| Field | Specification |
|---|---|
| Mfr No | |
| Species |
Viruses: Tested negative for ectromelia virus (mouse pox).
- cultureMedium: RPMI 1640, w: 2.0 mM stable Glutamine, w: 2.0 g/L NaHCO3 (Cytion article number 820700a)
- supplements: Supplement the medium with 10% FBS
- dissociationReagent: Accutase
- subculturing: Gather the suspension cells in a 15 ml tube and gently wash the adherent cells with PBS lacking calcium and magnesium (use 3-5 ml for T25 flasks and 5-10 ml for T75 flasks). Apply Accutase (1-2 ml for T25 flasks, 2.5 ml for T75 flasks) ensuring full coverage of the cell layer. Allow the cells to incubate at room temperature for 10 minutes. Following incubation, combine and centrifuge both the suspension and adherent cells. After centrifugation, carefully resuspend the cell pellet and transfer the cell suspension into new flasks containing fresh medium.
- seedingDensity: Start new cultures at 4 x 105 cells/ml. The cell density should not exceed 2 x 106 cells/ml.
- fluidRenewal: Every 3 to 4 days. Collect floating cells, centrifuge and add to the flask together with fresh medium.
- postThawRecovery: After thawing, plate the cells at 5 x 104 cells/cm2 and allow the cells to recover from the freezing process and to adhere for at least 48 hours.
- freezeMedium: As a cryopreservation medium, use complete growth medium (including FBS) + 10% DMSO for adequate post-thaw viability, or CM-1 (Cytion catalog number 800100), which includes optimized osmoprotectants and metabolic stabilizers to enhance recovery and reduce cryo-induced stress.