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Platelet disorders represent a very significant portion (directly or
indirectly) of human diseases. At present, clinical applications for
measurements of platelet number and function are a big portion of
clinical laboratory services. Flow cytometry, however, does not now
impact this area significantly. There are likely far more individuals who
will require platelet function assays than there have been, are, or will
ever need CD4 counts. Are we missing the boat (again?)? Perhaps.
We have been using flow cytometry to analyze human platelts for the past
two years (in collaboration with Dr. Jawed Fareed at Loyola University).
Our studies have centered on platelet activation, using platelet
surface expression of specific epitopes (CD62, CD63, gpIIbIIIa), and on
the interactions of platelets with other blood cells. I cannot speak on
the issue of platelet bound Ig, as our preliminary efforts in this area
were perhaps as frustrating as those commented upon by others. However,
the studies of platelet-blood cell interactions are likely to yield
important information, as this is probably the ONLY method to study these
interactions up "close and personal". We are now up to 3 color analysis,
not for the sake of doing more colors than anyone else, but to understand
the role of platelet-cell interactions in platelet activation.
For the sake of the discussion regarding platelet bound Ig, I would like
to share some information that may prove useful to others working in this
area. Our experience indicates that platelets do NOT like centrifugation,
they perfer 37 deg. C, and if handled at 4 deg C are likely to clump. We
do ALL our work with whole blood (at 37 deg C)- we see no reason to prepare
PRP, which increases the level of platelet activation (significantly!).
Like all "platelet" people, we have adapted "type A" behavior when harvesting
platelets- no tourniquet, NO VACUTAINERS, double syringe, and we use
recombinant hirudin as anticoagulant- this keeps platelets as quiet as
they will be for as long as possible. While our studies are aimed at
looking at platelet activation, I suspect that the platelet activation
state may impact on the binding of many proteins to platelets, including
antibodies, or passively, Ig.
While I recognize the problems involved in working with platelets (smart
flow people have ALWAYS gated out these buggie creatures), I would urge
those working in this area to carry on, and those watching from the
sidelines to jump in! This could be an important contribution by flow
cytometry to basic platelet physiology and clinical lab medicine. Go with
the flow.
Vince Shankey
Loyola University Medical Center
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