Sanford

J Slialtil

Clinical vation

and

pathological

occurs

and

vascular

disorders,

disease,

and

Therefore,

cilitate

identifying

therapeutic

including the

fulfill

migrate

from

the

platelet

platelets

factor

boxane

A2

helpful

(1

and

have

platelet

tiplatelet antibodies were rect detection of activated (2-4).

several

different

conversion

Use

ofthese

receptor, aggregation:

three

sequential

clinical

developed in whole

antibodies

aspects

of the (GP)

several

into

information

phases

by using

antibodies

ligand-binding site within ligand, or for neoepitopes

on

process: a competent

a process required for normal GP lIb-Ilia activation can

posed adhesive receptor-bound

an-

that permit the diblood by flow cy-

provides

lIb-Illa

biochemical

1) adhe-

platelet adhesion be dissected into specific

for the

ex-

GP IIb-IIIa, for the within GP lib or lila

be emphasized. platelet

preparation. to fix blood or after

during

some with

circumparafor-

Under samples

adding

monoclonal

antibody.

rather

than

Second,

is usually

conjugated

chromophore

for

polyclonal

preferred as detection reagents to minimize binding. Third. the activation-dependent to the

First,

activation

bodies are cific-antibody

antinonspeantibody

fluorescein

so that

antibody binding can be quantitated by measuring platelet-associated fluorescein fluorescence in the flow cytometer. When the simultaneous binding oftwo antibodies is assessed, the second antibody is conjugated to biotin, which is detected by flow cytometry after phycoerythrin-streptavidin is added. The use of conjugated antibodies

has

been

before

should

inadvertent

and sample be necessary studies,

maneuvers

collec-

points

to avoid

either

cell activation whole blood,

monoclonal

platelet-activation

maldehyde,

directly secondary

use.

and

drawing it may

of throm-

to sample

physical

surface,

recently platelets

ofglycoprotein

sion and

the

the

reliable released or urine:

markers

related

vein

by yeremain

plasma

metabolites

these

problems limited

1) activated

technical

be taken

whole-blood

is no

to the

Several

must

blood stances fa-

There

The most substances

in the and

understanding

might

care

evaluating

not be activated 3) platelets must

is completed. in vivo are

plasma.

on the basis of circulating acti-

of activation

measuring

at the activated

tometry

and

criteria:

measured

technical

processing

facilitate

changes

and

Although

).

throm-

activation.

must and

4, [3-thromboglobulin,

in research,

To

site

2) platelets manipulation,

stably activated until the assay markers of platelet activation activated

vascular

platelets

following

acti-

in several

peripheral

disorders

platelet

the

platelet

or coronary

of activated

thrombotic

to prevent

that is being sampled, nipuncture or in vitro

from

angina,

to assess thrombotic risk The ideal assay to detect

must

must

that

significance

angioplasty

detection

strategies

platelets

after

certain

platelets

indicate

unstable

and

good in vitro assay platelet dysfunction. vated

studies

is of pathophysiologic

stroke,

bolysis.

tion

thrombotic of using flow

likely

platelets

are

staining

with

ylated

antibodies eliminates the need for adding and for sample washing or centrifugation, to traumatize

platelets

in vitro. Fourth, when a two-color technique discriminated

from

red

an activation-independent

antibody,

flow-cytometer particles: thus,

such

as one

threshold only GP

that

and

induce

platelets is usually and

white

blood

platelet-specific recognizes

GP

microparticles) are analyzed. with a fluorescein-conjugated

dependent

to assess

antibody

activated including

flow cytometry platelets in only cardiopulmonary

cells

by

biotin-

lb. Then,

the

is set to detect phycoerythrin-positive lb-positive particles (that is, platelets

and platelet-derived stained simultaneously

To date,

unwanted

are analyzed in used in which

platelet

Platelets are activation-

activation.

has been

used

to detect

a small number ofclinical bypass and angioplasty

questions must be addressed before the technique wider scale: Can platelets activated in vivo routinely

circulating situations, (2). Several is used on a be detected

exposed during ligand binding; 2) exocytosis-related surface exposure of integral membrane proteins specific for either alpha, dense, or lysosomal platelet granules; 3) surface binding of pro-

by flow cytometry? What is the precise sensitivity and specificity of the flow-cytometric assay compared with existing plasma and urinary assays for platelet activation? Is one activation-dependent

teins, such as thrombospondin, released during platelet secretion; 4) development of platelet procoagulant activity, manifested by the surface expression of binding sites for coagulation factors Va and Villa on both activated platelets and platelet-derived

antibody suming

microparticles Flow whole

.min

(5).

cytometry blood

i Cliii

Nuir

and

‘From 2

can plasma

be

used or after

1992:56:789S-90S.

to study

platelet

separation

Printed in USA.

activation

of platelets

©

in from

1992 American

better than that platelets

Address

the Hospital reprint

pital of the University PA 19104.

another activated

for a particular clinical study? Asin a given disease process enter

of the University requests

to Si Shatill,

of Pennsylvania,

Society for Clinical Nutrition

of Pennsylvania,

Philadelphia.

Hematology-Oncology,

3400 Spruce

Street,

Hos-

Philadelphia,

789S

Downloaded from https://academic.oup.com/ajcn/article-abstract/56/4/789S/4715609 by University of Michigan user on 11 January 2019

Why is platelet activation useful for assessing risk? What are the advantages and limitations cytometry to measure platelet activation?1’2

790S

SHATTIL

the general circulation, how long do these platelets circulate? These questions are being addressed and willhave to be answered before flow cytometry gains widespread use in clinical studies of thrombotic disorders. 13 References

Blood

1990:75:128-38.

5. Sims Pi, Wiedmer T, Esmon of the platelet prothrombinase the platelet plasma membrane. defect in platelet procoagulant 17049-57.

CT, Weiss HJ, Shattil Si. Assembly complex is linked to vesiculation of Studies in Scott syndrome: an isolated activity. I Biol Chem l989;264:

Downloaded from https://academic.oup.com/ajcn/article-abstract/56/4/789S/4715609 by University of Michigan user on 11 January 2019

I. FitzGerald GA, Catella F, Oates IA. Eicosanoid biosynthesis in human cardiovascular disease. Hum Pathol 1987:18:248-52. 2. Abrams CA, Shattil Si. Immunological detection of activated platelets in clinical disorders. Thromb Haemost 199 l;65:467-73.

3. Shattil Si, Cunningham M, Hoxie JA. Detection of activated platelets in whole blood using activation-dependent monoclonal antibodies and flow cytometry. Blood 1987:70:307-15. 4. Abrams CS, Ellison N, Budzynski AZ, Shattil Si. Direct detection of activated platelets and platelet-derived microparticles in humans.

Why is platelet activation useful for assessing thrombotic risk? What are the advantages and limitations of using flow cytometry to measure platelet activation?

Sanford J Slialtil Clinical vation and pathological occurs and vascular disorders, disease, and Therefore, cilitate identifying therapeu...
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