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I am involved with a clinical lab, that mainly preforms
immunophenotyping on peripheral bloods, bone marrows, and lymph
nodes. I alos have a few questions that I would like to throw out to
this forum and get some opnions.
1). NH4CL Lysing. Curently we are not doing this. We are using
coulter lyse and ficol for bopne marrows. I think our scatters look
like hell, and I want to switch over to this type of lysing(NH4CL). My
question is wether or not to lyse before or after staining with the
antibody. I would like to lyse prior to adding antibody, so I can
channel each of the different types of specimens into one common
process after lysing. I know most labs doing flow currently are
staining than lysing. If I do prior lysing, what problems will I face,
and are any labs out there doing this?
2) A quick question about staining. Is it necessary to incubate on ice.
Where I trained this was the norm, and I was told that it allowed the
antibody to react better with antigens, and that it prevented capping.
However, when I came to this lab, they were not doing this. So my
question: is incubation at 4C necessary? Can it be done at room
temp. or in the frig.
thanks in advance. I am interested in getting any opnions on these
two questions. If I get any direct replys, I will post a summary.
thanks:
===
robert rainer
department of pathology
bowman gray school of medicine
winston-salem, n.c 27157
919-716-4311
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