How to seed mitotic cells after mitotic shake-off?

06 June 2017 1 8K Report

I want to synchronize my culture of H4-cells without using chemicals and decided to use the mitotic shake-off.

I followed this protocol for the mitotic shake-off: https://link.springer.com/protocol/10.1385%2F1-59259-646-0%3A11#page-1 (section 3.1)

During the collection process, they where hold on ice for 1 hour.

I got about 400.000 viable, mitotic cells and seeded them into a new flask but they would die within some hours. Does anybody know what to do?

Thanks in advance

Reza Hadavi

1- check temperature between 4-6 c exactly . before ice treatment , dead cells deleted by 120 G / 5 min / 5 c centrifugation.

2- you can omitted FBS from culture medium for 1 hour and then ready to test ( its an idea )

good luck

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