Gel voltages and times
huangz at biowww.net
Sun Feb 18 02:10:01 EST 2001
I have a question about protein transferation on PDVF membrane. I run mini
gel (same size as paul's 70x90x1.5mm) and do tank electronic transferation.
Previously we used 240 mA constant current for 1.5 hours or 140mA for 2
hours. The transfer is ok although it is higher than what recommended in
protocol books. Recently someone tell me that low current (say 60mA) for
overnight is better since the lower current decreased the risk of
destruction of antigen epitope. I tried once time and it seems I can get
stronger signal from the blot than before. But I am not sure if this is
because of above reason since I did only once time. Could anyone tell me the
best constant electric current for mini gel transferation?
Warren Gallin wrote:
> The important parameter is actually the voltage gradient,
> V/cm. The
> trick is, you have to measure the electrode-electrode distance,
> which is
> not necessarily the length of the gel. So, if you happen to be
> the two gels in the same submarine apparatus, you would not
> change the
> voltage at all. If you are using a smaller apparatus, the
> ratio of the
> inter-electrode distances is the relevant factor.
> Warren Gallin
> "Paul S. Brookes." wrote:
> > I have a real silly question about gel voltages.... Is the
> voltage you
> > apply across them determined by the length, or the
> > area?
> > The protocol I'm working from uses 120V, 12hrs, for a 140 x
> 140 x
> > 1.5mm gel - i.e. 210square mm cross-sectional area, 140mm
> > I have a 70 x 90 x 1.5mm mini-gel, i.e. 135 square mm area,
> > length.
> > * So, seeing as my gel's x-sectional area is 64% of the
> bigger one,
> > do I use 64% of 120V = 77V
> > * Or because the length is 50% do I use 50% of 120V = 60V
> > * And how long to run for? Does half the length = run for
> half the
> > time?
> > Is there a simple equation for working out voltages based on
> gel % and
> > dimensions?
> > Thanks in advence for any help.
> > Paul
> > _________________________________________
> > Dr. Paul S. Brookes. (brookes at uab.edu)
> > UAB Department of Pathology, G004 Volker Hall
> > 1670 University Blvd., Birmingham AL 35294 USA
> > Tel (001) 205 934 1915 Fax (001) 205 934 1775
> > http://peir.path.uab.edu/brookes
> > The quality of e-mails can go down as well as up
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