immune precipitation

[Dennis J. Templeton]

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?
>
>
You should be aware that some, even most, monoclonals don't bind directly
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.

good luck, 

dennis