Western blot and isoelectric point - (Jan/18/2012 )

Hi friends,

Continuing with my previous problem. I found that all my proteins have pI more than 9. I read from milipore site that in western blot the pI can affect the protein transfer to membrane.

Methanol strips the SDS from proteins and proteins are finally transferred to membrane - I was wondering whether really the pI will affect the SDS-PAGE followed by western blot OR the pI is imp only if use the proteins in native form ?


Any one faced problems with proteins with high pI

with sds present pI is not important. pI is important for native separations.

you can try a CAPS buffer at pH 11 for the transfer.

i am attaching a book from millipore and you can download a manual from ge healthcare from this link:

western blotting - priciples and methods