RE: beta galactosidase immunos
What are diluting your Ab in? what kind of AR are you doing? I had a
horrific background problem with my CWD's when I first started them up and
found that even though the AR for my CWD is something completely different,
the proteinase K digest was a very important step in reducing bg. Also,I
dilute my Ab in 0.25% BSA in 0.05M Tris. The combination of these two things
seems to work very well in checking the bg. I'm not saying you should do
these things in particular... you're dealing with an entirely different Ab
... but I am suggesting that you look at the little things in your reagents
because sometimes those little things make all the difference in the world.
They're so subtle, but so important.
Hope this was helpful.
Utah Veterinary Diagnostics Laboratories
Utah State University
phone: (435) 797-1891
From: Martin, Ronald [mailto:Ronald.Martin@umassmed.edu]
Sent: Monday, March 10, 2003 1:58 PM
Subject: beta galactosidase immunos
I am attempting beta gal immunos on mouse tissue and getting a lot of
background stain. I have tried 3% H202 as is and made up from 30% H2O2 in
methanol. I have also tried different antibody titration's from 1:200 up to
1:4000. I am redoing the 1:2000 and 1:3000 tomorrow but need some
suggestions on how to reduce the background staining. I also use the
recommended 3% goat serum.
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