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Problem with HEK stable cell line

HEK G418 Transfection

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#1 quentind

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Posted 21 November 2011 - 02:49 AM

Hi everybody,
I am a new user on this wonderful forum.
I just had a problem with a stable HEK line this week end.
I previously have had good transfection efficiency. I selected using 0.5 mg/ml G418 (established last year with a kill curve)
On Friday, after 10 days of selection, all the cells on my non-transfected dish were dead.
I was up to trypsinise my cells, but my chief told me to wait over the w/e.
yesterday, almost all my cells were detached, and attached in blocks...
I know that HEK are notorious for poor attachment to the surface...

I am starting a new selection. What can I change in my protocol?
Many thanks and sorry for this long message

#2 bob1

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Posted 22 November 2011 - 12:06 AM

It sounds like you just let them get over-confluent, try trypsinising them, they should be OK.





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