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Perth Haematology :: Dr Steven Ward

Platelet Function Tests


PFA-100 :: Platelet Function Analyser

 

An automated analyser assesses primary haemostasis under shear stress.

Two cartridges:

§  CEPI: collagen and epinephrine (adrenaline).

§  CADP: collagen and ADP

 

Blood sample is aspirated through the cartridge membrane – leading to platelet activation and aggregation.  The time to close the membrane aperture (closure time) is measured.

 

Aggregation depends on adequate platelet numbers, and platelet functional ability (vWF, platelets GP Ib, GP IIb/IIIa and fibrinogen).

 

Variables affecting results:

·         Collection Time: Assay within 4 hours of collection.

·         Haematocrit: Anaemia increases closure time (Hct <20%).

·         Platelet Count: Closure times increase as plate count<100.

·         Blood group/vWF levels

·         Drugs: Aspirin, NSAID, clopidogrel (variable)

·         Acquired platelet function defects: cardio-pulmonary bypass, liver disease, renal failure.

 

 

PFA-100 patterns

Disorder

Col/Epi

Col/ADP

Aspirin/NSAID

N

Clopidogrel

N/↑

N/↑

vWD

Glanzmann’s;  Bernard Soulier

Storage Pool Defect

N (or ↑)

Macrothrombocytopathy

N

Renal Failure, MPD, CABG

 


Platelet aggregation studies

 

Tests the ability of certain agonists (stimulants) to produce platelet clumping or aggregation.

Patterns of abnormality can help identify the type of platelet defect.

 

Common defect patterns:

Disorder

Defect

Plat count

Blood Film

ADP 2uM

ADP 5-10uM

Collagen 1ug/mL

AA 100uM

Adrenaline 10UM

Risto 1.2mg/ml

Aspirin/NSAID

 

Inhibit COX

N

N

1Y wave no 2y wave

1y
no 2y

absent

Absent

N

ADP antagonists (clopidogrel)

Inhib ADP receptor

N

N

Absent

Absent

N

N

N

N

Glanzmanns’s Thrombasthenia

GpIIb/IIIa receptor

N

N

Absent

Absent

Absent

Absent

Absent

N

Bernard Soulier Syndrome

GpIb-V-IX receptor

Giant plats

ę plat

N

N

N

N

N

Absent

vWD

 

N/

N

N

N

N

N

N

Gray Platelet Syndrome

α-granule defic

Gray plats

N/

N/

N/

N

N

N

Dense Granule Deficiency

δ-granules

N

N

N

Absent

N

No 2y wave

? 1Y only

VWD Type IIb

VWF protein

 

N

N

N

N

N

N  
0.5 risto

Platelet type vWD

Mutn GpIb receptor

 

N

N

N

N

N

N  
0.5 risto

Renal Failure

Acquired SPD

N/

 

N

N/

N/

N/

Liver Disease

 

N/

 

1Y only

1Y only

?

1Y only

N/

Afibrinogenaemia

 

 

 

Absent

Absent

Absent

 

Absent

 

Strong agonists: collagen, thrombin, TXA2 – directly induce plat agg, TX synthesis and plat gran secretion.

Weak agonists: ADP, Adren: Induce platn agg without secretion. (biphasic curve)

 

Agonists:

Agonist

Conc

Comment

ADP

LD:1–5 uM

HD 10uM

ADP binds ADP receptor on plat. Shape change – 1y agg. 2y wave = ADP from plat storage grans. LD –> 1y only. 2y wave inhib by aspirin and NSAIDs.

Clopidogrel inhibs P2Y1 receptor

Collagen

1, 4ug/mL

Collagen binds GpVI and GpIa/IIa inducing granule release, TX and GPIIb/IIIa activn. Lag phase.

Ristocetin

LD: 0.5mg/mL

HD: 1.5-5mg/mL

Ristocetin (not LD) cuases plats agglutination (not agg) via vWF abd GP1b-IX-V complex.

Adrenaline

5,10uM

Adrenaline blinds a2 recepotor on plat -- Ca release. Similar to ADP agg. 1y , 2y wave with ADP released from grans. 2y wave inhib by aspirin,. NSAIDs.  Variation in a receptor density – reduced agg.

Arachadonic Acid

500ug/mL

AA is precursor to TXA2. Via cyclo-oxygenase and thromboxane synthase.  TX is potent plat activator