bone marrow - loss of CD19+

Katarzyna Ruckemann-Dziurdzińska kruck at
Tue Jan 8 14:09:17 EST 2008

Dear All,
again I ask for Your help.
Has anyone experienced the following phenomenon?
I analyse bone marrows from leukaemic children - the bone marrow is
anticoagulated with preservative free heparine and precessed within an
hour from collection - first I perform "a shortened diagnostics" - whole
marrow blood is stained for 20 min at room temperature, RBCs lysed, sample
washed with PBS and analysed - I get for example 95% of CD10+, CD19+,
CD34+ cells.
The rest of the bone marrow is diluted with RPMI and separated over the
Histopaque gradient. After separation cells are incubated with the same
antibodies at 4 C for 30 minutes and CD19+/CD10+/CD34+ constitute 2-3% of
all the mononuclear cells! The same surface stained cells are washed
twice, fixed with paraformaldehyde, permeabilised and stained for
intracellular antigens - CD19+s are 70%.
What happens to CD19 antigen or CD19 positive cells over the separation?
What does Hispopaque do to CD19?
And why does it happen only to the some of BMs?

I will gratefully appreciate any of Your comments.

Kasia Dziurdzińska

Katarzyna Ruckemann-Dziurdzińska MD, PhD
Pathophysiology Dept
Medical University of Gdańsk
ul. Dębinki 7
80-809 Gdańsk

tel. +48 58 3491511
fax. +48 58 3491510

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