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
