Weird SDS-PAGE... HELP!!

Joe Stains JPS144 at psuvm.psu.edu
Mon May 8 12:24:52 EST 1995


Someone in my lab is running SDS-PAGE on a cell lysate.  However, when
they run the gel, the sample lanes spread out (longitudinally), pushing
the adjacent lanes out as well.  No one seems to have seen anything like
this before.  We've tried several things to reduce salt, getting rid of
any lysis buffer, decreasing sample load volume, but nothing seems to
work.  We've even decreased the gel speed to hopefully correct the
problem.

Among the steps we've done are: 1) run samples containing positive
control samples with added lysate buffer => no problems with the run 2)
ppt protein by a variety of methods and redissolving them in a weak Tris
sol'n (same conc'n as is in the 1 X buffer) => still run "wide" 3)
combined step two with microconcentrator to decrease volume, as well as
add more Tris soln as a "wash" to get rid of any residual salts

4) running gel at lower amp (20mA constant)

I just don't get it.  Has anyone experienced anything like this before?
We've checked all of her soltions.  They all seem fine.  The TEMED and
APS are fresh.  The acrylamide is less than one month old.  The weird
thing is her first gel ran fine, but has been unrepeateable since.  If
you could e-mail me with any suggestions, it would be greatly!
appreciated.  We are wasting so much time and materials trying to figure
this one out!  Thanks in advance.

Joe Stains jps144 at psuvm.psu.edu Penn State University



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