Protocol Online logo
Top : New Forum Archives (2009-): : SDS-PAGE and Western Blotting

Transfer problems - (Apr/08/2010 )

Hi everyone,
I'm trying to transfer my proteins from a 10-20% acrylamide gel to a nitrocellulose membrane in a semi-dry transfer cell from Bio-Rad.
My transfer buffer has 20 % MeOH and I'm transfering 30 minutes at 15 V, 5,5 mA/cm^2 (as recomended by BioRad).

I do transfer, but some of the protein still remains in the gel and some other cross the membrane!

I'd appreciate a lot some advice!!


Transfer longer. I use a 6-15% gradient gel and transfer for 4h at 200mA. Use a small size pore nitrocellulose if you're worried about transferring small proteins through.