Immunodetection of 40-50kDa proteins

Simon Mauch Simon.Mauch at uni-konstanz.de
Wed Jun 10 15:55:16 EST 1998


On Mon, 1 Jun 1998 12:55:05 -0700, "Jose C. Juarez"
<jcjuarez at ucdavis.edu> wrote:

>I had an antigen in the same region where the IP Ab would migrate. What
>helped in the end was titrating out the IP Ab so that I could get the
>background signal much lower than my positive controls. I think I ended up
>IPing with about 0.2 micrograms Ab per 500-1000 ug of cell lysates.
>


The method Frank uses seems quite good. 
Another (easy)  "method"  to seperate the antibody from your antigen
is the elution of your antigen with a buffer ommitting ß-ME or DTT
(e.g.  nonreducing SDS-PAGE loading buffer or 0.5 M acetic acid or
something like that). So the antibody will not get reduced and
migrates at 150 kD (in the case of IgG)...

Simon


Simon.Mauch at uni-konstanz.de



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