I tried to separate glycoproteins using Con A -Sepharose in an open
column. In the first step I equilibrated the column (1ml of ConA-Sepharose
from Sigma onto 1cmx 5cm sigma column) with lectin column buffer about 6 ml.
I found a lot of free Concanavalin A passing through the column.( 4
bands on 12%SDS-PAGE, silver stained) The first band (~24kDa) has a size of
about 3mm.X 5mm. for 5microlitres.
Is it usual that we will have that amount of free Con A out off Sepharose?
Without luck, I haven't got any extra bands in my elution fractions
with either 10mM Methyl-alpha-D- glucoside or 500mM
Methyl-alpha-D-mannoside. What went wrong?
All suggestions would be greatly appreciated.
Thank you very much in advance.
Mr Somchai Sangamnadech