RE: Tissue Culture help

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From:"Montague, Donna C" <> (by way of histonet)
To:histonet <>
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I dont have a web address for you but when I've experienced these problems
I've had to methodically check the factors involved in cell culture which
may contribute to a contamination problem.
1) Clean incubator!
	Even water jacketed incubators require that you place a dish of
STERILE water on the bottom shelf of the incubator with the cells. This must
be changed daily. Each month wipe the interior of the empty incubator with a
5 % solution of household lysol (the pink junk at the grocery store)5 ml /
100 ml and dont forget the door seals and the inside of the doors.
2) CO2 concentration.
	Dont laugh, but first make sure the caps on the tissue flasks are
vented (loose). The best way to assure this is to snugly close the cap after
feeding or innoculation the unscrew the cap one quarter to one half a turn
until the cap rattles easily. Of course this doesnt apply if your using
dishes. : ) Even with the modern incubators and their built in CO2 sensors
the actual concentration of the CO2 in the incubator should be checked
periodically. Several commercial products are available for you to check it
yourself and of course incubator manufacturers will gladly send a service
rep for a fee.
3) Fetal Calf Serum and other serum media supplements.
	Have your supplier verify that the lot of serum you are using is
mycoplasma free. If you can't open fresh supplements for your media (growth
factors, antibiotics, serum etc.) filter sterilize them using a 0.22 micron
sterile filter.
Hope this helps. Feel free to call or email with further questions.
Donna Montague, Research Associate
UAMS Orthopaedic Research
Little Rock, AR
(501) 686-8739
(501) 686-8987 FAX

-----Original Message-----
From: []
Sent: Tuesday, December 01, 1998 11:28 AM
Subject: Tissue Culture help

I realise that this may not be the right place for tissue culture but if
anyone out there knows of a web page or sight to post problems on tissue
culture.  We have a PI who's projects have come to an almost complete halt
because of dying cells.  Sorry I do not know much more about the problem
but if I could supply her with an address of some sort that they could go
to I'm sure it would be greatly appreciated.

Thanks in advance

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