bsh at MED.PITT.EDU
Mon Oct 5 08:42:11 EST 1992
> In a previous article, skspoidn at susssys1.reading.ac.uk () says:
> >I am trying to Immune precipitate a cellular protein
> >(CD4) with Mabs using protein A and S35 metabolically
> >labelled cells, and finding that the immune reaction is
> > not occuring, although antibodies are being precipitated.
> > Is there some aspect of the immune reaction and susequent
> > precipitation which is critical in this procedure?
> to PrA, I'd cite the classes here but I'd probably mix them up. We usually
> test a new mAb empirically by binding some to PrASepharose and eluting and
> staining for the IgG on a PAGE.
> If your mAb doesn't bind to PrA directly, not all is lost, as you can add
> rabbit anti-mouse IgG to sandwich the mAb to the PrA with the rabbit IgG
> that binds pretty efficiently. In my experience, the Rabbit anti-mouse
> serum doesn't affect the quality of the IP's very much.
One suggestion to overcome these problems. I had used this procedure
with some success dut did not need to explore in detail.
Can we not use the say anti-mouse immunoglobulins that are bound to a solid-
phase. Since these are polyclonal, I think it should be fairly easy for them
to bind as opposed to the limited binding epitope of ProteinA. I know two
suppliers one is Kirkegaard and Perry Lab at Gaithersburg Md and the other is
Pierce (Call 1-800-555-1212) for both Toll free numbers.
If somebody has tried this method, can you please add your comments?
Univ of Pittsburgh.
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