Place your coomassie stained PVDF piece in an Eppendorf tube containing 1ml
of methanol and 0.1% triethylamine. Invert several times. The blue colour
will rapidly leave the blot but your protein will not. Repeat more times if
necessary with fresh solution, then rinse once with methanol and dry.
Useful trick for destaining (and removing residual SDS) from blots for
sequence analysis.
Len
>Does anyone out there who performs enzymatic digests on PVDF bound samples
>have any solutions to the coomassie stain which accompanies the samples?
>
>I regularly receive samples in which the PVDF bound
>samples are grossly over-stained with coomassie blue. After performing the
>digest and taking the sample through the speed vac I get about
>20 micro-liters of
>dark blue sludge. This is hell on the PE ABI 173A microblotter and
>associated cHPLC, not to mention the chromatograms.
>
>Is there a de-stain step I could use that wouldn't affect the protein
>prior to the digestion? Is there a pre-column that might also be useful
>for removing the stain prior to the cHPLC column?
>
>Any thoughts on this are greatly appreciated!
>
>Best Regards,
>Suzanne Perry-Riehm
*********************************************************************
Dr Len C. Packman
Assistant Director of Research
Protein and Nucleic Acid Chemistry Facility
Department of Biochemistry
University of Cambridge
80 Tennis Court Road
Old Addenbrookes Site
Cambridge, CB2 1GA, UK
Tel: +44 (1223) 333639
FAX: +44 (1223) 766002
e-mail: lcp2@mole.bio.cam.ac.uk
Visit my WWW page at http://www.bio.cam.ac.uk/proj/adr/PNAC/pnac.html