Protocol for Subculturing the MCF-7 and MCF-7/VP cell lines
Cell culture solutions
Remember the cells are grown in complete Dulbecco’s Modified Eagle medium
(DMEM) that also contains 10% FCS+glutamine.
The cells require a mixture of Trypsin (0.5g/L), EDTA (197.16mg/L), and PBS (1L) that
are filter sterilized. This solution is used for adherent cell purposes during subculturing.
Adherent cell cultures
The cells in the T75 flask must have the present medium removed and following this, add
10ml of the Trypsin/EDTA mixture and wait for 10 minutes to detach cells. Place the
cells into the incubator during this time period.
Add 10ml of the complete DME medium into the flask and transfer the contents to a
centrifuge tube. Spin the cells at 1300-1500rpm for 5-7 minutes. Discard the supernatant
and resuspend the cells in 10-15ml of complete DMEM.
At this point, take 1ml sample of the cells and perform the cell viability analysis (Trypan
Blue dye exclusion assay) to get a density of approx. 1 x 105 cells. Once the concentration
of cells is established, take the respective volume of cells and dilute it with the
appropriate amount of complete DMEM in a new T75 flask.
NOTE: Remember to check the cells under the light microscope to see if the cells are
present before putting the cells into the incubator.