Slowly migrating EGF receptor in SDS-PAGE

John A. Newitt newitt at ncifcrf.gov
Mon Mar 27 13:30:06 EST 1995


In article <3l4q10$cj1 at abc.ksu.ksu.edu>, maraman at ksu.ksu.edu (Dayanidhi
Raman) wrote:

> Hi, I immunoadsorbed the EGF receptor and eluted it, dialyzed to remove
the     detergent in the presence of added lipid, assayed for tyrosine
kinase activity. The prep. was run on 7.5% SDS-PAGE after heating the
sample at 60C for 3 min.   in Laemmli's sample buffer. The autoradiogram
has all receptors at the very top of the gel. Need help desperately!.
> A

I forgot to mention also that, depending on the detergent you used in the
immunopurification, you may not have removed it by dialysis--are you SURE
you did.  For example, if you have a lot of Triton X-100 present in your
samples  the SDS in your sample buffer will form mixed micelles with the
nonionic detergent, leaving little SDS remaining to bind to and denature
your protein(s).

John A. Newitt  <newitt at ncifcrf.gov>
National Institutes of Health
Bethesda, Maryland  USA



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