Hello All,
Could someone recommend me a good protocol for SDS-PAGE and subsequent
transfer of proteins onto PVDF membrane for subsequent Edman sequencing?
Which PVDF membrane did you use, how did you deal with non-polymerized
acrylamide that can modify proteins and impair sequencing, what transfer
buffers worked for you best, etc. Should I use PAGE buffers different from
Tris-glycine (Borate?), and do I have to care about glycine on Page stage at
all? I know that CAPS buffer is a good alternative for Tris glycine transfer
buffer, but I am concerned that high pH of the buffer (pH 11) will cause
modification of side chains of amino acids. In other words, I good detailed
working protocol would be very much appreciated, or pointers to published
resources.
Thank you.
Emir
University of Chicago
Radiation Oncology
