AFT024 IRR

These cells have been growth-arrested by irradiation with 12,000 rads.  The cells will begin to deteriorate 2 weeks after plating and may no longer support the growth of cells. We recommend that you do not keep the cells in culture for longer than 2 weeks.

It is recommended that the feeder cells be plated 24 hours before use at 5 to 6 X 10⁶ cells/T75 in order to obtain a 100% confluent monolayer for stem cells growth. Once the feeder cells have attached, the culture medium can be changed to accommodate the cells to be supported.

1. Check all containers for leakage or breakage.
2. Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below ­-130°C, preferably in liquid nitrogen vapor, until ready for use.

Flasks do not need to be coated before plating MEFs.

1. Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water. 
2. Remove the vial from the water bath as soon as the contents are half way thawed (approximately 90 seconds), and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.
3. Transfer the vial’s contents plus 5 mL of complete medium (see below for recipe) to a 15 mL centrifuge tube. Use an additional 1 mL of medium to rinse the vial and transfer the liquid to the 15 mL tube. Add 4 mL of complete medium to bring the total volume to 10 ml.
4. Gently mix and pellet the cells by centrifugation @ 270 x g for 5 minutes.
5. Discard the supernatant, resuspend the cells in fresh growth medium (warm), and transfer to the appropriate size flask.
6. Incubate 37°C in a 5% CO₂ in air atmosphere.
7. Fluid change twice a week or when pH decreases. It is important to avoid excessive alkalinity of the medium during recovery of the cells.  It is suggested that, prior to the addition of the vial contents, the culture vessel containing the growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6).

Cells should be plated 24 hours before use as a feeder layer for ES cells and kept for no more than 7 days.

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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