Thawing 293FT cells

→ thaw the cells in prewarmed , complete medium without Geneticin

materials: - a vial of frozen cells from the liquid nitrogen - 37ºC waterbath - 75 cm2 flask - HS for 293 FT cells - pipetts

procedure:

1) remove the vial of frozen cells from liquid nitrogen and thaw quickly in a 37ºC waterbath 2) transfer the cells to a 75 cm2 flask containing 12 ml Hs for 293 FT cells w/o Geneticin 3) incubate the flask for 2 –4 hours to allow the cells to attach to the bottom of the flask 4) aspirate off the medium and replace with 12 ml of fresh, complete medium without Geneticin 5) incubate the cells over night 6) the next day, aspirate off the medium and replace with fresh, complete medium containing 500 µg/ ml Geneticin 7) incubate the cells and check them daily until the cells are 80 – 90% confluent 8) proceed to subculturing cells