How does the PFA‑100 (Platelet Function Analyzer) work and what is its role in evaluating a healthy 30‑year‑old woman with spontaneous bruising and possible von Willebrand disease?

PFA-100 (Platelet Function Analyzer): Mechanism and Clinical Role

How the PFA-100 Works

The PFA-100 simulates high shear stress conditions in vitro by aspirating citrated whole blood through a capillary tube toward a membrane coated with collagen plus either adenosine diphosphate (ADP) or epinephrine (EPI), measuring the time required for platelet plug formation to occlude a small aperture—this "closure time" (CT) reflects von Willebrand factor-dependent platelet adhesion and aggregation under flow conditions. 1, 2

Technical Specifications

• Blood volume required: Small volume of citrated whole blood 2
• Cartridge types: Two membrane options available 1, 3
  • Collagen/ADP (COL/ADP or CADP)
  • Collagen/epinephrine (COL/EPI or CEPI)
• Measured endpoint: Closure time in seconds until aperture occlusion 2, 3
• Normal reference ranges: 4
  • EPI-CT: 82-159 seconds (or 98-158 seconds per another study) 2
  • ADP-CT: 62.5-120.5 seconds (or 66-124 seconds per another study) 2

Mechanism of Action

The device forces blood through high shear stress conditions that activate platelets upon contact with the collagen-coated membrane, triggering adhesion via von Willebrand factor binding to platelet glycoprotein Ib/IX, followed by platelet aggregation that progressively occludes the aperture. 2, 3 The process mimics primary hemostasis in damaged blood vessels under arterial flow conditions. 5

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Role in Evaluating a 30-Year-Old Woman with Spontaneous Bruising and Possible VWD

Diagnostic Performance for von Willebrand Disease

The PFA-100 demonstrates superior sensitivity (87-88%) compared to bleeding time (65%) for detecting von Willebrand disease, making it a valuable screening tool in this clinical scenario. 3, 6

Sensitivity and Specificity Data

• Overall sensitivity for VWD: 87-88% with both cartridges 3
• Overall specificity: 80-82% 5
• Comparative advantage: Significantly better sensitivity than bleeding time (p < 0.01) with similar specificity 6
• Detection capability: All patients with confirmed VWD (types 1, 2A, 2B, and 3) showed prolonged closure times (≥200 seconds) with both cartridges 2

Important Limitations in This Clinical Context

However, the PFA-100 is not an effective screen for some types of VWD and milder platelet abnormalities, and many centers have decreased or ceased its use due to these limitations. 1

Specific Diagnostic Gaps

• False negatives occur in approximately 6% of cases, particularly with: 5
  • Type 1 VWD (mild cases)
  • Hermansky-Pudlak syndrome
  • Storage pool defects
  • Release defects
  • Macrothrombocytopenia
• Cannot detect: 1
  • Some VWD subtypes
  • Milder platelet function abnormalities
  • Factor XIII deficiency
  • Mild hemophilia (factor VIII or IX deficiencies)

Confounding Variables That Affect Results

Multiple patient characteristics can produce false positive or false negative results: 1

• Hematocrit levels (test requires adequate hematocrit) 1
• Platelet count (thrombocytopenia prolongs CT) 2
• Pregnancy
• Age
• Multisystem trauma
• Sepsis
• Medications (especially aspirin and NSAIDs) 4
• Time of blood collection (evening samples show significantly longer CT than morning samples, p = 0.002-0.004) 4

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Clinical Algorithm for This Patient

Step 1: Initial PFA-100 Testing

Perform PFA-100 with both COL/ADP and COL/EPI cartridges on morning blood sample (to avoid diurnal variation). 4

• If both CTs are normal: VWD is unlikely (high negative predictive value), but does not completely exclude mild VWD or platelet disorders 2, 6
• If either or both CTs are prolonged: Proceed to confirmatory testing 5

Step 2: Mandatory Follow-Up Testing

When PFA-100 shows abnormal results, specific von Willebrand testing must be performed because abnormal closure times occur in multiple conditions beyond VWD. 2, 6

Required confirmatory tests include: 1, 7

• VWF antigen (VWF:Ag)
• VWF activity (VWF:RCo or VWF:Act)
• Factor VIII level
• VWF multimer analysis (if initial VWF studies suggest type 2 variants)

Step 3: Platelet Function Assessment

If VWF studies are normal but clinical suspicion remains high, proceed to: 1, 7

• Light transmission aggregometry (LTA) with standard agonists (ADP, collagen, epinephrine, ristocetin)
• Flow cytometry using antibodies against GPIIb/IIIa (CD41), GPIIIa (CD61), GPIb (CD42b), and GPIb/IX (CD42a)
• Hematology consultation for interpretation 1

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Critical Pitfalls to Avoid

Pre-Analytical Errors

• Timing: Blood must be processed within 0.5 to 4 hours after collection 4
• Duplicate measurements: Single measurements have coefficients of variation of 5.7-7.1%; perform duplicates 4
• Medication history: Document aspirin, NSAIDs, and antiplatelet agents—even single-dose aspirin (100-500 mg) prolongs EPI-CT 4
• Sample collection time: Avoid evening samples due to significantly prolonged closure times 4

Interpretation Errors

The most dangerous pitfall is using PFA-100 as a definitive diagnostic test rather than a screening tool—abnormal results require comprehensive follow-up testing to establish the true clinical disorder. 2, 6

• Do not diagnose VWD based solely on PFA-100 results 2
• Do not exclude bleeding disorders based solely on normal PFA-100 (6% false negative rate) 5
• Do not use PFA-100 results to guide treatment decisions without confirmatory testing 1

Clinical Context Considerations

The American Academy of Pediatrics notes that assessment of PFA-100 results and need for further testing are best accomplished in consultation with a hematologist, reflecting the complexity of interpretation in real-world practice. 1 Given that many centers have decreased or ceased use of the PFA-100 due to its limitations, consider proceeding directly to specific VWF testing and platelet aggregometry in this 30-year-old woman with spontaneous bruising rather than relying on PFA-100 screening. 1