I am currently measuring the effects of a drug in a cell line. Unfortunately, one of the effects is that the cells detach from the flask and because of that I would like to know if they are dead or it metabolism is stopped.
I have tried mtt, and alabar blue but because the cells that interest me are floating in the media I can't measure directly them.
Propidium iodide (dead) and Calcein AM (live) may do the trick. Microscopy or FACS..
FACs using the Annexin-V/FitC assay for identifying living and dead populations works well, you might be able to couple a differential nuclear staining live dead assay to this. Here's a reference: Curr Cell Biochem. 2011;1(1):1-14 for the latter assay. PMID: 27042697
Hi,
the most simple and straightforward way would be to use a membrane integrity stain (e.g. PI or even trypan blue) as already suggested. Your already mentioned approaches should be adequate, too, just use the available protocols for suspension cells (usually employing several centrifugation steps) after collecting the detached cells instead of those for adherent cells.
However in most cases, cellular detachment from the flask is the direct result from cytotoxic effects, and they usually can be considered either already dead or in the (later) process of dying, frequently via apoptosis. Still, there are exceptions to this, e.g. compounds specifically compromising the cytoskeleton.
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