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#Place flask in incubator at 37 °C with a 5% CO2 atmosphere for at least 30 min to equilibrate the medium. (Note: if the medium is clearly pink and likely has a pH >7.6, remake the medium; all manipulations and additions to cells are done with sterile tissue culture technique.)
#Enter the bar code of the cells into the cell line GUI.
#Remove Petri dishes containing cultured RAW 264.7 cells from the incubator. Gently pass a cell lifter over the surface of the dish to dislodgeany adherent cells into the medium. Prior to transferring the cell suspension, collect all possible cells by tipping the Petri dish at a 20-degree angle and gently streaming the medium over the cells using a 10-ml pipette. Avoid creating bubbles by not taking up and expelling air with medium in the pipettes. Several dishes of cultured cells may be pooled for counting.
#Transfer the single cell suspension to the vessel containing the medium originally removed from the Petri dish.
#Centrifuge the tube of cells at 400 x g for 5 min at 25 °C.
