re-probing western blot with chemi-luminiscent substrate - (May/23/2007 )
I ran a western a few days ago, and had saved the blot at 4 degrees in a wet tissue. I tried to see if I get any signal with the X-ray film today, and there was none.
So I was wondering if anyone could tell me if there is any way to re-use the same blot again, maybe moisten it with PBS and try and re-probe it with the chemi- substrate...... could/does that work??
or is there some other way to do it, or do I have to strip the blot and try priming with antibody and then detect the proteins??
Any help will be appreciated.
Thanks,
Vik.
-Gallus gallus-
QUOTE (Gallus gallus @ May 23 2007, 08:23 PM)
I ran a western a few days ago, and had saved the blot at 4 degrees in a wet tissue. I tried to see if I get any signal with the X-ray film today, and there was none.
So I was wondering if anyone could tell me if there is any way to re-use the same blot again, maybe moisten it with PBS and try and re-probe it with the chemi- substrate...... could/does that work??
or is there some other way to do it, or do I have to strip the blot and try priming with antibody and then detect the proteins??
Any help will be appreciated.
Thanks,
Vik.
So I was wondering if anyone could tell me if there is any way to re-use the same blot again, maybe moisten it with PBS and try and re-probe it with the chemi- substrate...... could/does that work??
or is there some other way to do it, or do I have to strip the blot and try priming with antibody and then detect the proteins??
Any help will be appreciated.
Thanks,
Vik.
in the mean time, the peroxidase may be inactive; add the 2nd Ab tagged with peroxidase again w/o stripping; you may succeed
there should have been no dry phase after the last probing which would denaturize also the first Ab
-The Bearer-
Thanks!!
I did try and re-prime just with the secondary, but that did not work... i guess i will have to strip and re-probe with both!!
thanks!
-Gallus gallus-