Ponceau S staining problem...

Ellen M. Quardokus ellenmq at indiana.edu
Thu Nov 30 15:37:53 EST 2000


Neal,

We routinely Ponceau S stain our nitrocellulose protein blots.
I use the protocol found in the book called Antibodies: A laboratory manual
by Ed Harlow and David Lane.

Ponceau S is kind of an odd stain in that in order to *see* the staining,
you briefly stain (5-10 minutes) and then very briefly wash with water (just
until a little of the red comes off--5-10 seconds, dump off the water and
look, add more water to clear the background a bit, swish around and dump).

If you wash too long, it will *all* be removed since it is a completely
*reversible* stain which is why it is the only stain that can be used if you
want to subsequently use your blot for immunodetection.

Hope this helps.

Ellen

Ellen M. Quardokus
Research Associate
Indiana University
Dept. of Biology
1001 E. 3rd Street
Bloomington, Indiana 47405
Phone: (812) 855-7239

Neal Robert Melvin wrote:

> I've tried 2 or 3 different protocols for Ponceau staining of
> nitrocellulose membranes, and can't seem to get my proteins to stain...
> the subsequent westerns are fine, indicating that protein electrophoresis
> and blotting was successful... but all I ever see with Ponceau is the
> tracking dye showing up. This doesn't appear to be a matter of amount of
> proteins, as I've tried putting a lot on the membrane and still nothing.
> I'm using Bio-Rad NC if that is useful info... any suggestions??
>
> Neal






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