Hello to all, Does anybody have any suggestions regarding lysis of red blood cells contaminating buffy coat preparations? I'm inclined to use distilled water because I want to use the polymorphonuclear cells in a subsequent chemotaxis assay (so anything with formaldehyde in it is out of the question). I've tried a number of different approaches, but the buffy coat preparation is very thick and seems to be resistant to hypotonic shock (even after dilution in 0.85% saline or PBS, or prolonged incubation at low salt concentration). Separating out rbc's using density gradients (Ficoll-Iso/Hypaque, before or after attempted lysis) doesn't work at all, but I was thinking of using dextran to get them to clump together or something (sort of like anti red cell mAbs). I also have a couple of protocols using weak Tris buffers, vinager or ammonium, but I thought I'd ask the list because I want something that won't affect the function of the polymorphonuclear cells. So does anyone have experience with this? Should I just give up and use whole blood? (I wouldn't get as many cells that way though....) Thanks for the help, Al Sabirsh One of these grad students who know nothing about whats going on their test tubes. Now where did I put that cell lysis kit?... (sorry, couldn't resist that :-)
This archive was generated by hypermail 2b29 : Wed Apr 03 2002 - 11:53:17 EST