I'm running blue native gels (Schagger & von Jagow method) on mitochondrial
respiratory complexes, using a biorad mini-protean III apparatus. There
are 2 key problems.....
1) After about 4hrs my power pack cuts out with a "no load" error. It's
designed to register <10 mA as a no-load, so I'm assuming the current is
too low (even at 80 V). Question - what is the current at 80 V in a
blue-native gel? Is it safe to crack up the volts (say to 120-140) to
bring the current back up? I assume the temperature (cold-room) also plays
a role in the current being low?
2) The dye-front that runs left-right across the gel also extends beyond
the edges of the gel and looks like it bridges the spacers. This suggests
the edges of the gel are leaking. I've never had this problem with
SDS-PAGE gels before, and have even tried squeezing another glass plate in
the apparatus to make sure the clamps are really tight, and also silicon
grease on all the gaskets/surfaces to ensure a good seal. Is there
something in the buffers that decreases the surface tension such that the
gels leak more easily? N.B. This problem #2 is not the cause of problem
#1, as it would cause a short-circuit, which gives a different type of
error message on the power supply.
Any suggestions? Biorad's technical support answer was "what does blue
native mean?" !
Thanks
PSB
_________________________________________
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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