INFLUENCE

States

OF TIME,

TEMPERATURE

AND

CONCENTRAT I ON ON DOG PLATELET

D.

Brackett, C. F.

J.

Department Health

8, PP. 441-451, Pergamon Press,

Vol.

THROMBOSIS RESEARCH Printed in the United

of

Sciences

and

University

Center,

PLATELET

AGGREGAT I ON

Schaefer

Medicine,

1976

Inc.

Oklahoma

C.

G.

of

Oklahoma

City,

Gunn

Ok.

73190

(Received 23.12.1975; in revised form 7.2.1976. Accepted by Editor P.M. Mannucci)

ABSTRACT Previously in

encountered

laboratory

physical

and

factors

canine

variability

clinical that

platelets).

various

affect Valid

conditions

of

this

technique

tration 90

and

min

stable, in

ed

the

not

by

Choice

amplitude 21

after

temperatures. aggregation though,

platelet was

in rich

stronger

after

Comparing response

that

of

hr) by

30

short

term

freezing

at or

agents, significantly

equal weights of the resulting molar ADP

(30 uM EPI

vs

concen-

For the aggregation

first was

after

temperature

two affectwas

temperature

(37OC and 40°C)

average greater

amplitude than to

agents per volume concentration of

21

but

(EPI),

aggregation

room

body

the

Using

platelet

uM epinephrine

storage

the the

below.

EPI-induced

(OOC)

under

occurred

storage

(ADP).

aggregating to ADP was

in administering plasma (PRP),

platelet

common

calibrating

between

aggregability

induced

at

by

response. of

(one

diphosphate

storage

than

platelet

short-term

aggregation

greater

than

even

of

uM adenosine

significantly (20°C)

in of

found

of

(specifically,

described

aggregation

magnitude

study

responses

possible

method

was

responses

this

platelets

platelet

made

the

the

the

deterioration

vitro.

using

of

of

of

was

relationship

amplitude

venepuncture,

but

hr

a direct

after

study

aggregation

prompted

comparison

system

the

platelet

aggregability

this

aggregometer-recorder

in

investigations

of EPI of EPI

uM ADP).

INTRODUCTION

ometry to

The in vitro measurement of platelet (1) is enjoying an expanding usage.

clinical

logical

and and

experimental

biochemical

hematology

factors

In experimental genetically determined

hematology, bleeding

Very

about

little

is

known

canine

which

aggregation

in

increase

the dog disorders platelets 441

by

continuous

This technique is of the study of physical, or

decrease

is a useful (2,3,) and beyond

platelet

model for coagulation

their

reduced

turbidgreat value physioaggregation.

the study of mechanisms reactivity

(4).

443

ON DOG

when

compared

platelet rich

with

plasma

ability

present As

and

serious

of

this

study

are

from

venepuncture

fully

physical

in

(8),

Gaetano

be

used

factors

as

istered

de

for

equal

From If

needle. the of

the

sample

syringe ferral tube

blood or

to

cuvette.

All

placed

in

filled

directly

the

and

aggregometer. from

Centrifugation plasma the

amount

observed visibly When

of at

this

PRP

point

blood

whole

3500

blood in

RPM to

and

at

37°C

Since

by there

were

marked

from

G) the

epinephrine ml).

from

differences

or

allowed

produce

time

admin-

was

gently

stoppers

were

for

by

covered

used

blood

a

ratio

tilting.

plasma

with

the

the in

down

or

vein

blood, from

streaming

whole

limb clear

(.13M)

mixed

blood

always

a

to

free-flowing

immediately

for

in

10 min

highest

to

Trans-

the

side

was

freshly

cover

platelet

of

the

parafilm

produced

platelet

centrifuged the

point

plasma rich

at at

until

counts

were

water

in

1% ammonium No.

bath

the

red

G)

and

cells

had

coat had formed. removed using a except

method collected

oxalate The

poor for

when

of

the

effects

Brecher in self-

and

contained in amount and extent

in plasma turbidity was measured Aggregometer coupled to a chart While in the aggregometer, the a Teflon coated stir bar and was

heating in

5855).

platelet possible

825 RPM (55

which

and a buffy plasma was

a 37oC

the

yield

using the phase contrast for platelet counts was

temperature

(EPI),

a jugular

were

with

was

and kept studied.

diluted

2)

storage temperature. studied using two

needle.

(986 get

concentration,

citrate

(Becton-Dickinson,

a constant

of

aggregation

standardization have evolved

transferred

of platelet aggregation reflected by a change photometrically using a Chronolog 300 Platelet recorder (Texas instruments Servo-Riter I I). plasma was constantly stirred at 1200 RPM by held

7)

3.8% sodium

pipettes

amount of the platelet

were made or plasma

tubes

platelet

contact

Teflon

blood

reservoirs

platelet

This variwhich prompted

platelet

the that

careful

were

determine

reached,

and

by

cuvettes

to

was

dog

of

independent

of

blood

and

Adams).

the

required

I)

drawn

done

intervals

the

of

PRP.

6,

also

3) short-term factors was

of

was

always

The

attempt

same

calibration

immediately

required,

counts Blood

capillary

polyethylene

not

intravenous

Pasteur pipette temperature were

Platelet Cronkite (9). filling

an

2 min

cleared

siliconized of storage

at In

(PPP).

the

was drops

glassware the

samples

METHODS

containing

was

Clay

times,

AND

blood

did

9 parts

plasma

all

of

The

or

of

method preparation

and three

these

several

tube

(Siliclad,

at

ml

discarded.

(See

diphosphate (ADP) and volume of PRP (5 ug/O.5

after

a siliconized

siliconized tubes

30

the

comparability

were

test,

by

venepuncture

citrate

of

the

was

to

1 part

dogs

syringes

on

a method

investigated

aggregation

adenosine weights per

fasting

plastic

serial

aggregation.

valid

MATERIALS

into

observations in

below.

influenced

agents,

preliminary

questions

platelet

described

to

Aggregability ing

Our

variability

determinations

of

VoL.S,No.4

AGGREGATION

The aggregometer calibration blood handling and platelet

techniques

aggregat

multiple

study

must

measurements.

(5). marked

methodological

by

variation

The

in

parametric

suggested

sample

species

disclosed

(PRP)

raised

the

other

aggregation

PLATELET

device. the

photodensities

of

the

sil

iconized

04 DOG

cuvettes,

they

cuvette’s

orientation

Before the

optical 100

of water The

scale.

Changes

in

PRP baseline

b)

platelet

The

gently was

density amount

being been

the

and

or

then

recorded

tilting in

to

stabilize

in

this

each

trial

This

to

optical

by

between

density

of

the

a cuvette

of the recorder set at its to the 10 mv scale for the

the

in The

density

time

the

tube

the

aggregometer,

of 30

were

aggregometer.

in

difference

at the calibration

for

(expressed

optical

(the

of and

0.4

ml

and the

set,

millivolts critical

between optical

was

to the method.

pipetted

after

baseline

challenged At

of deflecreadings of

PRP and density

peak response the recording

PRP,

determine

if

consists

of

modification of

of

any

the PPP) between

and PRP

aggregating

into

the

optical

was

recorded

a cuvette. density

of

for

with the aggregating this time the temperature

spontaneous

platelet

PPP

5 separate of

count

from

the

the

on

same

series

above

the

1 min.

agent of

90 set 37’C

to

Duplicate

counts

were

the

accuracy

dilutions

The

final

To

of

Samples

increase

less

than

EPI

from

di lution

were

of

of

each

had

a final

at

of

ADP

each

of

or

the

twice

and

5 ug

storaqe 1 hr at EPI

these

were

used

were established The plasma used

the

Relationship quots of

the

by dividing to obtain from

of platelet PRP were tested

on

usual

each

of

30

test

of the

amount

of

varying

dilutions

the

platelet

means

platelet

by

the

with

platelet

were

aggregated in

diluted

and

the

counts

ADP

to

requiring

of

these

counts

from

PRP

PRP was

added

to

4

halved.

5 ug

cuvette

of of

either

ADP

PRP was

21

or

EPI.

uM while

uM. on platelet of O°C, 20°C, platelet

aqqreqation. 37OC, or

aggregation

in

4O’C.

Al iquots Five

PRP samples

temperatures. parameters procedure

derived

of

the

temperatures temperatures

Effects of centrifuqation of a standard centrifugation

was

in

each

5 different observers

subsequent

concentration

Influence of short-term of PRP were stored for amounts

70%,

reservoir

concentrations

of

occurred

experiments,

counts

trained

diluting

had

PRP was

produce

two

used.

of

PRP photodensity.

blood

71%, 56%, 45%, and 36%). “bl ind” by were done

the

shift

of were and

method.

(lOO%, dilutions

ment

calibrated

system.

study

Influence amounts

ug

was

difference

by the PRP from all storage temperatures. The aggregating in 0.1 ml of isotonic saline was streamed down the side EPI) into the PRP. The changes in photodensity during aggregation PPP base1 ines were recorded before for 5 min (See Fig. 1).

follmting

stored

the

aggregation.

in

study

in

aggregometer-recorder

5 ug

the

each

reached

after

with

that

system until

chart range turned up

PRP

platelet

response

placed

agent (ADP the cuvette

the

of

of

difference

placed

allowed

ink

so

same.

measurements.

PRP samples

after had

the full was then

and the same plasma were based upon this

PRP was All

black

(the

After

marked

the

aggregometer

to

aggregation

cuvette

and

always

aggregometer-recorder the

of

the

a)

density

a cuvette

optical

reflected

baseline agent)

the of

Ic’c3

AGGREGXTIOX

was

was equal amplification

aggregation

tion)

optical

photocell

experiment,

density mv

for the

amplification

distilled

platelet

matched to

each

adjusting of

were

PLATELET

blood

on

platelet yield. To test the reliability for obtaining PRP from dog blood, ratios

PRP platelet PRP platelet centrifuged

response to for platelet

counts count at

825

time elapsed aggregation

by whole for the RPM

(55

from with

G)

blood ratio for

blood 5 ug

platelet in each

counts. experi-

8 min. collection. of ADP and

Ali5 ug of

iii

04 DOG PLATELET

after the PRP and of blood collection.

EPI immediately hours from time ADP

dose-response

Immediately with

1.0

phasic

ug

(4

curve,

the

standard

ADP

dose

curve

threshold

after

uM ADP)

a

function

of

were

PPP were

of

PRP and

added

to

aliquots

portending

maximum response

for

the

until

the

amount

was

adjusted If

found.

the

aggregation,

standard

curve

prepared

time

PPP,

aggregation

level

was

as

preparation

the

AGGREGA,TIOX

from

I

I

0

I

I

5

collection.

used

not

obtained.

Thi_s

At

achieved the 4 hr

was

10 min

to

I

I

for

was

sufficient

2

I

blood

hour

starting mono-

experiment.

was

each

increasing doses of ADP of PRP until a fast-rising

duced to determine the lowest dose of ADP yielding determinations were carried out for approximately

1

and

intervals

produce

the

the

next

as the

standard

dose

was

re-

standard curve. These after blood collection.

I

I

3

I

I

4

J

5

MINUTES

FIG. Aggregation

responses

diphosphate ing agents curves The from

(ADP) and in PRP was

were

aligned

amplitude baseline

following

of

of

at

the

to

(EPI). ADP and

point

(0

aggregation

density

1

platelets

epinephrine 21 uM for

platelet

(PRP

dog

prior

equal weights (5 ug) of adenosine The final concentration of aggregatThe two super imposed 30 uM for EPI.

min) was

to

0 min)

where

ADP

measured to

curve

or

EPI

in

millivolts

peak

was (most

added of

to

the

extreme

reading

0 min).

RESULTS

Correlation platelet

of platelet concentration

count with in platelet

PRP baseline rich plasma

photodensity was reflected

PRP.

deflection

levels. The in photodensito-

ON DOG

metric

and

statistical

IO

experiments,

count

across

factor

(10)

which

was

the all

used

otherwise

AGGREG_ATION

partial

correlation

5 PRP dilutions to

partial

would

have

of

was

out

the

PRP photo-

0.65

(P

influence

spuriously

c.01). of

inflated

this

This

the

PRP

and

follow-

correlations. The

to

platelet technique

dilution ing

In

baselines.

density

PLXTELET

be

PRP baselines

2 groups

of

The

identical. across

influence of tion response

all platelet with

was

number increasing

(P

aggregation

amp1 itude

< .Ol)

aggregation

produced

2 al iquots

by

significantly

of with

induced

used

the

with

ADP

baseline

and

EPI

were

found

photodensities

for

the

0.98.

analysis

more

be

of

The

on aqqreqation. PRP concentration

produced was

to

correlation

dilutions

significant whether

the

partial

by

variance or

aggregation

in EPI.

(P

increases

2)

(11).

increases ADP

1 inear

(Fig.

The

platelet However,

e.02)

than

in

were

found

mean

increase

numbers

5 ug of did

5 ug

aggrega-

to

in

was

the

ADP of

be same

(21

EPI

uM) (30

uM)..

OA DP(5yg)

4.0

3.0

2.0

1.0

..

0

40 PRP

The

influence

of

platelet

Each point represents and epinephrine as line

to

Influence Similar over

the

peak

of platelet fluctuations course

age amplitude of response to 5 ug response induced

of

FIG. plasma

rich

in

millivolts.

rich plasma in platelet 5 hr

from

80

100

CONCENTRATION

the average the aggregating

response

60

the

2 concentration

(%I

on

the

aggregation

of 10 experiments using adenosine agents. Aggregation was measured The bars indicate S.E.M.

storaqe aggregation time

of

time blood

on aqqreqation occurred for both co1 lection

(Fig.

response. diphasphate from

responses. ADP and 3).

EPI

The

platelet aggregation was consistently greater (P c.01) the aggregat Furthermore, of ADP than to 5 ug of EPI . by either ADP or EPI decreased with time elapsed from

PRP base

aver-

in ion blood

OS DOG

col

lect

ion,

although

collection significant

groups

results

were

data

at

at

4

5 hr

analyzed

in hr

C.01)

c.05)

for

Dunnett’s

collection

occurred

aggregability

(P

(P

using

from

AGGREGATIOZ

recovery

decline

2 and

and

I hr

at

transient

This

only

treated the

a

time.

PLATELET

t

found

after

blood

the

ADP-treated for

as

the

3 hr from

approximately was

Test

serving

k-01.8,so.G

to

be

statistically

collection

for

EPI-

groups.

These last Comparisons (11) with

Planned comparison

standard.

I 5.0

4.0

_

3.0

2.0

1

1.0

(5pq) (5JJg)

OADP .EPI

1

I

I

0

60

I

120

I

I

180

240

300

MINUTES FIG. Influence sponse of

to

elapsed

time

adenosine

term

The

storaqe

a definite

more responses

ADP

dose-related

Fig. the

5 demonstrates time of collection.

necessary

to

on

when storage

Furthermore, able. ADP produced reliably

constant (1 .O t ion. Within

bars

or

on

indicate

influence

on

aggregation

5 ug of each of PRP for

aggregating 1 hr at room

as

These a maximal

ug or 4-12 - 3.0 the period from

a

function in

point

aqqreqation.

EPI-induced

change

average

Each

aggregation

re-

respresents

a mean

S.E.M.

after 1 hr storage greater aggregation

the

the

epinephrine.

aggregation than after after storage at O°C,

threshold

produce

3

venepuncture

temperature

effect

induced aggregation With epinephrine, in significantly the platelet

after

diphosphate

9 experiments.

Short had

of

platelet

at

and

Storage no

effect

agent was temperature

temperature on

the

ADP-

used (Fig. 4). (20°C) resulted

storage at 37OC (P c .05), while 37oC, and 40°C were undifferenti-

all temperatures responses than of

time

after

sensitivity

data demonstrate and monophasic until uM ADP) 100 to 200 min

did blood over

that the aggregation approximately after blood

but 20°C, 5 ug of

5 ug of EPI.

collection. a 4

smallest curve

hr

period

from

dose of ADP is relatively

90 min collection,

from collectwo to

OS DOG

L-Ol.S,SO.ir

ten

fold

increases

platelet to

in the It is

response.

produce

maximum

‘3 E

5.0 I-

began

to

to produce the the dose of ADP to baseline after

decrease

IADP

(sing)

mEPI

(Spd

r

W

4.0

::

maximal sufficient 200 min.

I

u) ;

AGGREGATIOS

dose of ADP were required interesting to note that

aggregation

Y

iii

PLATELET

3.0

z 0

;

2.0 I-

a 0 ;

I.0 I

-

W W a

0

STORAGE

TEMPERATURE

4

FIG. Mean

amplitude

of

four

temperatures

sine

diphosphate

of

yield

for or The

9 experiments.

Platelet in platelet

aggregation

storage of platelet PRP was aggregated with

hour.

epinephrine.

Each

bars

S.E.M.

(55

ranged from from 1.3 to

centrifuged

one

indicate

as a function of numbers was observed

centrifugation

counts ranged

after

G for

0.7 2.2)

repeatedly

bar

represents

8 min).

The

Less when

2 min

ratios

of

variability blood

periods

at

PRP in

from

rich plasma at 5 ug of either

the

centrifuqation parameters. in 10 experiments when

to 2.6. occurred for

rc)

55 G until

response

High using to

the

these

mean

whole

1 ity

parameters

blood

platelet the

in

variabi

fixed

same

one of adeno-

platelet

counts

(ratios

10 experiments PRP yield

was

was

ade-

(18),

time

quate.

DISCUSSION

In

vitro

from physical (Figs.

time.

venepuncture variable 3 and 5).

As

previously to

influencing The

observed

aggregation pH of

in

test

(in

platelet plasma

man

(12,13)

vitro

time)

aggregation has

been

shown

and

in

was

a

rabbit highly

determinations to

cause

variability

important in.dogs in

ON

4is

platelet

aggregation

enhances

platelet

To

minimize

by

keeping

I vessels

it

changing

aggregation

cannot

nature

of

The min

this

the

CO2

an

increase,

completely

response

in

a

rising

pH of contact

pH which

8.0

with

S i nce the pH of out as a contributing

ruled

curves,

results

extensive

(Fig.

\‘01.8,?io.4

aphysiological

(16,17).

responses

present

from

results

Although

mechanism

tion

tests

and

certainly

is

air

the

reached.

was

avoided

PRP was not factor in the

but due to the precautions seem to be other influencing

3), there

of

aggregation

since

platelet

aggregability this

conducted more

rebound during

temporal

pattern

phenomenon

be

constant

and

late

As of

stated

tests to is

taken

and

factors

suggested in

in

platelet

by

performed

vitro

and

be

(6),

Mannucci

3 hr

the

less

who

in

aggregation

in

after vitro,

consequently,

will

aggregability,

within

markedly

after

known,

in

be

decreases

baseline

not

period

change all

should

aggregability

returned

this

variable.

similar held

indicate

venepuncture

time.

the

Escaping until

covered

be

AGGREGATIOS

(13,17).

involved

90

(14,lS). alkalinity

measured the

PLATELET

aggregation

this al

DOG

aggrega

meaningful

reported

vitro

time

a should

investigations.

IO -

a :

g 8-

L

0

7-

z

6

Q

5’

a c3

4-

0 “0

3-

I

* I -

L’l”““““““““““’ 30

0

60

90

120 AFTER

MINUTES

(ADP)

of

time

required

to

after

venepuncture

produce

on

the

amount

the standard aggregation The 4 curves depict minimal venepuncture concentrations

for

was

the

aggregation

In

vitro

sufficient

temperatures.

to

produce

Platelet

210

240

VENEPUNCTURE

and Methods Sect ion). determined every IO min after ranged from I to 10 ug (final not

180

5

FIG. influence

150

standard

aggregability

of

of

adenosine

response effective

PRP from 4 to 40

in

diphosphate (See Materials doses of ADP

4 dogs. Doses of ADP uM). alO ug of ADP response.

the

dog

was

influenced

by

03 DOG

VOl.8,SO.4

short

term

storage

concentrated temperature

on

(18)

stronger

after

ture.

This

induced than

of

after

at is

very

similar

was

markedly

tures

at

body

found

in

the

after

storage

In

and

view

factor platelet

The agent

must

be

son

of

amount is

of

ADP

are

in

decreased

assure Aqqreqat

uM EPI;

volume

to

to

human

time reached

21

ADP

in

of

EPI

dogs

in

was

Extreme

taken

released

was the

of

can

be

ADP

The also range

be

it

the

can

cause

relationship found to be of platelet

Although measured yields

at

low

when were

whole

to

to

have

tempera-

optimal is

analysis

the

21

a

of

series

(23).

of

the

findings and

that PRP as

the the

uM of

was

response

in mean experiments platelet platelet Born

and

during

aggrega-

Cross

is

(21)

density

withdrawal

PRP.

and

PRP optical recording

and

(16).

concentration

and

aggregation

response

was no chang in aggregabi lo4 - 65 x 10 2 /ul). O’Brien reactivity as a function

independent

high concentrations obtained in this

com-

This using

may be used to shown for human

aggregation

platelet method.

30

platelet 2-4). when

optical

the

to

with

rich plasma concentration.

PRP baseline

uM of

biphasic

platelet

in

30

average,

obtained

If erythrocytes are lysed the resulting in a false PRP baseline

(8 x

vs

the

and of

typical aggregaof complete

affected

there

doses

result

amplitude of (See Figs.

the aggregometer Zucker (22) has

microhemolysis

comparilow

37oC

ADP on

The curve

uM EPI

of the

at

proved,

uM EPI.

30

of

when

care be taken to agent is added.

monophasic to

aggregatif

adenosine diphosphate final concentrations (21

30

the

that,

aggregometer

uM ADP

than

count

avoid

before

shown

molarities

response

between platelet linear. Thus, concentration

aggregability

the

controlled

(5 ug) of used, the

EPI,

platelets

suggested of dog

blood

greatly

(20) in

in of

platelet is count

al

photodensity plasma reflects

also unable to find differences numbers using the optical density

easily platelet

were

aggregability

aggregated

strictly

a maximal monophasic by the differences

similar

the

centrifugation

while

aggregometer and

a steep-sloped

the

The poor

the

weights ml) were

common

Rarely was is reflected and

must

dog

most

Since are 1 inearly related, determine the platelet

density

and

EPI

temperature other hand,

responses

37oC,

They also suggest that before the aggregating

differed

uM ADP was

plasma.

care

et

37OC

molarity

the

in

intervals

Although equal of PRP (0.5

Platelet concentration. pared with the platelet observation

stored

consistent

Praga

agents

fact

remains

kept

made.

aggregator

this

PRP be

higher

while Fig. I).

responses

the

aggregation.

the

response

aggregation or

tempera-

desired.

be

aggregating

aggregation, tion (See

where

temperature

PRP has

Despite

tion

to

aqents.

a stronger

also).

20°C

were

dog

as

time

platelet

per

two

EPI).

at

platelets

increasing

the

inq

platelets

body

results,

should

used,

epinephrine these

of are

that

O°C,

ADP-induced

was

at

at room On the

less

storage Con-

term

is

added

curves

of

the

I hr

for

(and

that

in

of

of

platelets

storage

results

storage

temperature

rabbit

after

present

after

human

of

than

room temperature is recommended storage of PRP. storage Obviously, controlled if comparative or quantitative

short

aggregation

ing

the

Use

aggregation. a similar effect was not investigated.

aggregation

to

when

these

for

which

revealed but this

OOC).

of

temperature

EPI-induced

temperature

stronger

inhibited

(IOOC

room

similar

was

storage

for have

ADP-induced

storage

(19)

might

439

AGGREGATIOX

aggregation,

that

finding

only

ADP

ADP- induced found

aggregation

O’Brien

be

temperature

solutions

stantine

PLATELET

of study

of

PRP concentration,

platelets using the

it

I ity

within

(7) of

is

are available. method of variable

was platelet

most High

OX DOG

4jO

centrifugation

time

richer

in

However

,

yield,

requires

less

may time

at

temperature

both

optimal

rapidly run

The and

This H.T.S. Heart

it

Jo

work

was

stored

be

carefully

thank

consistently this

method.

variabi

1 ity

this

method

since

be

W.

Dorothy

Ansten,

and

in

Heart

related storage

to

as

if

It PRP

M.D.

gave

by

for

Barbara

the

Latta

his

prepared

critical

and

review

David

Warner,

assistance.

Brain

Grant

and

suggested

(20°C-230C)

Blouin,

valuable

Association

the time

is

is

at room temperature venepuncture.

Hampton,

as

storage

the

to

PRP concen-

well

controlled.

obtained

Hall

supported

prerequisite

aggregometer,

aggregability,

Grady,

John

the

directly

plasma and

stored min from

is

using

influence

will

James

Hodge,

of

selected

90

Oklahoma

Research

may

described,

partially

5351-14,

PRP,

concentratration is

duration

is

to

Ann

of

necessary

aggregability response

the

within

wish

even

PRP sample

responses

Louise

Jessup,

of

since

aggregometer

authors

amounts

always obtained with time, despite greater

and

platelet

aggregation

techniques

ass istance.

Janet

of

should

the

the

preferable

the

aggregation

by in

be

of

which

were

attention.

Furthermore,

temperature

Required

blood,

knowledge

amplitude

ion.

that

and

AGGREGATIOX

centrifugation

often

comparison

the

trat

above.

whole

a constant

conclusion,

meaningful since

than

of

platelet

In

described

platelets use

PLATELET

Research

Fund,

OHA G69-82,

Gold

USPHS Fami ly

Fund. REFERENCES

1.

BORN,

2.

Quantitative

G.V.R. J.

platelets. BROCK,

W.E.,

R.G.

hemophilia: 3.

4.

5.

6

7.

8.

and

attempts

the

VI I

by

F. 26.

BUCKNER,

R.G. in

at

Congress

Ala

and

of

the

K.W.E.

MACMILLAN, in man and

D.C. and laboratory

new

therapy. World

Factors O’BRIEN, J.R. Acta Med. Stand. Suppl. What de GAETANO, G. measure? Discussion.

does

of

A

the

the

blood

WULZ. 76,

464,

Canine 1963.

optical

Tehran,

plasma

1971,

levels

of

of

edited No.

252,

factor

1967. study

Suppl.

ia,

of

platelet

aggregation

Haemorrh.

aggregation 525,

55,

platelet

24, test

385,

1970.

actually

1971. aggregation

test.

1971.

optical

Med.

C.E.

Medica,

Diathes.

platelet

Stand. the

43,

525,

on

comparative

optical

Med.

and Pathol.

Haemophil

124,

Thrombos.

of

MILLER, and J.S. MAYES. of factor VIII synthesis, Proceedings Haemoph (1 ia.

Excerpta

influence

influencing

Acta

In:

212,

aggregation

BIRD, Arch.

L.R. site

Amsterdam.

Acta

does

R.M.

Federation

A.K. SIM. animals.

the

1962.

colony.

C.G. GUNN, Genetics,

Denson.

What P.M. Discussion.

67,

HAMPTON, a

CNS J.W. HAMPTON. Amer. J. Physiol.

and

I,

BUCKNER, beagles:

into

162,

J.W. of

experimental

GUNN, C.G. VIII activity.

MANNUCC measure?

London.

Establishment

HAMPTON, J.W., Canine hemophilia

p.

investigations

Physiol.,

Stand.

platelet Suppl.

aggregation 525,

60,

test 1971.

actually

OX

vo1.8,xo.4

9.

BRECHER,

G.

and

platelets. 10.

HAK,

J.

WINER,

GOLDENFARB,

15.

17.

18.

reaction

ROGERS,

The

A.B.

DAY, from

and

human

ROSSI,

effect

Holt,

Experimental

G.R.

human

blood

Rinehart

and

New York:

Design.

COOPER. The determination Atherosclerosis.

normal

of

men.

A.S. DOUGLAS. interval after

ADP-

335,

12,

1970.

The relationship venepuncture.

133, 1974.

31, inhibitors,

contact

platelets.

Thrombos.

E.C.

and

G.

platelet-rich

J.R.

Platelet

study.

PRAGA, C.A. aggregation:

and

other

variables

Diathes.

in

Haemorrh.

the 28,

29,

ZUCKER,

HASLAM, erythrocytes diphosphate

Tests

R.J.

and

of Suppl.

D.C.B.

and platelets in plasma.

separating

in

the

Clin.

Med.

and the aggregation 205, 1075, 1965.

of

rabbit

Part

I I:

15, 452, Effect

of

studying

platelets

33, 648,

increase J.

by

Some

1975.

responsiveness

85, 300,

1975.

platelets

results

from

by

a new

1962.

temperature

on

ADP-induced

anti-aggregating

platelet

drugs.

Thrombos.

183, 1973.

22.

Haemorrh.

for

Haemorrh.

dependent

in

BORN, G.V.R. and M.J. CROSS. London. 168, 178, 1963. M.B.

Methods

induced

139, 1100, 1972.

Med.

Lab.

POGLIANI.

21.

Diathes.

time

aggregation

Biol.

ZUCKER. Diathes.

Pathol.

significance

Haemorrh.

platelet

epinephrine.

Cl in.

E.M. Its

human Exptl.

aggregation,

J.

and

to

Temperature Nature.

diphosphate.

of

A

LOUIS.

J.W.

pH on Sot.

and M.B. Thrombos.

plasma

CONSTANTINE,

O’BRIEN,

of Proc.

ADP.

H. HOLMSEN, and plasma.

H.J., blood

Diathes.

23.

of

1972.

method

20.

Haemorrh.

of

and

OGSTON, and and time

Proteolytic

M.B.

release

adenosine

19.

New York:

in

in

CORINA, D. aggregation

Diathes.

ZUCKER,

of

CATHEY,

aggregation

A. C., platelet

epinephrine

16.

Psychologists.

Principles

M.H.

P.B.,

platelet

WARLOW, between

393,

for

enumeration

1950.

1962.

11.

Thrombos. 14.

and

3, 365,

Physiol.

‘iJ 1

AGGREGXTIOK

Morphology

CRONKITE.

Statistical

B.J.

induced 13.

PLATELET

1966.

McGraw-Hi 12.

E.P. Appl.

Statistics

W.L.

Winston. 11.

DOG

The

platelet 42, MILLS.

aggregation

adhesion, 1,

of

blood

J.

platelets.

aggregation

and

release.

1970. The

in relation Biochem. J.

adenylate to

kinase the

103, 773,

of

degradation

1967.

human of

plasma, adenosine

Physiol.,

Thrombos.