klenchin at macc.wisc.edu (Dima Klenchin) wrote:
>In article <477q8p$102 at mserv1.dl.ac.uk>,
> LOGAND <logand at msdos.ensam.inra.fr> wrote:
>>>Can anyone provide or point to me towards a method to stain total protein
>>on western blots (nitrocellulose) after antibody detection of specific
>>antigen using alkaline phosphatase ?
>>AFTER will not work - there is no way to avoid blocking membrane for ab
>detection, which means your total protein is all over the blot.
>>BEFORE is easy. Ponceau S is reversible stain (one drawback - not vry
>>Stock: 2% solution in 30% TCA, 30% sulfosalicylic acid (store rt).
>>Staining: dilute 1:10 with water, stain for 10-20 min.
>Destaining: water or TBS. Then proceed with blocking for ab staining.
>>Hope it helps.
> / /\ Dima Klenchin
> / / \
> / / /\ \ klenchin at macc.wisc.edu> / / /\ \ \ tel. (608)262-4380
> / /_/__\ \ \ FAX (608)262-4570
>/________\ \ \
I don't do Westerns but I've gotten some flyers in the mail that may help. One is from an outfit called ZOiON Biotech, They say they've got a negative stain that stains only where there is no protein, no denaturation, the pattern stays after blotting and sensitivity like Coomasie. They give an 800 # of 1-800-430-5454. With this one I guess you could stain the gel do your Western and compare the gel to the blot.
The other is from an outfit called Molecular Probes, Inc. They say they've got Fluorescent PAGE stains with the sensitivity of silver stain and without interference with Westren blotting. I can't find the flyer but I did find the envelope with an address of P.O. Box 22010, Eugene, OR. 97402.
Hope this helps