Subculturing Jurkat Cell Line

Protocol

1. Transfer growing culture from T75 flask to a sterile 50 ml conical tube.
2. Centrifuge culture at ≤ 1,500 rpm for 2-3 minutes at room temperature.
3. Remove and discard 10 ml of old growth medium from above the cell pellet.
4. Add 10 ml of fresh complete growth medium and gently resuspend cell pellet by pipetting up and down 2-3 times.
5. Add 2 ml of the Jurkat cell suspension to a new sterile T75 flask containing 20 ml of complete growth medium and a final concentration of 1 mg/ml G418. For 75 cm² flasks, use approximately 10⁵ cells/flask and subculture every 3-4 days.
6. Place the culture vessel in a 5% CO₂, 37°C incubator.