Cytoplasmic marker for western blotting - (Aug/02/2007 )

Hi,
I have been doing membrane and cytoplasmic fractions to look at the recruitment of different proteins to the endosomes, and assessing this by western blotting. However I need a good cytoplasmic marker to assess the loading. So far I have tried Cbl, LDH and tubulin, but all of them show some bands in the membrane fraction as well (even though the fraction is pure).
Does anyone have any suggestions? (I am thinking of trying GAPDH).
Thanks!
Viola.

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Hi Viola,
I usually use one of the IkappB proteins (alpha or beta) to show how well my nuclear and cytoplasmic fractionation worked. I would think that GAPDH would work as well but not from experience.

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Hi! Can you tell me the procedure to isolate endosomes?

Thanks

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My method actually isolates all membranes (however I do get rid of the nuclei), so not specific to the endosomes. Are you still interested?

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yes, sure!! Can you send me the protocol? Many thanks!

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