Western Blotting of Membrane Proteins in the presence of SDS

Shaun D. Black SHAUN%JASON.DECNET at relay.the.net
Thu Apr 22 23:07:14 EST 1993


Dear Netters,
     I'd appreciate your experience as to electroblotting of membrane 
proteins on PVDF (polyvinyldenedifluoride) membranes.  We've tried a number 
of conditions but would be curious as to what others have found optimal for 
membrane proteins from SDS-PAGE -> PVDF.

     1. Buffers:  [50mM Tris / 380mM Glycine] vs [25mM Tris / 190mM Gly]
                  (or others?)

     2. SDS    :  0% vs 0.1% (or others?)

     3. Methanol: 20% required?

     4. Voltage / Time:  Is there an optimum or do membrane proteins simply
                         require more time?

     5. Is it important to blot right after SDS-PAGE or can gels be soaked
        in blotting buffer overnight before electrotransfer?

Thanks in advance.  Your thoughts are much appreciated.  Cheers.  -Shaun

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  = Shaun D. Black, PhD     | Internet:  shaun%jason.decnet at relay.the.net = 
  = Dept. of Biochemistry   | Bitnet:    shaun%jason.decnet at thenic.bitnet = 
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