Undifferentiated embryonic stem cell culture

For continuing undifferentiated embryonic stem cell cultures, usually different cell densities are plated, and 2 days later select those cultures with optimal morphology and density for subculture. Cultures should not contain cells with differentiated morphology density should be around 70-80% with ample evidence of exponential growth as observed by cell division in culture. This procedure allows that some variation in culture... 

Ouyang, A., Ng, R., and Yang, S.-T. 2007. Long-term culturing of undifferentiated embryonic stem cells in conditioned media and three-dimensional fibrous matrices without extracellular matrix coating. Stem Cells 25(2) 447-54. 

Complete medium for culture of the undifferentiated ES-D3 embryonic stem cell line Dulbecco s Modified Eagle s Medium (DMEM), 20% Fetal bovine serum (FBS), 1% Nonessential Amino Acids (NEAA) (Gibco, Gaithersburg, MD), 2 mM L-Glutamine, 1% 5,000 lU/ml Penicillin/5,000 pg/ml Streptomycin, 0.1 mM P-Mercaptoethanol. 

Embryonic stem cell The embryonic stem cell line is typically derived from a male agouti 129/terSv embryo. The cell line must be maintained with meticulous culture procedures to retain normal karyotype and an undifferentiated state. 

As long as embryonic stem cells in culture are grown under certain conditions, they can remain undifferentiated (unspecialized). But if cells are allowed to clump together to form embryoid bodies they begin to differentiate spontaneously into specific cell types (muscle cells, nerve cells, etc.). To generate cultures of specific types of differentiated cells, various growth factors are used in the culture media [45],... 

Human embryonic stem cells were first collected in 1998 by two different research teams. The cells obtained from the inner cell mass of the blastocyst (4- to 5-day embryo) are embryonic stem cells (ESC) in contrast, cells cultured from the primordial germ cells of 5- to 9-week fetuses are embryonic germ cells (EGC). In the laboratory, ES or EG cells can proliferate indefinitely in an undifferentiated state but can also be manipulated to become specialized or partially specialized cell types, a process known as directed differentiation. Both ES and EG cells are pluripotent, meaning they have the potential to develop into more than 200 different known cell types. This class of human stem cells holds the promise of being able to repair or replace cells or tissues that are damaged or destroyed by many of our most devastating diseases and disabilities. 

The EST has been developed with the aim to exploit the characteristics and differentiation potential of mouse embryonic stem cells (ES cells), established from the early embryo in 1981 [4, 5], ES cells are cultured in suspension to induce the formation of embryoid bodies, and afterwards they are transferred in 24-well dishes to allow attachment and differentiation in contracting cardiomyocytes. The toxicological endpoint is the inhibition of cardiac differentiation. In parallel a cytotoxicity test is performed on undifferentiated ES cells and a control somatic (fibroblast) cell line (3T3). The concentrations of testing chemicals that induce 50 % of differentiation inhibition (ID50) and 50 % cytotoxicity (IC50) in ES cells and 3T3 cells are inserted in a validated prediction model to classify the test chemical as non-embryotoxic, moderate, or strong embryotoxic [2, 6, 7], The validation of the method has been... 

Stem cells, whether derived from embryos, fetuses, or adults, can be simply defined as progeny of cells that are capable of differentiating into different lineages [152], Embryonic stem cells (ESCs) are isolated from the inner cell mass of blastocysts and have the ability to be cultured and maintained in an undifferentiated and pluripotent state, and directed to differentiate into all specific cell types [153,154], A variety of adult stem cells (often referred as progenitor or multipotent cells), including bone marrow-derived mesenchymal stem cells (MSCs), hematopoietic stem cells (HSCs), adipose-derived stem cells (ADSCs), and neutral stem cells (NSCs), have been found more committed but less pluripotent than ESCs. 

Cormier, J. T., Zur Nieden, N. I., Rancourt, D. E., and KaUos, M. S. 2006. Expansion of undifferentiated murine embryonic stem cells as aggregates in suspension culture bioreactors. Tissue Eng 12(11) 3233-45. 

Ameen, C., Strehl, R., Bjorquist, R, Lindahl, A., Hyllner, J., and Sartipy, R 2008. Human embryonic stem cells Current technologies and emerging industrial applications. Crit Rev Oncol Hematol, 65,54-80. Amit, M., Chebafh, J., Margulets, V., Laevsky, I., Miropolsky, Y., Shariki, K., Peri, M. et al. 2010. Suspension culture of undifferentiated human embryonic and induced pluripotent stem cells. Stem Cell Rev, 6,248-59. 

M. Amit, J. Chebath, V. Margulets, I. Laevsky, Y. Miropolsky, K. Shariki, M. Peri, I. Blais, G. Slutsky, M. Revel, and J. Itskovitz-Eldor, Suspension culture of undifferentiated human embryonic and induced pluripotent stem cells. Stem Cell Rev., 6 (2) 248-259, Jan. 2010. 

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