chemiluminescence detection/western blot

[Dan Oerther]

In article <D3r575.D3u at boss.cs.ohiou.edu>, janapati at ouvaxa.cats.ohiou.edu wrote:

> I am getting high background and multiple proteins are getting
labelled.  I am using monoclonal primary antibodies.  Any suggestions on
blocking agent or detection method.  

   I am using the Boehringer Manheim HRP kit quite easily with PVDF
membrane blocking for one hour with 1% milk (weight/vol) and 0.1% Tween -
20 (vol/vol) in Phosphate Buffered Saline (PBS).  Additionally, I carry
out the hybridizations in the same solutions.  Hope this helps.  By the
way, I also tried Zeta Probe Nitrocellulose membrane - doesn't block as
well, but seems to work okay if you increase to 5% milk (weight/vol).

Best, 

Dan

-- 
Dan Oerther
Departments of Biochemistry and Civil Engineering
Northwestern University, Evanston, IL

doerther at nwu.edu