2 replies to this topic

[Lee6948]

Hi everyone,

currently i'm trying to establish stable, chronic drug-resistant cancer cells.
but i'm struggling to generate single clones and pooled ones and i can't tell whther the drug-resistant cells are single or pooled. i disperately need a working protocol or method to generate those clones or a website or reference.
thanks a billion

                                                                       the frustrated rob

[bob1]

Usually what you do is dilute the cells to a point where you are likely to get a single cell per well of a 96 well plate, then seed as many plate as you need - you may need to seed 10 or more to get single clones effectively.  A;ternatively, you can seed them at a higher concentration in 24 or 6 well plates (i'd use 1000 cells per well for a 6 well) and look for the formation of colonies, which will usually be formed from one cell.
You also need to determine a concentration of the drug(s) that kills the non-drug resistant cells, but leaves the drug resistant alive.

[Lee6948]

Thank you so much

I'll try that