Plating with DMEM full media .

Warm DMEM full media, PBS, and Trypsin in the 37°C bead bath for 0h 30m 0s. Clean the working area by using 70% ethanol.

Sup out old media without touching cells.

Wash by adding 5mL PBS slowly, rinse, and rock back and forth.

Add 2mL-3mL trypsin (0.25%); keep in incubator for 0h 3m 0s.

Check under microscope if cells are detached, add 5mL media and transfer to a tube.

Spin 300x g for 0h 3m 0s.

Sup out and add 10mL fresh media & re-suspend cells gently and carefully.

Count cells density and split accordingly. 15,000 cells/ml for maintenance.

• (i) Usually 1.0-1.5 x10^4/ml cells for Biochem, and
• (ii) 0.5 x10^4/ml cells for IF.

Rest.

Replace with DMEM full media/Drug treat.

Drug treat if necessary /Harvesting.

Drug treat if necessary /Harvesting.