Subject: cells in culture

[Steve Crampton]

Hi Pow,
A lot of possibilities there.  I agree with Zhonglin's suggestion of the
coating.  Although coating is not necessary, precoating with 1% gelatin
seems to work well.  Also, were these 293s straight out of freezing?  From
the -80 or liquid nitrogen?  293s can last a few weeks, maybe months in
the -80 in DMSO, but after that, they'll die before they hit the plate,
maybe try a another batch.  Also the media is very critical.  We use DMEM
with 10% HI FBS.  And finally, I've found they are happier starting off at
a higher density.  Thaw 2 million in a T75 cm dish and they should be
fine.  Good luck!
Steve
Message: 3
Date: Wed, 24 Jan 2007 18:04:03 -0500
From: "Pow Joshi" <pow.joshi At gmail.com>
Subject: cells in culture
To: methods At magpie.bio.indiana.edu
Message-ID:
        <710764ea0701241504p3d9d1425v80f5cc2921e46cf9 At mail.gmail.com>
Content-Type: text/plain; charset=ISO-8859-1; format=flowed

Hi Folks,

this is sheer desperation: I was wondering if anyone would have any
idea why some cells (in this case HEK) are having problems attaching
to the culture plate, and then die a sad horrible death.... all
possible explanations are welcome including: "Pow, you've lost your
touch at Tissue culture, even if you've been doing it for half your
life". Thanks a bunch

pow

Steve Crampton
Molecular Biology & Biochemistry
University of California, Irvine
3113 McGaugh Hall
Irvine, CA 92697-3900
lab (949) 824-8143
fax (949) 824-8551
scrampto At uci.edu