Sorry to be posting so often lately, but..... With regard to lysing solutions. DO NOT refrigerate whole blood and expect the red cells to lyse properly. Some sort of irreversible matrix formation takes place (pure speculation based on observation) and most of the rbc's in the sample will not lyse without industrial strength chemicals, but then you are ripping apart wbc's also. Many times, if you share samples with a hematology lab, they will refrigerate the samples to preserve the rbc morphology (after 6 hours at RT, the MCV of the rbc's starts to climb). I only mention this as I have used BD's lysing solution for a very long time, and I have NEVER had it not adequately lyse 100 uL of peripheral anticoagulated (EDTA, Na Heparin, or ACD) whole blood when 2 mL of lysing solution is used UNLESS the sample was refrigerated. If you're having this problem and you're not drawing the samples yourself, you may want to check and see if they are refrigerated prior to transit?? As for NH4Cl, I have always seen CONSISTENT performance with this in non-refrigerated blood. The quirk is that it CONSISTENTLY leaves a pellet of rbc's. BUT, as NH4Cl does not shrink the cytoplasm of the cell (as lysing solutions with fixatives do), the lymphocytes are quite easy to resolve from the rbc's. Hope that helps somebody. I wish I knew this 5 years ago! -- Keith Bahjat Graduate Assistant University of Florida College of Medicine Gainesville, Florida kbahjat@ufl.edu
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