APS & TEMED in acid gels

Leonard Ornstein lenornst at pipeline.com
Mon Jan 27 15:56:37 EST 1997


Straight from (one of) the horses' mouths:

Louis' question has generated quite a thread; with a mixture of truth, 
half-truth and error.

I'm one of the two (or four, depending how you look at it) inventors of 
polyacrylamide electrophoresis; and it was B.J. Davis and I who first 
introduced the APS/TEMED, TEMED/Riboflavin and APS/TEMED/Riboflavin 
polymerization systems. Most of the details are available in Adobe 
Acrobat versions of our two original manuscripts, which can be 
downloaded from:

<http://www.pipeline.com/~lenornst/DiscElectrophoresis.html>  .

My paper also discusses the effects of degree of cross-linking on 
separation performance, Since, as already noted in this thread, the rate 
of initiation is dependent upon the free radical concentration, and this 
determines the number of chains that start to propagate, you ultimately 
get many short chains, at high rates of polymerization, and fewer very 
longer chains at lower rates. With high concentrations of BIS, this 
hardly changes pore size. But with low concentrations of BIS, the 
effective pore size can end up larger, and, as noted, in some cases the 
solution, though viscous, might not even gel.

In addition, with a poorly optimized catalyts/initiator system, the 
system may not consume all the monomer, and so the effective pore size 
can end up larger than with an optimized system. (Incompletely 
polymerized gels end up far-UV opaque, whereas completely polymerized 
gels are usually far-UV transparent.) 

Len Ornstein 

Louis Hom wrote:
> 
> So I'm running these HOAc-urea polyacrylamide gels (15% resolving, 4%
> stack) and each part takes >1hr to polymerize at the concentrations of APS
> and TEMED that I currently use.  Will I mess things up by increasing the
> APS & TEMED concentrations so that polymerizatin occurs more on the order
> of 30 minutes instead?  I won't be transferring or eluting the proteins
> from the gel for anything.
> --
> _______________________________________________________________________________
> Lou Hom >K '93
> lhom at nature.berkeley.edu
> http://www.ocf.berkeley.edu/~lhom



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