Western Blotting problems

Ian A. York iayork at panix.com
Mon Oct 16 11:30:30 EST 2000


In article <B610F15D.32A%ralf.koelling at uni-duesseldorf.de>,
Ralf Kölling  <ralf.koelling at uni-duesseldorf.de> wrote:
>
>Ponceau S. Instead of the normal protein banding pattern, we often see
>"clouds" of staining on the NC membrane ("smear blots" as we call it). It

I've very intermittently seen something like this, too.  I don't know what
causes it, but I'll give you my guess:  Overheating of the buffer.  My
guess is that if the buffer warms up a little too much it degasses, and a
bubble gets between the blot and the membrane.  Solution:  Reduce the
voltage and do longer transfers at lower current; keep the transfer in the
cold (4 oC, or ice pack, or both).

It's been a very intermittent problem with me, and so I haven't really
been able to test this very well, so I certainly won't make any
guarantees.  Good luck.

Ian 
-- 
    Ian York   (iayork at panix.com)  <http://www.panix.com/~iayork/>
    "-but as he was a York, I am rather inclined to suppose him a
     very respectable Man." -Jane Austen, The History of England






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