MLR. For each of the five unrelated MSC cultures, paired MLR assays of MSC allowed five days of post-thaw culture and MSC included in MLR assay immediately post-thawing were evaluated. We report no difference in the suppression of lymphocyte proliferation by CB-MSC that had undergone post-thaw culture and MSC not cultured post-thaw (p<0.0001. Also, there was no inter-donor variability between the lymphocyte suppressive properties of MSC harvested from the five different donors (p = 0.13. These findings suggest that cryopreserved CB-MSC may have clinical utility immediately upon thawing. One implication hereof is the possibility of using cryopreserved CB-MSC at third party locations without the need for cell culture equipment or competencies.

  13. Radiation adaptive response for the growth of cultured glial cells

    International Nuclear Information System (INIS)

    Suzuki, S.; Miura, Y.; Kano, M.; Toda, T.; Urano, S.

    2003-01-01

    Full text: To examine the molecular mechanism of radiation adaptive response (RAR) for the growth of cultured glial cells and to investigate the influence of aging on the response, glial cells were cultured from young and aged rats (1 month and 24 months old). RAR for the growth of glial cells conditioned with a low dose of X-rays and subsequently exposed to a high dose of X-rays was examined for cell number and BrdU incorporation. Involvement of the subcellular signaling pathway factors in RAR was investigated using their inhibitors, activators and mutated glial cells. RAR was observed in cells cultured from young rats, but was not in cells from aged rats. The inhibitors of protein kinase C (PKC) and DNA-dependent protein kinase (DNA-PK) or phosphatidylinositol 3-kinase (PI3K) suppressed RAR. The activators of PKC instead of low dose irradiation also caused RAR. Moreover, glial cells cultured from severe combined immunodeficiency (scid) mice (CB-17 scid) and ataxia-telangiectasia (AT) cells from AT patients showed no RAR. These results indicated that PKC, ATM, DNAPK and/or PI3K were involved in RAR for growth and BrdU incorporation of cultured glial cells and RAR decreased with aging. Proteomics data of glial cells exposed to severe stress of H 2 O 2 or X-rays also will be presented in the conference since little or no difference has not been observed with slight stress yet

  14. A Refined Culture System for Human Induced Pluripotent Stem Cell-Derived Intestinal Epithelial Organoids

    Directory of Open Access Journals (Sweden)

    Yu Takahashi

    2018-01-01

    Full Text Available Gut epithelial organoids are routinely used to investigate intestinal biology; however, current culture methods are not amenable to genetic manipulation, and it is difficult to generate sufficient numbers for high-throughput studies. Here, we present an improved culture system of human induced pluripotent stem cell (iPSC-derived intestinal organoids involving four methodological advances. (1 We adopted a lentiviral vector to readily establish and optimize conditioned medium for human intestinal organoid culture. (2 We obtained intestinal organoids from human iPSCs more efficiently by supplementing WNT3A and fibroblast growth factor 2 to induce differentiation into definitive endoderm. (3 Using 2D culture, followed by re-establishment of organoids, we achieved an efficient transduction of exogenous genes in organoids. (4 We investigated suspension organoid culture without scaffolds for easier harvesting and assays. These techniques enable us to develop, maintain, and expand intestinal organoids readily and quickly at low cost, facilitating high-throughput screening of pathogenic factors and candidate treatments for gastrointestinal diseases.

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

    NARCIS (Netherlands)

    Heo, Inha; Clevers, Hans

    2015-01-01

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

  16. Novel culturing platform for brain slices and neuronal cells

    DEFF Research Database (Denmark)

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

    2015-01-01

    In this paper we demonstrate a novel culturing system for brain slices and neuronal cells, which can control the concentration of nutrients and the waste removal from the culture by adjusting the fluid flow within the device. The entire system can be placed in an incubator. The system has been...... tested successfully with brain slices and PC12 cells. The culture substrate can be modified using metal electrodes and/or nanostructures for conducting electrical measurements while culturing and for better mimicking the in vivo conditions....

  17. Reducing electrocoagulation harvesting costs for practical microalgal biodiesel production.

    Science.gov (United States)

    Dassey, Adam J; Theegala, Chandra S

    2014-01-01

    Electrocoagulation has shown potential to be a primary microalgae harvesting technique for biodiesel production. However, methods to reduce energy and electrode costs are still necessary for practical application. Electrocoagulation tests were conducted on Nannochloris sp. and Dunaliella sp. using perforated aluminium and iron electrodes under various charge densities. Aluminium electrodes were shown to be more efficient than iron electrodes when harvesting both algal species. Despite the lower harvesting efficiency, however, the iron electrodes were more energy and cost efficient. Operational costs of less than $0.03/L oil were achieved when harvesting Nannochloris sp. with iron electrodes at 35% harvest efficiency, whereas aluminium electrodes cost $0.75/L oil with 42% harvesting efficiency. Increasing the harvesting efficiencies for both aluminium and iron electrodes also increased the overall cost per litre of oil, therefore lower harvesting efficiencies with lower energy inputs was recommended. Also, increasing the culturing salinity to 2 ppt sodium chloride for freshwater Nannochloris sp. was determined practical to improve the electrocoagulation energy efficiency despite a 25% reduction in cell growth.

  18. Reduced cadmium-induced cytotoxicity in cultured liver cells following 5-azacytidine pretreatment

    International Nuclear Information System (INIS)

    Waalkes, M.P.; Wilson, M.J.; Poirier, L.A.

    1985-01-01

    Recent work indicated that administration of the pyrimidine analog 5-azacytidine (AZA), either to cells in culture or to rats, results in an enhancement of expression of the metallothionein (MT) gene. Since MT is thought to play a central role in the detoxification of cadmium, the present study was designed to assess the effect of AZA pretreatment on cadmium cytotoxicity. Cultured rat liver cells in log phase of growth were first exposed to AZA (8 microM). Forty-eight hours later, cadmium was added. A modest increase in MT amounts over control was detected after AZA treatment alone. Cadmium alone resulted in a 10-fold increase in MT concentrations. The combination of AZA pretreatment followed by cadmium exposure caused a 23-fold increase in MT concentrations over control. Treatment with the DNA synthesis inhibitor hydroxyurea (HU) eliminated the enhancing effect of AZA pretreatment on cadmium induction of MT, indicating that cell division is required. AZA-pretreated cells were also harvested and incubated in suspension with cadmium for 0 to 90 min. AZA-pretreated cells showed marked reductions in cadmium-induced cytotoxicity as reflected by reduced intracellular potassium loss, glutamic-oxaloacetic transaminase loss, and lipid peroxidation following cadmium exposure. Results suggest that AZA pretreatment induces tolerance to cadmium cytotoxicity which appears to be due to an increased capacity to synthesize MT rather than high quantities of preexisting MT at the time of cadmium exposure

  19. Senescent vs. non-senescent cells in the human annulus in vivo: Cell harvest with laser capture microdissection and gene expression studies with microarray analysis

    Directory of Open Access Journals (Sweden)

    Ingram Jane A

    2010-01-01

    Full Text Available Abstract Background Senescent cells are well-recognized in the aging/degenerating human disc. Senescent cells are viable, cannot divide, remain metabolically active and accumulate within the disc over time. Molecular analysis of senescent cells in tissue offers a special challenge since there are no cell surface markers for senescence which would let one use fluorescence-activated cell sorting as a method for separating out senescent cells. Methods We employed a novel laser capture microdissection (LCM design to selectively harvest senescent and non-senescent annulus cells in paraffin-embedded tissue, and compared their gene expression with microarray analysis. LCM was used to separately harvest senescent and non-senescent cells from 11 human annulus specimens. Results Microarray analysis revealed significant differences in expression levels in senescent cells vs non-senescent cells: 292 genes were upregulated, and 321 downregulated. Genes with established relationships to senescence were found to be significantly upregulated in senescent cells vs. non-senescent cells: p38 (MPAK14, RB-Associated KRAB zinc finger, Discoidin, CUB and LCCL domain, growth arrest and DNA-damage inducible beta, p28ING5, sphingosine-1-phosphate receptor 2 and somatostatin receptor 3; cyclin-dependent kinase 8 showed significant downregulation in senescent cells. Nitric oxidase synthase 1, and heat shock 70 kDa protein 6, both of which were significantly down-regulated in senescent cells, also showed significant changes. Additional genes related to cytokines, cell proliferation, and other processes were also identified. Conclusions Our LCM-microarray analyses identified a set of genes associated with senescence which were significantly upregulated in senescent vs non-senescent cells in the human annulus. These genes include p38 MAP kinase, discoidin, inhibitor of growth family member 5, and growth arrest and DNA-damage-inducible beta. Other genes, including genes

  20. Stimulation and support of haemopoietic stem cell proliferation by irradiated stroma cell colonies in bone marrow cell culture in vitro

    International Nuclear Information System (INIS)

    Mori, K.J.; Izumi, Hiroko; Seto, Akira

    1981-01-01

    A culture system was established in which haemopoietic stem cells can undergo a recovery proliferation after a depletion of the stem cells, completely in vitro. To elucidate the source of the stimulatory factors, normal bone marrow cells were overlayed on top of the irradiated adherent 'stromal' cell colonies in the bone marrow cell culture. This stimulated the proliferation of haemopoietic stem cells in the cultured cells in suspension. The present results indicate that the stromal cells produce factors which stimulate stem cell proliferation. Whether the stimulation is evoked by direct cell-cell interactions or by humoral factors is as yet to be studied. (author)

  1. Monitoring Dynamic Interactions between Breast Cancer Cells and Human Bone Tissue in a Co-Culture Model

    Science.gov (United States)

    Contag, Christopher H.; Lie, Wen-Rong; Bammer, Marie C.; Hardy, Jonathan W.; Schmidt, Tobi L.; Maloney, William J.; King, Bonnie L.

    2015-01-01

    Purpose Bone is a preferential site of breast cancer metastasis and models are needed to study this process at the level of the microenvironment. We have used bioluminescence imaging (BLI) and multiplex biomarker immunoassays to monitor dynamic breast cancer cell behaviors in co-culture with human bone tissue. Procedures Femur tissue fragments harvested from hip replacement surgeries were co-cultured with luciferase-positive MDA-MB-231-fLuc cells. BLI was performed to quantify breast cell division and track migration relative to bone tissue. Breast cell colonization of bone tissues was assessed with immunohistochemistry. Biomarkers in co-culture supernatants were profiled with MILLIPLEX® immunoassays. Results BLI demonstrated increased MDA-MB-231-fLuc proliferation (pbones, and revealed breast cell migration toward bone. Immunohistochemistry illustrated MDA-MB-231-fLuc colonization of bone, and MILLIPLEX® profiles of culture supernatants suggested breast/bone crosstalk. Conclusions Breast cell behaviors that facilitate metastasis occur reproducibly in human bone tissue co-cultures and can be monitored and quantified using BLI and multiplex immunoassays. PMID:24008275

  2. Quantitative volumetric Raman imaging of three dimensional cell cultures

    Science.gov (United States)

    Kallepitis, Charalambos; Bergholt, Mads S.; Mazo, Manuel M.; Leonardo, Vincent; Skaalure, Stacey C.; Maynard, Stephanie A.; Stevens, Molly M.

    2017-03-01

    The ability to simultaneously image multiple biomolecules in biologically relevant three-dimensional (3D) cell culture environments would contribute greatly to the understanding of complex cellular mechanisms and cell-material interactions. Here, we present a computational framework for label-free quantitative volumetric Raman imaging (qVRI). We apply qVRI to a selection of biological systems: human pluripotent stem cells with their cardiac derivatives, monocytes and monocyte-derived macrophages in conventional cell culture systems and mesenchymal stem cells inside biomimetic hydrogels that supplied a 3D cell culture environment. We demonstrate visualization and quantification of fine details in cell shape, cytoplasm, nucleus, lipid bodies and cytoskeletal structures in 3D with unprecedented biomolecular specificity for vibrational microspectroscopy.

  3. Growth of melanocytes in human epidermal cell cultures

    International Nuclear Information System (INIS)

    Staiano-Coico, L.; Hefton, J.M.; Amadeo, C.; Pagan-Charry, I.; Madden, M.R.; Cardon-Cardo, C.

    1990-01-01

    Epidermal cell cultures were grown in keratinocyte-conditioned medium for use as burn wound grafts; the melanocyte composition of the grafts was studied under a variety of conditions. Melanocytes were identified by immunohistochemistry based on a monoclonal antibody (MEL-5) that has previously been shown to react specifically with melanocytes. During the first 7 days of growth in primary culture, the total number of melanocytes in the epidermal cultures decreased to 10% of the number present in normal skin. Beginning on day 2 of culture, bipolar melanocytes were present at a mean cell density of 116 +/- 2/mm2; the keratinocyte to melanocyte ratio was preserved during further primary culture and through three subpassages. Moreover, exposure of cultures to mild UVB irradiation stimulated the melanocytes to proliferate, suggesting that the melanocytes growing in culture maintained their responsiveness to external stimuli. When the sheets of cultured cells were enzymatically detached from the plastic culture flasks before grafting, melanocytes remained in the basal layer of cells as part of the graft applied to the patient

  4. Surface modified alginate microcapsules for 3D cell culture

    Science.gov (United States)

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

    2016-06-01

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

  5. Mammalian Cell Culture Clarification: A Case Study Using Chimeric Anti-CEA Monoclonal Antibodies

    Directory of Open Access Journals (Sweden)

    Mohamed Ali Abol Hassan

    2011-12-01

    Full Text Available The extracellular expression of monoclonal antibodies (mAbs in mammalian cell culture provides both opportunities and restrictions for the design of robust harvest and clarification operations. With advances in cell culture media and cell lines, it is now possible to achieve high titers of over 5 g/l for mAbs. However, Mammalian cells are sensitive to breakage due to shear stress that can result in release of proteases and other host cell proteins (HCPs which eventually affects product stability and purity. There is larger number of mAbs undergoing clinical development and it has placed significant importance on platform technologies of process development. Generally, Centrifugation and microfiltration are the primary harvest techniques used in the industry and depth filtration is also used as a step operation on clarification. This study compares the unit operations; centrifugation, microfiltration and depth filtration for maximum recovery of monoclonal antibodies. The results have shown that the depth filtration as more suitable operation for mammalian cell culture clarification since it gives 96% recovery of mAbs in comparison to centrifugation and microfiltration. ABSTRAK: Pengungkapan luar sel dari antibodi monoklon (monoclonal antibodies ((mAbs dalam kultur sel mamalia memberi ruang dan batasan terhadap reka bentuk penuaian yang cekap dan penerangan operasi. Dengan kemajuan dalam media sel kultur dan cell lines (produk yang berupa sel kekal yang digunakan untuk tujuan kajian biologi, kini adalah berkemungkinan untuk memperolehi titer tinggi melebihi 5g/l untuk mAbs [2]. Walaupun begitu, sel mamalia sensitif terhadap retakan disebabkan tegasan ricih yang menyebabkan pengeluaran protease dan hos sel protein yang lain, (host cell proteins (HCPs akhirnya mempengaruhi kestabilan dan keaslian produk. Terdapat mAbs dalam jumlah besar yang masih menjalani pembangunan klinikal dan sesungguhnya ini penting sebagai satu landasan teknologi dalam

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

    DEFF Research Database (Denmark)

    Ziolkowska, K.; Jedrych, E.; Kwapiszewski, R.

    2010-01-01

    In this paper, hybrid (PDMS/glass) microfluidic cell culture system (MCCS) integrated with the concentration gradient generator (CGG) is presented. PDMS gas permeability enabled cells' respiration in the fabricated microdevices and excellent glass hydrophilicity allowed successful cells' seeding...

  7. Application of cell co-culture system to study fat and muscle cells.

    Science.gov (United States)

    Pandurangan, Muthuraman; Hwang, Inho

    2014-09-01

    Animal cell culture is a highly complex process, in which cells are grown under specific conditions. The growth and development of these cells is a highly unnatural process in vitro condition. Cells are removed from animal tissues and artificially cultured in various culture vessels. Vitamins, minerals, and serum growth factors are supplied to maintain cell viability. Obtaining result homogeneity of in vitro and in vivo experiments is rare, because their structure and function are different. Living tissues have highly ordered complex architecture and are three-dimensional (3D) in structure. The interaction between adjacent cell types is quite distinct from the in vitro cell culture, which is usually two-dimensional (2D). Co-culture systems are studied to analyze the interactions between the two different cell types. The muscle and fat co-culture system is useful in addressing several questions related to muscle modeling, muscle degeneration, apoptosis, and muscle regeneration. Co-culture of C2C12 and 3T3-L1 cells could be a useful diagnostic tool to understand the muscle and fat formation in animals. Even though, co-culture systems have certain limitations, they provide a more realistic 3D view and information than the individual cell culture system. It is suggested that co-culture systems are useful in evaluating the intercellular communication and composition of two different cell types.

  8. Microfluidic bioreactors for culture of non-adherent cells

    DEFF Research Database (Denmark)

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

    2011-01-01

    Microfluidic bioreactors (μBR) are becoming increasingly popular for cell culture, sample preparation and analysis in case of routine genetic and clinical diagnostics. We present a novel μBR for non-adherent cells designed to mimic in vivo perfusion of cells based on diffusion of media through...

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

    DEFF Research Database (Denmark)

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

    1984-01-01

    Casein kinase II (CKII) activity is enhanced as much as 2-3 fold in established and 4-5-fold in transformed human cell lines when compared to that of fibroblasts and primary human tumour cell cultures where CKII activity never exceeded a basic level. The high activity of CKII in transformed cells...

  10. Serum-converted platelet lysate can substitute for fetal bovine serum in human mesenchymal stromal cell cultures.

    Science.gov (United States)

    Mojica-Henshaw, Mariluz P; Jacobson, Pam; Morris, Julie; Kelley, Linda; Pierce, Jan; Boyer, Michael; Reems, Jo-Anna

    2013-12-01

    Fetal bovine serum (FBS) is commonly used as a serum supplement for culturing human mesenchymal stromal cells (hMSCs). However, human cells grown in FBS, especially for extended periods, risk potential exposure to bovine immunogenic proteins and infectious agents. To address this issue, we investigated the ability of a novel human platelet serum supplement to substitute for FBS in hMSC cultures. Platelet lysate-serum (PL-serum) was converted from platelet lysate-plasma (PL-plasma) that was manufactured from pooled platelet-rich plasma (PRP) apheresis units. Growth factor levels and the number of residual intact platelets in PL-serum and PL-plasma were compared with enzyme-linked immunosorbent assays and flow cytometry, respectively. Proliferation responses of hMSCs cultured in PL-serum, PL-plasma, or FBS were assessed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, the immunophenotype of harvested hMSCs was evaluated by flow cytometry and tri-lineage differentiation potential was evaluated by assessing adipogenic, osteogenic and chondrogenic development. Selected growth factor levels in PL-serum were not significantly different from PL-plasma (P > 0.05). hMSC cultures supplemented with PL-serum had comparable growth kinetics to PL-plasma, and hMSC yields were consistently greater than with FBS. hMSCs harvested from cultures supplemented with PL-serum, PL-plasma or FBS had similar cell surface phenotypes and maintained tri-lineage differentiation potential. PL-serum, similar to PL-plasma, can substitute for FBS in hMSC cultures. Use of PL-serum, in contrast to PL-plasma, has an added advantage of not requiring addition of a xenogeneic source of heparin, providing a completely xeno-free culture medium. Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  11. Protein biosynthesis in cultured human hair follicle cells.

    Science.gov (United States)

    Weterings, P J; Vermorken, A J; Bloemendal, H

    1980-10-31

    A new technique has been used for culturing human keratinocytes. The cells grow on the basement membrane-like capsules of bovine lenses. Lens cells were removed from the capsules by rigid trypsinization. In order to exclude any contamination with remaining living cells the isolated capsules were irradiated with X-rays at a dose of 10,000 rad. In this way human epithelial cells can be brought in culture from individual hair follicles. Since feeder cells are not used in this culture technique, the biosynthesis of keratinocyte proteins can be studied in these cultures. The newly synthesized proteins can be separated into a water-soluble, a urea-soluble, and a urea-insoluble fraction. Product analysis has been performed on the first two fractions revealing protein patterns identical to those of intact hair follicles. Product analysis of the urea-soluble fractions of microdissected hair follicles shows that the protein pattern of the cultured keratinocytes resembles the protein pattern of the hair follicle sheath. Studies on the metabolism of benzo(a)pyrene revealed that the enzyme aryl hydrocarbon hydroxylase (AHH) is present in cultured hair follicle cells. A possible use of our culture system for eventual detection of inherited predisposition for smoking-dependent lung cancer is discussed.

  12. Control of fibronectin synthesis by rat granulosa cells in culture

    International Nuclear Information System (INIS)

    Skinner, M.K.; Dorrington, J.H.

    1984-01-01

    The secreted and cellular [ 35 S]methionine-radiolabeled proteins of cultured rat granulosa cells were separated by electrophoresis on sodium dodecylsulfate (SDS) polyacrylamide gradient slab gels. From 24 to 72 h of culture FSH increased the intensity of labeling of most of the secreted proteins. A 220,000-dalton protein, however, increased in intensity only in control cultures and became the major secreted protein after 72 h, comprising 20% of the total radiolabeled proteins. This protein was identified as fibronectin by immunoprecipitation. There was no increase in the secreted or cellular fibronectin in FSH- or testosterone- and insulin-treated cultures. These studies indicate that a component of extracellular matrix is a major secretory product of unstimulated immature granulosa cells. As hormones induce the differentiated functions of granulosa cells in culture, the secretion of fibronectin is inhibited

  13. Cytotoxicity of TSP in 3D Agarose Gel Cultured Cell.

    Directory of Open Access Journals (Sweden)

    Song-I Chun

    Full Text Available A reference reagent, 3-(trimethylsilyl propionic-2, 2, 3, 3-d4 acid sodium (TSP, has been used frequently in nuclear magnetic resonance (NMR and magnetic resonance spectroscopy (MRS as an internal reference to identify cell and tissue metabolites, and determine chemical and protein structures. This reference material has been exploited for the quantitative and dynamic analyses of metabolite spectra acquired from cells. The aim of this study was to evaluate the cytotoxicity of TSP on three-dimensionally, agarose gel, cultured cells.A human osteosarcoma cell line (MG-63 was selected, and cells were three dimensionally cultured for two weeks in an agarose gel. The culture system contained a mixture of conventional culture medium and various concentrations (0, 1, 3, 5, 7, 10, 20 30 mM of TSP. A DNA quantification assay was conducted to assess cell proliferation using Quant-iT PicoGreen dsDNA reagent and kit, and cell viability was determined using a LIVE/DEAD Viability/Cytotoxicity kit. Both examinations were performed simultaneously at 1, 3, 7 and 14 days from cell seeding.In this study, the cytotoxicity of TSP in the 3D culture of MG-63 cells was evaluated by quantifying DNA (cell proliferation and cell viability. High concentrations of TSP (from 10 to 30 mM reduced both cell proliferation and viability (to 30% of the control after one week of exposure, but no such effects were found using low concentrations of TSP (0-10 mM.This study shows that low concentrations of TSP in 3D cell culture medium can be used for quantitative NMR or MRS examinations for up to two weeks post exposure.

  14. Decreased hypertrophic differentiation accompanies enhanced matrix formation in co-cultures of outer meniscus cells with bone marrow mesenchymal stromal cells

    Science.gov (United States)

    2012-01-01

    Introduction The main objective of this study was to determine whether meniscus cells from the outer (MCO) and inner (MCI) regions of the meniscus interact similarly to or differently with mesenchymal stromal stem cells (MSCs). Previous study had shown that co-culture of meniscus cells with bone marrow-derived MSCs result in enhanced matrix formation relative to mono-cultures of meniscus cells and MSCs. However, the study did not examine if cells from the different regions of the meniscus interacted similarly to or differently with MSCs. Methods Human menisci were harvested from four patients undergoing total knee replacements. Tissue from the outer and inner regions represented pieces taken from one third and two thirds of the radial distance of the meniscus, respectively. Meniscus cells were released from the menisci after collagenase treatment. Bone marrow MSCs were obtained from the iliac crest of two patients after plastic adherence and in vitro culture until passage 2. Primary meniscus cells from the outer (MCO) or inner (MCI) regions of the meniscus were co-cultured with MSCs in three-dimensional (3D) pellet cultures at 1:3 ratio, respectively, for 3 weeks in the presence of serum-free chondrogenic medium containing TGF-β1. Mono-cultures of MCO, MCI and MSCs served as experimental control groups. The tissue formed after 3 weeks was assessed biochemically, histochemically and by quantitative RT-PCR. Results Co-culture of inner (MCI) or outer (MCO) meniscus cells with MSCs resulted in neo-tissue with increased (up to 2.2-fold) proteoglycan (GAG) matrix content relative to tissues formed from mono-cultures of MSCs, MCI and MCO. Co-cultures of MCI or MCO with MSCs produced the same amount of matrix in the tissue formed. However, the expression level of aggrecan was highest in mono-cultures of MSCs but similar in the other four groups. The DNA content of the tissues from co-cultured cells was not statistically different from tissues formed from mono-cultures of

  15. Microfluidic perfusion culture of human induced pluripotent stem cells under fully defined culture conditions.

    Science.gov (United States)

    Yoshimitsu, Ryosuke; Hattori, Koji; Sugiura, Shinji; Kondo, Yuki; Yamada, Rotaro; Tachikawa, Saoko; Satoh, Taku; Kurisaki, Akira; Ohnuma, Kiyoshi; Asashima, Makoto; Kanamori, Toshiyuki

    2014-05-01

    Human induced pluripotent stem cells (hiPSCs) are a promising cell source for drug screening. For this application, self-renewal or differentiation of the cells is required, and undefined factors in the culture conditions are not desirable. Microfluidic perfusion culture allows the production of small volume cultures with precisely controlled microenvironments, and is applicable to high-throughput cellular environment screening. Here, we developed a microfluidic perfusion culture system for hiPSCs that uses a microchamber array chip under defined extracellular matrix (ECM) and culture medium conditions. By screening various ECMs we determined that fibronectin and laminin are appropriate for microfluidic devices made out of the most popular material, polydimethylsiloxane (PDMS). We found that the growth rate of hiPSCs under pressure-driven perfusion culture conditions was higher than under static culture conditions in the microchamber array. We applied our new system to self-renewal and differentiation cultures of hiPSCs, and immunocytochemical analysis showed that the state of the hiPSCs was successfully controlled. The effects of three antitumor drugs on hiPSCs were comparable between microchamber array and 96-well plates. We believe that our system will be a platform technology for future large-scale screening of fully defined conditions for differentiation cultures on integrated microfluidic devices. © 2013 Wiley Periodicals, Inc.

  16. Quantitative volumetric Raman imaging of three dimensional cell cultures

    KAUST Repository

    Kallepitis, Charalambos

    2017-03-22

    The ability to simultaneously image multiple biomolecules in biologically relevant three-dimensional (3D) cell culture environments would contribute greatly to the understanding of complex cellular mechanisms and cell–material interactions. Here, we present a computational framework for label-free quantitative volumetric Raman imaging (qVRI). We apply qVRI to a selection of biological systems: human pluripotent stem cells with their cardiac derivatives, monocytes and monocyte-derived macrophages in conventional cell culture systems and mesenchymal stem cells inside biomimetic hydrogels that supplied a 3D cell culture environment. We demonstrate visualization and quantification of fine details in cell shape, cytoplasm, nucleus, lipid bodies and cytoskeletal structures in 3D with unprecedented biomolecular specificity for vibrational microspectroscopy.

  17. Adherence of Moraxella bovis to cell cultures of bovine origin.

    Science.gov (United States)

    Annuar, B O; Wilcox, G E

    1985-09-01

    The adherence of five strains of Moraxella bovis to cell cultures was investigated. M bovis adhered to cultures of bovine corneal epithelial and Madin-Darby bovine kidney cells but not to cell types of non-bovine origin. Both piliated and unpiliated strains adhered but piliated strains adhered to a greater extent than unpiliated strains. Antiserum against pili of one strain inhibited adherence of piliated strains but caused only slight inhibition of adherence to the unpiliated strains. Treatment of bacteria with magnesium chloride caused detachment of pili from the bacterial cell and markedly inhibited adherence of piliated strains but caused only slight inhibition of adherence by the unpiliated strains. The results suggested that adhesion of piliated strains to cell cultures was mediated via pili but that adhesins other than pili may be involved in the attachment of unpiliated strains of M bovis to cells.

  18. Core–shell heterostructured metal oxide arrays enable superior light-harvesting and hysteresis-free mesoscopic perovskite solar cells

    KAUST Repository

    Mahmood, Khalid; Swain, Bhabani Sankar; Amassian, Aram

    2015-01-01

    To achieve highly efficient mesoscopic perovskite solar cells (PSCs), the structure and properties of an electron transport layer (ETL) or material (ETM) have been shown to be of supreme importance. Particularly, the core-shell heterostructured mesoscopic ETM architecture has been recognized as a successful electrode design, because of its large internal surface area, superior light-harvesting efficiency and its ability to achieve fast charge transport. Here we report the successful fabrication of a hysteresis-free, 15.3% efficient PSC using vertically aligned ZnO nanorod/TiO2 shell (ZNR/TS) core-shell heterostructured ETMs for the first time. We have also added a conjugated polyelectrolyte polymer into the growth solution to promote the growth of high aspect ratio (AR) ZNRs and substantially improve the infiltration of the perovskite light absorber into the ETM. The PSCs based on the as-synthesized core-shell ZnO/TiO2 heterostructured ETMs exhibited excellent performance enhancement credited to the superior light harvesting capability, larger surface area, prolonged charge-transport pathways and lower recombination rate. The unique ETM design together with minimal hysteresis introduces core-shell ZnO/TiO2 heterostructures as a promising mesoscopic electrode approach for the fabrication of efficient PSCs. This journal is © The Royal Society of Chemistry.

  19. Long-term culture and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized mesenchymal cells.

    Science.gov (United States)

    Garba, Abubakar; Acar, Delphine D; Roukaerts, Inge D M; Desmarets, Lowiese M B; Devriendt, Bert; Nauwynck, Hans J

    2017-09-01

    Mesenchymal cells are multipotent stromal cells with self-renewal, differentiation and immunomodulatory capabilities. We aimed to develop a co-culture model for differentiating hematopoietic cells on top of immortalized mesenchymal cells for studying interactions between hematopoietic and mesenchymal cells, useful for adequately exploring the therapeutic potential of mesenchymal cells. In this study, we investigated the survival, proliferation and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized porcine bone marrow mesenchymal cells for a period of five weeks. Directly after collection, primary porcine bone marrow mesenchymal cells adhered firmly to the bottom of the culture plates and showed a fibroblast-like appearance, one week after isolation. Upon immortalization, porcine bone marrow mesenchymal cells were continuously proliferating. They were positive for simian virus 40 (SV40) large T antigen and the mesenchymal cell markers CD44 and CD55. Isolated red bone marrow cells were added to these immortalized mesenchymal cells. Five weeks post-seeding, 92±6% of the red bone marrow hematopoietic cells were still alive and their number increased 3-fold during five weekly subpassages on top of the immortalized mesenchymal cells. The red bone marrow hematopoietic cells were originally small and round; later, the cells increased in size. Some of them became elongated, while others remained round. Tiny dendrites appeared attaching hematopoietic cells to the underlying immortalized mesenchymal cells. Furthermore, weekly differential-quick staining of the cells indicated the presence of monoblasts, monocytes, macrophages and lymphocytes in the co-cultures. At three weeks of co-culture, flow cytometry analysis showed an increased surface expression of CD172a, CD14, CD163, CD169, CD4 and CD8 up to 37±0.8%, 40±8%, 41±4%, 23±3% and 19±5% of the hematopoietic cells, respectively. In conclusion, continuous mesenchymal cell

  20. Cell-cycle distributions and radiation responses of Chinese hamster cells cultured continuously under hypoxic conditions

    International Nuclear Information System (INIS)

    Tokita, N.; Carpenter, S.G.; Raju, M.R.

    1984-01-01

    Cell-cycle distributions were measured by flow cytometry for Chinese hamster (CHO) cells cultured continuously under hypoxic conditions. DNA histograms showed an accumulation of cells in the early S phase followed by a traverse delay through the S phase, and a G 2 block. During hypoxic culturing, cell viability decreased rapidly to less than 0.1% at 120 h. Radiation responses for cells cultured under these conditions showed an extreme radioresistance at 72 h. Results suggest that hypoxia induces a condition similar to cell synchrony which itself changes the radioresistance of hypoxic cells. (author)

  1. In vitro production of azadirachtin from cell suspension cultures of ...

    Indian Academy of Sciences (India)

    PRAKASH KUMAR G

    proven effective in the control of agricultural pests in an environmentally ..... Prakash G and Srivastava A K 2005 Statistical media optimization for cell growth and ... Juss. suspension cultures; Process Biochemistry 40 3795–3800. Prakash G ...

  2. Establishment of the callus and cell suspension culture of ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-10-05

    Oct 5, 2009 ... Full Length Research Paper. Establishment of the callus ... study provided an efficient way for E. angustifolia cell suspension culture to produce secondary metabolite. .... was also observed that in these treatments the stem.

  3. Enhancement of Diosgenin Production in Plantlet and Cell Cultures ...

    African Journals Online (AJOL)

    Enhancement of Diosgenin Production in Plantlet and Cell Cultures of Dioscorea zingiberensis by Palmarumycin C13 from the Endophytic fungus, Berkleasmium sp. Dzf12. Y Mou, K Zhou, D Xu, R Yu, J Li, C Yin, L Zhou ...

  4. Establishment of sorghum cell suspension culture system for ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-03-18

    Mar 18, 2008 ... Additionally, sorghum cell suspension cultures have been initiated from the friable ... proteomics technologies. The field of proteomics is .... air dried at room temperature and resuspended in 2 ml of urea buffer [9 M urea, 2 M ...

  5. Immunocytochemical characterization of explant cultures of human prostatic stromal cells

    NARCIS (Netherlands)

    A. Kooistra (Anko); A.M.J. Elissen (Arianne ); J.J. Konig (Josee); M. Vermey; Th.H. van der Kwast (Theo); J.C. Romijn (Johannes); F.H. Schröder (Fritz)

    1995-01-01

    textabstractThe study of stromal-epithelial interactions greatly depends on the ability to culture both cell types separately, in order to permit analysis of their interactions under defined conditions in reconstitution experiments. Here we report the establishment of explant cultures of human

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

    NARCIS (Netherlands)

    Huang, L.; van Loveren, C.; Ling, J.; Wei, X.; Crielaard, W.; Deng, D.M.

    2016-01-01

    Porphyromonas gingivalis is present as a biofilm at the sites of periodontal infections. The detachment of gingival epithelial cells induced by P. gingivalis biofilms was examined using planktonic cultures as a comparison. Exponentially grown planktonic cultures or 40-h biofilms were co-incubated

  7. Free-energy carriers in human cultured muscle cells

    NARCIS (Netherlands)

    Bolhuis, P. A.; de Zwart, H. J.; Ponne, N. J.; de Jong, J. M.

    1985-01-01

    Creatine phosphate (CrP), adenosine triphosphate (ATP), creatine kinase (CK), adenylate kinase (AK), protein, and DNA were quantified in human muscle cell cultures undergoing transition from dividing myoblasts to multinucleate myotubes. CrP is negligible in cultures grown in commonly applied media

  8. Radiation effects on cultured human lymphoid cells

    International Nuclear Information System (INIS)

    Johansson, L.; Nilsson, K.; Carlsson, J.; Larsson, B.; Jakobsson, P.

    1981-01-01

    The cloning efficiency of human normal and malignant lymphoid cells is usually low. Radiation effects in vitro on such cells can therefore not be analysed with conventional cloning. However, this problem can be circumscribed by using the growth extrapolation method. A panel of human leukemia-lymphoma cell-lines representing Epstein-Barr virus carrying lymphoblastoid cells of presumed non-neoplastic derivation and neoplastic T- and B-lymphocytes was used to test the efficiency of this method. The sensitivity to radiation could be determined for all these cell types. The growth extrapolation method gave generally the same result as conventional cloning demonstrated by comparison with one exceptional cell-line with capacity for cloning in agar. The sensitivity varied largely between the different cell types. A common feature was that none of the cell lines had a good capacity to accumulate sublethal radiation injury. (Auth.)

  9. Radiosensitivity of cultured insect cells: II. Diptera

    International Nuclear Information System (INIS)

    Koval, T.M.

    1983-01-01

    The radiosensitivity of five dipteran cell lines representing three mosquito genera and one fruit fly genus were examined. These lines are: (1) ATC-10, Aedes aegypti; (2) RU-TAE-14, Toxorhynchites amboinensis; (3) RU-ASE-2A, Anopheles stephensi; (4) WR69-DM-1, Drosophila melanogaster; and (5) WR69-DM-2, Drosophila melanogaster. Population doubling times for these lines range from approximately 16 to 48 hr. Diploid chromosome numbers are six for the mosquito cells and eight for the fruit fly cells D 0 values are 5.1 and 6.5 Gy for the Drosophila cell lines and 3.6, 6.2, and 10.2 Gy for the mosquito cell lines. The results of this study demonstrate that dipteran insect cells are a few times more resistant to radiation than mammalian cells, but not nearly as radioresistant as lepidopteran cells

  10. Impact of cell culture process changes on endogenous retrovirus expression.

    Science.gov (United States)

    Brorson, Kurt; De Wit, Christina; Hamilton, Elizabeth; Mustafa, Mehnaz; Swann, Patrick G; Kiss, Robert; Taticek, Ron; Polastri, Gian; Stein, Kathryn E; Xu, Yuan

    2002-11-05

    Cell culture process changes (e.g., changes in scale, medium formulation, operational conditions) and cell line changes are common during the development life cycle of a therapeutic protein. To ensure that the impact of such process changes on product quality and safety is minimal, it is standard practice to compare critical product quality and safety attributes before and after the changes. One potential concern introduced by cell culture process improvements is the possibility of increased endogenous retrovirus expression to a level above the clearance capability of the subsequent purification process. To address this, retrovirus expression was measured in scaled down and full production scaled Chinese hamster ovary (CHO) cell cultures of four monoclonal antibodies and one recombinant protein before and after process changes. Two highly sensitive, quantitative (Q)-PCR-based assays were used to measure endogenous retroviruses. It is shown that cell culture process changes that primarily alter media components, nutrient feed volume, seed density, cell bank source (i.e., master cell bank vs. working cell bank), and vial size, or culture scale, singly or in combination, do not impact the rate of retrovirus expression to an extent greater than the variability of the Q-PCR assays (0.2-0.5 log(10)). Cell culture changes that significantly alter the metabolic state of the cells and/or rates of protein expression (e.g., pH and temperature shifts, NaButyrate addition) measurably impact the rate of retrovirus synthesis (up to 2 log(10)). The greatest degree of variation in endogenous retrovirus expression was observed between individual cell lines (up to 3 log(10)). These data support the practice of measuring endogenous retrovirus output for each new cell line introduced into manufacturing or after process changes that significantly increase product-specific productivity or alter the metabolic state, but suggest that reassessment of retrovirus expression after other

  11. 5-Fluorouracil-induced apoptosis in cultured oral cancer cells.

    Science.gov (United States)

    Tong, D; Poot, M; Hu, D; Oda, D

    2000-03-01

    Chemotherapy is commonly used to treat advanced oral squamous cell carcinoma (SCC) and is known to kill cancer cells through apoptosis. Our hypothesis states that 5-fluorouracil (5FU) also kills cultured oral epithelial cells through programmed cell death or apoptosis. Cultured oral cancer cells were exposed to an optimum dose of 20 mg/ml of 5FU. Cells were analyzed for changes in cell cycle distribution and induction of cell death including apoptosis. Normal control, human papilloma virus-immortalized (PP), ATCC SCC cell line (CA1) and two primary oral SCC cell lines (CA3 and -4) were studied. Inhibition of apoptosis by a pan-caspase inhibitor was used. SYTO 11 flow cytometry showed increased apoptosis in all 5FU-treated cell cultures compared to untreated controls. The results show biological variation in apoptotic response. CA1 had the lowest apoptotic rate of the cancer cell lines at 1.5%. Next lowest was CA3, followed by CA4 and PP. In addition, alteration in the G1 and S phase fractions were found. Untreated CA1 showed 28% G1, 53% S compared to 43% G1, and 40% S of treated. We investigated the pathway of apoptosis using the pan-caspase inhibitor IDN-1529 by methylthiazolyl diphenyl tetrazolium bromide (MTT) colorimetric analysis. Results showed mild inhibition of cell death when cells were incubated with 50 microM IDN-1529 for 24 h. This suggests a probable caspase-dependent apoptotic pathway. In conclusion, our data suggest that 5FU induces oral cancer cell death through apoptosis and that biological variation exists between normal and cancer cells and between different types of cancer cells themselves. Our data indicate that cultures of a useful in vitro model for chemosensitivity assays are possible. Our results also suggest a caspase-dependent pathway for chemocytotoxicity in oral SCC.

  12. 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-te...

  13. Good Cell Culture Practice for stem cells and stem-cell-derived models.

    Science.gov (United States)

    Pamies, David; Bal-Price, Anna; Simeonov, Anton; Tagle, Danilo; Allen, Dave; Gerhold, David; Yin, Dezhong; Pistollato, Francesca; Inutsuka, Takashi; Sullivan, Kristie; Stacey, Glyn; Salem, Harry; Leist, Marcel; Daneshian, Mardas; Vemuri, Mohan C; McFarland, Richard; Coecke, Sandra; Fitzpatrick, Suzanne C; Lakshmipathy, Uma; Mack, Amanda; Wang, Wen Bo; Yamazaki, Daiju; Sekino, Yuko; Kanda, Yasunari; Smirnova, Lena; Hartung, Thomas

    2017-01-01

    The first guidance on Good Cell Culture Practice (GCCP) dates back to 2005. This document expands this to include aspects of quality assurance for in vitro cell culture focusing on the increasingly diverse cell types and culture formats used in research, product development, testing and manufacture of biotechnology products and cell-based medicines. It provides a set of basic principles of best practice that can be used in training new personnel, reviewing and improving local procedures, and helping to assure standard practices and conditions for the comparison of data between laboratories and experimentation performed at different times. This includes recommendations for the documentation and reporting of culture conditions. It is intended as guidance to facilitate the generation of reliable data from cell culture systems, and is not intended to conflict with local or higher level legislation or regulatory requirements. It may not be possible to meet all recommendations in this guidance for practical, legal or other reasons. However, when it is necessary to divert from the principles of GCCP, the risk of decreasing the quality of work and the safety of laboratory staff should be addressed and any conclusions or alternative approaches justified. This workshop report is considered a first step toward a revised GCCP 2.0.

  14. Granulocyte-colony stimulating factor (G-CSF)-primed, delayed marrow harvests as a source of hematopoietic stem and progenitor cells for allogeneic transplantation.

    Science.gov (United States)

    Phillips, G L; Davey, D D; Hale, G A; Marshall, K W; Munn, R K; Nath, R; Reece, D E; Van Zant, G

    1999-10-01

    We evaluated the ability of G-CSF to increase the number of hematopoietic stem cells obtained by "delayed" BM harvest for allogeneic transplantation. Five normal donors received G-CSF @ 10 mcg/kg/day x 5 followed by repeat PB and BM assays at day 6 and 16, and BM harvest at day 16. Stem cells were not increased in the BM at day 16. Five patients underwent BMT and engrafted at +10 to +19 days. While the tested strategy offers no intrinsic advantages, its potential cannot be evaluated fully without alternative timing and/or additional, "early acting" growth factors.

  15. Umbilical cord Wharton's jelly repeated culture system: a new device and method for obtaining abundant mesenchymal stem cells for bone tissue engineering.

    Directory of Open Access Journals (Sweden)

    Zhengqi Chang

    Full Text Available To date, various types of cells for seeding regenerative scaffolds have been used for bone tissue engineering. Among seed cells, the mesenchymal stem cells derived from human umbilical cord Wharton's jelly (hUCMSCs represent a promising candidate and hold potential for bone tissue engineering due to the the lack of ethical controversies, accessibility, sourced by non-invasive procedures for donors, a reduced risk of contamination, osteogenic differentiation capacities, and higher immunomodulatory capacity. However, the current culture methods are somewhat complicated and inefficient and often fail to make the best use of the umbilical cord (UC tissues. Moreover, these culture processes cannot be performed on a large scale and under strict quality control. As a result, only a small quantity of cells can be harvested using the current culture methods. To solve these problems, we designed and evaluated an UC Wharton's jelly repeated culture device. Using this device, hUCMSCs were obtained from the repeated cultures and their quantities and biological characteristics were compared. We found that using our culture device, which retained all tissue blocks on the bottom of the dish, the total number of obtained cells increased 15-20 times, and the time required for the primary passage was reduced. Moreover, cells harvested from the repeated cultures exhibited no significant difference in their immunophenotype, potential for multilineage differentiation, or proliferative, osteoinductive capacities, and final osteogenesis. The application of the repeated culture frame (RCF not only made full use of the Wharton's jelly but also simplified and specified the culture process, and thus, the culture efficiency was significantly improved. In summary, abundant hUCMSCs of dependable quality can be acquired using the RCF.

  16. Cell proliferation and radiosensitivity of cow lymphocytes in culture

    International Nuclear Information System (INIS)

    Modave, C.; Fabry, L.; Leonard, A.

    1982-01-01

    The harlequin-staining technique has been used to study, after PHA-stimulation, the cell proliferation of cow lymphocytes in culture and to assess the radiosensitivity in first mitosis cells. At the 48 h fixation time, only 34% of the cells are in first mitosis whereas 55% are already in second and 11% in third mitosis. The exposure of cow lymphocytes to 200 rad X-rays result in the production of 16% dicentric chromosomes in first mitosis cells [fr

  17. Topological defects control collective dynamics in neural progenitor cell cultures

    Science.gov (United States)

    Kawaguchi, Kyogo; Kageyama, Ryoichiro; Sano, Masaki

    2017-04-01

    Cultured stem cells have become a standard platform not only for regenerative medicine and developmental biology but also for biophysical studies. Yet, the characterization of cultured stem cells at the level of morphology and of the macroscopic patterns resulting from cell-to-cell interactions remains largely qualitative. Here we report on the collective dynamics of cultured murine neural progenitor cells (NPCs), which are multipotent stem cells that give rise to cells in the central nervous system. At low densities, NPCs moved randomly in an amoeba-like fashion. However, NPCs at high density elongated and aligned their shapes with one another, gliding at relatively high velocities. Although the direction of motion of individual cells reversed stochastically along the axes of alignment, the cells were capable of forming an aligned pattern up to length scales similar to that of the migratory stream observed in the adult brain. The two-dimensional order of alignment within the culture showed a liquid-crystalline pattern containing interspersed topological defects with winding numbers of +1/2 and -1/2 (half-integer due to the nematic feature that arises from the head-tail symmetry of cell-to-cell interaction). We identified rapid cell accumulation at +1/2 defects and the formation of three-dimensional mounds. Imaging at the single-cell level around the defects allowed us to quantify the velocity field and the evolving cell density; cells not only concentrate at +1/2 defects, but also escape from -1/2 defects. We propose a generic mechanism for the instability in cell density around the defects that arises from the interplay between the anisotropic friction and the active force field.

  18. Enhancement of Chlorella vulgaris harvesting via the electro-coagulation-flotation (ECF) method.

    Science.gov (United States)

    Wong, Y K; Ho, Y H; Leung, H M; Ho, K C; Yau, Y H; Yung, K K L

    2017-04-01

    This article explores the potential of using an electro-coagulation-flotation (ECF) harvester to allow flotation of microalgae cells for surface harvesting. A response surface methodology (RSM) model was used to optimize ECF harvesting by adjusting electrode plate material, electrode plate number, charge of the electrodes, electrolyte concentration, and pH value of the culture solution. The result revealed that three aluminum electrode plates (one anode and two cathodes), brine solution (8 g/L), and acidity (pH = 4) of culture solution (optimized ECF harvester) The highest flocculant concentration was measured at 2966 mg/L after 60 min and showed a 79.8 % increase of flocculation concentration. Such results can provide a basis for designing a large-scale microalgae harvester for commercial use in the future.

  19. Characterization of glucocerebrosidase in peripheral blood cells and cultured blastoid cells

    NARCIS (Netherlands)

    Aerts, J. M.; Heikoop, J.; van Weely, S.; Donker-Koopman, W. E.; Barranger, J. A.; Tager, J. M.; Schram, A. W.

    1988-01-01

    We have characterized glucocerebrosidase in various cell types of peripheral blood of control subjects and in cultured human blastoid cells. The intracellular level of glucocerebrosidase in cultured blastoid cells (10-30 nmol substrate hydrolyzed/h.mg protein) resembles closely values observed for

  20. EXPLANTATION OF MESANGIAL CELL HILLOCKS - A METHOD FOR OBTAINING HUMAN MESANGIAL CELLS IN CULTURE

    NARCIS (Netherlands)

    MULLER, EW; KIM, Y; MICHAEL, AF; VERNIER, RL; VANDERHEM, GK; VANDERWOUDE, FJ

    A simple method is presented for selective cell culture of human mesangial cells using explanatation of mesangial cell hillocks. Glomeruli which had been incubated with collagenase were explanted on plastic tissue culture flasks. Three to 6 weeks after explantation, a rapidly growing multilayer of

  1. Introducing Mammalian Cell Culture and Cell Viability Techniques in the Undergraduate Biology Laboratory.

    Science.gov (United States)

    Bowey-Dellinger, Kristen; Dixon, Luke; Ackerman, Kristin; Vigueira, Cynthia; Suh, Yewseok K; Lyda, Todd; Sapp, Kelli; Grider, Michael; Crater, Dinene; Russell, Travis; Elias, Michael; Coffield, V McNeil; Segarra, Verónica A

    2017-01-01

    Undergraduate students learn about mammalian cell culture applications in introductory biology courses. However, laboratory modules are rarely designed to provide hands-on experience with mammalian cells or teach cell culture techniques, such as trypsinization and cell counting. Students are more likely to learn about cell culture using bacteria or yeast, as they are typically easier to grow, culture, and manipulate given the equipment, tools, and environment of most undergraduate biology laboratories. In contrast, the utilization of mammalian cells requires a dedicated biological safety cabinet and rigorous antiseptic techniques. For this reason, we have devised a laboratory module and method herein that familiarizes students with common cell culture procedures, without the use of a sterile hood or large cell culture facility. Students design and perform a time-efficient inquiry-based cell viability experiment using HeLa cells and tools that are readily available in an undergraduate biology laboratory. Students will become familiar with common techniques such as trypsinizing cells, cell counting with a hemocytometer, performing serial dilutions, and determining cell viability using trypan blue dye. Additionally, students will work with graphing software to analyze their data and think critically about the mechanism of death on a cellular level. Two different adaptations of this inquiry-based lab are presented-one for non-biology majors and one for biology majors. Overall, these laboratories aim to expose students to mammalian cell culture and basic techniques and help them to conceptualize their application in scientific research.

  2. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Science.gov (United States)

    2010-04-01

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

  3. Culture of Iris Pigment Epithelial Cells on Expanded-Polytetrafluroethylene (ePTFE Substrates for the Treatment of Age-Related Macular Degeneration

    Directory of Open Access Journals (Sweden)

    S Nian

    2011-05-01

    Full Text Available Introduction: Transplantation of an intact differentiated retinal pigment epithelial (RPE cell layer may provide a means to treat Age-Related Macular Degeneration (AMD. However, harvesting RPE cells can be a technically complicated procedure. Our current work aimed to prepare intact differentiated iris pigment epithelial (IPE cell layers, which are easy to obtain and have the same embryonic origin and similar properties as RPE cells, on ePTFE substrates for transplantation purposes to rescue deteriorated photoreceptors in AMD. Methods: IPE cells isolated from rat eyes were seeded on different substrates, including fibronectin n-heptylamine (HA ePTFE substrates, HA ePTFE substrates, ePTFE substrates and fibronectin tissue culture polystyrene (TCPS as control. Cell number and morphology were assessed at each time interval. The formation of tight junction was examined by immunostaining of junction proteins. Results: An obvious increasing trend of cell number was observed in IPE cells on fibronectin n-heptylamine (HA ePTFE substrate, exhibiting heavy pigmentation and epithelial morphology. At Day 28, tight junction formation was indicated by cell-cell junctional proteins along cell borders. Conclusion: Harvested IPE cells cultured on fibronectin HA-ePTFE substrates can differentiate and form a cell monolayer that may be suitable for transplantation.

  4. Increased light harvesting in dye-sensitized solar cells with energy relay dyes

    KAUST Repository

    Hardin, Brian E.; Hoke, Eric T.; Armstrong, Paul B.; Yum, Jun-Ho; Comte, Pascal; Torres, Tomá s; Fré chet, Jean M. J.; Nazeeruddin, Md Khaja; Grä tzel, Michael; McGehee, Michael D.

    2009-01-01

    Conventional dye-sensitized solar cells have excellent charge collection efficiencies, high open-circuit voltages and good fill factors. However, dye-sensitized solar cells do not completely absorb all of the photons from the visible and near

  5. Aromatase inhibitor (anastrozole) affects growth of endometrioma cells in culture.

    Science.gov (United States)

    Badawy, Shawky Z A; Brown, Shereene; Kaufman, Lydia; Wojtowycz, Martha A

    2015-05-01

    To study the effects of aromatase inhibitor (anastrozole) on the growth and estradiol secretion of endometrioma cells in culture. Endometrioma cells are grown in vitro until maximum growth before used in this study. This was done in the research laboratory for tissue culture, in an academic hospital. Testosterone at a concentration of 10 μg/mL was added as a substrate for the intracellular aromatase. In addition, aromatase inhibitor was added at a concentration of 200 and 300 μg/mL. The effect on cell growth and estradiol secretion is evaluated using Student's t-test. The use of testosterone increased estradiol secretion by endometrioma cells in culture. The use of aromatase inhibitor significantly inhibited the growth of endometrioma cells, and estradiol secretion. Aromatase inhibitor (anastrozole) may be an effective treatment for endometriosis due to inhibition of cellular aromatase. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

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

    DEFF Research Database (Denmark)

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

    2011-01-01

    Standard cell culture plastic was surface modified by passive adsorption or covalent attachment of interleukin (IL)-4 and investigated for its ability to induce differentiation of human monocytes into mature dendritic cells, a process dose-dependently regulated by IL-4. Covalent attachment of IL-4...... in water instead of phosphate-buffered saline. Passively adsorbed IL-4 was observed to induce differentiation to dendritic cells, but analysis of cell culture supernatants revealed that leakage of IL-4 into solution could account for the differentiation observed. Covalent attachment resulted in bound IL-4...... at similar concentrations to the passive adsorption process, as measured by enzyme-linked immunosorbent assays, and the bound IL-4 did not leak into solution to any measurable extent during cell culture. However, covalently bound IL-4 was incapable of inducing monocyte differentiation. This may be caused...

  7. Determination of thymidine in serum used for cell culture media

    International Nuclear Information System (INIS)

    Schaer, J.C.; Maurer, U.; Schindler, R.

    1978-01-01

    Thymidine concentrations in serum used for cell culture media were determined with an assay based on isotope dilution. In this assay, incorporation of (3H)-thymidine into DNA of cultured cells was measured in the presence of 5 and 20% serum as a function of the concentration of unlabeled thymidine added to the medium. Thymidine concentrations were measured using horse serum as well as fetal calf serum in the culture media. Dialysis of serum resulted in a reduction of thymidine levels by factors of at least 10

  8. Efficient Secretion of Recombinant Proteins from Rice Suspension-Cultured Cells Modulated by the Choice of Signal Peptide.

    Science.gov (United States)

    Huang, Li-Fen; Tan, Chia-Chun; Yeh, Ju-Fang; Liu, Hsin-Yi; Liu, Yu-Kuo; Ho, Shin-Lon; Lu, Chung-An

    2015-01-01

    Plant-based expression systems have emerged as a competitive platform in the large-scale production of recombinant proteins. By adding a signal peptide, αAmy3sp, the desired recombinant proteins can be secreted outside transgenic rice cells, making them easy to harvest. In this work, to improve the secretion efficiency of recombinant proteins in rice expression systems, various signal peptides including αAmy3sp, CIN1sp, and 33KDsp have been fused to the N-terminus of green fluorescent protein (GFP) and introduced into rice cells to explore the efficiency of secretion of foreign proteins. 33KDsp had better efficiency than αAmy3sp and CIN1sp for the secretion of GFP from calli and suspension-cultured cells. 33KDsp was further applied for the secretion of mouse granulocyte-macrophage colony-stimulating factor (mGM-CSF) from transgenic rice suspension-cultured cells; approximately 76%-92% of total rice-derived mGM-CSF (rmGM-CSF) was detected in the culture medium. The rmGM-CSF was bioactive and could stimulate the proliferation of a murine myeloblastic leukemia cell line, NSF-60. The extracellular yield of rmGM-CSF reached 31.7 mg/L. Our study indicates that 33KDsp is better at promoting the secretion of recombinant proteins in rice suspension-cultured cell systems than the commonly used αAmy3sp.

  9. Callus and cell suspension cultures of carnation

    DEFF Research Database (Denmark)

    Engvild, Kjeld Christensen

    1972-01-01

    Callus cultures of carnation, Dianthus caryophyllus L. ev. G. J. Sim, were grown on a synthetic medium of half strength Murashige and Skoog salts, 3 % sucrose, 100 mg/l of myo-inositol, 0.5 mg/l each of thiamin, HCl, pyridoxin, HCl and nicotinic acid and 10 g/l agar. Optimal concentrations...

  10. Systematic microcarrier screening and agitated culture conditions improves human mesenchymal stem cell yield in bioreactors

    Science.gov (United States)

    Rafiq, Qasim A.; Coopman, Karen; Nienow, Alvin W.

    2016-01-01

    Abstract Production of human mesenchymal stem cells for allogeneic cell therapies requires scalable, cost‐effective manufacturing processes. Microcarriers enable the culture of anchorage‐dependent cells in stirred‐tank bioreactors. However, no robust, transferable methodology for microcarrier selection exists, with studies providing little or no reason explaining why a microcarrier was employed. We systematically evaluated 13 microcarriers for human bone marrow‐derived MSC (hBM‐MSCs) expansion from three donors to establish a reproducible and transferable methodology for microcarrier selection. Monolayer studies demonstrated input cell line variability with respect to growth kinetics and metabolite flux. HBM‐MSC1 underwent more cumulative population doublings over three passages in comparison to hBM‐MSC2 and hBM‐MSC3. In 100 mL spinner flasks, agitated conditions were significantly better than static conditions, irrespective of donor, and relative microcarrier performance was identical where the same microcarriers outperformed others with respect to growth kinetics and metabolite flux. Relative growth kinetics between donor cells on the microcarriers were the same as the monolayer study. Plastic microcarriers were selected as the optimal microcarrier for hBM‐MSC expansion. HBM‐MSCs were successfully harvested and characterised, demonstrating hBM‐MSC immunophenotype and differentiation capacity. This approach provides a systematic method for microcarrier selection, and the findings identify potentially significant bioprocessing implications for microcarrier‐based allogeneic cell therapy manufacture. PMID:26632496

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

    Science.gov (United States)

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

    2018-05-21

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

  12. Cell death in Tetrahymena thermophila: new observations on culture conditions.

    Science.gov (United States)

    Christensen, S T; Sørensen, H; Beyer, N H; Kristiansen, K; Rasmussen, L; Rasmussen, M I

    2001-01-01

    We previously suggested that the cell fate of the protozoan ciliate, Tetrahymena thermophila, effectively relates to a quorum-sensing mechanism where cell-released factors support cell survival and proliferation. The cells have to be present above a critical initial density in a chemically defined nutrient medium in order to release a sufficient level of these factors to allow a new colony to flourish. At a relatively high rate of metabolism and/or macromolecular synthesis and below this critical density, cells began to die abruptly within 30 min of inoculation, and this death took the form of an explosive disintegration lasting less than 50 milliseconds. The cells died at any location in the culture, and the frequency of cell death was always lower in well-filled vials than those with medium/air interface. Cell death was inhibited by the addition of Actinomycin D or through modifications of the culture conditions either by reducing the oxygen tension or by decreasing the temperature of the growth medium. In addition, plastic caps in well-filled vials release substances, which promote cell survival. The fate of low-density cultures is related to certain 'physical' conditions, in addition to the availability of oxygen within closed culture systems. Copyright 2001 Academic Press.

  13. Contributions of 3D Cell Cultures for Cancer Research.

    Science.gov (United States)

    Ravi, Maddaly; Ramesh, Aarthi; Pattabhi, Aishwarya

    2017-10-01

    Cancer cell lines have contributed immensely in understanding the complex physiology of cancers. They are excellent material for studies as they offer homogenous samples without individual variations and can be utilised with ease and flexibility. Also, the number of assays and end-points one can study is almost limitless; with the advantage of improvising, modifying or altering several variables and methods. Literally, a new dimension to cancer research has been achieved by the advent of 3Dimensional (3D) cell culture techniques. This approach increased many folds the ways in which cancer cell lines can be utilised for understanding complex cancer biology. 3D cell culture techniques are now the preferred way of using cancer cell lines to bridge the gap between the 'absolute in vitro' and 'true in vivo'. The aspects of cancer biology that 3D cell culture systems have contributed include morphology, microenvironment, gene and protein expression, invasion/migration/metastasis, angiogenesis, tumour metabolism and drug discovery, testing chemotherapeutic agents, adaptive responses and cancer stem cells. We present here, a comprehensive review on the applications of 3D cell culture systems for these aspects of cancers. J. Cell. Physiol. 232: 2679-2697, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  14. Suspension culture of pluripotent stem cells: effect of shear on stem cell fate.

    Science.gov (United States)

    Keller, Kevin C; Rodrigues, Beatriz; zur Nieden, Nicole I

    2014-01-01

    Despite significant promise, the routine usage of suspension cell culture to manufacture stem cell-derived differentiated cells has progressed slowly. Suspension culture is an innovative way of either expanding or differentiating cells and sometimes both are combined into a single bioprocess. Its advantages over static 2D culturing include a homogeneous and controllable culture environment and producing a large quantity of cells in a fraction of time. This feature makes suspension cell culture ideal for use in stem cell research and eventually ideal in the large-scale production of differentiated cells for regenerative medicine. Because of their tremendous differentiation capacities and unlimited growth properties, pluripotent stem cells (PSCs) in particular are considered potential sources for future cell-replacement therapies. Currently, expansion of PSCs is accomplished in 2D, which only permits a limited amount of cell growth per culture flask before cells need to be passaged. However, before stem cells can be applied clinically, several aspects of their expansion, such as directed growth, but also differentiation, need to be better controlled. This review will summarize recent advantages in suspension culture of PSCs, while at the same time highlighting current challenges.

  15. Culture conditions for bovine embryonic stem cell-like cells isolated from blastocysts after external fertilization

    OpenAIRE

    Jin, Muzi; Wu, Asga; Dorzhin, Sergei; Yue, Qunhua; Ma, Yuzhen; Liu, Dongjun

    2012-01-01

    Although isolation and characterization of embryonic stem cells have been successful in cattle, maintenance of bovine embryonic stem cells in culture remains difficult. In this study, we compared different methods of cell passaging, feeder cell layers and medium conditions for bovine embryonic stem cell-like cells. We found that a murine embryonic fibroblast feeder layer is more suitable for embryonic stem cell-like cells than bovine embryonic fibroblasts. When murine embryonic fibroblasts we...

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

    DEFF Research Database (Denmark)

    Rasmussen, Eva; Nicolaisen, G.M.

    1999-01-01

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

  17. Automation of 3D cell culture using chemically defined hydrogels.

    Science.gov (United States)

    Rimann, Markus; Angres, Brigitte; Patocchi-Tenzer, Isabel; Braum, Susanne; Graf-Hausner, Ursula

    2014-04-01

    Drug development relies on high-throughput screening involving cell-based assays. Most of the assays are still based on cells grown in monolayer rather than in three-dimensional (3D) formats, although cells behave more in vivo-like in 3D. To exemplify the adoption of 3D techniques in drug development, this project investigated the automation of a hydrogel-based 3D cell culture system using a liquid-handling robot. The hydrogel technology used offers high flexibility of gel design due to a modular composition of a polymer network and bioactive components. The cell inert degradation of the gel at the end of the culture period guaranteed the harmless isolation of live cells for further downstream processing. Human colon carcinoma cells HCT-116 were encapsulated and grown in these dextran-based hydrogels, thereby forming 3D multicellular spheroids. Viability and DNA content of the cells were shown to be similar in automated and manually produced hydrogels. Furthermore, cell treatment with toxic Taxol concentrations (100 nM) had the same effect on HCT-116 cell viability in manually and automated hydrogel preparations. Finally, a fully automated dose-response curve with the reference compound Taxol showed the potential of this hydrogel-based 3D cell culture system in advanced drug development.

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

    DEFF Research Database (Denmark)

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

    2005-01-01

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

  19. Cell culture supernatants for detection perforin ELISA

    African Journals Online (AJOL)

    Najwa

    2014-02-19

    Feb 19, 2014 ... Leukemia is a cancer originating in any of hematopoietic cell that tends to ... treatment of children (Borek and Jaskolski, 2001). The current study was .... which led to the best results at 48 h of exposure than after 72 h of cells ...

  20. Isolation of plasma membranes from cultured glioma cells and application to evaluation of membrane sphingomyelin turnover

    International Nuclear Information System (INIS)

    Cook, H.W.; Palmer, F.B.; Byers, D.M.; Spence, M.W.

    1988-01-01

    A rapid and reliable method for the isolation of plasma membranes and microsomes of high purity and yield from cultured glioma cells is described. The procedure involves disruption by N2 cavitation, preliminary separation by centrifugation in Tricine buffer, and final separation on a gradient formed from 40% Percoll at pH 9.3. Enzyme and chemical markers indicated greater than 60% yield with six- to eightfold enrichment for plasma membranes and greater than 25% yield with three- to fourfold enrichment for a microsomal fraction consisting mainly of endoplasmic reticulum. The final fractions were obtained with high reproducibility in less than 1 h from the time of cell harvesting. Application of this procedure to human fibroblasts in culture is assessed. The isolation procedure was applied to investigations of synthesis and turnover of sphingomyelin and phosphatidylcholine in plasma membranes of glioma cells following incubation for 4-24 h with [methyl- 3 H]choline. These studies indicated that radioactivity from phosphatidylcholine synthesized in microsomes from exogenous choline may serve as a precursor of the head-group of sphingomyelin accumulating in the plasma membrane

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

    DEFF Research Database (Denmark)

    Richter, W W; Issinger, O G

    1986-01-01

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

  2. Isolation and Characterization of Poliovirus in Cell Culture Systems.

    Science.gov (United States)

    Thorley, Bruce R; Roberts, Jason A

    2016-01-01

    The isolation and characterization of enteroviruses by cell culture was accepted as the "gold standard" by clinical virology laboratories. Methods for the direct detection of all enteroviruses by reverse transcription polymerase chain reaction, targeting a conserved region of the genome, have largely supplanted cell culture as the principal diagnostic procedure. However, the World Health Organization's Global Polio Eradication Initiative continues to rely upon cell culture to isolate poliovirus due to the lack of a reliable sensitive genetic test for direct typing of enteroviruses from clinical specimens. Poliovirus is able to infect a wide range of mammalian cell lines, with CD155 identified as the primary human receptor for all three seroytpes, and virus replication leads to an observable cytopathic effect. Inoculation of cell lines with extracts of clinical specimens and subsequent passaging of the cells leads to an increased virus titre. Cultured isolates of poliovirus are suitable for testing by a variety of methods and remain viable for years when stored at low temperature.This chapter describes general procedures for establishing a cell bank and routine passaging of cell lines. While the sections on specimen preparation and virus isolation focus on poliovirus, the protocols are suitable for other enteroviruses.

  3. Animal-cell culture in aqueous two-phase systems

    NARCIS (Netherlands)

    Zijlstra, G.M.

    1998-01-01

    In current industrial biotechnology, animal-cell culture is an important source of therapeutic protein products. The conventional animal-cell production processes, however, include many unit operations as part of the fermentation and downstream processing strategy. The research described in

  4. Endothelial cell cultures as a tool in biomaterial research

    NARCIS (Netherlands)

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

    1999-01-01

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

  5. Rapid and reliable identification of Gram-negative bacteria and Gram-positive cocci by deposition of bacteria harvested from blood cultures onto the MALDI-TOF plate.

    Science.gov (United States)

    Barnini, Simona; Ghelardi, Emilia; Brucculeri, Veronica; Morici, Paola; Lupetti, Antonella

    2015-06-18

    Rapid identification of the causative agent(s) of bloodstream infections using the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) methodology can lead to increased empirical antimicrobial therapy appropriateness. Herein, we aimed at establishing an easier and simpler method, further referred to as the direct method, using bacteria harvested by serum separator tubes from positive blood cultures and placed onto the polished steel target plate for rapid identification by MALDI-TOF. The results by the direct method were compared with those obtained by MALDI-TOF on bacteria isolated on solid media. Identification of Gram-negative bacilli was 100 % concordant using the direct method or MALDI-TOF on isolated bacteria (96 % with score > 2.0). These two methods were 90 % concordant on Gram-positive cocci (32 % with score > 2.0). Identification by the SepsiTyper method of Gram-positive cocci gave concordant results with MALDI-TOF on isolated bacteria in 87 % of cases (37 % with score > 2.0). The direct method herein developed allows rapid identification (within 30 min) of Gram-negative bacteria and Gram-positive cocci from positive blood cultures and can be used to rapidly report reliable and accurate results, without requiring skilled personnel or the use of expensive kits.

  6. Co-cultivation of fungal and microalgal cells as an efficient system for harvesting microalgal cells, lipid production and wastewater treatment.

    Directory of Open Access Journals (Sweden)

    Digby Wrede

    Full Text Available The challenges which the large scale microalgal industry is facing are associated with the high cost of key operations such as harvesting, nutrient supply and oil extraction. The high-energy input for harvesting makes current commercial microalgal biodiesel production economically unfeasible and can account for up to 50% of the total cost of biofuel production. Co-cultivation of fungal and microalgal cells is getting increasing attention because of high efficiency of bio-flocculation of microalgal cells with no requirement for added chemicals and low energy inputs. Moreover, some fungal and microalgal strains are well known for their exceptional ability to purify wastewater, generating biomass that represents a renewable and sustainable feedstock for biofuel production. We have screened the flocculation efficiency of the filamentous fungus A. fumigatus against 11 microalgae representing freshwater, marine, small (5 µm, large (over 300 µm, heterotrophic, photoautotrophic, motile and non-motile strains. Some of the strains are commercially used for biofuel production. Lipid production and composition were analysed in fungal-algal pellets grown on media containing alternative carbon, nitrogen and phosphorus sources contained in wheat straw and swine wastewater, respectively. Co-cultivation of algae and A. fumigatus cells showed additive and synergistic effects on biomass production, lipid yield and wastewater bioremediation efficiency. Analysis of fungal-algal pellet's fatty acids composition suggested that it can be tailored and optimised through co-cultivating different algae and fungi without the need for genetic modification.

  7. Co-cultivation of fungal and microalgal cells as an efficient system for harvesting microalgal cells, lipid production and wastewater treatment.

    Science.gov (United States)

    Wrede, Digby; Taha, Mohamed; Miranda, Ana F; Kadali, Krishna; Stevenson, Trevor; Ball, Andrew S; Mouradov, Aidyn

    2014-01-01

    The challenges which the large scale microalgal industry is facing are associated with the high cost of key operations such as harvesting, nutrient supply and oil extraction. The high-energy input for harvesting makes current commercial microalgal biodiesel production economically unfeasible and can account for up to 50% of the total cost of biofuel production. Co-cultivation of fungal and microalgal cells is getting increasing attention because of high efficiency of bio-flocculation of microalgal cells with no requirement for added chemicals and low energy inputs. Moreover, some fungal and microalgal strains are well known for their exceptional ability to purify wastewater, generating biomass that represents a renewable and sustainable feedstock for biofuel production. We have screened the flocculation efficiency of the filamentous fungus A. fumigatus against 11 microalgae representing freshwater, marine, small (5 µm), large (over 300 µm), heterotrophic, photoautotrophic, motile and non-motile strains. Some of the strains are commercially used for biofuel production. Lipid production and composition were analysed in fungal-algal pellets grown on media containing alternative carbon, nitrogen and phosphorus sources contained in wheat straw and swine wastewater, respectively. Co-cultivation of algae and A. fumigatus cells showed additive and synergistic effects on biomass production, lipid yield and wastewater bioremediation efficiency. Analysis of fungal-algal pellet's fatty acids composition suggested that it can be tailored and optimised through co-cultivating different algae and fungi without the need for genetic modification.

  8. Differentiation of oligodendrocyte progenitor cells from dissociated monolayer and feeder-free cultured pluripotent stem cells.

    Science.gov (United States)

    Yamashita, Tomoko; Miyamoto, Yuki; Bando, Yoshio; Ono, Takashi; Kobayashi, Sakurako; Doi, Ayano; Araki, Toshihiro; Kato, Yosuke; Shirakawa, Takayuki; Suzuki, Yutaka; Yamauchi, Junji; Yoshida, Shigetaka; Sato, Naoya

    2017-01-01

    Oligodendrocytes myelinate axons and form myelin sheaths in the central nervous system. The development of therapies for demyelinating diseases, including multiple sclerosis and leukodystrophies, is a challenge because the pathogenic mechanisms of disease remain poorly understood. Primate pluripotent stem cell-derived oligodendrocytes are expected to help elucidate the molecular pathogenesis of these diseases. Oligodendrocytes have been successfully differentiated from human pluripotent stem cells. However, it is challenging to prepare large amounts of oligodendrocytes over a short amount of time because of manipulation difficulties under conventional primate pluripotent stem cell culture methods. We developed a proprietary dissociated monolayer and feeder-free culture system to handle pluripotent stem cell cultures. Because the dissociated monolayer and feeder-free culture system improves the quality and growth of primate pluripotent stem cells, these cells could potentially be differentiated into any desired functional cells and consistently cultured in large-scale conditions. In the current study, oligodendrocyte progenitor cells and mature oligodendrocytes were generated within three months from monkey embryonic stem cells. The embryonic stem cell-derived oligodendrocytes exhibited in vitro myelinogenic potency with rat dorsal root ganglion neurons. Additionally, the transplanted oligodendrocyte progenitor cells differentiated into myelin basic protein-positive mature oligodendrocytes in the mouse corpus callosum. This preparative method was used for human induced pluripotent stem cells, which were also successfully differentiated into oligodendrocyte progenitor cells and mature oligodendrocytes that were capable of myelinating rat dorsal root ganglion neurons. Moreover, it was possible to freeze, thaw, and successfully re-culture the differentiating cells. These results showed that embryonic stem cells and human induced pluripotent stem cells maintained in a

  9. Radiosensitivity of cultured insect cells: I. Lepidoptera

    International Nuclear Information System (INIS)

    Koval, T.M.

    1983-01-01

    The radiosensitivity of five lepidopteran insect cell lines representing five different genera has been investigated. These lines are: (1) TN-368, Trichoplusia ni; (2) IPLB-SF-1254, Spodoptera frugiperda; (3) IPLB-1075, Heliothis zea; (4) MRRL-CHl, clone GVl, Manduca sexta; and (5) IAL-PID2, Plodia interpunctella. The cell lines grew at different rates and had population doubling times that ranged from 19 to 52 hr. All of the lines are highly heteroploid and have approximate chromosome numbers near or above 100. The chromosomes are very small. All of the lines are extremely radioresistant; cell populations are able to recover from 260 kVp X-ray exposures up to and including 400 Gy, the highest dose examined. Cell survival curves were obtainable for only the TN-368 and IPLB-SF-1254 lines. The TN-368 cells displayed a biphasic survival response with D 0 , d/sub q/, and n values of 65.7 and 130.2 Gy, 9.0 and -36.1 Gy, and 1.2 and 0.8, respectively, for the steep and shallow portions of the curve. The IPLB-SF-1254 cells had a D 0 of 63.9 Gy. D/sub q/ of 19.0 Gy, and n value of 1.4. These studies provide definitive evidence of the radioresistance of lepidopteran cells, and suggest that this radioresistance is a characteristic of lepidopteran insects

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

    DEFF Research Database (Denmark)

    Hellsten, Ylva; Frandsen, Ulrik

    1997-01-01

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

  11. 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......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...

  12. Comparative characteristics of spermatozoa harvested and cryopreserved in culture and cryoprotectant media with or without donor serum proteins.

    Science.gov (United States)

    Ata'Allah, Ghofraan A; Adenan, Noor Azmi Mat; Razali, Nuguelis; Palaniappan, Kannappan; Saad, Rosliza; Idris, Siti Khadijah; Kanniah, Krishnan; Ali, Jaffar

    2017-06-01

    The objectives of this study is to evaluate the efficacy of protein-free media in the preparation, holding and crypreservation of spermatazoa for use in ART. Normozoospermic semen samples (N=71) were used to compare the effects of media on the survival and quality of spermatozoa when washed and cultured with different media with and without added proteins at 4°C, 15°C, 22°C and 37°C for 0, 4-7 and 24h. Survival and quality of spermatozoa were assessed after freeze-thaw with synthetic cryoprotectant with and without proteins. Ethics/IRB approval was obtained (Ref. 1073.52). Spermatozoa parameters were similar in all media after washing and culture for 24h. Post-thaw survival and quality of spermatozoa was not significantly different 24h after thawing of samples frozen in all cryoprotectant medium. In conclusion synthetic protein-free culture and cryoprotectant media are equal in efficacy to protein-containing media in culture and cryopreservation of spermatozoa . Use of these synthetic media are anticipated to significantly reduce the risk, potentially associated with conventional protein-containing media, of transmission of disease and possibly harmful undeclared proteins to the patient, baby and the healtcare worker. Synthetic media also ensure consistency of quality between batches of media. Copyright © 2017 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  13. Culture and Characterization of Circulating Endothelial Progenitor Cells in Patients with Renal Cell Carcinoma.

    Science.gov (United States)

    Gu, Wenyu; Sun, Wei; Guo, Changcheng; Yan, Yang; Liu, Min; Yao, Xudong; Yang, Bin; Zheng, Junhua

    2015-07-01

    Although emerging evidence demonstrates increased circulating endothelial progenitor cells in patients with solid tumors, to our knowledge it is still unknown whether such cells can be cultured from patients with highly angiogenic renal cell carcinoma. We cultured and characterized circulating endothelial progenitor cells from patients with renal cell carcinoma. The circulating endothelial progenitor cell level (percent of CD45(-)CD34(+) VEGF-R2(+) cells in total peripheral blood mononuclear cells) was quantified in 47 patients with renal cell carcinoma and 40 healthy controls. Peripheral blood mononuclear cells were then isolated from 33 patients with renal cell carcinoma and 30 healthy controls to culture and characterize circulating endothelial progenitor cells. The circulating endothelial progenitor cell level was significantly higher in patients with renal cell carcinoma than in healthy controls (0.276% vs 0.086%, p cells first emerged significantly earlier in patient than in control preparations (6.72 vs 14.67 days, p culture success rate (87.8% vs 40.0% of participants) and the number of colonies (10.06 vs 1.83) were significantly greater for patients than for controls (each p cell level correlated positively with the number of patient colonies (r = 0.762, p Cells cultured from patients and controls showed a similar growth pattern, immunophenotype, ability to uptake Ac-LDL and bind lectin, and form capillary tubes in vitro. However, significantly more VEGF-R2(+) circulating endothelial progenitor cells were found in preparations from patients with renal cell carcinoma than from healthy controls (21.1% vs 13.4%, p cell colonies, a higher cell culture success rate and more colonies were found for patients with renal cell carcinoma than for healthy controls. Results indicate the important significance of VEGF-R2(+) circulating endothelial progenitors in patients with renal cell carcinoma. Copyright © 2015 American Urological Association Education and Research

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

    International Nuclear Information System (INIS)

    Fox, I.H.; Bertorini, T.; Palmieri, G.M.A.; Shefner, R.

    1986-01-01

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

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

    DEFF Research Database (Denmark)

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

    2011-01-01

    in water instead of phosphate-buffered saline. Passively adsorbed IL-4 was observed to induce differentiation to dendritic cells, but analysis of cell culture supernatants revealed that leakage of IL-4 into solution could account for the differentiation observed. Covalent attachment resulted in bound IL-4...... at similar concentrations to the passive adsorption process, as measured by enzyme-linked immunosorbent assays, and the bound IL-4 did not leak into solution to any measurable extent during cell culture. However, covalently bound IL-4 was incapable of inducing monocyte differentiation. This may be caused...

  16. Cloning higher plants from aseptically cultured tissues and cells

    Science.gov (United States)

    Krikorian, A. D.

    1982-01-01

    A review of aseptic culture methods for higher plants is presented, which focuses on the existing problems that limit or prevent the full realization of cloning plants from free cells. It is shown that substantial progress in clonal multiplication has been made with explanted stem tips or lateral buds which can be stimulated to produce numerous precocious axillary branches. These branches can then be separated or subdivided and induced to root in order to yield populations of genetically and phenotypically uniorm plantlets. Similarly, undifferentiated calluses can sometimes be induced to form shoots and/or roots adventitiously. Although the cell culture techniques required to produce somatic embryos are presently rudimentary, steady advances are being made in learning how to stimulate formation of somatic or adventive embryos from totipotent cells grown in suspension cultures. It is concluded that many problems exist in the producing and growing of totipotent or morphogenetically competent cell suspensions, but the potential benefits are great.

  17. Miniature Bioreactor System for Long-Term Cell Culture

    Science.gov (United States)

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

    2010-01-01

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

  18. Upright nanopyramid structured cover glass with light harvesting and self-cleaning effects for solar cell applications

    International Nuclear Information System (INIS)

    Amalathas, Amalraj Peter; Alkaisi, Maan M

    2016-01-01

    In this paper, we demonstrate the effect of upright nanopyramid (UNP) structured cover glass with light harvesting and self-cleaning functions on the device performance of monocrystalline Si solar cells. The UNP structures were fabricated on the surface of the glass substrate by simple, high throughput and low cost UV nanoimprint lithography, using a Si master mold with inverted nanopyramid (INP) structures. The diffuse transmittance and haze ratio values were significantly increased for UNP patterned glass, especially in the wavelength range 300–600 nm compared to the bare glass; this implies that antireflection and strong light scattering are due to the UNP structures. By replacing a bare cover glass with UNP patterned glass, the power conversion efficiency of the monocrystalline Si solar cell was substantially enhanced by about 10.97%; this is mainly due to the increased short-circuit current density J SC of 32.39 mA cm −2 compared to the reference cell with bare cover glass (i.e. J SC   =  31.60 mA cm −2 ). In addition, unlike the bare cover glass (i.e. θ CA ∼ 36°), the fluorinated UNP structured cover glass exhibited a hydrophobic surface with a water contact angle ( θ CA ) of ∼132° and excellent self-cleaning of dust particles by rolling down water droplets. (paper)

  19. Carbon-Electrode-Tailored All-Inorganic Perovskite Solar Cells To Harvest Solar and Water-Vapor Energy.

    Science.gov (United States)

    Duan, Jialong; Hu, Tianyu; Zhao, Yuanyuan; He, Benlin; Tang, Qunwei

    2018-05-14

    Moisture is the worst enemy for state-of-the-art perovskite solar cells (PSCs). However, the flowing water vapor within nanoporous carbonaceous materials can create potentials. Therefore, it is a challenge to integrate water vapor and solar energies into a single PSC device. We demonstrate herein all-inorganic cesium lead bromide (CsPbBr 3 ) solar cells tailored with carbon electrodes to simultaneously harvest solar and water-vapor energy. Upon interfacial modification and plasma treatment, the bifunctional PSCs yield a maximum power conversion efficiency up to 9.43 % under one sun irradiation according to photoelectric conversion principle and a power output of 0.158 μW with voltage of 0.35 V and current of 0.45 μA in 80 % relative humidity through the flowing potentials at the carbon/water interface. The initial efficiency is only reduced by 2 % on exposing the inorganic PSC with 80 % humidity over 40 days. The successful realization of physical proof-of-concept multi-energy integrated solar cells provides new opportunities of maximizing overall power output. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. The Evolution of Polystyrene as a Cell Culture Material.

    Science.gov (United States)

    Lerman, Max J; Lembong, Josephine; Muramoto, Shin; Gillen, Greg; Fisher, John P

    2018-04-10

    Polystyrene (PS) has brought in vitro cell culture from its humble beginnings to the modern era, propelling dozens of research fields along the way. This review discusses the development of the material, fabrication, and treatment approaches to create the culture material. However, native PS surfaces poorly facilitate cell adhesion and growthin vitro. To overcome this, liquid surface deposition, energetic plasma activation, and emerging functionalization methods transform the surface chemistry. This review seeks to highlight the many potential applications of the first widely accepted polymer growth surface. Although the majority of in vitro research occurs on 2D surfaces, the importance of 3D culture models cannot be overlooked. Here the methods to transition PS to specialized 3D culture surfaces are also reviewed. Specifically, casting, electrospinning, 3D printing, and microcarrier approaches to shift PS to a 3D culture surface are highlighted. The breadth of applications of the material makes it impossible to highlight every use, but the aim remains to demonstrate the versatility and potential as both a general and custom cell culture surface. The review concludes with emerging scaffolding approaches and, based on the findings, presents our insights on the future steps for PS as a tissue culture platform.

  1. 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.

  2. A novel three-dimensional cell culture method enhances antiviral drug screening in primary human cells.

    Science.gov (United States)

    Koban, Robert; Neumann, Markus; Daugs, Aila; Bloch, Oliver; Nitsche, Andreas; Langhammer, Stefan; Ellerbrok, Heinz

    2018-02-01

    Gefitinib is a specific inhibitor of the epidermal growth factor receptor (EGFR) and FDA approved for treatment of non-small cell lung cancer. In a previous study we could show the in vitro efficacy of gefitinib for treatment of poxvirus infections in monolayer (2D) cultivated cell lines. Permanent cell lines and 2D cultures, however, are known to be rather unphysiological; therefore it is difficult to predict whether determined effective concentrations or the drug efficacy per se are transferable to the in vivo situation. 3D cell cultures, which meanwhile are widely distributed across all fields of research, are a promising tool for more predictive in vitro investigations of antiviral compounds. In this study the spreading of cowpox virus and the antiviral efficacy of gefitinib were analyzed in primary human keratinocytes (NHEK) grown in a novel 3D extracellular matrix-based cell culture model and compared to the respective monolayer culture. 3D-cultivated NHEK grew in a polarized and thus a more physiological manner with altered morphology and close cell-cell contact. Infected cultures showed a strongly elevated sensitivity towards gefitinib. EGFR phosphorylation, cell proliferation, and virus replication were significantly reduced in 3D cultures at gefitinib concentrations which were at least 100-fold lower than those in monolayer cultures and well below the level of cytotoxicity. Our newly established 3D cell culture model with primary human cells is an easy-to-handle alternative to conventional monolayer cell cultures and previously described more complex 3D cell culture systems. It can easily be adapted to other cell types and a broad spectrum of viruses for antiviral drug screening and many other aspects of virus research under more in vivo-like conditions. In consequence, it may contribute to a more targeted realization of necessary in vivo experiments. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Energy Harvesting From River Sediment Using a Microbial Fuel Cell: Preliminary Results

    Directory of Open Access Journals (Sweden)

    Philippe Namour

    2014-05-01

    Full Text Available We have built a sedimentary fuel cell or Sediment Microbial Fuel Cell (SMFC. The device works on the principle of microbial fuel cells by exploiting directly the energy contained in sedimentary organic matter. It converts in electricity the sediment potential, thanks to microorganisms able to waste electrons from their metabolism directly to a solid anode instead of their natural electron acceptors, such as oxygen or nitrate. The sediment microbial fuel cell was made of a non-corrodible anode (graphite buried in anoxic sediments layer and connected via an electrical circuit to a cathode installed in surface water. We present the first results of laboratory sedimentary fuel cell and a prototype installed in the river.

  4. Hypoxic contraction of cultured pulmonary vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Murray, T.R.; Chen, L.; Marshall, B.E.; Macarak, E.J.

    1990-01-01

    The cellular events involved in generating the hypoxic pulmonary vasoconstriction response are not clearly understood, in part because of the multitude of factors that alter pulmonary vascular tone. The goal of the present studies was to determine if a cell culture preparation containing vascular smooth muscle (VSM) cells could be made to contract when exposed to a hypoxic atmosphere. Cultures containing only fetal bovine pulmonary artery VSM cells were assessed for contractile responses to hypoxic stimuli by two methods. In the first, tension forces generated by cells grown on a flexible growth surface (polymerized polydimethyl siloxane) were manifested as wrinkles and distortions of the surface under the cells. Wrinkling of the surface was noted to progressively increase with time as the culture medium bathing the cells was made hypoxic (PO2 approximately 25 mmHg). The changes were sometimes reversible upon return to normoxic conditions and appeared to be enhanced in cells already exhibiting evidence of some baseline tone. Repeated passage in culture did not diminish the hypoxic response. Evidence for contractile responses to hypoxia was also obtained from measurements of myosin light chain (MLC) phosphorylation. Conversion of MLC to the phosphorylated species is an early step in the activation of smooth muscle contraction. Lowering the PO2 in the culture medium to 59 mmHg caused a 45% increase in the proportion of MLC in the phosphorylated form as determined by two-dimensional gel electrophoresis. Similarly, cultures preincubated for 4 h with 32P and then exposed to normoxia or hypoxia for a 5-min experimental period showed more than twice as much of the label in MLCs of the hypoxic cells

  5. Animal-cell culture media: History, characteristics, and current issues.

    Science.gov (United States)

    Yao, Tatsuma; Asayama, Yuta

    2017-04-01

    Cell culture technology has spread prolifically within a century, a variety of culture media has been designed. This review goes through the history, characteristics and current issues of animal-cell culture media. A literature search was performed on PubMed and Google Scholar between 1880 and May 2016 using appropriate keywords. At the dawn of cell culture technology, the major components of media were naturally derived products such as serum. The field then gradually shifted to the use of chemical-based synthetic media because naturally derived ingredients have their disadvantages such as large batch-to-batch variation. Today, industrially important cells can be cultured in synthetic media. Nevertheless, the combinations and concentrations of the components in these media remain to be optimized. In addition, serum-containing media are still in general use in the field of basic research. In the fields of assisted reproductive technologies and regenerative medicine, some of the medium components are naturally derived in nearly all instances. Further improvements of culture media are desirable, which will certainly contribute to a reduction in the experimental variation, enhance productivity among biopharmaceuticals, improve treatment outcomes of assisted reproductive technologies, and facilitate implementation and popularization of regenerative medicine.

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

    International Nuclear Information System (INIS)

    Meca C, E. A.; Bourel, V.; Notcovich, C.; Duran, H.

    2015-10-01

    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 (Al 2 O 3 :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 cm 3 . 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)

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

    Science.gov (United States)

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

    2012-08-01

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

  8. Isolation and Culture of Postnatal Stem Cells from Deciduous Teeth

    OpenAIRE

    Olávez, Daniela; Facultad de Odontología Universidad de Los Andes; Salmen, Siham; Instituto de Inmunología Clínica, Universidad de Los Andes.; Padrón, Karla; Facultad de Odontología. Univerisdad de Los Andes.; Lobo, Carmine; Facultad de Odontología. Univerisdad de Los Andes.; Díaz, Nancy; Facultad de Odontología, Universidad de Los Andes.; Berrueta, Lisbeth; Doctora en Inmunología por Instituto Venezolano de Investigaciones Científicas (IVIC). Instituto de Inmunología Clínica, Facultad de Medicina, Universidad de Los Andes, Venezuela.; Solorzanio, Eduvigis; Facultad de Odontología, Universidad de Los Andes.

    2014-01-01

    Background: Currently, degenerative diseases represent a public health problem; therefore, the development and implementation of strategies to fully or partially recover of damaged tissues has a special interest in the biomedical field. Therapeutic strategies based on mesenchymal stem cells transplantation from dental pulp have been proposed as an alternative. Purpose: To develop a mesenchymal stem cells culture isolated from dental pulp of deciduous teeth. Methods: The mesenchymal stem cells...

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

    International Nuclear Information System (INIS)

    Hoffmann, M.E.; Meneghini, R.

    1982-01-01

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

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

    DEFF Research Database (Denmark)

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

    2010-01-01

    Microfluidic biochips offer a promising alternative to a conventional biochemical laboratory. There are two technologies for the microfluidic biochips: droplet-based and flow-based. In this paper we are interested in flow-based microfluidic biochips, where the liquid flows continuously through pre......-defined micro-channels using valves and pumps. We present an approach to the system-level modeling and simulation of a cell culture microfluidic biochip called ProCell, Programmable Cell Culture Chip. ProCell contains a cell culture chamber, which is envisioned to run 256 simultaneous experiments (viewed...

  11. Induction of pluripotent stem cells from fibroblast cultures.

    Science.gov (United States)

    Takahashi, Kazutoshi; Okita, Keisuke; Nakagawa, Masato; Yamanaka, Shinya

    2007-01-01

    Clinical application of embryonic stem (ES) cells faces difficulties regarding use of embryos, as well as tissue rejection after implantation. One way to circumvent these issues is to generate pluripotent stem cells directly from somatic cells. Somatic cells can be reprogrammed to an embryonic-like state by the injection of a nucleus into an enucleated oocyte or by fusion with ES cells. However, little is known about the mechanisms underlying these processes. We have recently shown that the combination of four transcription factors can generate ES-like pluripotent stem cells directly from mouse fibroblast cultures. The cells, named induced pluripotent stem (iPS) cells, can be differentiated into three germ layers and committed to chimeric mice. Here we describe detailed methods and tips for the generation of iPS cells.

  12. Efforts to develop a cultured sponge cell line: revisiting an intractable problem.

    Science.gov (United States)

    Grasela, James J; Pomponi, Shirley A; Rinkevich, Buki; Grima, Jennifer

    2012-01-01

    Residents of the marine environment, sponges (Porifera) have the ability to produce organic compounds known as secondary metabolites, which are not directly involved in the normal growth, development, or reproduction of an organism. Because of their sessile nature, the production of these bioactive compounds has been interpreted as a functional adaptation to serve in an important survival role as a means to counter various environmental stress factors such as predation, overgrowth by fouling organisms, or competition for limited space. Regardless of the reasons for this adaptation, a variety of isolated compounds have already proven to demonstrate remarkable anticancer, fungicidal, and antibiotic properties. A major obstacle to the isolation and production of novel compounds from sponges is the lack of a large, reliable source of sponge material. Sponge collection from the sea would be environmentally detrimental to the already stressed and sparse sponge populations. Sponge production in an aquaculture setting might appear to be an ideal alternative but would also be cost-ineffective and sponge growth is extremely slow. A third approach involves the development of a sponge cell culture system capable of producing the necessary cell numbers to harvest for research purposes as well as for the eventual commercial-scale production of promising bioactive compounds. Unfortunately, little progress has been made in this direction other than the establishment of temporary cultures containing aggregates and fragments of cells. One impediment toward successful sponge cell culture might be ascribed to a lack of published knowledge of failed methodologies, and thus, time and effort is wasted on continued reinvention of the same methods and procedures. Consequently, we have undertaken here to chart some of our unsuccessful research efforts, our methodology, and results to provide the sponge research community with knowledge to assist them to better avoid taking the same failed

  13. Periodically arranged colloidal gold nanoparticles for enhanced light harvesting in organic solar cells

    DEFF Research Database (Denmark)

    Mirsafaei, Mina; Fernandes Cauduro, André Luis; Kunstmann-Olsen, Casper

    2016-01-01

    Although organic solar cells show intriguing features such as low-cost, mechanical flexibility and light weight, their efficiency is still low compared to their inorganic counterparts. One way of improving their efficiency is by the use of light-trapping mechanisms from nano- or microstructures......, which makes it possible to improve the light absorption and charge extraction in the device’s active layer. Here, periodically arranged colloidal gold nanoparticles are demonstrated experimentally and theoretically to improve light absorption and thus enhance the efficiency of organic solar cells....... Surface-ordered gold nanoparticle arrangements are integrated at the bottom electrode of organic solar cells. The resulting optical interference and absorption effects are numerically investigated in bulk hetero-junction solar cells based on the Finite-Difference Time-Domain (FDTD) and Transfer Matrix...

  14. Perfusion cell culture decreases process and product heterogeneity in a head-to-head comparison with fed-batch.

    Science.gov (United States)

    Walther, Jason; Lu, Jiuyi; Hollenbach, Myles; Yu, Marcella; Hwang, Chris; McLarty, Jean; Brower, Kevin

    2018-05-30

    In this study, we compared the impacts of fed-batch and perfusion platforms on process and product attributes for IgG1- and IgG4-producing cell lines. A "plug-and-play" approach was applied to both platforms at bench scale, using commercially available basal and feed media, a standard feed strategy for fed-batch, and ATF filtration for perfusion. Product concentration in fed-batch was 2.5 times greater than perfusion, while average productivity in perfusion was 7.5 times greater than fed-batch. PCA revealed more variability in the cell environment and metabolism during the fed-batch run. LDH measurements showed that exposure of product to cell lysate was 7-10 times greater in fed-batch. Product analysis shows larger abundances of neutral species in perfusion, likely due to decreased bioreactor residence times and extracellular exposure. The IgG1 perfusion product also had higher purity and lower half-antibody. Glycosylation was similar across both culture modes. The first perfusion harvest slice for both product types showed different glycosylation than subsequent harvests, suggesting that product quality lags behind metabolism. In conclusion, process and product data indicate that intra-lot heterogeneity is decreased in perfusion cultures. Additional data and discussion is required to understand the developmental, clinical and commercial implications, and in what situations increased uniformity would be beneficial. This article is protected by copyright. All rights reserved.

  15. Surface Acoustic Waves (SAW-Based Biosensing for Quantification of Cell Growth in 2D and 3D Cultures

    Directory of Open Access Journals (Sweden)

    Tao Wang

    2015-12-01

    Full Text Available Detection and quantification of cell viability and growth in two-dimensional (2D and three-dimensional (3D cell cultures commonly involve harvesting of cells and therefore requires a parallel set-up of several replicates for time-lapse or dose–response studies. Thus, developing a non-invasive and touch-free detection of cell growth in longitudinal studies of 3D tumor spheroid cultures or of stem cell regeneration remains a major unmet need. Since surface acoustic waves (SAWs permit mass loading-based biosensing and have been touted due to their many advantages including low cost, small size and ease of assembly, we examined the potential of SAW-biosensing to detect and quantify cell growth. Herein, we demonstrate that a shear horizontal-surface acoustic waves (SH-SAW device comprising two pairs of resonators consisting of interdigital transducers and reflecting fingers can be used to quantify mass loading by the cells in suspension as well as within a 3D cell culture platform. A 3D COMSOL model was built to simulate the mass loading response of increasing concentrations of cells in suspension in the polydimethylsiloxane (PDMS well in order to predict the characteristics and optimize the design of the SH-SAW biosensor. The simulated relative frequency shift from the two oscillatory circuit systems (one of which functions as control were found to be concordant to experimental data generated with RAW264.7 macrophage and A549 cancer cells. In addition, results showed that SAW measurements per se did not affect viability of cells. Further, SH-SAW biosensing was applied to A549 cells cultured on a 3D electrospun nanofiber scaffold that generate tumor spheroids (tumoroids and the results showed the device's ability to detect changes in tumor spheroid growth over the course of eight days. Taken together, these results demonstrate the use of SH-SAW device for detection and quantification of cell growth changes over time in 2D suspension cultures and in

  16. Surface Acoustic Waves (SAW)-Based Biosensing for Quantification of Cell Growth in 2D and 3D Cultures.

    Science.gov (United States)

    Wang, Tao; Green, Ryan; Nair, Rajesh Ramakrishnan; Howell, Mark; Mohapatra, Subhra; Guldiken, Rasim; Mohapatra, Shyam Sundar

    2015-12-19

    Detection and quantification of cell viability and growth in two-dimensional (2D) and three-dimensional (3D) cell cultures commonly involve harvesting of cells and therefore requires a parallel set-up of several replicates for time-lapse or dose-response studies. Thus, developing a non-invasive and touch-free detection of cell growth in longitudinal studies of 3D tumor spheroid cultures or of stem cell regeneration remains a major unmet need. Since surface acoustic waves (SAWs) permit mass loading-based biosensing and have been touted due to their many advantages including low cost, small size and ease of assembly, we examined the potential of SAW-biosensing to detect and quantify cell growth. Herein, we demonstrate that a shear horizontal-surface acoustic waves (SH-SAW) device comprising two pairs of resonators consisting of interdigital transducers and reflecting fingers can be used to quantify mass loading by the cells in suspension as well as within a 3D cell culture platform. A 3D COMSOL model was built to simulate the mass loading response of increasing concentrations of cells in suspension in the polydimethylsiloxane (PDMS) well in order to predict the characteristics and optimize the design of the SH-SAW biosensor. The simulated relative frequency shift from the two oscillatory circuit systems (one of which functions as control) were found to be concordant to experimental data generated with RAW264.7 macrophage and A549 cancer cells. In addition, results showed that SAW measurements per se did not affect viability of cells. Further, SH-SAW biosensing was applied to A549 cells cultured on a 3D electrospun nanofiber scaffold that generate tumor spheroids (tumoroids) and the results showed the device's ability to detect changes in tumor spheroid growth over the course of eight days. Taken together, these results demonstrate the use of SH-SAW device for detection and quantification of cell growth changes over time in 2D suspension cultures and in 3D cell

  17. 40 CFR 798.5300 - Detection of gene mutations in somatic cells in culture.

    Science.gov (United States)

    2010-07-01

    ... cells in culture. 798.5300 Section 798.5300 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY....5300 Detection of gene mutations in somatic cells in culture. (a) Purpose. Mammalian cell culture... selected by resistance to ouabain. (2) Description. Cells in suspension or monolayer culture are exposed to...

  18. Impact of starting material (fresh versus cryopreserved marrow) on mesenchymal stem cell culture.

    Science.gov (United States)

    Kaplan, Alesia; Sackett, Katie; Sumstad, Darin; Kadidlo, Dianne; McKenna, David H

    2017-09-01

    Mesenchymal stem cells (MSCs) continue to be investigated in multiple clinical trials as potential therapy for different disorders. There is ongoing controversy surrounding the clinical use of cryopreserved versus fresh MSCs. However, little is known about how cryopreservation affects marrow as starting material. The growth kinetics of MSC cultures derived from fresh versus cryopreserved marrow were compared. Data were reviewed on the growth kinetics of MSCs derived from fresh versus cryopreserved marrow of nine donors. Marrow harvested from each donor was separated into four aliquots (one fresh and three cryopreserved for culture). Data on the date of mononuclear cell cryopreservation/thaw, MSC counts at Passages 1 and 2, MSC doubling, MSC fold expansion, viability (of mononuclear cells and final MSCs), and on flow cytometry markers of mononuclear cells and final MSCs were analyzed for the fresh and cryopreserved marrow groups. In total, 21 MSC lots (seven fresh and 14 cryopreserved) were obtained. The average age of cryopreserved mononuclear cell product was 295 days (range, 18-1241 days). There were no significant differences between MSC numbers at Passage 1 (p = 0.1), final MSC numbers (p = 0.5), MSC doubling (p = 0.7), or MSC fold expansion (p = 0.7). A significant difference was observed in viability by flow cytometry for both mononuclear cells (p = 0.002) and final MSCs (p = 0.009), with higher viability in the fresh marrow group. This study demonstrates that MSCs derived from cryopreserved marrow have the same growth characteristics as fresh marrow-derived MSCs. Further studies are needed to explore potential differences in clinical efficacy. © 2017 AABB.

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

    Science.gov (United States)

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

    2016-01-01

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

  20. HUMAN CELLS IN CULTURE: REVISlTED*

    African Journals Online (AJOL)

    advantages, e.g. the generation time is reduced to about. 1/10000 that of the ... or less reflects the cellular biology of the donor tissut:'Y .... X-linked. Autosomal recessive. Autosomal recessive. Autosomal recessive mothers of affected males, however, show that only 50% of the cell population is defective, which furnishes an.

  1. Cultural relativism: maintenance of genomic imprints in pluripotent stem cell culture systems.

    Science.gov (United States)

    Greenberg, Maxim Vc; Bourc'his, Déborah

    2015-04-01

    Pluripotent stem cells (PSCs) in culture have become a widely used model for studying events occurring during mammalian development; they also present an exciting avenue for therapeutics. However, compared to their in vivo counterparts, cultured PSC derivatives have unique properties, and it is well established that their epigenome is sensitive to medium composition. Here we review the specific effects on genomic imprints in various PSC types and culture systems. Imprinted gene regulation is developmentally important, and imprinting defects have been associated with several human diseases. Therefore, imprint abnormalities in PSCs may have considerable consequences for downstream applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

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

    Science.gov (United States)

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

    1988-01-01

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

  3. Establishment of Cancer Stem Cell Cultures from Human Conventional Osteosarcoma.

    Science.gov (United States)

    Palmini, Gaia; Zonefrati, Roberto; Mavilia, Carmelo; Aldinucci, Alessandra; Luzi, Ettore; Marini, Francesca; Franchi, Alessandro; Capanna, Rodolfo; Tanini, Annalisa; Brandi, Maria Luisa

    2016-10-14

    The current improvements in therapy against osteosarcoma (OS) have prolonged the lives of cancer patients, but the survival rate of five years remains poor when metastasis has occurred. The Cancer Stem Cell (CSC) theory holds that there is a subset of tumor cells within the tumor that have stem-like characteristics, including the capacity to maintain the tumor and to resist multidrug chemotherapy. Therefore, a better understanding of OS biology and pathogenesis is needed in order to advance the development of targeted therapies to eradicate this particular subset and to reduce morbidity and mortality among patients. Isolating CSCs, establishing cell cultures of CSCs, and studying their biology are important steps to improving our understanding of OS biology and pathogenesis. The establishment of human-derived OS-CSCs from biopsies of OS has been made possible using several methods, including the capacity to create 3-dimensional stem cell cultures under nonadherent conditions. Under these conditions, CSCs are able to create spherical floating colonies formed by daughter stem cells; these colonies are termed "cellular spheres". Here, we describe a method to establish CSC cultures from primary cell cultures of conventional OS obtained from OS biopsies. We clearly describe the several passages required to isolate and characterize CSCs.

  4. Cytotoxicity of extracts of spices to cultured cells.

    Science.gov (United States)

    Unnikrishnan, M C; Kuttan, R

    1988-01-01

    The cytotoxicity of the extracts from eight different spices used in the Indian diet was determined using Dalton's lymphoma ascites tumor cells and human lymphocytes in vitro and Chinese Hamster Ovary cells and Vero cells in tissue culture. Alcoholic extracts of the spices were found to be more cytotoxic to these cells than their aqueous extracts. Alcoholic extracts of several spices inhibited cell growth at concentrations of 0.2-1 mg/ml in vitro and 0.12-0.3 mg/ml in tissue culture. Ginger, pippali (native to India; also called dried catkins), pepper, and garlic showed the highest activity followed by asafetida, mustard, and horse-gram (native to India). These extracts also inhibited the thymidine uptake into DNA.

  5. Isolation of Lysosomes from Mammalian Tissues and Cultured Cells.

    Science.gov (United States)

    Aguado, Carmen; Pérez-Jiménez, Eva; Lahuerta, Marcos; Knecht, Erwin

    2016-01-01

    Lysosomes participate within the cells in the degradation of organelles, macromolecules, and a wide variety of substrates. In any study on specific roles of lysosomes, both under physiological and pathological conditions, it is advisable to include methods that allow their reproducible and reliable isolation. However, purification of lysosomes is a difficult task, particularly in the case of cultured cells. This is mainly because of the heterogeneity of these organelles, along with their low number and high fragility. Also, isolation methods, while disrupting plasma membranes, have to preserve the integrity of lysosomes, as the breakdown of their membranes releases enzymes that could damage all cell organelles, including themselves. The protocols described below have been routinely used in our laboratory for the specific isolation of lysosomes from rat liver, NIH/3T3, and other cultured cells, but can be adapted to other mammalian tissues or cell lines.

  6. Optimization of human corneal endothelial cell culture: density dependency of successful cultures in vitro.

    Science.gov (United States)

    Peh, Gary S L; Toh, Kah-Peng; Ang, Heng-Pei; Seah, Xin-Yi; George, Benjamin L; Mehta, Jodhbir S

    2013-05-03

    Global shortage of donor corneas greatly restricts the numbers of corneal transplantations performed yearly. Limited ex vivo expansion of primary human corneal endothelial cells is possible, and a considerable clinical interest exists for development of tissue-engineered constructs using cultivated corneal endothelial cells. The objective of this study was to investigate the density-dependent growth of human corneal endothelial cells isolated from paired donor corneas and to elucidate an optimal seeding density for their extended expansion in vitro whilst maintaining their unique cellular morphology. Established primary human corneal endothelial cells were propagated to the second passage (P2) before they were utilized for this study. Confluent P2 cells were dissociated and seeded at four seeding densities: 2,500 cells per cm2 ('LOW'); 5,000 cells per cm2 ('MID'); 10,000 cells per cm2 ('HIGH'); and 20,000 cells per cm2 ('HIGH(×2)'), and subsequently analyzed for their propensity to proliferate. They were also subjected to morphometric analyses comparing cell sizes, coefficient of variance, as well as cell circularity when each culture became confluent. At the two lower densities, proliferation rates were higher than cells seeded at higher densities, though not statistically significant. However, corneal endothelial cells seeded at lower densities were significantly larger in size, heterogeneous in shape and less circular (fibroblastic-like), and remained hypertrophic after one month in culture. Comparatively, cells seeded at higher densities were significantly homogeneous, compact and circular at confluence. Potentially, at an optimal seeding density of 10,000 cells per cm2, it is possible to obtain between 10 million to 25 million cells at the third passage. More importantly, these expanded human corneal endothelial cells retained their unique cellular morphology. Our results demonstrated a density dependency in the culture of primary human corneal endothelial

  7. Lingual Epithelial Stem Cells and Organoid Culture of Them

    Directory of Open Access Journals (Sweden)

    Hiroko Hisha

    2016-01-01

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

  8. Bridging the gap between cell culture and live tissue

    Directory of Open Access Journals (Sweden)

    Stefan Przyborski

    2017-11-01

    Full Text Available Traditional in vitro two-dimensional (2-D culture systems only partly imitate the physiological and biochemical features of cells in their original tissue. In vivo, in organs and tissues, cells are surrounded by a three-dimensional (3-D organization of supporting matrix and neighbouring cells, and a gradient of chemical and mechanical signals. Furthermore, the presence of blood flow and mechanical movement provides a dynamic environment (Jong et al., 2011. In contrast, traditional in vitro culture, carried out on 2-D plastic or glass substrates, typically provides a static environment, which, however is the base of the present understanding of many biological processes, tissue homeostasis as well as disease. It is clear that this is not an exact representation of what is happening in vivo and the microenvironment provided by in vitro cell culture models are significantly different and can cause deviations in cell response and behaviour from those distinctive of in vivo tissues. In order to translate the present basic knowledge in cell control, cell repair and regeneration from the laboratory bench to the clinical application, we need a better understanding of the cell and tissue interactions. This implies a detailed comprehension of the natural tissue environment, with its organization and local signals, in order to more closely mimic what happens in vivo, developing more physiological models for efficient in vitro systems. In particular, it is imperative to understand the role of the environmental cues which can be mainly divided into those of a chemical and mechanical nature.

  9. 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...

  10. Power Harvesting from Human Serum in Buckypaper-Based Enzymatic Biofuel Cell

    Energy Technology Data Exchange (ETDEWEB)

    Güven, Güray, E-mail: gguven@ginerinc.com [Giner, Inc., Newton, MA (United States); Şahin, Samet [Pennsylvania State University College of Medicine, Hershey, PA (United States); Güven, Arcan [Chemical Engineering and Advanced Materials, Merz Court, Newcastle University, Newcastle upon Tyne (United Kingdom); Yu, Eileen H., E-mail: gguven@ginerinc.com [Pennsylvania State University College of Medicine, Hershey, PA (United States)

    2016-02-16

    The requirement for a miniature, high density, long life, and rechargeable power source is common to a vast majority of microsystems, including the implantable devices for medical applications. A model biofuel cell system operating in human serum has been studied for future applications of biomedical and implantable medical devices. Anodic and cathodic electrodes were made of carbon nanotube-buckypaper modified with PQQ-dependent glucose dehydrogenase and laccase, respectively. Modified electrodes were characterized electrochemically and assembled in a biofuel cell setup. Power density of 16.12 μW cm{sup −2} was achieved in human serum for lower than physiological glucose concentrations. Increasing the glucose concentration and biofuel cell temperature caused an increase in power output leading up to 49.16 μW cm{sup −2}.

  11. Increased light harvesting in dye-sensitized solar cells with energy relay dyes

    KAUST Repository

    Hardin, Brian E.

    2009-06-21

    Conventional dye-sensitized solar cells have excellent charge collection efficiencies, high open-circuit voltages and good fill factors. However, dye-sensitized solar cells do not completely absorb all of the photons from the visible and near-infrared domain and consequently have lower short-circuit photocurrent densities than inorganic photovoltaic devices. Here, we present a new design where high-energy photons are absorbed by highly photoluminescent chromophores unattached to the titania and undergo Förster resonant energy transfer to the sensitizing dye. This novel architecture allows for broader spectral absorption, an increase in dye loading, and relaxes the design requirements for the sensitizing dye. We demonstrate a 26% increase in power conversion efficiency when using an energy relay dye (PTCDI) with an organic sensitizing dye (TT1). We estimate the average excitation transfer efficiency in this system to be at least 47%. This system offers a viable pathway to develop more efficient dye-sensitized solar cells.

  12. A single-cell and feeder-free culture system for monkey embryonic stem cells.

    Science.gov (United States)

    Ono, Takashi; Suzuki, Yutaka; Kato, Yosuke; Fujita, Risako; Araki, Toshihiro; Yamashita, Tomoko; Kato, Hidemasa; Torii, Ryuzo; Sato, Naoya

    2014-01-01

    Primate pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), hold great potential for research and application in regenerative medicine and drug discovery. To maximize primate PSC potential, a practical system is required for generating desired functional cells and reproducible differentiation techniques. Much progress regarding their culture systems has been reported to date; however, better methods would still be required for their practical use, particularly in industrial and clinical fields. Here we report a new single-cell and feeder-free culture system for primate PSCs, the key feature of which is an originally formulated serum-free medium containing FGF and activin. In this culture system, cynomolgus monkey ESCs can be passaged many times by single-cell dissociation with traditional trypsin treatment and can be propagated with a high proliferation rate as a monolayer without any feeder cells; further, typical PSC properties and genomic stability can be retained. In addition, it has been demonstrated that monkey ESCs maintained in the culture system can be used for various experiments such as in vitro differentiation and gene manipulation. Thus, compared with the conventional culture system, monkey ESCs grown in the aforementioned culture system can serve as a cell source with the following practical advantages: simple, stable, and easy cell maintenance; gene manipulation; cryopreservation; and desired differentiation. We propose that this culture system can serve as a reliable platform to prepare primate PSCs useful for future research and application.

  13. Aragonite precipitation by "proto-polyps" in coral cell cultures.

    Directory of Open Access Journals (Sweden)

    Tali Mass

    Full Text Available The mechanisms of coral calcification at the molecular, cellular and tissue levels are poorly understood. In this study, we examine calcium carbonate precipitation using novel coral tissue cultures that aggregate to form "proto-polyps". Our goal is to establish an experimental system in which calcification is facilitated at the cellular level, while simultaneously allowing in vitro manipulations of the calcifying fluid. This novel coral culturing technique enables us to study the mechanisms of biomineralization and their implications for geochemical proxies. Viable cell cultures of the hermatypic, zooxanthellate coral, Stylophora pistillata, have been maintained for 6 to 8 weeks. Using an enriched seawater medium with aragonite saturation state similar to open ocean surface waters (Ω(arag~4, the primary cell cultures assemble into "proto-polyps" which form an extracellular organic matrix (ECM and precipitate aragonite crystals. These extracellular aragonite crystals, about 10 µm in length, are formed on the external face of the proto-polyps and are identified by their distinctive elongated crystallography and X-ray diffraction pattern. The precipitation of aragonite is independent of photosynthesis by the zooxanthellae, and does not occur in control experiments lacking coral cells or when the coral cells are poisoned with sodium azide. Our results demonstrate that proto-polyps, aggregated from primary coral tissue culture, function (from a biomineralization perspective similarly to whole corals. This approach provides a novel tool for investigating the biophysical mechanism of calcification in these organisms.

  14. Feasibility and Efficiency of Human Bone Marrow Stromal Cell Culture with Allogeneic Platelet Lysate-Supplementation for Cell Therapy against Stroke

    Directory of Open Access Journals (Sweden)

    Chengbo Tan

    2016-01-01

    Full Text Available Currently, there is increasing interest in human bone marrow stromal cells (hBMSCs as regeneration therapy against cerebral stroke. The aim of the present study was to evaluate the feasibility and validity of hBMSC cultures with allogeneic platelet lysates (PLs. Platelet concentrates (PC were harvested from healthy volunteers and made into single donor-derived PL (sPL. The PL mixtures (mPL were made from three different sPL. Some growth factors and platelet cell surface antigens were detected by enzyme-linked immunosorbent assay (ELISA. The hBMSCs cultured with 10% PL were analyzed for their proliferative potential, surface markers, and karyotypes. The cells were incubated with superparamagnetic iron oxide (SPIO agents and injected into a pig brain. MRI and histological analysis were performed. Consequently, nine lots of sPL and three mPL were prepared. ELISA analysis showed that PL contained adequate growth factors and a particle of platelet surface antigens. Cell proliferation capacity of PLs was equivalent to or higher than that of fetal calf serum (FCS. No contradiction in cell surface markers and no chromosomal aberrations were found. The MRI detected the distribution of SPIO-labeled hBMSCs in the pig brain. In summary, the hBMSCs cultured with allogeneic PL are suitable for cell therapy against stroke.

  15. Monitoring Tumour Cell Purge by Long Term Marrow Culture in Acute Leukemia

    International Nuclear Information System (INIS)

    El-Masry, M.; Hashem, T. M.

    2001-01-01

    Purging of leukemic cells from bone marrow harvested for autologous bone marrow transplantation (ABMT) remains a challenge. This work aimed at evaluating the efficacy of long-term marrow culture (LTMC) on purging leukemic progenitors in acute leukemia. Design and methods: We planned to study the presence of immunoglobulin heavy (lgH) chain gene rearrangements by polymerase chain reaction (PCR) at diagnosis for bone marrow of 23 patients with acute leukemia. LTMC was performed only for patients who showed positive IgH chain gene monoclonality at diagnosis. The efficiency of purge was evaluated by PCR for monoclonal IgH chain gene on weekly basis of LTMC. Results: Of the 23 studied cases, 18 (78.26%) showed positive clonal IgH chain gene at diagnosis. LTMC study showed that 6/]8 (33.33%), 3/18 (16.67%),7/18 (38.89%) and 2/18 (11.11 %) underwent complete purging of the leukemic progenitors at the first, second, third and fourth weeks of culture, respectively. Follow up could be performed for 14 positive ALL cases after induction of remission; 12/14 (85.7%) showed minimal residual disease (MRD) while only two cases did not show MRD. Complete purging of the latter two cases by LTMC occurred on the second and third weeks of culture. Conclusion: LTMC is a useful and successful method for leukemic cell purging. LTMC should be undertaken at initial diagnosis and on an individual basis. Each case should be dealt with solely to determine at which week of culture complete purging could be obtained for subsequent autologous grafting of the purged marrow

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

    International Nuclear Information System (INIS)

    Fan, Ping; He, Lan; Pu, Dan; Lv, Xiaohong; Zhou, Wenxu; Sun, Yining; Hu, Nan

    2011-01-01

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

  17. Identification of differences in gene expression in primary cell cultures of human endometrial epithelial cells and trophoblast cells following their interaction

    DEFF Research Database (Denmark)

    Høgh, Mette; Islin, Henrik; Møller, Charlotte

    2006-01-01

    The interaction between the cell types was simulated in vitro by growing primary cell cultures of human endometrial epithelial cells and trophoblast cells together (co-culture) and separately (control cultures). Gene expression in the cell cultures was compared using the Differential Display method and confirmed...

  18. Modelling response of glycolysis in S. cerevisiae cells harvested at diauxic shift.

    NARCIS (Netherlands)

    Albers, E.; Bakker, B.M.; Gustafsson, L.

    2002-01-01

    The response of glycolysis to exposure of glucose in non-growing S. cerevisiae cells from diauxic shift was monitored. The result was compared to a kinetic model of glycolysis with branches to glycogen, trehalose, glycerol, and succinate. Experimental data at steady-state concentrations of

  19. High-Throughput Cancer Cell Sphere Formation for 3D Cell Culture.

    Science.gov (United States)

    Chen, Yu-Chih; Yoon, Euisik

    2017-01-01

    Three-dimensional (3D) cell culture is critical in studying cancer pathology and drug response. Though 3D cancer sphere culture can be performed in low-adherent dishes or well plates, the unregulated cell aggregation may skew the results. On contrary, microfluidic 3D culture can allow precise control of cell microenvironments, and provide higher throughput by orders of magnitude. In this chapter, we will look into engineering innovations in a microfluidic platform for high-throughput cancer cell sphere formation and review the implementation methods in detail.

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

    Directory of Open Access Journals (Sweden)

    Sergei V. Jargin

    2012-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-08-15

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

  2. Transfection in perfused microfluidic cell culture devices: A case study.

    Science.gov (United States)

    Raimes, William; Rubi, Mathieu; Super, Alexandre; Marques, Marco P C; Veraitch, Farlan; Szita, Nicolas

    2017-08-01

    Automated microfluidic devices are a promising route towards a point-of-care autologous cell therapy. The initial steps of induced pluripotent stem cell (iPSC) derivation involve transfection and long term cell culture. Integration of these steps would help reduce the cost and footprint of micro-scale devices with applications in cell reprogramming or gene correction. Current examples of transfection integration focus on maximising efficiency rather than viable long-term culture. Here we look for whole process compatibility by integrating automated transfection with a perfused microfluidic device designed for homogeneous culture conditions. The injection process was characterised using fluorescein to establish a LabVIEW-based routine for user-defined automation. Proof-of-concept is demonstrated by chemically transfecting a GFP plasmid into mouse embryonic stem cells (mESCs). Cells transfected in the device showed an improvement in efficiency (34%, n = 3) compared with standard protocols (17.2%, n = 3). This represents a first step towards microfluidic processing systems for cell reprogramming or gene therapy.

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

    International Nuclear Information System (INIS)

    Zhao, Fei; Severson, Paul; Pacheco, Samantha; Futscher, Bernard W.; Klimecki, Walter T.

    2013-01-01

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

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

    DEFF Research Database (Denmark)

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

    The brain is the center of the nervous system, where serious neurodegenerative diseases such as Parkinson’s, Alzheimer’s and Huntington’s are products of functional loss in the neural cells (1). Typical techniques used to investigate these diseases lack precise control of the cellular surroundings......, in addition to isolating the neural tissue from nutrient delivery and to creating unwanted gradients (2). This means that typical techniques used to investigate neurodegenerative diseases cannot mimic in vivo conditions, as closely as desired. We have developed a novel microfluidic system for culturing PC12...... cells, neuronal cells, astrocytes cultures and brain slices. The microfluidic system provides efficient nutrient delivery, waste removal, access to oxygen, fine control over the neurochemical environment and access to modern microscopy. Additionally, the setup consists of an in vitro culturing...

  5. Production of betalaines by Myrtillocactus cell cultures. Passage from heterotrophic state to autotrophic state with Asparagus cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Bulard, C; Mary, J; Chaumont, D; Gudin, C

    1982-11-01

    Myrtillocactus tissue cultures are grown from the epicotyl of young plantlets. With an appropriate growing medium it is possible, after transfer of fragments of these cultures to a liquid environment, to obtain dissociation and proliferation of cells. The production of betalaic pigments is induced in solid surroundings by adjustement of the growing medium composition and can be maintained in a liquid environment. The multiplication of pigmented cells in suspension may thus be obtained. The conversion of Asparagus cell suspensions from the heterotrophic state (use of lactose as source of carbon) to the autotrophic state (carbon supplied by CO/sub 2/) is obtained by a gradual reduction in the sugar concentration of the medium combined with a rise in the CO/sub 2/ content of the gas mixture atmosphere injected into the cultivator. The passage to the autotrophic state of a Myrtillocactus suspension would enable the production conditions of a metabolite (Betalaine) to be studied by micro-algae culture techniques.

  6. Resonant cavity enhanced light harvesting in flexible thin-film organic solar cells

    KAUST Repository

    Sergeant, Nicholas P.

    2013-04-24

    Dielectric/metal/dielectric (DMD) electrodes have the potential to significantly increase the absorption efficiency and photocurrent in flexible organic solar cells. We demonstrate that this enhancement is attributed to a broadband cavity resonance. Silver-based semitransparent DMD electrodes with sheet resistances below 10 ohm/sq. are fabricated on flexible polyethylene terephthalate (PET) substrates in a high-throughput roll-to-roll sputtering tool. We carefully study the effect of the semitransparent DMD electrode (here composed of ZnxSnyOz/Ag/InxSn yOz) on the optical device performance of a copper phthalocyanine (CuPc)/fullerene (C60) bilayer cell and illustrate that a resonant cavity enhanced light trapping effect dominates the optical behavior of the device. © 2013 Optical Society of America.

  7. Temperature Field Analysis for PZT Pyroelectric Cells for Thermal Energy Harvesting

    Directory of Open Access Journals (Sweden)

    Chi-Yuan Lee

    2011-11-01

    Full Text Available This paper proposes the idea of etching PZT to improve the temperature variation rate of a thicker PZT sheet in order to enhance the energy conversion efficiency when used as pyroelectric cells. A partially covered electrode was proven to display a higher output response than a fully covered electrode did. A mesh top electrode monitored the temperature variation rate and the electrode area. The mesh electrode width affected the distribution of the temperature variation rate in a thinner pyroelectric material. However, a pyroelectric cell with a thicker pyroelectric material was beneficial in generating electricity pyroelectrically. The PZT sheet was further etched to produce deeper cavities and a smaller electrode width to induce lateral temperature gradients on the sidewalls of cavities under homogeneous heat irradiation, enhancing the temperature variation rate.

  8. Temperature field analysis for PZT pyroelectric cells for thermal energy harvesting.

    Science.gov (United States)

    Hsiao, Chun-Ching; Ciou, Jing-Chih; Siao, An-Shen; Lee, Chi-Yuan

    2011-01-01

    This paper proposes the idea of etching PZT to improve the temperature variation rate of a thicker PZT sheet in order to enhance the energy conversion efficiency when used as pyroelectric cells. A partially covered electrode was proven to display a higher output response than a fully covered electrode did. A mesh top electrode monitored the temperature variation rate and the electrode area. The mesh electrode width affected the distribution of the temperature variation rate in a thinner pyroelectric material. However, a pyroelectric cell with a thicker pyroelectric material was beneficial in generating electricity pyroelectrically. The PZT sheet was further etched to produce deeper cavities and a smaller electrode width to induce lateral temperature gradients on the sidewalls of cavities under homogeneous heat irradiation, enhancing the temperature variation rate.

  9. Solar harvesting by a heterostructured cell with built-in variable width quantum wells

    Science.gov (United States)

    Brooks, W.; Wang, H.; Mil'shtein, S.

    2018-02-01

    We propose cascaded heterostructured p-i-n solar cells, where inside of the i-region is a set of Quantum Wells (QWs) with variable thicknesses to enhance absorption of different photonic energies and provide quick relaxation for high energy carriers. Our p-i-n heterostructure carries top p-type and bottom n-type 11.3 Å thick AlAs layers, which are doped by acceptors and donor densities up to 1019/cm3. The intrinsic region is divided into 10 segments where each segment carries ten QWs of the same width and the width of the QWs in each subsequent segment gradually increases. The top segment consists of 10 QWs with widths of 56.5Å, followed by a segment with 10 wider QWs with widths of 84.75Å, followed by increasing QW widths until the last segment has 10 QWs with widths of 565Å, bringing the total number of QWs to 100. The QW wall height is controlled by alternating AlAs and GaAs layers, where the AlAs layers are all 11.3Å thick, throughout the entire intrinsic region. Configuration of variable width QWs prescribes sets of energy levels which are suitable for absorption of a wide range of photon energies and will dissipate high electron-hole energies rapidly, reducing the heat load on the solar cell. We expect that the heating of the solar cell will be reduced by 8-11%, enhancing efficiency. The efficiency of the designed solar cell is 43.71%, the Fill Factor is 0.86, the density of short circuit current (ISC) will not exceed 338 A/m2 and the open circuit voltage (VOC) is 1.51V.

  10. Development of biologically modified anodes for energy harvesting using microbial fuel cells

    Science.gov (United States)

    Sumner, James J.; Ganguli, Rahul; Chmelka, Brad

    2012-06-01

    Biological fuel cells hold promise as an alternative energy source to batteries for unattended ground sensor applications due to the fact that they can be extremely long lived. This lifetime can be extended over batteries by scavenging fuel from the deployed environment. Microbial fuel cells (MFC) are one class of such sources that produce usable energy from small organic compounds (i.e. sugars, alcohols, organic acids, and biopolymers) which can be easily containerized or scavenged from the environment. The use of microorganisms as the anodic catalysts is what makes these systems unique from other biofuel cell designs. One of the main drawbacks of engineering a sensor system powered by an MFC is that power densities and current flux are extremely low in currently reported systems. The power density is limited by the mass transfer of the fuel source to the catalyst, the metabolism of the microbial catalysts and the electron transfer from the organism to the anode. This presentation will focus on the development of a new style of microbially-modified anodes which will increase power density to a level where a practical power source can be engineered. This is being achieved by developing a three dimensional matrix as an artificial, conductive biofilm. These artificial biofilms will allow the capture of a consortium of microbes designed for efficient metabolism of the available fuel source. Also it will keep the microbes close to the electrode allowing ready access by fuel and providing a low resistance passage of the liberated electrons from fuel oxidation.

  11. Biocompatibility of Tygon® tubing in microfluidic cell culture.

    Science.gov (United States)

    Jiang, Xiao; Jeffries, Rex E; Acosta, Miguel A; Tikunov, Andrey P; Macdonald, Jeffrey M; Walker, Glenn M; Gamcsik, Michael P

    2015-02-01

    Growth of the MDA-MB-231 breast cancer cell line in microfluidic channels was inhibited when culture media was delivered to the channels via microbore Tygon® tubing. Culture media incubated within this tubing also inhibited growth of these cells in conventional 96-well plates. These detrimental effects were not due to depletion of critical nutrients due to adsorption of media components onto the tubing surface. A pH change was also ruled out as a cause. Nuclear magnetic resonance spectroscopy of the cell growth media before and after incubation in the tubing confirmed no detectable loss of media components but did detect the presence of additional unidentified signals in the aliphatic region of the spectrum. These results indicate leaching of a chemical species from microbore Tygon® tubing that can affect cell growth in microfluidic devices.

  12. Efficient dye-sensitized solar cells from curved silicate microsheet caged TiO2 photoanodes. An avenue of enhancing light harvesting

    International Nuclear Information System (INIS)

    Wang, Zubin; Tang, Qunwei; He, Benlin; Chen, Haiyan; Yu, Liangmin

    2015-01-01

    Graphical abstract: - Highlights: • Curved silicate microsheets are incorporated with TiO 2 for light harvesting in DSSC • The optical matching between silicate and TiO 2 is superior to light reflection. • The curved silicate can hinder the recombination reaction of electrons with I 3 − . • The DSSC with TiO 2 /curved silicate photoanode shows an efficiency of 9.22% - Abstract: Enhancement of light harvesting has been a persistent objective for elevating dye excitation and therefore power conversion efficiency of dye-sensitized solar cells (DSSCs). Here we launch a strategy of markedly enhancing light harvesting by caging TiO 2 nanoparticles with curved silica microsheets. The results show that the strategy is versatile in suppressing the recombination reaction of electrons with I 3 − species in liquid electrolyte. Due to the superior reflective behaviors of curved silica microsheets, an optimal efficiency of 9.22% is recorded under simulated air mass 1.5 global sunlight on the DSSC in comparison with 6.51% and 7.51% from pristine TiO 2 and planar silicate microsheet incorporated TiO 2 photoanode based solar cells, respectively. This strategy is also believed to be applicable to other solar cells such as perovskite solar cells and quantum dot-sensitized solar cells.

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

    Directory of Open Access Journals (Sweden)

    Hanley Edward N

    2000-10-01

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

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

    International Nuclear Information System (INIS)

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

  15. T cell resistance to activation by dendritic cells requires long-term culture in simulated microgravity

    Science.gov (United States)

    Bradley, Jillian H.; Stein, Rachel; Randolph, Brad; Molina, Emily; Arnold, Jennifer P.; Gregg, Randal K.

    2017-11-01

    Immune impairment mediated by microgravity threatens the success of space exploration requiring long-duration spaceflight. The cells of most concern, T lymphocytes, coordinate the host response against microbial and cancerous challenges leading to elimination and long-term protection. T cells are activated upon recognition of specific microbial peptides bound on the surface of antigen presenting cells, such as dendritic cells (DC). Subsequently, this engagement results in T cell proliferation and differentiation into effector T cells driven by autocrine interleukin-2 (IL-2) and other cytokines. Finally, the effector T cells acquire the weaponry needed to destroy microbial invaders and tumors. Studies conducted on T cells during spaceflight, or using Earth-based culture systems, have shown reduced production of cytokines, proliferation and effector functions as compared to controls. This may account for the cases of viral reactivation events and opportunistic infections associated with astronauts of numerous missions. This work has largely been based upon the outcome of T cell activation by stimulatory factors that target select T cell signaling pathways rather than the complex, signaling events related to the natural process of antigen presentation by DC. This study tested the response of an ovalbumin peptide-specific T cell line, OT-II TCH, to activation by DC when the T cells were cultured 24-120 h in a simulated microgravity (SMG) environment generated by a rotary cell culture system. Following 72 h culture of T cells in SMG (SMG-T) or control static (Static-T) conditions, IL-2 production by the T cells was reduced in SMG-T cells compared to Static-T cells upon stimulation by phorbol 12-myristate 13-acetate (PMA) and ionomycin. However, when the SMG-T cells were stimulated with DC and peptide, IL-2 was significantly increased compared to Static-T cells. Such enhanced IL-2 production by SMG-T cells peaked at 72 h SMG culture time and decreased thereafter. When

  16. T cell resistance to activation by dendritic cells requires long-term culture in simulated microgravity.

    Science.gov (United States)

    Bradley, Jillian H; Stein, Rachel; Randolph, Brad; Molina, Emily; Arnold, Jennifer P; Gregg, Randal K

    2017-11-01

    Immune impairment mediated by microgravity threatens the success of space exploration requiring long-duration spaceflight. The cells of most concern, T lymphocytes, coordinate the host response against microbial and cancerous challenges leading to elimination and long-term protection. T cells are activated upon recognition of specific microbial peptides bound on the surface of antigen presenting cells, such as dendritic cells (DC). Subsequently, this engagement results in T cell proliferation and differentiation into effector T cells driven by autocrine interleukin-2 (IL-2) and other cytokines. Finally, the effector T cells acquire the weaponry needed to destroy microbial invaders and tumors. Studies conducted on T cells during spaceflight, or using Earth-based culture systems, have shown reduced production of cytokines, proliferation and effector functions as compared to controls. This may account for the cases of viral reactivation events and opportunistic infections associated with astronauts of numerous missions. This work has largely been based upon the outcome of T cell activation by stimulatory factors that target select T cell signaling pathways rather than the complex, signaling events related to the natural process of antigen presentation by DC. This study tested the response of an ovalbumin peptide-specific T cell line, OT-II TCH, to activation by DC when the T cells were cultured 24-120 h in a simulated microgravity (SMG) environment generated by a rotary cell culture system. Following 72 h culture of T cells in SMG (SMG-T) or control static (Static-T) conditions, IL-2 production by the T cells was reduced in SMG-T cells compared to Static-T cells upon stimulation by phorbol 12-myristate 13-acetate (PMA) and ionomycin. However, when the SMG-T cells were stimulated with DC and peptide, IL-2 was significantly increased compared to Static-T cells. Such enhanced IL-2 production by SMG-T cells peaked at 72 h SMG culture time and decreased thereafter

  17. Cell-free DNA in a three-dimensional spheroid cell culture model

    DEFF Research Database (Denmark)

    Aucamp, Janine; Calitz, Carlemi; Bronkhorst, Abel J.

    2017-01-01

    Background Investigating the biological functions of cell-free DNA (cfDNA) is limited by the interference of vast numbers of putative sources and causes of DNA release into circulation. Utilization of three-dimensional (3D) spheroid cell cultures, models with characteristics closer to the in vivo...... cultures can serve as effective, simplified in vivo-simulating “closed-circuit” models since putative sources of cfDNA are limited to only the targeted cells. In addition, cfDNA can also serve as an alternative or auxiliary marker for tracking spheroid growth, development and culture stability. Biological...... significance 3D cell cultures can be used to translate “closed-circuit” in vitro model research into data that is relevant for in vivo studies and clinical applications. In turn, the utilization of cfDNA during 3D culture research can optimize sample collection without affecting the stability of the growth...

  18. A Novel Counter Sheet-flow Sandwich Cell Culture Device for Mammalian Cell Growth in Space

    Science.gov (United States)

    Sun, Shujin; Gao, Yuxin; Shu, Nanjiang; Tang, Zemei; Tao, Zulai; Long, Mian

    2008-08-01

    Cell culture and growth in space is crucial to understand the cellular responses under microgravity. The effects of microgravity were coupled with such environment restrictions as medium perfusion, in which the underlying mechanism has been poorly understood. In the present work, a customer-made counter sheet-flow sandwich cell culture device was developed upon a biomechanical concept from fish gill breathing. The sandwich culture unit consists of two side chambers where the medium flow is counter-directional, a central chamber where the cells are cultured, and two porous polycarbonate membranes between side and central chambers. Flow dynamics analysis revealed the symmetrical velocity profile and uniform low shear rate distribution of flowing medium inside the central culture chamber, which promotes sufficient mass transport and nutrient supply for mammalian cell growth. An on-orbit experiment performed on a recovery satellite was used to validate the availability of the device.

  19. Cultured meat from stem cells: challenges and prospects.

    Science.gov (United States)

    Post, Mark J

    2012-11-01

    As one of the alternatives for livestock meat production, in vitro culturing of meat is currently studied. The generation of bio-artificial muscles from satellite cells has been ongoing for about 15 years, but has never been used for generation of meat, while it already is a great source of animal protein. In order to serve as a credible alternative to livestock meat, lab or factory grown meat should be efficiently produced and should mimic meat in all of its physical sensations, such as visual appearance, smell, texture and of course, taste. This is a formidable challenge even though all the technologies to create skeletal muscle and fat tissue have been developed and tested. The efficient culture of meat will primarily depend on culture conditions such as the source of medium and its composition. Protein synthesis by cultured skeletal muscle cells should further be maximized by finding the optimal combination of biochemical and physical conditions for the cells. Many of these variables are known, but their interactions are numerous and need to be mapped. This involves a systematic, if not systems, approach. Given the urgency of the problems that the meat industry is facing, this endeavor is worth undertaking. As an additional benefit, culturing meat may provide opportunities for production of novel and healthier products. Copyright © 2012 Elsevier Ltd. All rights reserved.

  20. 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.

  1. Enrichment of skin-derived neural precursor cells from dermal cell populations by altering culture conditions.

    Science.gov (United States)

    Bayati, Vahid; Gazor, Rohoullah; Nejatbakhsh, Reza; Negad Dehbashi, Fereshteh

    2016-01-01

    As stem cells play a critical role in tissue repair, their manipulation for being applied in regenerative medicine is of great importance. Skin-derived precursors (SKPs) may be good candidates for use in cell-based therapy as the only neural stem cells which can be isolated from an accessible tissue, skin. Herein, we presented a simple protocol to enrich neural SKPs by monolayer adherent cultivation to prove the efficacy of this method. To enrich neural SKPs from dermal cell populations, we have found that a monolayer adherent cultivation helps to increase the numbers of neural precursor cells. Indeed, we have cultured dermal cells as monolayer under serum-supplemented (control) and serum-supplemented culture, followed by serum free cultivation (test) and compared. Finally, protein markers of SKPs were assessed and compared in both experimental groups and differentiation potential was evaluated in enriched culture. The cells of enriched culture concurrently expressed fibronectin, vimentin and nestin, an intermediate filament protein expressed in neural and skeletal muscle precursors as compared to control culture. In addition, they possessed a multipotential capacity to differentiate into neurogenic, glial, adipogenic, osteogenic and skeletal myogenic cell lineages. It was concluded that serum-free adherent culture reinforced by growth factors have been shown to be effective on proliferation of skin-derived neural precursor cells (skin-NPCs) and drive their selective and rapid expansion.

  2. Permanently Hypoxic Cell Culture Yields Rat Bone Marrow Mesenchymal Cells with Higher Therapeutic Potential in the Treatment of Chronic Myocardial Infarction

    Directory of Open Access Journals (Sweden)

    Yihua Liu

    2017-11-01

    Full Text Available Background: The mismatch between traditional in vitro cell culture conditions and targeted chronic hypoxic myocardial tissue could potentially hamper the therapeutic effects of implanted bone marrow mesenchymal stem cells (BMSCs. This study sought to address (i the extent of change to BMSC biological characteristics in different in vitro culture conditions and (ii the effectiveness of permanent hypoxic culture for cell therapy in treating chronic myocardial infarction (MI in rats. Methods: rat BMSCs were harvested and cultured in normoxic (21% O2, n=27 or hypoxic conditions (5% O2, n=27 until Passage 4 (P4. Cell growth tests, flow cytometry, and Bio-Plex assays were conducted to explore variations in the cell proliferation, phenotype, and cytokine expression, respectively. In the in vivo set-up, P3-BMSCs cultured in normoxia (n=6 or hypoxia (n=6 were intramyocardially injected into rat hearts that had previously experienced 1-month-old MI. The impact of cell therapy on cardiac segmental viability and hemodynamic performance was assessed 1 month later by 2-Deoxy-2[18F]fluoro-D-glucose (18F-FDG positron emission tomography (PET imaging and pressure-volume catheter, respectively. Additional histomorphological examinations were conducted to evaluate inflammation, fibrosis, and neovascularization. Results: Hypoxic preconditioning significantly enhanced rat BMSC clonogenic potential and proliferation without altering the multipotency. Different profiles of inflammatory, fibrotic, and angiogenic cytokine secretion were also documented, with a marked correlation observed between in vitro and in vivo proangiogenic cytokine expression and tissue neovessels. Hypoxic-preconditioned cells presented a beneficial effect on the myocardial viability of infarct segments and intrinsic contractility. Conclusion: Hypoxic-preconditioned BMSCs were able to benefit myocardial perfusion and contractility, probably by modulating the inflammation and promoting

  3. Permanently Hypoxic Cell Culture Yields Rat Bone Marrow Mesenchymal Cells with Higher Therapeutic Potential in the Treatment of Chronic Myocardial Infarction.

    Science.gov (United States)

    Liu, Yihua; Yang, Xiaoxi; Maureira, Pablo; Falanga, Aude; Marie, Vanessa; Gauchotte, Guillaume; Poussier, Sylvain; Groubatch, Frederique; Marie, Pierre-Yves; Tran, Nguyen

    2017-01-01

    The mismatch between traditional in vitro cell culture conditions and targeted chronic hypoxic myocardial tissue could potentially hamper the therapeutic effects of implanted bone marrow mesenchymal stem cells (BMSCs). This study sought to address (i) the extent of change to BMSC biological characteristics in different in vitro culture conditions and (ii) the effectiveness of permanent hypoxic culture for cell therapy in treating chronic myocardial infarction (MI) in rats. rat BMSCs were harvested and cultured in normoxic (21% O2, n=27) or hypoxic conditions (5% O2, n=27) until Passage 4 (P4). Cell growth tests, flow cytometry, and Bio-Plex assays were conducted to explore variations in the cell proliferation, phenotype, and cytokine expression, respectively. In the in vivo set-up, P3-BMSCs cultured in normoxia (n=6) or hypoxia (n=6) were intramyocardially injected into rat hearts that had previously experienced 1-month-old MI. The impact of cell therapy on cardiac segmental viability and hemodynamic performance was assessed 1 month later by 2-Deoxy-2[18F]fluoro-D-glucose (18F-FDG) positron emission tomography (PET) imaging and pressure-volume catheter, respectively. Additional histomorphological examinations were conducted to evaluate inflammation, fibrosis, and neovascularization. Hypoxic preconditioning significantly enhanced rat BMSC clonogenic potential and proliferation without altering the multipotency. Different profiles of inflammatory, fibrotic, and angiogenic cytokine secretion were also documented, with a marked correlation observed between in vitro and in vivo proangiogenic cytokine expression and tissue neovessels. Hypoxic-preconditioned cells presented a beneficial effect on the myocardial viability of infarct segments and intrinsic contractility. Hypoxic-preconditioned BMSCs were able to benefit myocardial perfusion and contractility, probably by modulating the inflammation and promoting angiogenesis. © 2017 The Author(s). Published by S. Karger AG

  4. Mesenchymal stem cells enhance the metastasis of 3D-cultured hepatocellular carcinoma cells

    International Nuclear Information System (INIS)

    Liu, Chang; Liu, Yang; Xu, Xiao-xi; Guo, Xin; Sun, Guang-wei; Ma, Xiao-jun

    2016-01-01

    Accumulating evidences have demonstrated that mesenchymal stem cells (MSC) could be recruited to the tumor microenvironment. Umbilical cord mesenchymal stem cells (UCMSC) were attractive vehicles for delivering therapeutic agents against cancer. Nevertheless, the safety of UCMSC in the treatment of tumors including hepatocellular carcinoma (HCC) was still undetermined. In this study, an in vitro co-culture system was established to evaluate the effect of UCMSC on the cell growth, cancer stem cell (CSC) characteristics, drug resistance, metastasis of 3D-cultured HCC cells, and the underlying mechanism was also investigated. It was found that after co-cultured with UCMSC, the metastatic ability of 3D-cultured HCC cells was significantly enhanced as indicated by up-regulation of matrix metalloproteinase (MMP), epithelial-mesenchymal transition (EMT)-related genes, and migration ability. However, cell growth, drug resistance and CSC-related gene expression of HCC cells were not affected by UCMSC. Moreover, EMT was reversed, MMP-2 expression was down-regulated, and migration ability of HCC cell was significantly inhibited when TGF-β receptor inhibitor SB431542 was added into the co-culture system. Therefore, these data indicated that UCMSC could significantly enhance the tumor cell metastasis, which was due to the EMT of HCC cells induced by TGF-β. The online version of this article (doi:10.1186/s12885-016-2595-4) contains supplementary material, which is available to authorized users

  5. Microalgae harvesting techniques: A review.

    Science.gov (United States)

    Singh, Gulab; Patidar, S K

    2018-07-01

    Microalgae with wide range of commercial applications have attracted a lot of attention of the researchers in the last few decades. However, microalgae utilization is not economically sustainable due to high cost of harvesting. A wide range of solid - liquid separation techniques are available for microalgae harvesting. The techniques include coagulation and flocculation, flotation, centrifugation and filtration or a combination of various techniques. Despite the importance of harvesting to the economics and energy balance, there is no universal harvesting technique for microalgae. Therefore, this review focuses on assessing technical, economical and application potential of various harvesting techniques so as to allow selection of an appropriate technology for cost effectively harvesting of microalgae from their culture medium. Various harvesting and concentrating techniques of microalgae were reviewed to suggest order of suitability of the techniques for four main microalgae applications i.e biofuel, human and animal food, high valued products, and water quality restoration. For deciding the order of suitability, a comparative analysis of various harvesting techniques based on the six common criterions (i.e biomass quality, cost, biomass quantity, processing time, species specific and toxicity) has been done. Based on the order of various techniques vis-a-vis various criteria and preferred order of criteria for various applications, order of suitability of harvesting techniques for various applications has been decided. Among various harvesting techniques, coagulation and flocculation, centrifugation and filtration were found to be most suitable for considered applications. These techniques may be used alone or in combination for increasing the harvesting efficiency. Copyright © 2018 Elsevier Ltd. All rights reserved.

  6. Effects of viscoelastic ophthalmic solutions on cell cultures

    Directory of Open Access Journals (Sweden)

    Madhavan Hajib

    1998-01-01

    Full Text Available The development of mild but significant inflammation probably attributable to viscoelastic ophthalmic solutions in cataract surgery was recently brought to the notice of the authors, and hence a study of the effects of these solutions available in India, on cell cultures was undertaken. We studied the effects of 6 viscoelastic ophthalmic solutions (2 sodium hyaluronate designated as A and B, and 4 hydroxypropylmethylcellulose designated as C, D, E and F on HeLa, Vero and BHK-21 cell lines in tissue culture microtitre plates using undiluted, 1:10 and 1:100 dilutions of the solutions, and in cover slip cultures using undiluted solutions. Phase contrast microscopic examination of the solutions was also done to determine the presence of floating particles. The products D and F produced cytotoxic changes in HeLa cell line and these products also showed the presence of floating particles under phase contrast microscopy. Other products did not have any adverse effects on the cell lines nor did they show floating particles. The viscoelastic ophthalmic pharmaceutical products designated D and F have cytotoxic effects on HeLa cell line which appears to be a useful cell line for testing these products for their toxicity. The presence of particulate materials in products D and F indicates that the methods used for purification of the solution are not effective.

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

    International Nuclear Information System (INIS)

    Simenc, Janez; Lipnik-Stangelj, Metoda

    2012-01-01

    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

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

    International Nuclear Information System (INIS)

    Dai, R.X.; Djakiew, D.; Dym, M.

    1987-01-01

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

  9. Apple derived cellulose scaffolds for 3D mammalian cell culture.

    Directory of Open Access Journals (Sweden)

    Daniel J Modulevsky

    Full Text Available There are numerous approaches for producing natural and synthetic 3D scaffolds that support the proliferation of mammalian cells. 3D scaffolds better represent the natural cellular microenvironment and have many potential applications in vitro and in vivo. Here, we demonstrate that 3D cellulose scaffolds produced by decellularizing apple hypanthium tissue can be employed for in vitro 3D culture of NIH3T3 fibroblasts, mouse C2C12 muscle myoblasts and human HeLa epithelial cells. We show that these cells can adhere, invade and proliferate in the cellulose scaffolds. In addition, biochemical functionalization or chemical cross-linking can be employed to control the surface biochemistry and/or mechanical properties of the scaffold. The cells retain high viability even after 12 continuous weeks of culture and can achieve cell densities comparable with other natural and synthetic scaffold materials. Apple derived cellulose scaffolds are easily produced, inexpensive and originate from a renewable source. Taken together, these results demonstrate that naturally derived cellulose scaffolds offer a complementary approach to existing techniques for the in vitro culture of mammalian cells in a 3D environment.

  10. Somatic embryogenesis in cell cultures of Glycine species.

    Science.gov (United States)

    Gamborg, O L; Davis, B P; Stahlhut, R W

    1983-08-01

    This report describes the development of procedures for the production of somatic embryos in cell cultures of Glycine species including soybean. The conditions for callus induction and initiation of rapidly growing cell suspension cultures were defined. Methods for inducing embryogenesis were tested on 16 lines of several Glycine species and cultivars of soybean. The SB-26 Culture of a G. soja gave the best results and was used in the experiments. Embryogenesis required the presence of picloram or 2,4-D. AMO 1618, CCC, PP-333 and Ancymidol enhanced the embryogenesis frequency. Plants of the G. soja (SB-26) were grown to maturity from seed-derived shoot tips. Characteristics of the plants are discussed.

  11. Radiation transformation in differentiated human cells in culture

    International Nuclear Information System (INIS)

    Mothersill, C.; Seymour, C.; Moriarty, M.; Malone, J.; Byrne, P.; Hennessy, T.

    1986-01-01

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

  12. 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.

  13. The bioconversion process of deoxypodophyllotoxin with Linum flavum cell cultures

    NARCIS (Netherlands)

    Koulman, A; Beekman, A.C; Pras, N.; Quax, Wim

    2003-01-01

    The in vitro cell suspension culture of Linum flavum is able to convert high amounts of the 2,7'-cyclolignan deoxypodophyllotoxin into 6-methoxypodophyllotoxin 7-O-glucoside. We studied this conversion in detail by monitoring the intermediates and side-products after feeding different concentrations

  14. Using Haworthia Cultured Cells as an Aid in Teaching Botany

    Science.gov (United States)

    Majumdar, Shyamal K.; Castellano, John M.

    1977-01-01

    Callus induction from species of Haworthia can be done quickly in the laboratory with minimal equipment to study tissue dedifferentiation and cellular redifferentiation. It is shown that the cultured cell can also be used to study and evaluate the effects of various mutagens, carcinogens, and pesticides in controlled environments. (Author/MA)

  15. 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

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

  16. Plant Cell Cultures as Source of Cosmetic Active Ingredients

    Directory of Open Access Journals (Sweden)

    Ani Barbulova

    2014-04-01

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

  17. Establishment of the callus and cell suspension culture of ...

    African Journals Online (AJOL)

    The objective of this work was the optimization of the conditions of callus and cell suspension culture of Elaeagnus angustifolia for the production of condensed tannins. The effects of different conditions on the callus growth and the production of condensed tannins were researched. The leaf tissue part of E. angustifolia was ...

  18. Ultrastructure of cells of Ulmus americana cultured in vitro and exposed to the culture filtrate of Ceratocystis ulmi

    Science.gov (United States)

    Paula M. Pijut; R. Daniel Lineberger; Subhash C. Domir; Jann M. Ichida; Charles R. Krause

    1990-01-01

    Calli of American elm susceptible and resistant to Dutch elm disease were exposed to a culture filtrate of a pathogenic isolate of Ceratocystis ulmi. Cells from untreated tissue exhibited typical internal composition associated with healthy, actively growing cells. All cells exposed to culture filtrate showed appreciable ultrastructural changes....

  19. Glycyrrhiza glabra (Linn.) and Lavandula officinalis (L.) cell suspension cultures-based biotransformation of β-artemether.

    Science.gov (United States)

    Patel, Suman; Gaur, Rashmi; Upadhyaya, Mohita; Mathur, Archana; Mathur, Ajay K; Bhakuni, Rajendra S

    2011-07-01

    The biotransformation of β-artemether (1) by cell suspension cultures of Glycyrrhiza glabra and Lavandula officinalis is reported here for the first time. The major biotransformed product appeared as a grayish-blue color spot on thin-layer chromatography (TLC) with transparent crystal-like texture. Based on its infrared (IR) and (1)H nuclear magnetic resonance (NMR) spectra, the product was characterized as a tetrahydrofuran (THF)-acetate derivative (2). The highest conversion efficiencies of 57 and 60% were obtained when 8-9-day-old cell suspensions of G. glabra and L. officinalis were respectively fed with 4-7 mg of compound 1 in 40 ml of medium per culture and the cells were harvested after 2-5 days of incubation. The addition of compound 1 at the beginning of the culture cycle caused severe growth depression in a dose-dependent manner, resulting in poor bioconversion efficiency of ~25% at 2-5 mg/culture dose only.

  20. Lethal impacts of cigarette smoke in cultured tobacco cells

    Directory of Open Access Journals (Sweden)

    Kawano Tomonori

    2011-07-01

    Full Text Available Abstract Background In order to understand and generalize the toxic mechanism of cigarette smoke in living cells, comparison of the data between animal systems and other biological system such as microbial and plant systems is highly beneficial. Objective By employing the tobacco cells as model materials for cigarette smoke toxicity assay, the impacts of the combustion by-products such as nitrogen oxides could be highlighted as the toxic impacts of the plant-derived endogenous chemicals could be excluded in the plant cells. Methods Cigarette smoke-induced cell death was assessed in tobacco cell suspension cultures in the presence and absence of pharmacological inhibitors. Results Cigarette smoke was effective in induction of cell death. The smoke-induced cell death could be partially prevented by addition of nitric oxide (NO scavenger, suggesting the role for NO as the cell death mediator. Addition of NO donor to tobacco cells also resulted in development of partial cell death further confirming the role of NO as cell death mediator. Members of reactive oxygen species and calcium ion were shown to be protecting the cells from the toxic action of smoke-derived NO.

  1. Evaluation of the radiosensitivity of acute myeloblastic leukaemia progenitor cells by culture methods exploring self-renewal. Evaluation de la radiosensibilite des progeniteurs de leucemie aigue myeloblastique par des methodes de culture explorant ou non l'autorenouvellement

    Energy Technology Data Exchange (ETDEWEB)

    Cowen, D; Richaud, P; Landriau, S; Lagarde, P; Gualde, N [Fondation Bergonie, 33 - Bordeaux (France); Boiron, J M [Hopital du Haut-Leveque, 33 - Pessac (France); Mahon, F X; Belloc, F [Hopital Regional, 33 - Bordeaux (France); Reiffers, J [Hopital du Haut-Leveque, 33 - Pessac (France) Hopital Regional, 33 - Bordeaux (France)

    1993-01-01

    The progenitor cells of acute myeloblastic leukaemia (AML) are usually cultured in methylcellulose which selects for terminal dividing cells. Suspension cultures have been developed because they reflect self-renewal: the exponential growth of the progenitors of AML cultured in suspension is due to self-renewal. We have compared the radiosensitivity of the progenitors of AML grown either in methylcellulose alone or first in suspension for 7 days before being plated in methylcellulose. Cells were harvested from leukaemic bone marrows at the moment of diagnosis. The myeloblastic lineage of the colonies was assessed by morphological, cytochemical and immunophenotypic analysis and by the use of growth factors which do not stimulate T-lymphocytes. The cell-cycle distribution of leukaemic blasts was comparable for all the samples. This method enabled aggressive leukaemias to be selected. The radiosensitivity showed wide variations from one patient to another (Do ranging from 0.35 to 2.6 Gy) whichever culture method used. The progenitor cells capable of self-renewal were more radiosensitive (Mean Do 0.9[+-]0.4 Gy) than terminal dividing cells (Mean Do = 1.35[+-]0.5 Gy). In two cases, a shoulder was found in the initial part of the cell-survival curves of cells capable of self-renewal. The shape of the curves was better fitted by the linear quadratic model with very low values of [alpha]/[beta], suggesting a reduced antileukaemic effect in case of fractionation.

  2. Porcine platelet lysate as a supplement for animal cell culture

    Science.gov (United States)

    Aldén, Anna; Gonzalez, Lorena; Persson, Anna; Christensson, Kerstin; Holmqvist, Olov

    2007-01-01

    A novel supplementation of cell growth media based on a porcine platelet lysate was developed for culture of animal-derived cells. The platelet lysate was produced from porcine blood and contained lysate of platelets and plasma components. It showed satisfactory microbiological integrity and it carried only low amount of endotoxins (platelet lysate supported well proliferation of Vero (African green monkey transformed kidney epithelial cells), Chinese hamster ovary (CHO) and hybridoma cells comparable to fetal bovine serum (FBS). Platelet lysate shows promise as a viable choice over FBS as it can be produced in large quantities, high lot-to-lot consistency and with an attractive price structure. Furthermore it is a strong alternative to FBS for ethical reasons. It is expected that it can be used as a general supplementation for most animal cells for research studies on the proliferation of cells and their expression of products. PMID:19002989

  3. Anthocyanines as light harvesters in the dye-sensitized TiO2 solar cell

    International Nuclear Information System (INIS)

    Sokolsky, M.; Kaiser, M.; Cirak, J.; Kusko, M.

    2011-01-01

    In this paper anthocyanine extracted from blackberry was used instead of widely used dyes based on Ru and N3 complexes such as N3, N719 or 'black dye', on which one of the highest efficiencies where measured (10.0 % to 11 %). DSSC were successfully fabricated using anthocyanine dye extracted from blackberries. The open circuit voltage of 419.0 mV, short circuit current of 380.40 μA, fill factor of 41.2 % and efficiency of 0.0164 % were evaluated. The cell shows degradation in performance over time of the exponential type with a drop in the open circuit voltage to 406 mV in 15 minutes. (authors)

  4. 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.

  5. Mechanism for multiplicity of steady states with distinct cell concentration in continuous culture of mammalian cells.

    Science.gov (United States)

    Yongky, Andrew; Lee, Jongchan; Le, Tung; Mulukutla, Bhanu Chandra; Daoutidis, Prodromos; Hu, Wei-Shou

    2015-07-01

    Continuous culture for the production of biopharmaceutical proteins offers the possibility of steady state operations and thus more consistent product quality and increased productivity. Under some conditions, multiplicity of steady states has been observed in continuous cultures of mammalian cells, wherein with the same dilution rate and feed nutrient composition, steady states with very different cell and product concentrations may be reached. At those different steady states, cells may exhibit a high glycolysis flux with high lactate production and low cell concentration, or a low glycolysis flux with low lactate and high cell concentration. These different steady states, with different cell concentration, also have different productivity. Developing a mechanistic understanding of the occurrence of steady state multiplicity and devising a strategy to steer the culture toward the desired steady state is critical. We establish a multi-scale kinetic model that integrates a mechanistic intracellular metabolic model and cell growth model in a continuous bioreactor. We show that steady state multiplicity exists in a range of dilution rate in continuous culture as a result of the bistable behavior in glycolysis. The insights from the model were used to devise strategies to guide the culture to the desired steady state in the multiple steady state region. The model provides a guideline principle in the design of continuous culture processes of mammalian cells. © 2015 Wiley Periodicals, Inc.

  6. Characterization of primary human mammary epithelial cells isolated and propagated by conditional reprogrammed cell culture.

    Science.gov (United States)

    Jin, Liting; Qu, Ying; Gomez, Liliana J; Chung, Stacey; Han, Bingchen; Gao, Bowen; Yue, Yong; Gong, Yiping; Liu, Xuefeng; Amersi, Farin; Dang, Catherine; Giuliano, Armando E; Cui, Xiaojiang

    2018-02-20

    Conditional reprogramming methods allow for the inexhaustible in vitro proliferation of primary epithelial cells from human tissue specimens. This methodology has the potential to enhance the utility of primary cell culture as a model for mammary gland research. However, few studies have systematically characterized this method in generating in vitro normal human mammary epithelial cell models. We show that cells derived from fresh normal breast tissues can be propagated and exhibit heterogeneous morphologic features. The cultures are composed of CK18, desmoglein 3, and CK19-positive luminal cells and vimentin, p63, and CK14-positive myoepithelial cells, suggesting the maintenance of in vivo heterogeneity. In addition, the cultures contain subpopulations with different CD49f and EpCAM expression profiles. When grown in 3D conditions, cells self-organize into distinct structures that express either luminal or basal cell markers. Among these structures, CK8-positive cells enclosing a lumen are capable of differentiation into milk-producing cells in the presence of lactogenic stimulus. Furthermore, our short-term cultures retain the expression of ERα, as well as its ability to respond to estrogen stimulation. We have investigated conditionally reprogrammed normal epithelial cells in terms of cell type heterogeneity, cellular marker expression, and structural arrangement in two-dimensional (2D) and three-dimensional (3D) systems. The conditional reprogramming methodology allows generation of a heterogeneous culture from normal human mammary tissue in vitro . We believe that this cell culture model will provide a valuable tool to study mammary cell function and malignant transformation.

  7. Imprint lithography provides topographical nanocues to guide cell growth in primary cortical cell culture

    NARCIS (Netherlands)

    Xie, S.; Luttge, R.

    2014-01-01

    In this paper, we describe a technology platform to study the effect of nanocues on the cell growth direction in primary cortical cell culture. Topographical cues to cells are provided using nanoscale features created by Jet and Flash Imprint Lithography, coated with polyethylenimine. We

  8. Engineering systems for the generation of patterned co-cultures for controlling cell-cell interactions.

    Science.gov (United States)

    Kaji, Hirokazu; Camci-Unal, Gulden; Langer, Robert; Khademhosseini, Ali

    2011-03-01

    Inside the body, cells lie in direct contact or in close proximity to other cell types in a tightly controlled architecture that often regulates the resulting tissue function. Therefore, tissue engineering constructs that aim to reproduce the architecture and the geometry of tissues will benefit from methods of controlling cell-cell interactions with microscale resolution. We discuss the use of microfabrication technologies for generating patterned co-cultures. In addition, we categorize patterned co-culture systems by cell type and discuss the implications of regulating cell-cell interactions in the resulting biological function of the tissues. Patterned co-cultures are a useful tool for fabricating tissue engineered constructs and for studying cell-cell interactions in vitro, because they can be used to control the degree of homotypic and heterotypic cell-cell contact. In addition, this approach can be manipulated to elucidate important factors involved in cell-matrix interactions. Patterned co-culture strategies hold significant potential to develop biomimetic structures for tissue engineering. It is expected that they would create opportunities to develop artificial tissues in the future. This article is part of a Special Issue entitled Nanotechnologies - Emerging Applications in Biomedicine. 2010 Elsevier B.V. All rights reserved.

  9. The major bovine mastitis pathogens have different cell tropisms in cultures of bovine mammary gland cells

    NARCIS (Netherlands)

    Lammers, A.; Vorstenbosch, van C.J.; Erkens, J.H.F.; Smith, H.E.

    2001-01-01

    We previously showed that Staphylococcus aureus cells adhered mainly to an elongated cell type, present in cultures of bovine mammary gland cells. Moreover. we showed that this adhesion was mediated by binding to fibronectin. The same in vitro model was used here, to study adhesion of other

  10. MEMS-based dynamic cell-to-cell culture platforms using electrochemical surface modifications

    International Nuclear Information System (INIS)

    Chang, Jiyoung; Lin, Liwei; Yoon, Sang-Hee; Mofrad, Mohammad R K

    2011-01-01

    MEMS-based biological platforms with the capability of both spatial placements and time releases of living cells for cell-to-cell culture experiments have been designed and demonstrated utilizing electrochemical surface modification effects. The spatial placement is accomplished by electrochemical surface modification of substrate surfaces to be either adhesive or non-adhesive for living cells. The time control is achieved by the electrical activation of the selective indium tin oxide co-culture electrode to allow the migration of living cells onto the electrode to start the cell-to-cell culture studies. Prototype devices have a three-electrode design with an electrode size of 50 × 50 µm 2 and the separation gaps of 2 µm between them. An electrical voltage of −1.5 V has been used to activate the electrodes independently and sequentially to demonstrate the dynamic cell-to-cell culture experiments of NIH 3T3 fibroblast and Madin Darby canine kidney cells. As such, this MEMS platform could be a basic yet versatile tool to characterize transient cell-to-cell interactions

  11. Cell culture density affects the proliferation activity of human adipose tissue stem cells.

    Science.gov (United States)

    Kim, Dae Seong; Lee, Myoung Woo; Ko, Young Jong; Chun, Yong Hoon; Kim, Hyung Joon; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee

    2016-01-01

    In this study, we investigated the effect of cell density on the proliferation activity of human mesenchymal stem cells (MSCs) derived from adipose tissue (AT-MSCs) over time in culture. Passage #4 (P4) and #12 (P12) AT-MSCs from two donors were plated at a density of 200 (culture condition 1, CC1) or 5000 (culture condition 2, CC2) cells cm(-2) . After 7 days of incubation, P4 and P12 AT-MSCs cultured in CC1 were thin and spindle-shaped, whereas those cultured in CC2 had extensive cell-to-cell contacts and an expanded cell volume. In addition, P4 and P12 AT-MSCs in CC1 divided more than three times, while those in CC2 divided less than once on average. Flow cytometric analysis using 5(6)-carboxyfluorescein diacetate N-succinimidyl ester dye showed that the fluorescence intensity of AT-MSCs was lower in CC1 than in CC2. Furthermore, expression of proliferation-associated genes, such as CDC45L, CDC20A and KIF20A, in P4 AT-MSCs was higher in CC1 than in CC2, and this difference was also observed in P12 AT-MSCs. These data demonstrated that cell culture density affects the proliferation activity of MSCs, suggesting that it is feasible to design a strategy to prepare suitable MSCs using specific culture conditions. Copyright © 2016 John Wiley & Sons, Ltd.

  12. Improved Cell Culture Method for Growing Contracting Skeletal Muscle Models

    Science.gov (United States)

    Marquette, Michele L.; Sognier, Marguerite A.

    2013-01-01

    An improved method for culturing immature muscle cells (myoblasts) into a mature skeletal muscle overcomes some of the notable limitations of prior culture methods. The development of the method is a major advance in tissue engineering in that, for the first time, a cell-based model spontaneously fuses and differentiates into masses of highly aligned, contracting myotubes. This method enables (1) the construction of improved two-dimensional (monolayer) skeletal muscle test beds; (2) development of contracting three-dimensional tissue models; and (3) improved transplantable tissues for biomedical and regenerative medicine applications. With adaptation, this method also offers potential application for production of other tissue types (i.e., bone and cardiac) from corresponding precursor cells.

  13. [Primary culture of cat intestinal epithelial cell and construction of its cDNA library].

    Science.gov (United States)

    Ye, L; Gui-Hua, Z; Kun, Y; Hong-Fa, W; Ting, X; Gong-Zhen, L; Wei-Xia, Z; Yong, C

    2017-04-12

    Objective To establish the primary cat intestinal epithelial cells (IECs) culture methods and construct the cDNA library for the following yeast two-hybrid experiment, so as to screen the virulence interaction factors among the final host. Methods The primary cat IECs were cultured by the tissue cultivation and combined digestion with collagenase XI and dispase I separately. Then the cat IECs cultured was identified with the morphological observation and cyto-keratin detection, by using goat anti-cyto-keratin monoclonal antibodies. The mRNA of cat IECs was isolated and used as the template to synthesize the first strand cDNA by SMART™ technology, and then the double-strand cDNAs were acquired by LD-PCR, which were subsequently cloned into the plasmid PGADT7-Rec to construct yeast two-hybrid cDNA library in the yeast strain Y187 by homologous recombination. Matchmaker™ Insert Check PCR was used to detect the size distribution of cDNA fragments after the capacity calculation of the cDNA library. Results The comparison of the two cultivation methods indicated that the combined digestion of collagenase XI and dispase I was more effective than the tissue cultivation. The cat IECs system of continuous culture was established and the cat IECs with high purity were harvested for constructing the yeast two-hybrid cDNA library. The library contained 1.1×10 6 independent clones. The titer was 2.8×10 9 cfu/ml. The size of inserted fragments was among 0.5-2.0 kb. Conclusion The yeast two-hybrid cDNA library of cat IECs meets the requirements of further screen research, and this study lays the foundation of screening the Toxoplasma gondii virulence interaction factors among the cDNA libraries of its final hosts.

  14. Biotechnological production of caffeic acid derivatives from cell and organ cultures of Echinacea species.

    Science.gov (United States)

    Murthy, Hosakatte Niranjana; Kim, Yun-Soo; Park, So-Young; Paek, Kee-Yoeup

    2014-09-01

    Caffeic acid derivatives (CADs) are a group of bioactive compounds which are produced in Echinacea species especially Echinacea purpurea, Echinacea angustifolia, and Echinacea pallida. Echinacea is a popular herbal medicine used in the treatment of common cold and it is also a prominent dietary supplement used throughout the world. Caffeic acid, chlorogenic acid (5-O-caffeoylquinic acid), caftaric acid (2-O-caffeoyltartaric acid), cichoric acid (2, 3-O-dicaffeoyltartaric acid), cynarin, and echinacoside are some of the important CADs which have varied pharmacological activities. The concentrations of these bioactive compounds are species specific and also they vary considerably with the cultivated Echinacea species due to geographical location, stage of development, time of harvest, and growth conditions. Due to these reasons, plant cell and organ cultures have become attractive alternative for the production of biomass and caffeic acid derivatives. Adventitious and hairy roots have been induced in E. pupurea and E. angustifolia, and suspension cultures have been established from flask to bioreactor scale for the production of biomass and CADs. Tremendous progress has been made in this area; various bioprocess methods and strategies have been developed for constant high-quality productivity of biomass and secondary products. This review is aimed to discuss biotechnological methods and approaches employed for the sustainable production of CADs.

  15. 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...... not succeed. Cultures grew exponentially on a shaker at 27°C in the light. Their doubling times varied from 1.1 days on 2,4–D (10–6M) or NAA (10−5M)+ 1 g/1 casein hydrolysate to 2.7 days on BAP (3 × 10−7M) and 5.1 days on supraoptimal levels of 2,4-D (10−5M). Cultures grew on NH4+-N alone (from ammonium...... malate) or on NO3−-N alone. Dry weight yield was proportional to the amount of nitrate-N added (47 mg/mg N). Filtered suspension cultures containing single cells (plating cultures) could be grown in agar in petri dishes when NAA or 2,4-D were used as growth substances. Cells grew at densities above 500...

  16. Trophic Effects of Mesenchymal Stem Cells in Chondrocyte Co-Cultures are Independent of Culture Conditions and Cell Sources

    NARCIS (Netherlands)

    Wu, Ling; Prins, H.J.; Helder, M.; van Blitterswijk, Clemens; Karperien, Hermanus Bernardus Johannes

    2012-01-01

    Earlier, we have shown that the increased cartilage production in pellet co-cultures of chondrocytes and bone marrow-derived mesenchymal stem cells (BM-MSCs) is due to a trophic role of the MSC in stimulating chondrocyte proliferation and matrix production rather than MSCs actively undergoing

  17. Trophic effects of mesenchymal stem cells in chondrocyte co-cultures are independent of culture conditions and cell sources

    NARCIS (Netherlands)

    Wu, L.; Prins, H.J.; Helder, M.N.; van Blitterswijk, C.A.; Karperien, M.

    2012-01-01

    Earlier, we have shown that the increased cartilage production in pellet co-cultures of chondrocytes and bone marrow-derived mesenchymal stem cells (BM-MSCs) is due to a trophic role of the MSC in stimulating chondrocyte proliferation and matrix production rather than MSCs actively undergoing

  18. The Effect of Spaceflight on Bone Cell Cultures

    Science.gov (United States)

    Landis, William J.

    1999-01-01

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

  19. Infrared Harvesting Colloidal Quantum Dot Solar Cell Based on Multi-scale Disordered Electrodes

    KAUST Repository

    Tian, Yi

    2015-06-23

    Colloidal quantum dot photovoltaics (CQDPV) offer a big potential to be a renewable energy source due to low cost and tunable band-gap. Currently, the certified power conversion efficiency of CQDPV has reached 9.2%. Compared to the 31% theoretical efficiency limit of single junction solar cells, device performances have still have a large potential to be improved. For photovoltaic devices, a classical way to enhance absorption is to increase the thickness of the active layers. Although this approach can improve absorption, it reduces the charge carriers extraction efficiency. Photo-generated carriers, in fact, are prone to recombine within the defects inside CQD active layers. In an effort to solve this problem, we proposed to increase light absorption from a given thickness of colloidal quantum dot layers with the assistance of disorder. Our approach is to develop new types of electrodes with multi-scale disordered features, which localize energy into the active layer through plasmonic effects. We fabricated nanostructured gold substrates by electrochemical methods, which allow to control surface disorder as a function of deposition conditions. We demonstrated that the light absorption from 600 nm to 800 nm is impressively enhanced, when the disorder of the nanostructured surface increases. Compared to the planar case, the most disorder case increased 65% light absorption at the wavelength of λ = 700nm in the 100 nm PbS film. The average absorption enhancement across visible and infrared region in 100 nm PbS film is 49.94%. By developing a photovoltaic module, we measured a dramatic 34% improvement in the short-circuit current density of the device. The power conversion efficiency of the tested device in top-illumination configuration showed 25% enhancement.

  20. Alginate as a cell culture substrate for growth and differentiation of human retinal pigment epithelial cells.

    Science.gov (United States)

    Heidari, Razeih; Soheili, Zahra-Soheila; Samiei, Shahram; Ahmadieh, Hamid; Davari, Maliheh; Nazemroaya, Fatemeh; Bagheri, Abouzar; Deezagi, Abdolkhalegh

    2015-03-01

    The purpose of this study was to evaluate retinal pigment epithelium (RPE) cells' behavior in alginate beads that establish 3D environment for cellular growth and mimic extracellular matrix versus the conventional 2D monolayer culture. RPE cells were encapsulated in alginate beads by dripping alginate cell suspension into CaCl2 solution. Beads were suspended in three different media including Dulbecco's modified Eagle's medium (DMEM)/F12 alone, DMEM/F12 supplemented with 10 % fetal bovine serum (FBS), and DMEM/F12 supplemented with 30 % human amniotic fluid (HAF). RPE cells were cultivated on polystyrene under the same conditions as controls. Cell phenotype, cell proliferation, cell death, and MTT assay, immunocytochemistry, and real-time RT-PCR were performed to evaluate the effect of alginate on RPE cells characteristics and integrity. RPE cells can survive and proliferate in alginate matrixes. Immunocytochemistry analysis exhibited Nestin, RPE65, and cytokeratin expressions in a reasonable number of cultured cells in alginate beads. Real-time PCR data demonstrated high levels of Nestin, CHX10, RPE65, and tyrosinase gene expressions in RPE cells immobilized in alginate when compared to 2D monolayer culture systems. The results suggest that alginate can be used as a reliable scaffold for maintenance of RPE cells' integrity and in vitro propagation of human retinal progenitor cells for cell replacement therapies in retinal diseases.

  1. Cell chirality: emergence of asymmetry from cell culture.

    Science.gov (United States)

    Wan, Leo Q; Chin, Amanda S; Worley, Kathryn E; Ray, Poulomi

    2016-12-19

    Increasing evidence suggests that intrinsic cell chirality significantly contributes to the left-right (LR) asymmetry in embryonic development, which is a well-conserved characteristic of living organisms. With animal embryos, several theories have been established, but there are still controversies regarding mechanisms associated with embryonic LR symmetry breaking and the formation of asymmetric internal organs. Recently, in vitro systems have been developed to determine cell chirality and to recapitulate multicellular chiral morphogenesis on a chip. These studies demonstrate that chirality is indeed a universal property of the cell that can be observed with well-controlled experiments such as micropatterning. In this paper, we discuss the possible benefits of these in vitro systems to research in LR asymmetry, categorize available platforms for single-cell chirality and multicellular chiral morphogenesis, and review mathematical models used for in vitro cell chirality and its applications in in vivo embryonic development. These recent developments enable the interrogation of the intracellular machinery in LR axis establishment and accelerate research in birth defects in laterality.This article is part of the themed issue 'Provocative questions in left-right asymmetry'. © 2016 The Author(s).

  2. Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

    Directory of Open Access Journals (Sweden)

    Magnusson Magnus K

    2010-07-01

    Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

  3. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    International Nuclear Information System (INIS)

    Zhang, Fenxi; Hong, Yan; Liang, Wenmei; Ren, Tongming; Jing, Suhua; Lin, Juntang

    2012-01-01

    Highlights: ► Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). ► Presence of SCs dramatically increased proliferation and migration of UCMSCs. ► Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of “nurse” cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  4. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Fenxi, E-mail: fxzhang0824@gmail.com [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Hong, Yan; Liang, Wenmei [Department of Histology and Embryology, Guiyang Medical University, Guizhou 550004, People' s Republic of China (China); Ren, Tongming [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Jing, Suhua [ICU Center, The Third Hospital of Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Lin, Juntang [Stem Cell Center, Xinxiang Medical University, Henan 453003, People' s Republic of China (China)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). Black-Right-Pointing-Pointer Presence of SCs dramatically increased proliferation and migration of UCMSCs. Black-Right-Pointing-Pointer Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of 'nurse' cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  5. Blue-light filtering alters angiogenic signaling in human retinal pigmented epithelial cells culture model.

    Science.gov (United States)

    Vila, Natalia; Siblini, Aya; Esposito, Evangelina; Bravo-Filho, Vasco; Zoroquiain, Pablo; Aldrees, Sultan; Logan, Patrick; Arias, Lluis; Burnier, Miguel N

    2017-11-02

    Light exposure and more specifically the spectrum of blue light contribute to the oxidative stress in Age-related macular degeneration (AMD). The purpose of the study was to establish whether blue light filtering could modify proangiogenic signaling produced by retinal pigmented epithelial (RPE) cells under different conditions simulating risk factors for AMD. Three experiments were carried out in order to expose ARPE-19 cells to white light for 48 h with and without blue light-blocking filters (BLF) in different conditions. In each experiment one group was exposed to light with no BLF protection, a second group was exposed to light with BLF protection, and a control group was not exposed to light. The ARPE-19 cells used in each experiment prior to light exposure were cultured for 24 h as follows: Experiment 1) Normoxia, Experiment 2) Hypoxia, and Experiment 3) Lutein supplemented media in normoxia. The media of all groups was harvested after light exposure for sandwich ELISA-based assays to quantify 10 pro-angiogenic cytokines. A significant decrease in angiogenin secretion levels and a significant increase in bFGF were observed following light exposure, compared to dark conditions, in both normoxia and hypoxia conditions. With the addition of a blue light-blocking filter in normoxia, a significant increase in angiogenin levels was observed. Although statistical significance was not achieved, blue light filters reduce light-induced secretion of bFGF and VEGF to near normal levels. This trend is also observed when ARPE-19 cells are grown under hypoxic conditions and when pre-treated with lutein prior to exposure to experimental conditions. Following light exposure, there is a decrease in angiogenin secretion by ARPE-19 cells, which was abrogated with a blue light - blocking filter. Our findings support the position that blue light filtering affects the secretion of angiogenic factors by retinal pigmented epithelial cells under normoxic, hypoxic, and lutein

  6. Aging and senescence of skin cells in culture

    DEFF Research Database (Denmark)

    Rattan, Suresh

    2015-01-01

    Studying age-related changes in the physiology, biochemistry, and molecular biology of isolated skin cell populations in culture has greatly expanded the understanding of the fundamental aspects of skin aging. The three main cell types that have been studied extensively with respect to cellular...... aging in vitro are dermal fibroblasts, epidermal keratinocytes, and melanocytes. Serial subcultivation of normal diploid skin cells can be performed only a limited number of times, and the emerging senescent phenotype can be categorized into structural, physiological, biochemical, and molecular...... phenotypes, which can be used as biomarkers of cellular aging in vitro. The rate and phenotype of aging are different in different cell types. There are both common features and specific features of aging of skin fibroblasts, keratinocytes, melanocytes, and other cell types. A progressive accumulation...

  7. Propagation and isolation of ranaviruses in cell culture

    DEFF Research Database (Denmark)

    Ariel, Ellen; Nicolajsen, Nicole; Christophersen, Maj-Britt

    2009-01-01

    The optimal in vitro propagation procedure for a panel of ranavirus isolates and the best method for isolation of Epizootic haematopoietic necrosis virus (EHNV) from organ material in cell-culture were investigated. The panel of ranavirus isolates included: Frog virus 3 (FV3), Bohle iridovirus (BIV......), Pike-perch iridovirus (PPIV), European catfish virus (ECV), European sheatfish virus (ESV), EHNV, Doctor fish virus (DFV), Guppy virus 6 (GF6), short-finned eel virus (SERV) and Rana esculenta virus Italy 282/102 (REV 282/102). Each isolate was titrated in five cell lines: bluegill fry (BF-2......), epithelioma papulosum cyprini (EPC), chinook salmon embryo (CHSE-214) rainbow trout gonad (RTG-2) and fathead minnow (FHM), and incubated at 10, 15, 20, 24 and 28 °C for two weeks. BF-2, EPC and CHSE-214 cells performed well and titers obtained in the three cell lines were similar, whereas FHM and RTG-2 cells...

  8. 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

  9. Neuronal differentiation of hair-follicle-bulge-derived stem cells co-cultured with mouse cochlear modiolus explants.

    Directory of Open Access Journals (Sweden)

    Timo Schomann

    Full Text Available Stem-cell-based repair of auditory neurons may represent an attractive therapeutic option to restore sensorineural hearing loss. Hair-follicle-bulge-derived stem cells (HFBSCs are promising candidates for this type of therapy, because they (1 have migratory properties, enabling migration after transplantation, (2 can differentiate into sensory neurons and glial cells, and (3 can easily be harvested in relatively high numbers. However, HFBSCs have never been used for this purpose. We hypothesized that HFBSCs can be used for cell-based repair of the auditory nerve and we have examined their migration and incorporation into cochlear modiolus explants and their subsequent differentiation. Modiolus explants obtained from adult wild-type mice were cultured in the presence of EF1α-copGFP-transduced HFBSCs, constitutively expressing copepod green fluorescent protein (copGFP. Also, modiolus explants without hair cells were co-cultured with DCX-copGFP-transduced HFBSCs, which demonstrate copGFP upon doublecortin expression during neuronal differentiation. Velocity of HFBSC migration towards modiolus explants was calculated, and after two weeks, co-cultures were fixed and processed for immunohistochemical staining. EF1α-copGFP HFBSC migration velocity was fast: 80.5 ± 6.1 μm/h. After arrival in the explant, the cells formed a fascicular pattern and changed their phenotype into an ATOH1-positive neuronal cell type. DCX-copGFP HFBSCs became green-fluorescent after integration into the explants, confirming neuronal differentiation of the cells. These results show that HFBSC-derived neuronal progenitors are migratory and can integrate into cochlear modiolus explants, while adapting their phenotype depending on this micro-environment. Thus, HFBSCs show potential to be employed in cell-based therapies for auditory nerve repair.

  10. A microfluidic cell culture array with various oxygen tensions.

    Science.gov (United States)

    Peng, Chien-Chung; Liao, Wei-Hao; Chen, Ying-Hua; Wu, Chueh-Yu; Tung, Yi-Chung

    2013-08-21

    Oxygen tension plays an important role in regulating various cellular functions in both normal physiology and disease states. Therefore, drug testing using conventional in vitro cell models under normoxia often possesses limited prediction capability. A traditional method of setting an oxygen tension in a liquid medium is by saturating it with a gas mixture at the desired level of oxygen, which requires bulky gas cylinders, sophisticated control, and tedious interconnections. Moreover, only a single oxygen tension can be tested at the same time. In this paper, we develop a microfluidic cell culture array platform capable of performing cell culture and drug testing under various oxygen tensions simultaneously. The device is fabricated using an elastomeric material, polydimethylsiloxane (PDMS) and the well-developed multi-layer soft lithography (MSL) technique. The prototype device has 4 × 4 wells, arranged in the same dimensions as a conventional 96-well plate, for cell culture. The oxygen tensions are controlled by spatially confined oxygen scavenging chemical reactions underneath the wells using microfluidics. The platform takes advantage of microfluidic phenomena while exhibiting the combinatorial diversities achieved by microarrays. Importantly, the platform is compatible with existing cell incubators and high-throughput instruments (liquid handling systems and plate readers) for cost-effective setup and straightforward operation. Utilizing the developed platform, we successfully perform drug testing using an anti-cancer drug, triapazamine (TPZ), on adenocarcinomic human alveolar basal epithelial cell line (A549) under three oxygen tensions ranging from 1.4% to normoxia. The developed platform is promising to provide a more meaningful in vitro cell model for various biomedical applications while maintaining desired high throughput capabilities.

  11. Evaluation of the osteogenic differentiation of gingiva-derived stem cells grown on culture plates or in stem cell spheroids: Comparison of two- and three-dimensional cultures.

    Science.gov (United States)

    Lee, Sung-Il; Ko, Youngkyung; Park, Jun-Beom

    2017-09-01

    Three-dimensional cell culture systems provide a convenient in vitro model for the study of complex cell-cell and cell-matrix interactions in the absence of exogenous substrates. The current study aimed to evaluate the osteogenic differentiation potential of gingiva-derived stem cells cultured in two-dimensional or three-dimensional systems. To the best of our knowledge, the present study is the first to compare the growth of gingiva-derived stem cells in monolayer culture to a three-dimensional culture system with microwells. For three-dimensional culture, gingiva-derived stem cells were isolated and seeded into polydimethylsiloxane-based concave micromolds. Alkaline phosphatase activity and alizarin red S staining assays were then performed to evaluate osteogenesis and the degree of mineralization, respectively. Stem cell spheroids had a significantly increased level of alkaline phosphatase activity and mineralization compared with cells from the two-dimensional culture. In addition, an increase in mineralized deposits was observed with an increase in the loading cell number. The results of present study indicate that gingiva-derived stem cell spheroids exhibit an increased osteogenic potential compared with stem cells from two-dimensional culture. This highlights the potential of three-dimensional culture systems using gingiva-derived stem cells for regenerative medicine applications requiring stem cells with osteogenic potential.

  12. Differentiation of Human Pluripotent Stem Cells into Functional Endothelial Cells in Scalable Suspension Culture

    Directory of Open Access Journals (Sweden)

    Ruth Olmer

    2018-05-01

    Full Text Available Summary: Endothelial cells (ECs are involved in a variety of cellular responses. As multifunctional components of vascular structures, endothelial (progenitor cells have been utilized in cellular therapies and are required as an important cellular component of engineered tissue constructs and in vitro disease models. Although primary ECs from different sources are readily isolated and expanded, cell quantity and quality in terms of functionality and karyotype stability is limited. ECs derived from human induced pluripotent stem cells (hiPSCs represent an alternative and potentially superior cell source, but traditional culture approaches and 2D differentiation protocols hardly allow for production of large cell numbers. Aiming at the production of ECs, we have developed a robust approach for efficient endothelial differentiation of hiPSCs in scalable suspension culture. The established protocol results in relevant numbers of ECs for regenerative approaches and industrial applications that show in vitro proliferation capacity and a high degree of chromosomal stability. : In this article, U. Martin and colleagues show the generation of hiPSC endothelial cells in scalable cultures in up to 100 mL culture volume. The generated ECs show in vitro proliferation capacity and a high degree of chromosomal stability after in vitro expansion. The established protocol allows to generate hiPSC-derived ECs in relevant numbers for regenerative approaches. Keywords: hiPSC differentiation, endothelial cells, scalable culture

  13. Single molecule microscopy in 3D cell cultures and tissues.

    Science.gov (United States)

    Lauer, Florian M; Kaemmerer, Elke; Meckel, Tobias

    2014-12-15

    From the onset of the first microscopic visualization of single fluorescent molecules in living cells at the beginning of this century, to the present, almost routine application of single molecule microscopy, the method has well-proven its ability to contribute unmatched detailed insight into the heterogeneous and dynamic molecular world life is composed of. Except for investigations on bacteria and yeast, almost the entire story of success is based on studies on adherent mammalian 2D cell cultures. However, despite this continuous progress, the technique was not able to keep pace with the move of the cell biology community to adapt 3D cell culture models for basic research, regenerative medicine, or drug development and screening. In this review, we will summarize the progress, which only recently allowed for the application of single molecule microscopy to 3D cell systems and give an overview of the technical advances that led to it. While initially posing a challenge, we finally conclude that relevant 3D cell models will become an integral part of the on-going success of single molecule microscopy. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Calcium exchange, structure, and function in cultured adult myocardial cells

    International Nuclear Information System (INIS)

    Langer, G.A.; Frank, J.S.; Rich, T.L.; Orner, F.B.

    1987-01-01

    Cells digested from adult rat heart and cultured for 14 days demonstrate all the structural elements, in mature form, associated with the process of excitation-contraction (EC) coupling. The transverse tubular (TT) system is well developed with an extensive junctional sarcoplasmic reticulum (JSR). In nonphosphate-containing buffer contraction of the cells is lost as rapidly as zero extracellular Ca concentration ([Ca] 0 ) solution is applied and a negative contraction staircase is produced on increase of stimulation frequency. Structurally and functionally the cells have the characteristics of adult cells in situ. 45 Ca exchange and total 45 Ca measurement in N-2-hydroxyethylpiperazine N'-2-ethanesulfonic acid (HEPES)-buffered perfusate define three components of cellular Ca: 1) a rapidly exchangeable component accounting for 36% of total Ca, 2) a slowly exchangeable component (t/sub 1/2/ 53 min) accounting for 7% total Ca, and 3) the remaining 57% cellular Ca is inexchangeable (demonstrates no significant exchange within 60 min). The slowly exchangeable component can be increased 10-fold within 60 min by addition of phosphate to the perfusate. The Ca distribution and exchange characteristics are little different from those of 3-day cultures of neonatal rat heart previously studied. The results suggest that the cells are representative of adult cells in situ and that both sarcolemmal-bound and sarcoplasmic reticular Ca contribute to the component of Ca that is rapidly exchangeable

  15. Diffusion chamber culture of mouse bone marrow cells, (1)

    International Nuclear Information System (INIS)

    Sigeta, Chiharu; Tanaka, Kimio; Kawakami, Masahito; Takahashi, Hiroshi; Ohkita, Takeshi

    1980-01-01

    Mouse bone marrow cells were cultured in diffusion chambers (DC) implanted in the peritoneal cavity of host mice. Host mice were subjected to (1) irradiation ( 60 Co 800 rad) and/or (2) phenylhydrazine induced anemia and then receiving irradiation ( 60 Co 600 rad). After culture periods of 3-7 days, the total number of cells in DC was increased. A marked increase in DC is due to the proliferation of granulocyte series. When host mice were subjected to anemia and irradiation, the start of cell proliferation in DC was delay about two days. On the whole, anemia and irradiation host reduced a little cell growth in DC. The number of immature granulocytes grown in DC in irradiated hosts or anemia and irradiated hosts increased and reached a plateu at day 5. During the plateu period, the proportions between immature and mature granulocytes in DC were kept constantly. The number of macrophages showed a two-phase increasing. Erythroid cells and lymphocytes rapidly disappeared from the chambers during 3 days. The number of erythroid cells was not significantly influenced even in anemia and irradiation hosts. (author)

  16. Clonal variation in proliferation rate of cultures of GPK cells.

    Science.gov (United States)

    Riley, P A; Hola, M

    1981-09-01

    Pedigrees of twenty-six clones of a line of keratocytes derived from guinea-pig ear epidermis (GPK cells) were analysed from time-lapse film. The mean interdivision time (IDT) for the culture was 1143 +/- 215 (SD) min. The mean generation rates (mean reciprocal interdivision times) of clones varied over a range of 3.93--10.2 x 10(-4)/min and the standard deviation of the clonal mean generation rates was 16.8% of the average value. Transient intraclonal variations in IDT due to mitoses in a plane perpendicular to the substratum were observed. The data were also analysed on the basis of cell location in sixteen equal zones (quadrats) of the filmed area. The mean generation rate of quadrats was 8.73 x 10(-4)/min (SD = 4.9%). The spatial distribution showed some clustering of cells. The mean local density of the clones (2.25 +/- 0.62 cells/10(-4) cm2) was significantly higher than the quadrat density (1.76 +/- 0.8 cells/10(-4) cm2). There was no significant correlation between clonal density and mean generation rates, whereas for quadrats a significant negative correlation was found (P = 2.7%). The results support the proposition that cell lineage is the major determinant of the proliferation rate of subconfluent cultures.

  17. Vulnerability of cultured canine lung tumor cells to NK cell-mediated cytolysis

    International Nuclear Information System (INIS)

    Haley, P.J.; Kohr, J.M.; Kelly, G.; Muggenburg, B.A.; Guilmette, B.A.

    1988-01-01

    Five cell lines, designated as canine lung epithelial cell (CLEP), derived from radiation induced canine lung tumors and canine thyroid adeno-carcinoma (CTAC) cells were compared for their susceptibility to NK cell-mediated cytolysis using peripheral blood lymphocytes from normal, healthy Beagle dogs as effector cells. Effector cells and chromium 51 radiolabeled target cells were incubated for 16 h at ratios of 12.5:1, 25:1, 50:1, and 100:1. Increasing cytolysis was observed for all cell lines as the effector-to-target-cell ratios increased from 12.5:1 to 100:1. The percent cytotoxicity was significantly less for all lung tumor cell lines as compared to CTAC at the 100:1 ratio. One lung tumor cell line, CLEP-9, had 85% of the lytic vulnerability of the CTAC cell line and significantly greater susceptibility to NK cell-mediated lysis than all of the other lung tumor cell lines. Susceptibility to NK cell cytolysis did not correlate with in vivo malignant behavior of the original tumor. These data suggest that cultured canine lung tumor cells are susceptible to NK cell cytolytic activity in vitro and that at least one of these cell lines (CLEP-9) is a candidate for substitution of the standard canine NK cell target, CTAC, in NK cell assays. The use of lung tumor cells in NK cell assays may provide greater insight into the control of lung tumors by immune mechanisms. (author)

  18. Alginate/PEG based microcarriers with cleavable crosslinkage for expansion and non-invasive harvest of human umbilical cord blood mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Chunge [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Qian, Yufeng [Department of Chemistry and Biochemistry, University of Texas at Austin, 2500 Speedway, Austin, TX 78712 (United States); Zhao, Shuang [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Yin, Yuji, E-mail: yinyuji@tju.edu.cn [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Li, Junjie, E-mail: li41308@tju.edu.cn [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Department of Advanced Interdisciplinary Studies, Institute of Basic Medical Sciences and Tissue Engineering Research Center, Academy of Military Medical Sciences, No. 27, Taiping Road, Beijing 100850 (China)

    2016-07-01

    Porous microcarriers are increasingly used to expand and harvest stem cells. Generally, the cells are harvested via proteolytic enzyme treatment, which always leads to damages to stem cells. To address this disadvantage, a series of alginate/PEG (AL/PEG) semi-interpenetrating network microcarriers are prepared in this study. In this AL/PEG system, the chemically cross-linked alginate networks are formed via the reaction between carboxylic acid group of alginate and di-terminated amine groups of cystamine. PEG is introduced to modulate the degradation of microcarriers, which does not participate in this cross-linked reaction, while it interpenetrates in alginate network via physical interactions. In addition, chitosan are coated on the surface of AL/PEG to improve the mechanical strength via the electrostatic interactions. Biocompatible fibronectin are also coated on these microcarriers to modulate the biological behaviors of cells seeded in microcarriers. Results suggest that the size of AL/PEG microcarriers can be modulated via adjusting the contents and molecular weight of PEG. Moreover, the microcarriers are designed to be degraded with cleavage of disulfide crosslinkage. By changing the type and concentration of reductant, the ratio of AL to PEG, and the magnitude of chitosan coating, the degradation ability of AL/PEG microcarriers can be well controlled. In addition, AL/PEG microcarriers can support the attachment and proliferation of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs). More importantly, the expanded hUCB-MSCs can be detached from microcarriers after addition of reductant, which indeed reduce the cell damage caused by proteolytic enzyme treatment. Therefore, it is convinced that AL/PEG based microcarriers will be a promising candidate for large-scale expansion of hUCB-MSCs. - Graphical abstract: Alginate/PEG IPN microcarriers can support the attachment and expansion of hUCB-MSCs. More importantly, the expanded cells can be harvested

  19. Alginate/PEG based microcarriers with cleavable crosslinkage for expansion and non-invasive harvest of human umbilical cord blood mesenchymal stem cells

    International Nuclear Information System (INIS)

    Li, Chunge; Qian, Yufeng; Zhao, Shuang; Yin, Yuji; Li, Junjie

    2016-01-01

    Porous microcarriers are increasingly used to expand and harvest stem cells. Generally, the cells are harvested via proteolytic enzyme treatment, which always leads to damages to stem cells. To address this disadvantage, a series of alginate/PEG (AL/PEG) semi-interpenetrating network microcarriers are prepared in this study. In this AL/PEG system, the chemically cross-linked alginate networks are formed via the reaction between carboxylic acid group of alginate and di-terminated amine groups of cystamine. PEG is introduced to modulate the degradation of microcarriers, which does not participate in this cross-linked reaction, while it interpenetrates in alginate network via physical interactions. In addition, chitosan are coated on the surface of AL/PEG to improve the mechanical strength via the electrostatic interactions. Biocompatible fibronectin are also coated on these microcarriers to modulate the biological behaviors of cells seeded in microcarriers. Results suggest that the size of AL/PEG microcarriers can be modulated via adjusting the contents and molecular weight of PEG. Moreover, the microcarriers are designed to be degraded with cleavage of disulfide crosslinkage. By changing the type and concentration of reductant, the ratio of AL to PEG, and the magnitude of chitosan coating, the degradation ability of AL/PEG microcarriers can be well controlled. In addition, AL/PEG microcarriers can support the attachment and proliferation of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs). More importantly, the expanded hUCB-MSCs can be detached from microcarriers after addition of reductant, which indeed reduce the cell damage caused by proteolytic enzyme treatment. Therefore, it is convinced that AL/PEG based microcarriers will be a promising candidate for large-scale expansion of hUCB-MSCs. - Graphical abstract: Alginate/PEG IPN microcarriers can support the attachment and expansion of hUCB-MSCs. More importantly, the expanded cells can be harvested

  20. Peptide Hydrogelation and Cell Encapsulation for 3D Culture of MCF-7 Breast Cancer Cells

    Science.gov (United States)

    Sun, Xiuzhi S.; Nguyen, Thu A.

    2013-01-01

    Three-dimensional (3D) cell culture plays an invaluable role in tumor biology by providing in vivo like microenviroment and responses to therapeutic agents. Among many established 3D scaffolds, hydrogels demonstrate a distinct property as matrics for 3D cell culture. Most of the existing pre-gel solutions are limited under physiological conditions such as undesirable pH or temperature. Here, we report a peptide hydrogel that shows superior physiological properties as an in vitro matrix for 3D cell culture. The 3D matrix can be accomplished by mixing a self-assembling peptide directly with a cell culture medium without any pH or temperature adjustment. Results of dynamic rheological studies showed that this hydrogel can be delivered multiple times via pipetting without permanently destroying the hydrogel architecture, indicating the deformability and remodeling ability of the hydrogel. Human epithelial cancer cells, MCF-7, are encapsulated homogeneously in the hydrogel matrix during hydrogelation. Compared with two-dimensional (2D) monolayer culture, cells residing in the hydrogel matrix grow as tumor-like clusters in 3D formation. Relevant parameters related to cell morphology, survival, proliferation, and apoptosis were analyzed using MCF-7 cells in 3D hydrogels. Interestingly, treatment of cisplatin, an anti-cancer drug, can cause a significant decrease of cell viability of MCF-7 clusters in hydrogels. The responses to cisplatin were dose- and time-dependent, indicating the potential usage of hydrogels for drug testing. Results of confocal microscopy and Western blotting showed that cells isolated from hydrogels are suitable for downstream proteomic analysis. The results provided evidence that this peptide hydrogel is a promising 3D cell culture material for drug testing. PMID:23527204

  1. Ultra-thin Polyethylene glycol Coatings for Stem Cell Culture

    Science.gov (United States)

    Schmitt, Samantha K.

    Human mesenchymal stem cells (hMSCs) are a widely accessible and a clinically relevant cell type that are having a transformative impact on regenerative medicine. However, current clinical expansion methods can lead to selective changes in hMSC phenotype resulting from relatively undefined cell culture surfaces. Chemically defined synthetic surfaces can aid in understanding stem cell behavior. In particular we have developed chemically defined ultra-thin coatings that are stable over timeframes relevant to differentiation of hMSCs (several weeks). The approach employs synthesis of a copolymer with distinct chemistry in solution before application to a substrate. This provides wide compositional flexibility and allows for characterization of the orthogonal crosslinking and peptide binding groups. Characterization is done in solution by proton NMR and after crosslinking by X-ray photoelectron spectroscopy (XPS). The solubility of the copolymer in ethanol and low temperature crosslinking, expands its applicability to plastic substrates, in addition to silicon, glass, and gold. Cell adhesive peptides, namely Arg-Gly-Asp (RGD) fragments, are coupled to coating via different chemistries resulting in the urethane, amide or the thioester polymer-peptide bonds. Development of azlactone-based chemistry allowed for coupling in water at low peptide concentrations and resulted in either an amide or thioester bonds, depending on reactants. Characterization of the peptide functionalized coating by XPS, infrared spectroscopy and cell culture assays, showed that the amide linkages can present peptides for multiple weeks, while shorter-term presentation of a few days is possible using the more labile thioester bond. Regardless, coatings promoted initial adhesion and spreading of hMSCs in a peptide density dependent manner. These coatings address the following challenges in chemically defined cell culture simultaneously: (i) substrate adaptability, (ii) scalability over large areas

  2. Tumor necrosis factor (cachetin) decreases adipose cell differentiation in primary cell culture

    International Nuclear Information System (INIS)

    Martin, R.J.; Jones, D.D.; Jewell, D.E.; Hausman, G.J.

    1986-01-01

    Cachetin has been shown to effect gene product expression in the established adipose cell line 3T3-L1. Expression of messenger RNA for lipoprotein lipase is suppressed in cultured adipocytes. The purpose of this study was to determine the effect of Cachetin on adipose cell differentiation in primary cell culture. Stromalvascular cells obtained from the inguinal fat pad of 4-5 week old Sprague-Dawley rats were grown in culture for two weeks. During the proliferative growth phase all cells were grown on the same medium and labelled with 3 H-thymidine. Cachetin treatment (10 -6 to 10 -10 M) was initiated on day 5, the initial phase of preadipocyte differentiation. Adipocytes and stromal cells were separated using density gradient, and 3 H-thymidine was determined for both cell types. Thymidine incorporation into adipose cells was decreased maximally (∼ 50%) at 10 -10 M. Stromalvascular cells were not influenced at any of the doses tested. Adipose cell lipid content as indicated by oil red-O staining was decreased by Cachetin. Esterase staining by adipose cells treated with Cachetin was increased indicating an increase in intracellular lipase. These studies show that Cachetin has specific effects on primary adipose cell differentiation

  3. Isolation of human monocytes by double gradient centrifugation and their differentiation to macrophages in teflon-coated cell culture bags.

    Science.gov (United States)

    Menck, Kerstin; Behme, Daniel; Pantke, Mathias; Reiling, Norbert; Binder, Claudia; Pukrop, Tobias; Klemm, Florian

    2014-09-09

    Human macrophages are involved in a plethora of pathologic processes ranging from infectious diseases to cancer. Thus they pose a valuable tool to understand the underlying mechanisms of these diseases. We therefore present a straightforward protocol for the isolation of human monocytes from buffy coats, followed by a differentiation procedure which results in high macrophage yields. The technique relies mostly on commonly available lab equipment and thus provides a cost and time effective way to obtain large quantities of human macrophages. Briefly, buffy coats from healthy blood donors are subjected to a double density gradient centrifugation to harvest monocytes from the peripheral blood. These monocytes are then cultured in fluorinated ethylene propylene (FEP) Teflon-coated cell culture bags in the presence of macrophage colony-stimulating factor (M-CSF). The differentiated macrophages can be easily harvested and used for subsequent studies and functional assays. Important methods for quality control and validation of the isolation and differentiation steps will be highlighted within the protocol. In summary, the protocol described here enables scientists to routinely and reproducibly isolate human macrophages without the need for cost intensive tools. Furthermore, disease models can be studied in a syngeneic human system circumventing the use of murine macrophages.

  4. Micro fluidic System for Culturing and Monitoring of Neuronal Cells and Tissue

    DEFF Research Database (Denmark)

    Bakmand, Tanya; Waagepetersen, Helle S.

    The aim of this Ph.D. project was to combine experience within cell and tissue culturing, electrochemistry and microfabrication in order to develop an in vivo-like fluidic culturing platform, challenging the traditional culturing methods. The first goal was to develope a fluidic system for cultur...... with mass production. The last part of this thesis also includes perspectives on how to expand the latest designed device to facilitate culturing of tissue and co-culturing of cells....

  5. Radiation-induced bystander effects in cultured human stem cells.

    Directory of Open Access Journals (Sweden)

    Mykyta V Sokolov

    2010-12-01

    Full Text Available The radiation-induced "bystander effect" (RIBE was shown to occur in a number of experimental systems both in vitro and in vivo as a result of exposure to ionizing radiation (IR. RIBE manifests itself by intercellular communication from irradiated cells to non-irradiated cells which may cause DNA damage and eventual death in these bystander cells. It is known that human stem cells (hSC are ultimately involved in numerous crucial biological processes such as embryologic development; maintenance of normal homeostasis; aging; and aging-related pathologies such as cancerogenesis and other diseases. However, very little is known about radiation-induced bystander effect in hSC. To mechanistically interrogate RIBE responses and to gain novel insights into RIBE specifically in hSC compartment, both medium transfer and cell co-culture bystander protocols were employed.Human bone-marrow mesenchymal stem cells (hMSC and embryonic stem cells (hESC were irradiated with doses 0.2 Gy, 2 Gy and 10 Gy of X-rays, allowed to recover either for 1 hr or 24 hr. Then conditioned medium was collected and transferred to non-irradiated hSC for time course studies. In addition, irradiated hMSC were labeled with a vital CMRA dye and co-cultured with non-irradiated bystander hMSC. The medium transfer data showed no evidence for RIBE either in hMSC and hESC by the criteria of induction of DNA damage and for apoptotic cell death compared to non-irradiated cells (p>0.05. A lack of robust RIBE was also demonstrated in hMSC co-cultured with irradiated cells (p>0.05.These data indicate that hSC might not be susceptible to damaging effects of RIBE signaling compared to differentiated adult human somatic cells as shown previously. This finding could have profound implications in a field of radiation biology/oncology, in evaluating radiation risk of IR exposures, and for the safety and efficacy of hSC regenerative-based therapies.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1984-01-01

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

  7. Isolation and culture of Celosia cristata L cell suspension protoplasts

    Directory of Open Access Journals (Sweden)

    Retno Mastuti

    2003-06-01

    Full Text Available Developmental competence of Celosia cristata L. cell suspension-derived protoplasts was investigated. The protoplasts were isolatedfrom 3- to 9-d old cultures in enzyme solution containing 2% (w/v Cellulase YC and 0.5% (w/v Macerozyme R-10 which was dissolvedin washing solution (0.4 M mannitol and 10 mM CaCl2 at pH 5.6 for 3 hours. The highest number of viable protoplasts was releasedfrom 5-d old culture of a homogenous cell suspension. Subsequently, three kinds of protoplast culture media were simultaneously examinedwith four kinds of concentration of gelling agent. Culturing the protoplasts on KM8p medium solidified with 1.2% agarose significantlyenhanced plating efficiency as well as microcolony formation. Afterwards, the microcalli actively proliferated into friable watery calluswhen they were subcultured on MS medium supplemented with 0.3 mg/l 2,4-D and 1.0 mg/l kinetin. Although the plant regenerationfrom the protoplasts-derived calli has not yet been obtained, the reproducible developmental step from protoplasts to callus in thisstudy may facilitate the establishment of somatic hybridization using C. cristata as one parent.

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

    Science.gov (United States)

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

    2012-01-01

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

  9. [Biological characteristics of mesenchymal stem cell and hematopoietic stem cell in the co-culture system].

    Science.gov (United States)

    Wei, Wei; Xu, Chao; Ye, Zhi-Yong; Huang, Xiao-Jun; Yuan, Jia-En; Ma, Tian-Bao; Lin, Han-Biao; Chen, Xiu-Qiong

    2016-10-25

    The aim of the present study was to obtain the qualified hematopoietic stem/progenitor cells (HSC/HPC) and human umbilical cord-mesenchymal stem cells (MSC) in vitro in the co-culture system. Cord blood mononuclear cells were separated from umbilical cord blood by Ficoll lymphocyte separation medium, and then CD34 + HSC was collected by MACS immunomagnetic beads. The selected CD34 + HSC/HPC and MSC were transferred into culture flask. IMDM culture medium with 15% AB-type cord plasma supplemented with interleukin-3 (IL-3), IL-6, thrombopoietin (TPO), stem cell factor (SCF) and FMS-like tyrosine kinase 3 ligand (Flt-3L) factors were used as the co-culture system for the amplification of HSC/HPC and MSC. The cellular growth status and proliferation on day 6 and 10 after co-culture were observed by using inverted microscope. The percentage of positive expression of CD34 in HSC/HPC, as well as the percentages of positive expressions of CD105, CD90, CD73, CD45, CD34 and HLA-DR in the 4 th generation MSC, was tested by flow cytometry. Semisolid colony culture was used to test the HSC/HPC colony forming ability. The osteogenic, chondrogenesis and adipogenic ability of the 4 th generation MSC were assessed. The karyotype analysis of MSC was conducted by colchicines. The results demonstrated that the HSC/HPC of co-culture group showed higher ability of amplification, CFU-GM and higher CD34 + percentage compared with the control group. The co-cultured MSC maintained the ability to differentiate into bone cells, fat cells and chondrocytes. And the karyotype stability of MSC remained normal. These results reveal that the appropriate co-culture system for MSC and HSC is developed, and via this co-culture system we could gain both two kinds of these cells. The MSCs under the co-culture system maintain the biological characteristics. The CFU-GM ability, cell counting and the flow cytometry results of HSC/HPC under the co-culture system are conform to the criterion, showing that

  10. Hydrolyzed fish proteins modulates both inflammatory and antioxidant gene expression as well as protein expression in a co culture model of liver and head kidney cells isolated from Atlantic salmon (Salmo salar).

    Science.gov (United States)

    Holen, Elisabeth; He, Juyun; Araujo, Pedro; Seliussen, Jørgen; Espe, Marit

    2016-07-01

    Hydrolyzed fish proteins (H-pro) contain high concentrations of free amino acids and low molecular peptides that potentially may benefit fish health. The following study aimed to test whether the water-soluble phase of H-pro could attenuate lipopolysaccharide (LPS) provoked inflammation in liver cells and head kidney cells isolated from Atlantic salmon. Cells were grown as mono cultures or co cultures to assess possible crosstalk between immune cells and metabolic cells during treatments. Cells were added media with or without H-pro for 2 days before LPS exposure and harvested 24 h post LPS exposure. Respective cells without H-pro and LPS were used as controls. H-pro alone could affect expression of proteins directly as H-pro increased catalase protein expression in head kidney- and liver cells, regardless of culturing methods and LPS treatment. Leukotriene B4 (LTB4) production was also increased by H-pro in head kidney cells co cultured with liver cells. H-pro increased LPS induced interleukin 1β (IL-1β) transcription in liver cells co cultured with head kidney cells. All cultures of head kidney cells showed a significant increase in IL-1β transcription when treated with H-pro + LPS. H-pro decreased caspase-3 transcription in liver cells cultured co cultured with head kidney cells. Peroxisome proliferator activated receptor α (PPAR α) was upregulated, regardless of treatment, in liver cells co cultured with head kidney cells clearly showing that culturing method alone affected gene transcription. H-pro alone and together with LPS as an inflammation inducer, affect both antioxidant and inflammatory responses. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Enhanced chondrocyte culture and growth on biologically inspired nanofibrous cell culture dishes.

    Science.gov (United States)

    Bhardwaj, Garima; Webster, Thomas J

    2016-01-01

    Chondral and osteochondral defects affect a large number of people in which treatment options are currently limited. Due to its ability to mimic the natural nanofibrous structure of cartilage, this current in vitro study aimed at introducing a new scaffold, called XanoMatrix™, for cartilage regeneration. In addition, this same scaffold is introduced here as a new substrate onto which to study chondrocyte functions. Current studies on chondrocyte functions are limited due to nonbiologically inspired cell culture substrates. With its polyethylene terephthalate and cellulose acetate composition, good mechanical properties and nanofibrous structure resembling an extracellular matrix, XanoMatrix offers an ideal surface for chondrocyte growth and proliferation. This current study demonstrated that the XanoMatrix scaffolds promote chondrocyte growth and proliferation as compared with the Corning and Falcon surfaces normally used for chondrocyte cell culture. The XanoMatrix scaffolds also have greater hydrophobicity, three-dimensional surface area, and greater tensile strength, making them ideal candidates for alternative treatment options for chondral and osteochondral defects as well as cell culture substrates to study chondrocyte functions.

  12. Cell division requirement for activation of murine leukemia virus in cell culture by irradiation

    International Nuclear Information System (INIS)

    Otten, J.A.; Quarles, J.M.; Tennant, R.W.

    1976-01-01

    Actively dividing cultures of AKR mouse cells were exposed to relatively low dose-rates of γ radiation and tested for activation of endogenous leukemia viruses. Efficient and reproducible induction of virus was obtained with actively dividing cells, but cultures deprived of serum to inhibit cell division before and during γ irradiation were not activated, even when medium with serum was added immediately after irradiation. These results show that cell division was required for virus induction but that a stable intermediate similar to the state induced by halogenated pyrimidines was not formed. In actively dividing AKR cell cultures, virus activation appeared to be proportional to the dose of γ radiation; the estimated frequency of activation was 1-8 x 10 - 5 per exposed cell and the efficiency of activation was approximately 0.012 inductions per cell per rad. Other normal primary and established mouse cell cultures tested were not activated by γ radiation. The requirement of cell division for radiation and chemical activation may reflect some common mechanism for initiation of virus expression

  13. Mesenchymal stem cells cultured on magnetic nanowire substrates

    Science.gov (United States)

    Perez, Jose E.; Ravasi, Timothy; Kosel, Jürgen

    2017-02-01

    Stem cells have been shown to respond to extracellular mechanical stimuli by regulating their fate through the activation of specific signaling pathways. In this work, an array of iron nanowires (NWs) aligned perpendicularly to the surface was fabricated by pulsed electrodepositon in porous alumina templates followed by a partial removal of the alumina to reveal 2-3 μm of the NWs. This resulted in alumina substrates with densely arranged NWs of 33 nm in diameter separated by 100 nm. The substrates were characterized by scanning electron microscopy (SEM) energy dispersive x-ray analysis and vibrating sample magnetometer. The NW array was then used as a platform for the culture of human mesenchymal stem cells (hMSCs). The cells were stained for the cell nucleus and actin filaments, as well as immuno-stained for the focal adhesion protein vinculin, and then observed by fluorescence microscopy in order to characterize their spreading behavior. Calcein AM/ethidium homodimer-1 staining allowed the determination of cell viability. The interface between the cells and the NWs was studied using SEM. Results showed that hMSCs underwent a re-organization of actin filaments that translated into a change from an elongated to a spherical cell shape. Actin filaments and vinculin accumulated in bundles, suggesting the attachment and formation of focal adhesion points of the cells on the NWs. Though the overall number of cells attached on the NWs was lower compared to the control, the attached cells maintained a high viability (>90%) for up to 6 d. Analysis of the interface between the NWs and the cells confirmed the re-organization of F-actin and revealed the adhesion points of the cells on the NWs. Additionally, a net of filopodia surrounded each cell, suggesting the probing of the array to find additional adhesion points. The cells maintained their round shape for up to 6 d of culture. Overall, the NW array is a promising nanostructured platform for studying and influencing h

  14. Mesenchymal stem cells cultured on magnetic nanowire substrates

    KAUST Repository

    Perez, Jose E.

    2016-12-28

    Stem cells have been shown to respond to extracellular mechanical stimuli by regulating their fate through the activation of specific signaling pathways. In this work, an array of iron nanowires (NWs) aligned perpendicularly to the surface was fabricated by pulsed electrodepositon in porous alumina templates followed by a partial removal of the alumina to reveal 2-3 μm of the NWs. This resulted in alumina substrates with densely arranged NWs of 33 nm in diameter separated by 100 nm. The substrates were characterized by scanning electron microscopy (SEM) energy dispersive x-ray analysis and vibrating sample magnetometer. The NW array was then used as a platform for the culture of human mesenchymal stem cells (hMSCs). The cells were stained for the cell nucleus and actin filaments, as well as immuno-stained for the focal adhesion protein vinculin, and then observed by fluorescence microscopy in order to characterize their spreading behavior. Calcein AM/ethidium homodimer-1 staining allowed the determination of cell viability. The interface between the cells and the NWs was studied using SEM. Results showed that hMSCs underwent a re-organization of actin filaments that translated into a change from an elongated to a spherical cell shape. Actin filaments and vinculin accumulated in bundles, suggesting the attachment and formation of focal adhesion points of the cells on the NWs. Though the overall number of cells attached on the NWs was lower compared to the control, the attached cells maintained a high viability (>90%) for up to 6 d. Analysis of the interface between the NWs and the cells confirmed the re-organization of F-actin and revealed the adhesion points of the cells on the NWs. Additionally, a net of filopodia surrounded each cell, suggesting the probing of the array to find additional adhesion points. The cells maintained their round shape for up to 6 d of culture. Overall, the NW array is a promising nanostructured platform for studying and influencing h

  15. New 2-arylbenzofuran metabolite from cell cultures of Morus alba.

    Science.gov (United States)

    Zhang, De-Wu; Tao, Xiao-Yu; Yu, Li-Yan; Dai, Jun-Gui

    2015-01-01

    A new 2-arylbenzofuran compound, 5-dehydroxy-moracin U (1), along with 10 known compounds (2-11), were isolated from cell cultures of Morus alba. Their structures were elucidated on the basis of extensive spectroscopic analyses. The anti-inflammatory activity assay of 1-8 showed that 2 and 8 exhibited significant inhibitory effect on LPS-induced NO production with the values of 76.4% and 98.7% at 10(- 5) M, respectively.

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

    Science.gov (United States)

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

    2017-08-01

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

  17. Photodamage of the cells in culture sensitized with bilirubin

    Science.gov (United States)

    Kozlenkova, O. A.; Plavskaya, L. G.; Mikulich, A. V.; Leusenko, I. A.; Tretyakova, A. I.; Plavskii, V. Yu

    2016-08-01

    It has been shown that exposure to radiation of LED sources of light with an emission band maximum at about 465 and 520 nm having substantially identical damaging effects on animal cells in culture, that are in a logarithmic growth phase and preincubated with pigment. Photobiological effect is caused by photodynamic processes involving singlet oxygen generated by triplet excited sensitizer. Mono-exponential type dependence of cell survival on the energy dose indicates that it is bilirubin that acts as a sensitizer but not its photoproducts. The inclusion of bilirubin in the cells, where it is primarily localized in the mitochondria cells, it is accompanied by multiple amplification photochemical stability compared to pigment molecules bound with albumin

  18. Neural differentiation of adipose-derived stem cells by indirect co-culture with Schwann cells

    Directory of Open Access Journals (Sweden)

    Li Xiaojie

    2009-01-01

    Full Text Available To investigate whether adipose-derived stem cells (ADSCs could be subject to neural differentiation induced only by Schwann cell (SC factors, we co-cultured ADSCs and SCs in transwell culture dishes. Immunoassaying, Western blot analysis, and RT-PCR were performed (1, 3, 7, 14 d and the co-cultured ADSCs showed gene and protein expression of S-100, Nestin, and GFAP. Further, qRT-PCR disclosed relative quantitative differences in the above three gene expressions. We think ADSCs can undergo induced neural differentiation by being co-cultured with SCs, and such differentia­tions begin 1 day after co-culture, become apparent after 7 days, and thereafter remain stable till the 14th day.

  19. Computerized microfluidic cell culture using elastomeric channels and Braille displays.

    Science.gov (United States)

    Gu, Wei; Zhu, Xiaoyue; Futai, Nobuyuki; Cho, Brenda S; Takayama, Shuichi

    2004-11-09

    Computer-controlled microfluidics would advance many types of cellular assays and microscale tissue engineering studies wherever spatiotemporal changes in fluidics need to be defined. However, this goal has been elusive because of the limited availability of integrated, programmable pumps and valves. This paper demonstrates how a refreshable Braille display, with its grid of 320 vertically moving pins, can power integrated pumps and valves through localized deformations of channel networks within elastic silicone rubber. The resulting computerized fluidic control is able to switch among: (i) rapid and efficient mixing between streams, (ii) multiple laminar flows with minimal mixing between streams, and (iii) segmented plug-flow of immiscible fluids within the same channel architecture. The same control method is used to precisely seed cells, compartmentalize them into distinct subpopulations through channel reconfiguration, and culture each cell subpopulation for up to 3 weeks under perfusion. These reliable microscale cell cultures showed gradients of cellular behavior from C2C12 myoblasts along channel lengths, as well as differences in cell density of undifferentiated myoblasts and differentiation patterns, both programmable through different flow rates of serum-containing media. This technology will allow future microscale tissue or cell studies to be more accessible, especially for high-throughput, complex, and long-term experiments. The microfluidic actuation method described is versatile and computer programmable, yet simple, well packaged, and portable enough for personal use.

  20. Cell cultures for schistosomes - Chances of success or wishful thinking?

    Science.gov (United States)

    Quack, T; Wippersteg, V; Grevelding, C G

    2010-08-01

    Due to their worldwide importance for human and animal health, schistosomes are in the focus of national and international research activities. Their aims are to elucidate the genome, the transcriptome, the proteome and the glycome of schistosomes with the expectation to understand the biology of these blood flukes and to identify new candidate antigens for the development of a vaccine, or target molecules for the design of novel pharmaceutical compounds. All of these efforts have delivered a vast amount of information about the genetic equipment of schistosomes. In the emerging era of post-genomic research, however, methods and tools are necessary to interpret all available data and to characterise molecules of interest in more detail. In addition to transgenesis, it is generally accepted that cell lines for schistosomes are among the requirements to overcome present research limitations. In our commentary the prospect of establishing cell cultures for schistosomes is discussed. To this end we summarise the comprehensive endeavours made in the past regarding the establishment of invertebrate cell lines pointing to critical parameters that should be considered when making new attempts towards schistosome cell culturing. Furthermore, based on preliminary data with pilot-character, we discuss recent advances indicating the possibility of overcoming existing restrictions with respect to the 'immortalisation' of cells by oncogenes. Copyright 2010 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

  1. Agent-Based Computational Modeling of Cell Culture ...

    Science.gov (United States)

    Quantitative characterization of cellular dose in vitro is needed for alignment of doses in vitro and in vivo. We used the agent-based software, CompuCell3D (CC3D), to provide a stochastic description of cell growth in culture. The model was configured so that isolated cells assumed a “fried egg shape” but became increasingly cuboidal with increasing confluency. The surface area presented by each cell to the overlying medium varies from cell-to-cell and is a determinant of diffusional flux of toxicant from the medium into the cell. Thus, dose varies among cells for a given concentration of toxicant in the medium. Computer code describing diffusion of H2O2 from medium into each cell and clearance of H2O2 was calibrated against H2O2 time-course data (25, 50, or 75 uM H2O2 for 60 min) obtained with the Amplex Red assay for the medium and the H2O2-sensitive fluorescent reporter, HyPer, for cytosol. Cellular H2O2 concentrations peaked at about 5 min and were near baseline by 10 min. The model predicted a skewed distribution of surface areas, with between cell variation usually 2 fold or less. Predicted variability in cellular dose was in rough agreement with the variation in the HyPer data. These results are preliminary, as the model was not calibrated to the morphology of a specific cell type. Future work will involve morphology model calibration against human bronchial epithelial (BEAS-2B) cells. Our results show, however, the potential of agent-based modeling

  2. CDB-4124 does not cause apoptosis in cultured fibroid cells.

    Science.gov (United States)

    Roeder, Hilary; Jayes, Friederike; Feng, Liping; Leppert, Phyllis C

    2011-09-01

    Selective progesterone receptor modulators (SPRMs), such as asoprisnil (J867) and ulipristal (CDB-2914), have been shown to reduce fibroid volume in vivo and to induce apoptosis in vitro. CDB-4124 (telapristone), a SPRM with different side groups, also reduced fibroid volume in vivo, and we hypothesized that this SPRM would also cause apoptosis in cultured fibroid cells. Immortalized, progesterone receptor-positive fibroid cells, known to be capable of apoptosis, were grown to 80% confluence in serum-containing media. Cells were then treated for 48 hours in serum-free media with 0, 10, 100, or 1000 nmol/L CDB-4124. Actinomycin-D and staurosporine were used as positive controls to induce apoptosis. Apoptosis was quantified using a TUNEL-fluorescein kit. Images were captured with a widefield-fluorescence microscope and analyzed using MetaMorph image analysis software. To validate results, Western blots of total cell lysates were probed for cleaved caspase-3 (c-CASP3). Experiments were repeated 3 times using independent cell batches. Analysis of 19 712 nuclei indicated 14.8% ± 10.9% (mean ± SEM), 8.4% ± 4.6%, 8.2% ± 4.7%, and 9.3% ± 6.3% apoptosis in 0, 10, 100, and 1000 nmol/L CDB-4124-treated cells, respectively. There was no evidence of elevated c-CASP3 over vehicle control after treatment with CDB-4124. CDB-4124 did not significantly induce apoptosis in cultured fibroid cells under the conditions described suggesting apoptosis may not be the main pathway responsible for CDB-4124-induced fibroid shrinkage. Variations in SPRM biological effects may be due to differences in fibroid source cells, binding kinetics, or extracellular matrix characteristics, and can be exploited in further investigations of the mechanisms of action of SPRMs in fibroid biology.

  3. Light transfer in agar immobilized microalgae cell cultures

    Science.gov (United States)

    Kandilian, Razmig; Jesus, Bruno; Legrand, Jack; Pilon, Laurent; Pruvost, Jérémy

    2017-09-01

    This paper experimentally and theoretically investigates light transfer in agar-immobilized cell cultures. Certain biotechnological applications such as production of metabolites secreted by photosynthetic microorganisms require cells to be immobilized in biopolymers to minimize contamination and to facilitate metabolite recovery. In such applications, light absorption by cells is one of the most important parameters affecting cell growth or metabolite productivity. Modeling light transfer therein can aid design and optimize immobilized-cell reactors. In this study, Parachlorella kessleri cells with areal biomass concentrations ranging from 0.36 to 16.9 g/m2 were immobilized in 2.6 mm thick agar gels. The average absorption and scattering cross-sections as well as the scattering phase function of P. kessleri cells were measured. Then, the absorption and transport scattering coefficients of the agar gel were determined using an inverse method based on the modified two-flux approximation. The forward model was used to predict the normal-hemispherical transmittance and reflectance of the immobilized-cell films accounting for absorption and scattering by both microalgae and the agar gel. Good agreement was found between the measured and predicted normal-hemispherical transmittance and reflectance provided absorption and scattering by agar were taken into account. Moreover, good agreement was found between experimentally measured and predicted mean rate of photon absorption. Finally, optimal areal biomass concentration was determined to achieve complete absorption of the incident radiation.

  4. Flow field measurements in the cell culture unit

    Science.gov (United States)

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

    2002-01-01

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

  5. Nonviral Gene Delivery of Growth and Differentiation Factor 5 to Human Mesenchymal Stem Cells Injected into a 3D Bovine Intervertebral Disc Organ Culture System

    Directory of Open Access Journals (Sweden)

    Christian Bucher

    2013-01-01

    Full Text Available Intervertebral disc (IVD cell therapy with unconditioned 2D expanded mesenchymal stem cells (MSC is a promising concept yet challenging to realize. Differentiation of MSCs by nonviral gene delivery of growth and differentiation factor 5 (GDF5 by electroporation mediated gene transfer could be an excellent source for cell transplantation. Human MSCs were harvested from bone marrow aspirate and GDF5 gene transfer was achieved by in vitro electroporation. Transfected cells were cultured as monolayers and as 3D cultures in 1.2% alginate bead culture. MSC expressed GDF5 efficiently for up to 21 days. The combination of GDF5 gene transfer and 3D culture in alginate showed an upregulation of aggrecan and SOX9, two markers for chondrogenesis, and KRT19 as a marker for discogenesis compared to untransfected cells. The cells encapsulated in alginate produced more proteoglycans expressed in GAG/DNA ratio. Furthermore, GDF5 transfected MCS injected into an IVD papain degeneration organ culture model showed a partial recovery of the GAG/DNA ratio after 7 days. In this study we demonstrate the potential of GDF5 transfected MSC as a promising approach for clinical translation for disc regeneration.

  6. Feeder Cell Type Affects the Growth of In Vitro Cultured Bovine Trophoblast Cells

    Directory of Open Access Journals (Sweden)

    Islam M. Saadeldin

    2017-01-01

    Full Text Available Trophectoderm cells are the foremost embryonic cells to differentiate with prospective stem-cell properties. In the current study, we aimed at improving the current approach for trophoblast culture by using granulosa cells as feeders. Porcine granulosa cells (PGCs compared to the conventional mouse embryonic fibroblasts (MEFs were used to grow trophectoderm cells from hatched bovine blastocysts. Isolated trophectoderm cells were monitored and displayed characteristic epithelial/cuboidal morphology. The isolated trophectoderm cells expressed mRNA of homeobox protein (CDX2, cytokeratin-8 (KRT8, and interferon tau (IFNT. The expression level was higher on PGCs compared to MEFs throughout the study. In addition, primary trophectoderm cell colonies grew faster on PGCs, with a doubling time of approximately 48 hrs, compared to MEFs. PGCs feeders produced a fair amount of 17β-estradiol and progesterone. We speculated that the supplementation of sex steroids and still-unknown factors during the trophoblasts coculture on PGCs have helped to have better trophectoderm cell’s growth than on MEFs. This is the first time to use PGCs as feeders to culture trophectoderm cells and it proved superior to MEFs. We propose PGCs as alternative feeders for long-term culture of bovine trophectoderm cells. This model will potentially benefit studies on the early trophoblast and embryonic development in bovines.

  7. 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.

  8. Culturing bone marrow cells with dexamethasone and ascorbic acid improves osteogenic cell sheet structure.

    Science.gov (United States)

    Akahane, M; Shimizu, T; Kira, T; Onishi, T; Uchihara, Y; Imamura, T; Tanaka, Y

    2016-11-01

    To assess the structure and extracellular matrix molecule expression of osteogenic cell sheets created via culture in medium with both dexamethasone (Dex) and ascorbic acid phosphate (AscP) compared either Dex or AscP alone. Osteogenic cell sheets were prepared by culturing rat bone marrow stromal cells in a minimal essential medium (MEM), MEM with AscP, MEM with Dex, and MEM with Dex and AscP (Dex/AscP). The cell number and messenger (m)RNA expression were assessed in vitro, and the appearance of the cell sheets was observed after mechanical retrieval using a scraper. β-tricalcium phosphate (β-TCP) was then wrapped with the cell sheets from the four different groups and subcutaneously implanted into rats. After mechanical retrieval, the osteogenic cell sheets from the MEM, MEM with AscP, and MEM with Dex groups appeared to be fragmented or incomplete structures. The cell sheets cultured with Dex/AscP remained intact after mechanical retrieval, without any identifiable tears. Culture with Dex/AscP increased the mRNA and protein expression of extracellular matrix proteins and cell number compared with those of the other three groups. More bridging bone formation was observed after transplantation of the β-TCP scaffold wrapped with cell sheets cultured with Dex/AscP, than in the other groups. These results suggest that culture with Dex/AscP improves the mechanical integrity of the osteogenic cell sheets, allowing retrieval of the confluent cells in a single cell sheet structure. This method may be beneficial when applied in cases of difficult tissue reconstruction, such as nonunion, bone defects, and osteonecrosis.Cite this article: M. Akahane, T. Shimizu, T. Kira, T. Onishi, Y. Uchihara, T. Imamura, Y. Tanaka. Culturing bone marrow cells with dexamethasone and ascorbic acid improves osteogenic cell sheet structure. Bone Joint Res 2016;5:569-576. DOI: 10.1302/2046-3758.511.BJR-2016-0013.R1. © 2016 Akahane et al.

  9. Distribution of AAV8 particles in cell lysates and culture media changes with time and is dependent on the recombinant vector

    Directory of Open Access Journals (Sweden)

    Bryan A Piras

    2016-01-01

    Full Text Available With clinical trials ongoing, efficient clinical production of adeno-associated virus (AAV to treat large numbers of patients remains a challenge. We compared distribution of AAV8 packaged with Factor VIII (FVIII in cell culture media and lysates on days 3, 5, 6, and 7 post-transfection and found increasing viral production through day 6, with the proportion of viral particles in the media increasing from 76% at day 3 to 94% by day 7. Compared to FVIII, AAV8 packaged with Factor IX and Protective Protein/Cathepsin A vectors demonstrated a greater shift from lysate towards media from day 3 to 6, implying that particle distribution is dependent on recombinant vector. Larger-scale productions showed that the ratio of full-to-empty AAV particles is similar in media and lysate, and that AAV harvested on day 6 post-transfection provides equivalent function in mice compared to AAV harvested on day 3. This demonstrates that AAV8 production can be optimized by prolonging the duration of culture post-transfection, and simplified by allowing harvest of media only, with disposal of cells that contain 10% or less of total vector yield. Additionally, the difference in particle distribution with different expression cassettes implies a recombinant vector-dependent processing mechanism which should be taken into account during process development.

  10. Is cell culture a risky business? Risk analysis based on scientist survey data.

    Science.gov (United States)

    Shannon, Mark; Capes-Davis, Amanda; Eggington, Elaine; Georghiou, Ronnie; Huschtscha, Lily I; Moy, Elsa; Power, Melinda; Reddel, Roger R; Arthur, Jonathan W

    2016-02-01

    Cell culture is a technique that requires vigilance from the researcher. Common cell culture problems, including contamination with microorganisms or cells from other cultures, can place the reliability and reproducibility of cell culture work at risk. Here we use survey data, contributed by research scientists based in Australia and New Zealand, to assess common cell culture risks and how these risks are managed in practice. Respondents show that sharing of cell lines between laboratories continues to be widespread. Arrangements for mycoplasma and authentication testing are increasingly in place, although scientists are often uncertain how to perform authentication testing. Additional risks are identified for preparation of frozen stocks, storage and shipping. © 2015 UICC.

  11. External nitrogen input affects pre- and post-harvest cell wall composition but not the enzymatic saccharification of wheat straw

    DEFF Research Database (Denmark)

    Baldwin, Laetitia Andrée; Glazowska, Sylwia Emilia; Mravec, Jozef

    2017-01-01

    Wheat is one of the most important crops for food and feed and its straw is a potential feedstock for biorefinery purposes. Nitrogen (N) is an essential input factor in wheat agriculture but no information is available on how it affects straw composition during maturation and at harvest....... To investigate this, we conducted a large scale field experiment in which wheat plants were cultivated at three levels of externally applied N. The plants were harvested at different stages of maturation, spanning green straw at heading (ear emergence) to fully yellow straw at final maturity. Defined parts...

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

    International Nuclear Information System (INIS)

    Hay, R.J.; Phillips, T.; Thompson, A.; Vilner, L.; Cleland, M.; Tchaw-ren Chen; Zabrenetzky, V.

    1999-01-01

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

  13. Leucine supplementation attenuates macrophage foam-cell formation: Studies in humans, mice, and cultured macrophages.

    Science.gov (United States)

    Grajeda-Iglesias, Claudia; Rom, Oren; Hamoud, Shadi; Volkova, Nina; Hayek, Tony; Abu-Saleh, Niroz; Aviram, Michael

    2018-02-05

    Whereas atherogenicity of dietary lipids has been largely studied, relatively little is known about the possible contribution of dietary amino acids to macrophage foam-cell formation, a hallmark of early atherogenesis. Recently, we showed that leucine has antiatherogenic properties in the macrophage model system. In this study, an in-depth investigation of the role of leucine in macrophage lipid metabolism was conducted by supplementing humans, mice, or cultured macrophages with leucine. Macrophage incubation with serum obtained from healthy adults supplemented with leucine (5 g/d, 3 weeks) significantly decreased cellular cholesterol mass by inhibiting the rate of cholesterol biosynthesis and increasing cholesterol efflux from macrophages. Similarly, leucine supplementation to C57BL/6 mice (8 weeks) resulted in decreased cholesterol content in their harvested peritoneal macrophages (MPM) in relation with reduced cholesterol biosynthesis rate. Studies in J774A.1 murine macrophages revealed that leucine dose-dependently decreased cellular cholesterol and triglyceride mass. Macrophages treated with leucine (0.2 mM) showed attenuated uptake of very low-density lipoproteins and triglyceride biosynthesis rate, with a concurrent down-regulation of diacylglycerol acyltransferase-1, a key enzyme catalyzing triglyceride biosynthesis in macrophages. Similar effects were observed when macrophages were treated with α-ketoisocaproate, a key leucine metabolite. Finally, both in vivo and in vitro leucine supplementation significantly improved macrophage mitochondrial respiration and ATP production. The above studies, conducted in human, mice, and cultured macrophages, highlight a protective role for leucine attenuating macrophage foam-cell formation by mechanisms related to the metabolism of cholesterol, triglycerides, and energy production. © 2018 BioFactors, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

  14. Bags versus flasks: a comparison of cell culture systems for the production of dendritic cell-based immunotherapies.

    Science.gov (United States)

    Fekete, Natalie; Béland, Ariane V; Campbell, Katie; Clark, Sarah L; Hoesli, Corinne A

    2018-04-19

    In recent years, cell-based therapies targeting the immune system have emerged as promising strategies for cancer treatment. This review summarizes manufacturing challenges related to production of antigen presenting cells as a patient-tailored cancer therapy. Understanding cell-material interactions is essential because in vitro cell culture manipulations to obtain mature antigen-producing cells can significantly alter their in vivo performance. Traditional antigen-producing cell culture protocols often rely on cell adhesion to surface-treated hydrophilic polystyrene flasks. More recent commercial and investigational cancer immunotherapy products were manufactured using suspension cell culture in closed hydrophobic fluoropolymer bags. The shift to closed cell culture systems can decrease risks of contamination by individual operators, as well as facilitate scale-up and automation. Selecting closed cell culture bags over traditional open culture systems entails different handling procedures and processing controls, which can affect product quality. Changes in culture vessels also entail changes in vessel materials and geometry, which may alter the cell microenvironment and resulting cell fate decisions. Strategically designed culture systems will pave the way for the generation of more sophisticated and highly potent cell-based cancer vaccines. As an increasing number of cell-based therapies enter the clinic, the selection of appropriate cell culture vessels and materials becomes a critical consideration that can impact the therapeutic efficacy of the product, and hence clinical outcomes and patient quality of life. © 2018 The Authors Transfusion published by Wiley Periodicals, Inc. on behalf of AABB.

  15. Effects of 2,3-iminosqualene in cultured cells

    International Nuclear Information System (INIS)

    Popjak, G.; Meenan, A.; Nes, W.D.

    1987-01-01

    2,3-Iminosqualene added to culture media 10 ug/ml) of rat hepatoma (H4-II-E-C3) or Chinese hamster ovary (CHO) cells irreversibly inactivates the squalene-oxide: lanosterol cyclase, but it does not inhibit general polyprenyl synthesis either from [ 14 C]acetate or [ 14 C]mevalonate. Isq added to lipoprotein-containing media of H4 cells causes in 24 hr an over twofold rise in HMG-CoA reductase and abolishes the repressive effect of mevalonate (MVA) on the reductase. H4 cells synthesize from [2- 14 C]-MVA labelled squalene, squalene-2,3-oxide, squalene-2,3-22,23-dioxide, but very little sterol. The conversion of MVA to these polyprenyls in the presence of Isq is as efficient as its conversion to squalene and cholesterol in control cells. They conclude that the repressor of HMG-CoA reductase derived from MVA is a sterol - whatever might be the nature of that sterol - and not a nonsteroidal derivative of MVA metabolism. H4 cells exposed to Isq in lipid-depleted media die in 48-72 hr, but can be rescued by LDL, but not by free cholesterol or MVA. CHO cells are more resistant than H4 cells and succumb only after 8-9 days' exposure to Isq

  16. Establishment of automated culture system for murine induced pluripotent stem cells

    Directory of Open Access Journals (Sweden)

    Koike Hiroyuki

    2012-11-01

    Full Text Available Abstract Background Induced pluripotent stem (iPS cells can differentiate into any cell type, which makes them an attractive resource in fields such as regenerative medicine, drug screening, or in vitro toxicology. The most important prerequisite for these industrial applications is stable supply and uniform quality of iPS cells. Variation in quality largely results from differences in handling skills between operators in laboratories. To minimize these differences, establishment of an automated iPS cell culture system is necessary. Results We developed a standardized mouse iPS cell maintenance culture, using an automated cell culture system housed in a CO2 incubator commonly used in many laboratories. The iPS cells propagated in a chamber uniquely designed for automated culture and showed specific colony morphology, as for manual culture. A cell detachment device in the system passaged iPS cells automatically by dispersing colonies to single cells. In addition, iPS cells were passaged without any change in colony morphology or expression of undifferentiated stem cell markers during the 4 weeks of automated culture. Conclusions Our results show that use of this compact, automated cell culture system facilitates stable iPS cell culture without obvious effects on iPS cell pluripotency or colony-forming ability. The feasibility of iPS cell culture automation may greatly facilitate the use of this versatile cell source for a variety of biomedical applications.

  17. Digital microfluidics for automated hanging drop cell spheroid culture.

    Science.gov (United States)

    Aijian, Andrew P; Garrell, Robin L

    2015-06-01

    Cell spheroids are multicellular aggregates, grown in vitro, that mimic the three-dimensional morphology of physiological tissues. Although there are numerous benefits to using spheroids in cell-based assays, the adoption of spheroids in routine biomedical research has been limited, in part, by the tedious workflow associated with spheroid formation and analysis. Here we describe a digital microfluidic platform that has been developed to automate liquid-handling protocols for the formation, maintenance, and analysis of multicellular spheroids in hanging drop culture. We show that droplets of liquid can be added to and extracted from through-holes, or "wells," and fabricated in the bottom plate of a digital microfluidic device, enabling the formation and assaying of hanging drops. Using this digital microfluidic platform, spheroids of mouse mesenchymal stem cells were formed and maintained in situ for 72 h, exhibiting good viability (>90%) and size uniformity (% coefficient of variation <10% intraexperiment, <20% interexperiment). A proof-of-principle drug screen was performed on human colorectal adenocarcinoma spheroids to demonstrate the ability to recapitulate physiologically relevant phenomena such as insulin-induced drug resistance. With automatable and flexible liquid handling, and a wide range of in situ sample preparation and analysis capabilities, the digital microfluidic platform provides a viable tool for automating cell spheroid culture and analysis. © 2014 Society for Laboratory Automation and Screening.

  18. Cell thickness of UV absorption by the cell: relation to UV action spectrum shift in mammalian cells in culture

    International Nuclear Information System (INIS)

    Sakharov, V.H.; Voronkova, L.N.; Blokhin, A.V.

    1985-01-01

    By means of reconstruction of series half - thin transverse sections the three - dimensional morphometry of SPEV cells for a series of their specific states in culture is performed: for exponential growth in a monolayer, in a merged monolayer, in the mitosis phase, for giant cells and suspension cells. In the monolayer the cell thickness in its central part depended mainly on the nucleus thickness and in average changed but slightly despite a wide range of changes in volumes of nuclei and cells and their density in culture. The cell thickness has noticeably increased in mitosis. For the above states of cells UV radiation absorption spectra are determined. It is shown that a certain shift of action spectrus of death of mammalian cells as compared with that for bacterial cell can be a seguence of selfshielding and not differences in the nature of active chromophores

  19. Progress toward forecasting product quality and quantity of mammalian cell culture processes by performance-based modeling.

    Science.gov (United States)

    Schmidberger, Timo; Posch, Christoph; Sasse, Alexandra; Gülch, Carina; Huber, Robert

    2015-01-01

    The production of biopharmaceuticals requires highly sophisticated, complex cell based processes. Once a process has been developed, acceptable ranges for various control parameters are typically defined based on process characterization studies often comprising several dozens of small scale bioreactor cultivations. A lot of data is generated during these studies and usually only the information needed to define acceptable ranges is processed in more detail. Making use of the wealth of information contained in such data sets, we present here a methodology that uses performance data (such as metabolite profiles) to forecast the product quality and quantity of mammalian cell culture processes based on a toolbox of advanced statistical methods. With this performance based modeling (PBM) the final product concentration and 12 quality attributes (QAs) for two different biopharmaceutical products were predicted in daily intervals throughout the main stage process. The best forecast was achieved for product concentration in a very early phase of the process. Furthermore, some glycan isoforms were predicted with good accuracy several days before the bioreactor was harvested. Overall, PBM clearly demonstrated its capability of early process endpoint prediction by only using commonly available data, even though it was not possible to predict all QAs with the desired accuracy. Knowing the product quality prior to the harvest allows the manufacturer to take counter measures in case the forecasted quality or quantity deviates from what is expected. This would be a big step towards real-time release, an important element of the FDA's PAT initiative. © 2015 American Institute of Chemical Engineers.

  20. Glycosylation-mediated phenylpropanoid partitioning in Populus tremuloides cell cultures

    Directory of Open Access Journals (Sweden)

    Babst Benjamin A

    2009-12-01

    Full Text Available Abstract Background Phenylpropanoid-derived phenolic glycosides (PGs and condensed tannins (CTs comprise large, multi-purpose non-structural carbon sinks in Populus. A negative correlation between PG and CT concentrations has been observed in several studies. However, the molecular mechanism underlying the relationship is not known. Results Populus cell cultures produce CTs but not PGs under normal conditions. Feeding salicyl alcohol resulted in accumulation of salicins, the simplest PG, in the cells, but not higher-order PGs. Salicin accrual reflected the stimulation of a glycosylation response which altered a number of metabolic activities. We utilized this suspension cell feeding system as a model for analyzing the possible role of glycosylation in regulating the metabolic competition between PG formation, CT synthesis and growth. Cells accumulated salicins in a dose-dependent manner following salicyl alcohol feeding. Higher feeding levels led to a decrease in cellular CT concentrations (at 5 or 10 mM, and a negative effect on cell growth (at 10 mM. The competition between salicin and CT formation was reciprocal, and depended on the metabolic status of the cells. We analyzed gene expression changes between controls and cells fed with 5 mM salicyl alcohol for 48 hr, a time point when salicin accumulation was near maximum and CT synthesis was reduced, with no effect on growth. Several stress-responsive genes were up-regulated, suggestive of a general stress response in the fed cells. Salicyl alcohol feeding also induced expression of genes associated with sucrose catabolism, glycolysis and the Krebs cycle. Transcript levels of phenylalanine ammonia lyase and most of the flavonoid pathway genes were reduced, consistent with down-regulated CT synthesis. Conclusions Exogenous salicyl alcohol was readily glycosylated in Populus cell cultures, a process that altered sugar utilization and phenolic partitioning in the cells. Using this system, we

  1. Cell in situ zymography: an in vitro cytotechnology for localization of enzyme activity in cell culture.

    Science.gov (United States)

    Chhabra, Aastha; Jaiswal, Astha; Malhotra, Umang; Kohli, Shrey; Rani, Vibha

    2012-09-01

    In situ zymography is a unique technique for detection and localization of enzyme-substrate interactions majorly in histological sections. Substrate with quenched fluorogenic molecule is incorporated in gel over which tissue sections are mounted and then incubated in buffer. The enzymatic activity is observed in the form of fluorescent signal. With the advancements in the field of biological research, use of in vitro cell culture has become very popular and holds great significance in multiple fields including inflammation, cancer, stem cell biology and the still emerging 3-D cell cultures. The information on analysis of enzymatic activity in cell lines is inadequate presently. We propose a single-step methodology that is simple, sensitive, cost-effective, and functional to perform and study the 'in position' activity of enzyme on substrate for in vitro cell cultures. Quantification of enzymatic activity to carry out comparative studies on cells has also been illustrated. This technique can be applied to a variety of enzyme classes including proteases, amylases, xylanases, and cellulases in cell cultures.

  2. A Cell Culture Approach to Optimized Human Corneal Endothelial Cell Function

    Science.gov (United States)

    Bartakova, Alena; Kuzmenko, Olga; Alvarez-Delfin, Karen; Kunzevitzky, Noelia J.; Goldberg, Jeffrey L.

    2018-01-01

    Purpose Cell-based therapies to replace corneal endothelium depend on culture methods to optimize human corneal endothelial cell (HCEC) function and minimize endothelial-mesenchymal transition (EnMT). Here we explore contribution of low-mitogenic media on stabilization of phenotypes in vitro that mimic those of HCECs in vivo. Methods HCECs were isolated from cadaveric donor corneas and expanded in vitro, comparing continuous presence of exogenous growth factors (“proliferative media”) to media without those factors (“stabilizing media”). Identity based on canonical morphology and expression of surface marker CD56, and function based on formation of tight junction barriers measured by trans-endothelial electrical resistance assays (TEER) were assessed. Results Primary HCECs cultured in proliferative media underwent EnMT after three to four passages, becoming increasingly fibroblastic. Stabilizing the cells before each passage by switching them to a media low in mitogenic growth factors and serum preserved canonical morphology and yielded a higher number of cells. HCECs cultured in stabilizing media increased both expression of the identity marker CD56 and also tight junction monolayer integrity compared to cells cultured without stabilization. Conclusions HCECs isolated from donor corneas and expanded in vitro with a low-mitogenic media stabilizing step before each passage demonstrate more canonical structural and functional features and defer EnMT, increasing the number of passages and total canonical cell yield. This approach may facilitate development of HCEC-based cell therapies. PMID:29625488

  3. In vitro cell culture lethal dose submitted to gamma radiation

    Energy Technology Data Exchange (ETDEWEB)

    Moreno, Carolina S.; Rogero, Sizue O.; Rogero, Jose Roberto [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)], e-mail: carolina_sm@hotmail.com; Ikeda, Tamiko I.; Cruz, Aurea S. [Instituto Adolfo Lutz, Sao Paulo, SP (Brazil)

    2009-07-01

    The present study was designed to evaluate the in vitro effect of gamma radiation in cell culture of mouse connective tissue exposed to different doses of gamma radiation and under several conditions. The cell viability was analyzed by neutral red uptake methodology. This assay was developed for establish a methodology to be used in the future in the study of resveratrol radioprotection. Resveratrol (3,4',5- trihydroxystilbene), a phenolic phytoalexin that occurs naturally in some spermatophytes, such as grapevines, in response to injury as fungal infections and exposure to ultraviolet light. In the wines this compound is found at high levels and is considered one of the highest antioxidant constituents. The intense antioxidant potential of resveratrol provides many pharmacological activities including cardioprotection, chemoprevention and anti-tumor effects. Our results demonstrated that {sup 60}Co gamma radiation lethal dose (LD50) on NCTC clone 929 cells was about 340Gy. (author)

  4. In vitro cell culture lethal dose submitted to gamma radiation

    International Nuclear Information System (INIS)

    Moreno, Carolina S.; Rogero, Sizue O.; Rogero, Jose Roberto; Ikeda, Tamiko I.; Cruz, Aurea S.

    2009-01-01

    The present study was designed to evaluate the in vitro effect of gamma radiation in cell culture of mouse connective tissue exposed to different doses of gamma radiation and under several conditions. The cell viability was analyzed by neutral red uptake methodology. This assay was developed for establish a methodology to be used in the future in the study of resveratrol radioprotection. Resveratrol (3,4',5- trihydroxystilbene), a phenolic phytoalexin that occurs naturally in some spermatophytes, such as grapevines, in response to injury as fungal infections and exposure to ultraviolet light. In the wines this compound is found at high levels and is considered one of the highest antioxidant constituents. The intense antioxidant potential of resveratrol provides many pharmacological activities including cardioprotection, chemoprevention and anti-tumor effects. Our results demonstrated that 60 Co gamma radiation lethal dose (LD50) on NCTC clone 929 cells was about 340Gy. (author)

  5. Sulphur XANES Analysis of Cultured Human Prostate Cancer Cells

    International Nuclear Information System (INIS)

    Kwiatek, W.M.; Podgorczyk, M.; Paluszkiewicz, Cz.; Balerna, A.; Kisiel, A.

    2008-01-01

    Prostate cancer is one of the most commonly diagnosed cancers in men throughout the world. It is believed that changes to the structure of protein binding sites, altering its metabolism, may play an important role in carcinogenesis. Sulphur, often present in binding sites, can influence such changes through its chemical speciation. Hence there is a need for precise investigation of coordination environment of sulphur. X-ray absorption near edge structure spectroscopy offers such possibility. Cell culture samples offer histologically well defined areas of good homogeneity, suitable for successful and reliable X-ray absorption near edge structure analysis. This paper presents sulphur speciation data collected from three different human prostate cancer cell lines (PC-3, LNCaP and DU-145). Sulphur X-ray absorption near edge structure analysis was performed on K-edge structure. The spectra of cells were compared with those of cancerous tissue and with organic substances as well as inorganic compounds. (authors)

  6. Centering Single Cells in Microgels via Delayed Crosslinking Supports Long-Term 3D Culture by Preventing Cell Escape

    NARCIS (Netherlands)

    Kamperman, Tom; Henke, Sieger; Visser, Claas Willem; Karperien, Marcel; Leijten, Jeroen

    2017-01-01

    Single-cell-laden microgels support physiological 3D culture conditions while enabling straightforward handling and high-resolution readouts of individual cells. However, their widespread adoption for long-term cultures is limited by cell escape. In this work, it is demonstrated that cell escape is

  7. Optimization of Storage Temperature for Cultured ARPE-19 Cells

    Directory of Open Access Journals (Sweden)

    Lara Pasovic

    2013-01-01

    Full Text Available Purpose. The establishment of future retinal pigment epithelium (RPE replacement therapy is partly dependent on the availability of tissue-engineered RPE cells, which may be enhanced by the development of suitable storage methods for RPE. This study investigates the effect of different storage temperatures on the viability, morphology, and phenotype of cultured RPE. Methods. ARPE-19 cells were cultured under standard conditions and stored in HEPES-buffered MEM at nine temperatures (4°C, 8°C, 12°C, 16°C, 20°C, 24°C, 28°C, 32°C, and 37°C for seven days. Viability and phenotype were assessed by a microplate fluorometer and epifluorescence microscopy, while morphology was analyzed by scanning electron microscopy. Results. The percentage of viable cells preserved after storage was highest in the 16°C group (48.7%±9.8%; P<0.01 compared to 4°C, 8°C, and 24°C–37°C; P<0.05 compared to 12°C. Ultrastructure was best preserved at 12°C, 16°C, and 20°C. Expression of actin, ZO-1, PCNA, caspase-3, and RPE65 was maintained after storage at 16°C compared to control cells that were not stored. Conclusion. Out of nine temperatures tested between 4°C and 37°C, storage at 12°C, 16°C, and 20°C was optimal for maintenance of RPE cell viability, morphology, and phenotype. The preservation of RPE cells is critically dependent on storage temperature.

  8. Low temperature and Daphnia-associated infochemicals promote colony formation of Scenedesmus obliquus and its harvesting.

    Science.gov (United States)

    Zhu, Xuexia; Yang, Jingwen; Zhang, Xingxing; Zhang, Lu; Wang, Xiaojun; Huang, Yuan; Yang, Zhou

    2017-01-01

    To explore the combined effects of temperature and Daphnia-associated infochemicals on colony formation of Scenedesmus obliquus to faciliate harvesting the algal biomass. A three-parameter modified Gaussian model fitted the changes of the number of cells per particle in S. obliquus induced by Daphnia culture filtrate well under any temperature. Decreases in temperature enhanced the induced-colony formation of Scenedesmus. The maximum colony size at 15-25 °C was significantly larger than those at 30-35 °C. An additional 1 or 2 days at low temperature was needed to reach the maximum colony size, which indicates the best harvest time for algal biomass. Induced-colony formation of Scenedesmus by Daphnia culture filtrate at 15-25 °C is recommended to settle algal cells. This condition facilitates harvesting the biomass.

  9. Evaluation of a dental pulp-derived cell sheet cultured on amniotic membrane substrate.

    Science.gov (United States)

    Honjo, Ken-ichi; Yamamoto, Toshiro; Adachi, Tetsuya; Amemiya, Takeshi; Mazda, Osam; Kanamura, Narisato; Kita, Masakazu

    2015-01-01

    Mesenchymal stem cells (MSC) are transplanted for periodontal tissue regeneration, and the periodontal ligament (PDL) is regenerated using a cultured cell sheet. This cultured cell sheet is prepared using PDL-derived cells, growth factors, and amniotic membrane (AM). Dental pulp (DP)-derived cells can be easily obtained from extracted wisdom teeth, proliferate rapidly, and are less susceptible to bacterial infection than PDL-derived cells. Thus, to prepare a novel cell sheet, DP-derived cells were cultured on AM as a culture substrate for immunohistochemical examination. Wisdom teeth extracted from three adults were cut along the cement-enamel border. DP tissue was collected, minced, and primarily cultured. After three or four passage cultures, DP-derived cells were cultured on AM, followed by hematoxylin-eosin (H-E) and immunofluorescence staining. DP-derived cells cultured on AM formed a layered structure. Cells positive for vimentin, Ki-67, ZO-1, desmoplakin, CD29, 44, 105 or 146, STRO-1, collagen IV or VII or laminin 5 or α5 chain were localized. DP-derived cells proliferated on AM, while retaining the properties of DP, which allowed the cultured cell sheet to be prepared. In addition, the cultured cell sheet contained MSC, which suggests its potential application in periodontal tissue regeneration.

  10. Quantitation of DNA repair in brain cell cultures: implications for autoradiographic analysis of mixed cell populations

    International Nuclear Information System (INIS)

    Dambergs, R.; Kidson, C.

    1979-01-01

    Quantitation of DNA repair in the mixed cell population of mouse embryo brain cultures has been assessed by autoradiographic analysis of unscheduled DNA synthesis following UV-irradiation. The proportion of labelled neurons and the grain density over neuronal nuclei were both less than the cor