SDS-PAGE problem

[Cornelius Fritsch]

It may sound funny to you, but I once had the same problem. It resulted from
putting the spacers too far to the inside of the gel. Therefore the clamps
which I used to secure the gel to the stand exerted pressure well outside
the spacers. And funny as it may sound, this was enough to cause the glass
plates (although they were rather thick) to bend slightly, allowing the
sample to kind of "fall in between gel and plate".

Please let us know whether you found a workaround to your problems.

Cornelius



"Steve Nothwehr" <nothwehrs at missouri.edu> schrieb im Newsbeitrag
news:67f3539b.0403040727.32c0ad91 at posting.google.com...
> Hello All,
>
> For the last few weeks we have been experiencing a problem with our
> SDS-PAGE gel system (a typical vertical gel system from CBS scientific
> for running 18 cm long gels).  What happens is that for the first part
> of the run all looks well and we see a nice tight bromophenol blue dye
> front and good separation of our pre-stained standards.  Then about
> half-way through the run the dye front becomes more diffuse and
> streaky.  Also we observe the gel slipping down between the plates in
> the direction of the bottom buffer reservoir.  When we detect the
> proteins from such gels the bands appear streaky and diffuse as if
> there was a non-uniform electrical field through the gel.  Generally,
> the plates behave a bit like they have been siliconized (even though
> we have not knowingly treated  them with anything out of the
> ordinary).
>
> We doubt if the gel reagents are a problem because we use the same
> reagents on our mini SDS-PAGE gel system and we never experience any
> problems.  Almost all of the reagents have been re-made since we began
> experiencing the problem.  In addition,  the problem with our large
> SDS-PAGE system is intermittent.  This made me suspect the plates.  We
> typically wash our plates in warm sudsy water,  rinse with distilled
> water,  drip dry, and wipe down with 95% ethanol with a kim-wipe. We
> tried soaking our gel plates in 2 M NaOH for 2 hr followed by washing
> to get rid of any foreign substances but this didn't solve the
> problem.  Recently we bought all new gel plates and for a while the
> problem seemed solved but just yesterday we had a gel run poorly once
> again.  The problem doesn't seem to correlate with using a particular
> rig or position on the rig (each rig has 2 positions).
>
> I have been running SDS gels for 20 years (9 years with the CBS
> system) and have never experienced a problem like this.  We are
> desperate.  If you have any ideas please let me know.  Thanks.
>
> Steve