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Vy high % PA gel drying

Song Tan tan at aeolus.vmsmail.ethz.ch
Sun Jul 10 14:14:09 EST 1994


In article <2vnfj4$pm7 at usenet.INS.CWRU.Edu> evw2 at po.CWRU.Edu (Eric V. Wong)
writes:
                 
>Here is the situation. I am using a modified Tris-tricine gel 
>system to resolve small proteolytic peptide fragments, and I get
>excellent resolution using a 15-30% gradient. The problem is,
>I can't seem to dry the darned thing without it shattering all
>to heck. I usually fix in 50% MeOH/ 10% acetic acid, and have
>tried adding 10% glycerol to the fix, but that doesn't seem to
>do it. I have no idea what I did *right* that first time: I 
>have fiddled with different gel dryers, different temps, etc.

My guess would be that your bisacrylamide:acrylamide cross-linking ratio is too
high.

It's very difficult to dry >15% acrylamide gels with >1:30 bisacrylamide:
acrylamide cross-linking ratios (approximate numbers only).  I know that I can
dry 15% acrylamide with 1:40 bis:acrylamide, but if I try 15% acrylamide with
1:20 bis:acrylamide, the chance of the gel cracking during drying increases
dramatically (but as you've observed, such a gel can be dried without cracking
occasionally).

Put another way:  high concentration acrylamide gels with high bis:acrylamide
ratios are very brittle.  Reducing the bis:acrylamide ratio makes them less
brittle.  Conversely, low concentration acrylamide gels with low bis:acrylamide
ratios are too jelly-like.  Increasing the bis:acrylamide ratio makes these low
concentration acrylamide gels more rigid and easier to handle.  

If you're not doing so already, try decreasing the bis:acrylamide ratio to 1:60
or 1:80 or perhaps even lower.  With luck, this change won't alter the
resolution of your gels.

Best regards,

Song Tan
Institute for Molecular Biology and Biophysics
ETH-Honggerberg (Swiss Federal Institute of Technology)
8093 Zurich, Switzerland
email:  tan at aeolus.vmsmail.ethz.ch




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