On Wed, 9 Jul 1997, John Ladasky wrote: > Unlike monoclonal antibodies, PI has a fairly high off rate. So > having excess stain in the solution helps to keep all of the binding sites > of the DNA saturated, thus facilitating DNA quantitation. If you just want > to distinguish live cells from dead cells, you can probably get away with > washing away the excess PI. But not for long -- after a few hours the dead > cells would not fluoresce significantly more than the live ones. Hmm, is this true for unwashed cells as well? I've been using a one-step hypotonic lysing buffer that contains PI (50 ug/ml), but I've been varying the time (up to several hours) before reading on the FACS scanner. As such, would the PI saturate all nuclei, giving a reduced amount of hypodiploid fluoresence and sending everything up to the G0/G1 peak (thus giving abnormally low levels of apoptosis)? Ryan. _/ \__/ \__/ \__/ \__/ \__/ \__/ \__/rhung@vcn.bc.ca__/ \__/ \__/ \_Apoptosis=programmed cell death/ \__/ \rwhung@netinfo.ubc.ca \__/ \__ _/ --you can't live without it!/ \__/ \http://www.vcn.bc.ca/people/rhung \__/ \__/ \__/ \__/ \__/ \__/ \__/ \My words Copyright (C) 1997 \__
This archive was generated by hypermail 2b29 : Wed Apr 03 2002 - 11:49:55 EST