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ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
The cells spontaneously differentiate into embryonic structures in the absence of a feeder layer or conditioned medium.
Undifferentiated cells can be genetically modified by gene targeting techniques.
The cells spontaneously differentiate into embryonic structures in the absence of a feeder layer or conditioned medium. They can be maintained in the undifferentiated state by frequent subculture (every 2 to 3 days) on confluent feeder layers (STO cells) arrested with Mitomycin-C (see ATCC 56-X.2; MITC- STO cells).
Fibroblast-like feeder layer cells are present in the ampules sent by ATCC.
The base medium for this cell line is Mouse ES Cell Basal Medium (ATCC SCRR-2011). To make the complete medium add the following components to 500 mL base medium and mix by swirling gently:
Complete Growth Medium for Mouse ES Cells is stable for 14 days when stored at 2°C to 8°C.
56-X.2 cells should be seeded one day prior to use.
The medium to use when initiating the feeder layer is DMEM with 10% FBS. This medium is prepared by aseptically combining:
56 mL FBS (ATCC 30-2020)
500 mL DMEM (ATCC 30-2002)
Initiation of CRL-1934 Cell Culture
Subculture before the CRL-1934 colonies are close to or touching each other. The CRL-1934 cells should never become 100% confluent (although the 56-X.2 feeder cells may be 100% confluent). Attached cells are subcultured using 0.25% Trypsin 0.53 mM EDTA (ATCC 30-2101). The action of the 0.25% Trypsin – 0.53mM EDTA (ATCC 30-2101) is halted by adding culture medium to the detached cells. A split ratio of 1:3 to 1:10 is used when subculturing.
The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid. Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.
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Doetschman TC, et al. The in vitro development of blastocyst-derived embryonic stem cell lines: formation of visceral yolk sac, blood islands and myocardium. J. Embryol. Exp. Morphol. 87: 27-45, 1985. PubMed: 3897439
Williams RL, et al. Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells. Nature 336: 684-687, 1988. PubMed: 3143916
Gossler A, et al. Transgenesis by means of blastocyst-derived embryonic stem cell lines. Proc. Natl. Acad. Sci. USA 83: 9065-9069, 1986. PubMed: 3024164
Doetschman T, et al. Targeted mutation of the Hprt gene in mouse embryonic stem cells. Proc. Natl. Acad. Sci. USA 85: 8583-8587, 1988. PubMed: 3186749