Labelling CHO cells with S35

[Neuropathology Laboratory]

> From: P1034294 at CSDVAX.CSD.UNSW.EDU.AU (DAVID MISZTAL)
> Newsgroups: bionet.molbio.proteins
> Subject: Labelling CHO cells with S35
> Date: 18 May 1995 20:31:44 -0700
> 
> 	Has anyone out there had much experience with labelling
> Chinese Hamster Ovary cells with 35S-labelled cystein and methionine?
> I am wanting to do short labelling times (15 minutes) with CHO cells
> but I have been unable to obtain consistent results in my labelling.
> What I want to know is do most people pre-incubate CHO cells in a
> cysteine and methionine-free medium before adding label? (in order
> to reduce intracellular pools of these two amino acids)  If so,
> how long is a "safe" time without any interuption to protein-synthsis
> and hence the viability of the cell?
> 	Also- is such a pre-incubation step necessary for long labelling times?
> 
> 	Thanks in advance,
> 	David Misztal
> 	Dept. Biotechnology
> 	University New South Wales
> 	Australia

YES!   You have to "starve" the cells from 20 min to 45 min prior to
labeling in Met minus medium.  Also you may want to use 1% DIALYZED calf
serum in your labeling medium or serum free medium for labeling.  We
routinely starve cells for 30 min even for 3 h labeling.

Gopal Thinakaran.
Gopal at welchlink.welch.jhu.edu