We have a user trying to use this live-dead kit to stain cells in a layer of Matrigel, then image with a Keyence microscope LIVE/DEAD™ Viability/Cytotoxicity Kit, for mammalian cells 1 kit | Buy Online
His staining doesn’t work, reading the manual and using my mk1 brain I suspect the stains simply don’t penetrate the matrigel (he adds the stain on top of matrigel layer for 30 minutes). The attached manual also mentions that serum might “activate” the calcein before it ever makes it into cells, which is another possibility (not sure if matrigel could do that, or if there’s serum stuck in the matrigel, etc; his media had serum in it).
Our current idea is to liquify Matrigel with cold PBS, centrifuge the organoids out (washing will also get rid of FBS), then stain. I think it’s better to stay at 4C and stain for an hour or two, but he will also try doing staining at 37C for 30 minutes and see if that is better.
Do you have any other advice? Thank you!