I have to extract DNA from 1.5 ml of fresh cancer cells (fibroblasts cultured in MEM), but before that i need to count the cells. I need to do both on the same day. My question is how long can i store cells in MEM in an eppendorf tube while i count cells on the side? Can i place the eppendorf back in incubator for the time being?
Note: I will be using 1.5 ml cell culture for DNA extraction and 100 micro litre from it will be used to count cells.
Hello Sabawoon Nisar,
Please use 100ul from the cell suspension for counting and centrifuge the remaining culture (1.4ml) at 700-800 rpm to pellet down the cells. Discard the supernatant and store the cell pellet in ice until you start the DNA extraction protocol. Cell counting should not take more than 10 mins.
Best.
I agree with Aakif Inayat's answer. The best way for counting to use the trypan blue 1% dye exclusion test. Doing this you can discriminate between living and dead cells. The former actively accumulate dye molecules in the cytoplasm, while the latter are not stained.
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