I want to measure contractility of the beating single cell and measure cell size.
Cardiomyocytes die after reseeding.
Dear Darshini,
yes it is possible to do so. You will of course lose some cells in the digestion/splitting process so pls calculate cell numbers accordingly. In our standard protocol we trypsinize the cells for 5min followed by harvesting and seeding them in RPMI+B27 medium supplemented with 20% FBS and Thiazovivin/Rocki to promote survival. Depending upon condition of the hiPSC-CMs, you might have to give these cells some time to recover (normally 3 days) after splitting. Hope this helps. Good luck!
I did it and it works fine with me.
You may need to include RevitalCell (1:100) for 24 h when reseeding them. You also should expect a loss between 30-50% as these cardiomyocytes are very sensitive to the passaging.
The protocol is like:
Before anything coat the new culture plates with Matrigel (0.3 ml in 48 ml DPBS) for 30-60 min.
you can read more at:
Article Electroconductive Biohybrid Hydrogel for Enhanced Maturation...
Article Nanofibrous Composite with Tailorable Electrical and Mechani...
https://pubs.rsc.org/en/content/articlelanding/2019/bm/c9bm00434c#!divAbstract
Thank you kaveh for your detailed answer, do you also experience the cells more in a clump form that it doesn't resuspend well. The clump of cells for me seems like a muscle mass which do not resuspend well and just float in the media. Do you have any suggestion how to separate them or how to make its suspension??. I tried using cell strainer and even used needle to separate but no success here.
I am trying this myself now, is the replating back on matrigel or is another coating better at this time?
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