Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.

DR-Wildtype cells, CRL-2977, are available for use as a control.

These cells lack the transcription factor CHOP, a key mediator of cell death in the unfolded protein response (UPR).

CHOP is a major regulator of the endoplasmic reticulum UPR and as such, these CHOP-KO cells are useful for studying the mechanisms of UPR and integrated stress response.

The mouse embryonic fibroblast (MEF) cell line, CHOP-KO-DR (CRL-2979), was established from a 13.5 day-old CHOP-/- mouse embryo by SV-40 immortalization.

C/EBP homologous protein (CHOP)/growth arrest and DNA damage-inducible gene 153 (GADD153), not expressed

C/EBP homologous protein (CHOP)/growth arrest and DNA damage-inducible gene 153 (GADD153), not expressed

Volumes used in this protocol are for 75 cm² flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

1. Remove and discard culture medium.
2. Briefly rinse the cell layer with Ca++/Mg++ free Dulbeccos phosphate-buffered saline (D-PBS) or 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.
3. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
   Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
5. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 2 X 10³ to 3 X 10³ viable cells/cm² is recommended.
6. Incubate cultures at 37°C.

Subcultivation ratio: A subcultivation ratio of 1:10 to 1:30 twice weekly is recommended.

Medium renewal: Every 2 to 3 days

Freeze medium: fetal bovine serum, 90%; DMSO, 10%

Storage temperature: liquid nitrogen vapor phase

Temperature: 37°C

Atmosphere: air, 95%; carbon dioxide (CO₂), 5%