I tried sorting human iPSC by flow cytometry, but lost a lot of cells during staining or transfer of cells from tubes to epis or so. Any suggestions to prevent this? I know that some people coat their tubes and culture ware before they put their cells in. Would that be the case for iPSC too?
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Chemically defined conditions for human iPSC derivation and culture
Guokai Chen, Daniel R Gulbranson, Zhonggang Hou, Jennifer M Bolin, Victor Ruotti, Mitchell D Probasco, Kimberly Smuga-Otto, Sara E Howden, Nicole R Diol, Nicholas E Propson, Ryan Wagner, Garrett O Lee, Jessica Antosiewicz-Bourget, Joyce M C Teng & James A Thomson
This is not specific to iPSCs, but in general, I do the following to avoid losing precious cells during staining:
1) Make sure you have some BSA (e.g. 0.5%) or serum in the buffer you use for staining and washing
2) For spinning cells down in 1.5 ml microcentrifuge tubes, I centrifuge for half the time with the hinge in the cap pointing towards the middle of the rotor, then I rotate the tube 180 degrees (hinge facing the outer edge of the rotor) and spin for the remaining time. This makes the pellet more compact, instead of having cells stuck along the tube wall.
Hi
I would appreciate any suggestions about the cell surface marker specific for iPSC (mostly iPSC Motor neurons) staining and FACS sorting.
Thanks,
Manoj
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