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A Practical Guide to Laboratory Haemostasis

 
  

Platelet Function Testing: PFA-100



Introduction

The PFA-100 is a system for analysing platelet function in which citrated whole blood is aspirated at high shear rates through disposable cartridges containing an aperture within a membrane coated with either collagen and epinephrine (CEPI) or collagen and ADP (CADP). These agonists induce platelet adhesion, activation and aggregation leading to rapid occlusion of the aperture and cessation of blood flow termed the closure time (CT). This LINK will take you to an animation that will explain this process in more detail.

Advantages of the PFA-100 include:
Only small volumes of citrated venous blood [800µL) are needed and so the test is useful for investigating platelet function in children.
Can be used by non-skilled personnel and is both rapid and automated
The PFA-100 was designed as a screen to detect problems with primary haemostasis and in part to replace the bleeding time and in this respect it is better standardised.
Measurement of platelet function at high shear [physiological] rates whereas LTA measures platelet function at low shear rates i.e. less physiological.
Relatively insensitive to clotting factor deficiencies
High negative predictive value – i.e. if the PFA-100 gives a normal result then with some exceptions primary haemostasis is intact [Exceptions: SPD, Primary Secretion Defects, mild Type 1 VWD]

 

A number of variables have been shown to affect the results obtained with the PFA-100 and these include:

Variable Effect
Citrate concentration Laboratories must use a fixed citrate concentration
Collection time How the sample was collected and transported to the lab. The tests must be performed within 4 hours of collection
Haematocrit Closure times increase progressively with decreases in haematocrit.  Conversely, CTs are shortened in the neonate due to their higher haematocrit.
Platelet count Closure times increase progressively as the platelet counts falls below 100 x 109/L
Blood Group and VWF levels Closure times correlate inversely with plasma VWF activity levels and may be increased in blood group O patients for the same reason [Blood group O individuals have lower VWF levels.]
Drugs: COX inhibitors such as aspirin and NSAIDs usually prolong the closure time of the CEPI cartridge but not the CADP cartridge. 

The effects of ADP receptor blockers such as Clopidogrel is unpredictable.

Inhibition of the GpIIb/IIIa receptor is associated with a significant prolongation of both cartridges.
Acquired platelet function defects Cardio-pulmonary bypass
Liver disease
Uraemia
Some foods Generation of false positives particularly with the CEPI cartridge. This is due probably to ingested foods or drugs.  An abnormal PFA-100 result is not, therefore, diagnostic. 


Aspirin Resistance: The PFA-100 is often used to establish the presence of absence of aspirin resistance. The frequency of aspirin resistance is unknown, but estimates range from 5-60%. The mechanism of aspirin resistance is unknown but proposed mechanisms include poor patient compliance, poor aspirin absorption, increased platelet hypersensitivity to agonists, increased COX activity, and polymorphisms in the Gp IIIa receptor and the COX enzyme.  Aspirin resistance appears to be dose related in some patients and may be overcome with higher doses.

Interpretation

The following table summarises some of the abnormalities that have been reported with the PFA-100.

Disorder
CT Collagen-ADP
CT Collagen-EPI
Normal
N
N
Aspirin and NSAIDs
N
ADP receptor disorders including the use of Clopidogrel
N or ↑
N or ↑
BSS
GTT
VWD
Platelet-Type VWD
Dense Granule Deficiency
N or ↑
N or ↑
Primary Secretion Defects
N or ↑
N or ↑
Gray Platelet Syndrome
MYH9-related Disorders
N
Scott Syndrome
N
N
MDS
N or ↑
N or ↑
Liver Disease
↑ [possibly as a result of ↓Hb]
↑ [possibly as a result of ↓Hb]
Uraemia
↑ [possibly as a result of ↓Hb]
↑ [possibly as a result of ↓Hb]


Reference Ranges

Reported reference ranges for closure times are:

  • 78 - 199 seconds for the CEPI cartridge
  • 55 - 137 seconds for the CADP cartridge

What Test Next

1. The PFA-100 has a high negative predictive value i.e. if the PFA-100 gives a normal result then with some exceptions primary haemostasis is intact [Exceptions: SPD, Primary Secretion Defects, mild Type 1 VWD] and so may obviate further screening of platelet function.

2. If the PFA-100 is abnormal then formal platelet aggregation testing and platelet nucleotide assays may be required to establish the underlying cause.

Data Interpretation

Click HERE to go to the Data Interpretation Exercises.