can i use EDTA instead of TEMED for SDS PAGE ?
Dr Engelbert Buxbaum
(by engelbert_buxbaum from hotmail.com)
Tue Jun 3 08:19:46 EST 2008
Am 30.05.2008, 20:50 Uhr, schrieb DK <dk from no.email.thankstospam.net>:
> Yeah, it's one of the biggest ironies of science techniques. The actual
> buffer system is what is properly referred to as Ornstein-Davis
> system, developed some time in 1960s. Leonard Ornstein, who also
> shares a credit for developing acrylamide gels to begin with (!), used
> to occasionally post here a while ago. All Laemmli did was to add SDS
> to this buffer system - with, as we know, quite spectacular results.
> All the pertinent details and refs can be found by digging at Ornstein's
> web page: http://www.pipeline.com/~lenornst/
> Sic transit gloria mundi. Hardly anyone these days knows his name.
> Then again, it's not like Tiselius is a household name either.
Well, the Ornstein & Davis paper introduced DISK electrophoresis (based on
work by several other authors, going back to Kohlrausch's 1897 paper on
what now we call isotachophoresis). The concentrating effect of
DISK-electrophoresis - or rather the narrow bands resulting from it - are
only one reason for the success of the Laemli method. The other is that
since SDS (and some other detergents like CTAB) bind to protein in an
approximately constant ratio of 1 detergent to 3 amino acids. Thus all
proteins have almost the same charge to weight ratio and experience the
same accellerating force in a field. Also, because of the denaturing
effect of SDS and BME all proteins have roughly the same shape. So in
Laemmli's method of the 3 factors that determine electrophoretic mobility
(size, shape and charge) only size remains. This makes the gels
interpretable, and that constitutes considerable progress.
By the way, although in Laemmli's 1970 Nature paper the discussion of the
method is indeed rather brief and limited to a figure legend, Laemmli &
Favre (J. Mol. Biol. 80 (1973) 575-599) gives more info and I have argued
that it is that paper that should be cited for the method.
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