Dear Sarah, 2 thoughts. Or maybe more I suck at counting: 1. One time (OK maybe 2 times) when I ran a burst test I diluted the lysing 10X stock in PBS instead of DI water (no hypotonic solution, no lysing). 2. Young patients (<2 weeks old) and preterm babies with frequent transfusions are IMPOSSIBLE to lyse due to their evilly sturdy RBC's 3. Certain drugs, such as iron chelating agents in Thalassemia patients will hinder lysis. Solution: 1. ensure the right hypotonicity of the lysing solution 2. make sure it's not dilluted out by an increase in sample volume for whatever reason 3. extend the time of lysis by 1/2, then 2X if that doesnt work 4. try lysing in a waterbath, 37oC works wonders 5. try a different lysing solution 6. if you have enough blood and are good with the technique - separate the lymphs over ficoll, then lyse. This is really evil to the cells and even RBC free preparations look crappy. This doesn't work in thalassemia patients their RBC's float on ficol with the rest of the pack. Hope you solve your problem! if you do please post the answer. Maciej __________________________________________________ Do You Yahoo!? Talk to your friends online with Yahoo! Messenger. http://im.yahoo.com
This archive was generated by hypermail 2b29 : Sat Mar 10 2001 - 19:31:05 EST