[western blotting] Abcam's ab9484 b-actin loading control vs casein blocking

[Pow Joshi]

On 8 Dec 2006 02:48:57 -0800, Lechu <lech_kaczmarczyk At yahoo.com> wrote:
> Hi all,
>
> Abcam strongly reccommend to block ab9484 b-actin loading control
> antibody (ab9484) with BSA instead of milk. Does anyone know what milk
> component is most likely to cause problems with this ab? I need to
> block this antibody with casein, hence my question.


so far I know milk protein gives problems with phosphorylation ....
that is if you plan to detect phosphorylation of proteins, you may
have a large background with phospho specific antibodies .... in this
case, where you're using GAPDH, the data sheet says that there's a
decrease in signal .....
now, if I were to give you a pie in the sky hypothesis, which would be
a plausible explanation for this decrease in activity:
a. you're using a monoclonal ... ergo: single epitope
b. the milk casein kinase phosphorylates this epitope
c ergo, your ab will not bind to it very well.
 ,,, probably if you had a polyclonal, you could get away with milk blocking.

best
pow


>
> best, Lech
>
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