Cys-Met free media?
smatzno at IECR02.IECR.MUKOGAWA-U.AC.JP
Mon Sep 27 05:59:22 EST 1999
We now start to try pulse-chase protein labelling for the evaluation of
the clearance of intracellular protein in skeletal myoblasts. Our
protocol is following:
1. Cells are plated onto 60mm dish. 24hr later medium is changed to DM
and culture for subsequent 24 hr.
2. Then cells are preincubated with Cys and Met-free medium for 4-6 hrs
3. Replace medium by fresh Cys and Met-free medium with 35S-Cys,Met (0.3
mCi/mL) and incubate for the time intervals required.
4. Wash cells with PBS, then lyse cells with Tris buffer containing 0.1%
Triton X-100 and some protein stabilizers.
5. Protein is collected by immunoprecipitation and analyze SDS-PAGE and
My problem is how obtaining Cys-Met free medium. In some catalog
(Gibco, Flow etc) I can't find Cys-Met free DMEM: only Met-free DMEM is
found. Does anyone know where I can purchase Cys-Met free DMEM, or we
can use only Met-free DMEM for this experiment?
Any help wold be gratefully appreciated.
Sumio Matzno, Ph.D.: First Department of Biochemistry
School of Pharmaceutical Sciences, Mukogawa Women's University
smatzno at mwu.mukogawa-u.ac.jp (lab) smatzno at sannet.ne.jp (home)
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