Hello everyone,
Which is the best way to resuspend a culture of adult rat ventricular cardiomyocytes without damaging the cells integrity?
This is what I am trying to do. I isolate ventricular cardiomyocytes (Langendorff). I then plate them on laminin-coated petri dishes. I leave them in colture for 2 days before testing a treatment. After the treatment, I (want to) resuspend the cells, pellet them and (using a kit) extract the cytosolic proteins on one side and the membrane proteins on the other. So, the cells need to be intact when resuspended.
Thanks
Dario
I don't have a precise hint for how to resuspend gently, but if the problem also involves not losing your cells during washing steps, we can discuss our new split centrifugal force supernatant removal technology. www.siliconbiosystems.com/vrnxt
We can remove 99% of supernatant form a cell pellet without losing any cell. And is very gentle with cells. I don't know if this may fit your protocol. Let me know if you want to know more or try it.
ciao
raimo
I'm not familiar with your cell line or the exact protocol for the kit you mentioned but, when I want to be gentle while bringing my cells to suspension, I use PBS-EDTA instead of Trypsin. You tend to get small clusters of cells (instead of a single cell suspension), conserving more of you membrane proteins.
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