Biotinylation

[piglet1]

one question about biotinylation. After biotinyled with BIOTIN, and then lysated cells in RIPA buffer and then added strepavidin -beads for 4 hours at 4c, RUN SDS-PAGE gel, probed with the antibody I am interested, should I reprobe the blot with tubulin to proof my experient work or not??
Thank you

[JackBean]

what you mean by reprobe with tubulin?

[piglet1]

Normally we just detect the target proteinafter SDS-PAGE, I mean should we reprobe with anti-tubulin or actin to claim the proteins pulled down with strepavidin beads are only cell memberance fractions.

thank you

[JackBean]

I see. yeah, you could use some controls, by which you should know they will/not be there

[piglet1]

do you know any classic cytoplasmic protein marker beside Tubulin or actin.

thank you

[JackBean]

I would guess some metabolic enzymes, like from glycolysis