|Complete Growth Medium
||RPMI-1640 +0.1mM NEAA +1mM Sodium Pyruvate +2mM L-glutamine +10% FBS +1% P/S
||Remove and discard culture medium. Briefly rinse the cell layer with DPBS solution to remove all traces of serum that contains trypsin inhibitor.
Add 1.0 to 2.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 2 to 3 minutes). Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
Add 4.0 to 6.0 mL of complete growth medium and aspirate cells by gently pipetting. Add appropriate aliquots of the cell suspension to new culture vessels.
||Every 2 to 3 days
||Freeze medium: 60% Basal medium+30% FBS+10% DMSO
Storage temperature: Liquid nitrogen vapor phase
||Atmosphere: Air, 95%; CO2, 5%