resolution on SDS-PAGE

Ron Tate rtate at bmb-fs1.biochem.okstate.edu
Wed Feb 28 10:35:45 EST 1996


R. Woodward wrote:
> 
> Dear All
> 
> I have two polypeptides of similar size (between 52 and 55KD) which I want to be able to resolve by SDS-PAGE. Both species are 
radiolabelled(35SMet) and the sm
> dried gel for too shorter period then I only see the lower band, on the other
> hand if I expose for too long then the signal from the lower band obscures
> the larger band. I want to be able to maximise the resolution of the two
> bands on SDS-PAGE without having to resort to 2D-gels. Any Ideas as to the
> best gel conditions to employ (I've tried 7.5,10 and 12.5% gels so far,
> what would a gradient gel do for me?)
> 
> Thanks Robert
> 
> R. Woodward
> Email rw200 at cus.cam.ac.uk

Robert,
Try a "Double Inverted Gradient" gel, I found this in Biotechniques, the 
authors claim clear separation of proteins differing as little as 0.5 
kDa. The bottom half of teh gel is an inverted gradient from higher 
concentration to lower and the top is a normal gradient.  I think I'm 
getting ready to try it but haven't had the time yet. The reference is 
_Biotechniques_ vol.16, #2, Feb. 94, pp270.  (If you don't have access 
to Biotechniques you can request a copy of the article from their web 
page.
it is at http://www.biotechniques.com, select the Biotechniques link and 
you will find a search page, I just tried it entering only an author, 
Zardoya, and I got the article.)
Hope this helps
Happy Hunting
Ron Tate
OSU BMB



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