Culture Method: Complete Growth Medium: MEMα (Gibco 12571-063) +10% FBS+ Penicillin/Streptomycin Subculturing: Volumes are given for a 25cm2 flask. Increase or decrease the amount of dissociation medium needed proportionally for culture vessels of other sizes. 1. Remove and discard culture medium. 2. Briefly rinse the cell layer with 0.25%(w/v) Trypsin-EDTA solution to remove all traces of serum that contains trypsin inhibitor. 3. Add 1.0 to 2.0mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed. 4. Add 6.0 to 8.0mL of complete growth medium and aspirate cells by gently pipetting. 5. Add appropriate aliquots of the cell suspension to new culture vessels. 6. Incubate cultures at 37°C, 5% CO2. Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:6 is recommended Cryopreservation: Freeze medium: Complete growth medium supplemented with 10% (v/v) DMSO Storage temperature: liquid nitrogen vapor phase