I wouldn't use actin, as it can be found in the nucleus. I usually use and anti-alpha tubulin antibody (Sigma T9026) as a cytoplasmic marker with good success, and I don't typically see a  double band.

Besides tubulina and actin, i'm only seeing cofilin (20kDa).

Hi everyone

I actually use the tubulin antibody you mentioned Cristopher. And what I saw is that some times I have a slight separation between both bands and depends on the cells I've used. Thanks for the advice.

Thanks Korrodi for the answer, do you have some spare aliquot you can send me for me to test? ;) 

Unfortunately no :P

I agree that most actin and actin related proteins are found in the nucleus as well. ERK1/2 should be exclusively cytosolic if you can use it. I have used COX4 which is  a mitochondrial protein both as loading control and confirm that I have pure nuclear fraction.

Try GAPDH

LDHA is a good one.