Blocking o/n after adding secondary antibody (Western blot)

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TaintedCherub
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Blocking o/n after adding secondary antibody (Western blot)

Post by TaintedCherub » Tue Jun 26, 2007 11:47 pm

Hi,

I have a major problem with a western - I've applied primary and now secondary antibodies, but I've just discovered that the door code to the dark room has changed, everyone's gone home and I'm basically locked out. If I put the membranes in block overnight will a signal still be there tomorrow?

Hopefully yours,

Chris

blcr11
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Post by blcr11 » Mon Jul 02, 2007 8:09 am

I suppose by now it's too late for any comments to be useful. I guess I would have preferred to leave it in 2nd Ab, but I imagine it was already too late for that when you discovered your predicament. If your Ab reactions are reasonably stable, you should have been OK going another 8 hrs in blocking solution. So how'd it come out?

konstantin
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Post by konstantin » Wed Jul 04, 2007 2:29 am

yes It mostly depens on what protein you are detecting.
personally, I had some of that problem before, but I did not seal in blocking solution, just 20 TBS-T is enough. good luck

TaintedCherub
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Post by TaintedCherub » Thu Oct 15, 2009 11:34 am

It worked fine, thanks!

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