Lymph Node Prep

Jan Nelson j.nelson at auckland.ac.nz
Fri Mar 22 12:00:18 EST 1996


We have been following the discussion about lymph node preps with 
interest as we test a lot of FNAs although don't process whole nodes as 
immunophenotyping is performed on sections by immunoenzyme analysis.

We usually receive FNAs on the day of collection so don't have a problem 
with dead cells- collection into tissue culture medium rather than 
saline or PBS helped to preserve the morphology of cells and allow them 
to survive the Cytocentrifuge. We don't ficoll but if blood stained, 
lyse the specimen (standard ammonium chloride method) as this also 
produces better cytospins.
Obviuously the cell numbers vary enormously but if sufficient cells we 
run through the flow but don't check viabiliity. We lyse first with a BD 
formalin/water protocol and there is usually little debris and the CD45 
gives us an idea about the quality of the cells.
For any neophyte FNA testers,the potential pitfall of analysing by flow 
is ensuring that one is gating on the tumour population. Checking the 
morphology of the cells is essential as sometimes the tumour cells are a 
minority  eg. 10%-20% with a background of T cells; therefore  gating on 
the lymphoid population produces spurious conclusions. If the tumour 
cells can't be confidently identified in the flow graph we check the 
lineage and for light chain restriction by APAAP immunoenzyme. 
Jan Nelson
Department of Molecular Medicine
Auckland University School of Medicine
Private Bag, Auckland NZ
Phone:   (+64)(9)373-7599 ext 6381
Fax:     (+64)(9)373-7492
E-mail:  j.nelson at auckland.ac.nz



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