Hello all,
Does anyone have a rough estimate (or has anyone actually determined) an
average dead/delay volume between buffers run on an AKTA Prime FPLC? We are
attempting to overexpress/isolate a smaller His-tagged transmembrane protein,
and require running several detergent buffers in succession over the column.
This obviously creates fractions that are just dead volume/ddH2O between
buffers, and we would like to narrow in on where to look for the protein prior
to analyzing the fractions (i.e. how many fractions can we discard?). Should
we run ddH2O and then the first mL into 'waste' before running each buffer over
the column (and collecting fractions)? Just not used to this system yet!
Thank you!
Jim
