Cell Envelope Proteins Smeared (View forum version)



Sarewarren

Posted 22 February 2012 - 09:46 AM

Hello,

I am attempted to do a western on cell envelope protein I have isolated from P. aeruginosa, however the lower proteins always come out smeared. I have ran gels that are 10%, 12% and 15%. I store my protein in -20 with PMSF, I run at 80V for 30 min then 120 for 2.5 hours. My ladder is always clear. I have remade all of my buffers and have changed my sonication time and amplitude. I am at a loss.

Any input would be appreciated.

Thank-you

mdfenko

Posted 22 February 2012 - 01:04 PM

the smearing could be due to post-translational modifications on the protein (eg glycosylation, lipidation).

it could also be caused by components in the sample's buffer.