Culturing HEK 293 Cells



Starting from a Kryotube:
Prewarm the medium at 37°C and fill two culture flasks (25 ml for 150 cm2, 5 ml for 25 cm2). Rapidly thaw the cells (at 37°C) and distribute them in two concentrations in the flasks.
Change the medium after 12 hrs or once the cells have attached.

Splitting Cells:
Aspirate the medium from the flask. Wash the cells carefully with PBS to remove residual medium. Add 1-2ml of Trypsin Solution (equilibrated at RT) to the flask (150 cm2) and incubate at 37°C until cells have detached (1-2 minutes). Prepare a new flask with fresh medium. Block trypsinization by adding a few ml of medium. Take a fraction of the cell solution and inocculate the new flask. Typically, when splitting confluent HEK 293 cells in a 1:10 ratio, confluency is reached again after 2-3 days.


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