Hi everybody,
I have cultivated the C4-2 cell line in double charcoal stripped serum containing medium. A lot of the cells do not attach to the bottom of the flask. If someone has experience using he dextran coated charcoal stripped serum containing medium can explain do the cells attach to the bottom of flask like normal serum or not?
Since I have to refresh the medium after cultivation of the cells in double charcoal stripped containing serum , a lot of floated cells in this medium would be aspirated after refreshing the medium so the number of attached cells to the bottom of flask is important for my next experiment I would appreciate it if you have suggestion to solve this problem
This procedure is used to deplete the serum of estrogen and possibly other steroids and other small molecules. See e.g. here. https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma/Product_Information_Sheet/c6197pis.pdf
If it doesn't interfere with your experiment, attachment of cells might be improved by coating the dishes with gelatin, collagen, poly-lysine or fibronectin.
If you're just concerned about cell numbers, collecting the floating cells by centrifugation might do the trick, combined with counting alive and dead cells.
The difference is that the first medium contains serum that is depleted of androgen sex hormone. In the second case, the dextran coated charcoal is present in the medium. In both cases, the medium is depleted of androgens. Therefore, C4-2 cells, that are androgen independent should be able to grow. The lack of attachment could be due to over trypsinization of cells during subculture. One could collect the medium and centrifuge to collect the floating cells to add back to the flask (so the loss of cells is minimized due to medium change).
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