Whole blood platelet assays

Dan Smith DJS at ALLP.COM
Tue Feb 18 14:42:00 EST 1997



{From: owner-cyto-sendout
{To: Cytometry Mailing List
{Subject: Whole blood platelet assays
{Date: Friday, February 14, 1997 1:23PM
{
{
{Greetings,
{
{I'd be interested in corresponding with anyone who does flow cytometric
{studies
{on platelets using a whole blood methodology.  My short-range interest is
{specifically related to platelet-associated immunglobulin assays, but if 
you
{have experience with platelet activation assays, I'm sure your insights 
would
{be valuable.
{
{Please correspond by e-mail initially.  If you send a phone number, I'd be
{glad
{to call you at your convenience, if you prefer that medium.
{
{TIA
{
{Marc Langweiler
{marcl at dartmouth.edu
{
 --------------------------------------------
On Feb.21,1996:
Frederic I. Preffer wrote:
i would like to take a look at human platelets. do those of you with
experience use a whole blood technique, or density gradient to enrich? Any
other tips appreciated.
`````````````````````````````````````````````````
We tend to do a fair amount with platelets so we use multiple techniques.
We use whole blood for all in vivo platelet studies and for corresponding in 

vitro experiments to get a "snapshot" of platelet receptor and ligand
expression, platelet activation by expression of P-selectin, CD63, or
granulophysin, and platelet adhesion to leukocytes. We have had good luck
with this approach since nearly all of the platelet antigens that we study
are stable after fixation and with the right marker combination, the
antibodies are highly specific for platelets.

 For functional studies (serotonin release, Ca flux, RNA content, etc) in
which we want to study all platelets, we usually purify platelets since
there is always significant  binding to leukocytes when platelets are
resting and a whole lot more leukocyte binding with platelet activation
(EDTA will block activated platelet-leukocyte adhesion but it tends to
disrupt receptors that you want to study). Depending upon the application,
we use either arabinogalactan gradients to isolate close to 100% of
peripheral blood platelets from a sample or density centrifugation for
platelet-rich plasma (which is less stringent but much less of a hassle).

I'd be happy to discuss specific applications and methods.
Harv Rinder
Ph. 203-785-4231
Fx. 203-737-4111


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