What is the recommended workup for a patient with suspected polycythemia vera?

Polycythemia Vera Workup

Initial Laboratory Testing

Begin with JAK2 V617F mutation testing and serum erythropoietin (EPO) level simultaneously—these two tests form the diagnostic cornerstone and should be ordered immediately upon suspicion. 1, 2, 3

First-Line Tests (Order Immediately)

• JAK2 V617F mutation (exon 14): Present in >95% of PV cases, providing the highest diagnostic sensitivity and specificity 1, 4, 3, 5
• Serum EPO level: Low or inappropriately normal EPO has >90% specificity for PV, though sensitivity is only 64-70% 1, 2, 6
• Complete blood count with differential and red cell indices: Confirm sustained hemoglobin ≥18.5 g/dL (men) or ≥16.5 g/dL (women), and assess for thrombocytosis (53% of cases) and leukocytosis (49% of cases) 4, 3
• Iron studies (ferritin, serum iron, transferrin saturation): Iron deficiency can mask true erythrocytosis by suppressing hemoglobin into the normal range despite underlying PV 1, 4

Critical Interpretation Points

• Low EPO (<2 U/L) + elevated hemoglobin → Strongly suggests PV; proceed to bone marrow biopsy 1, 2
• Normal EPO (2-12 U/L) → Does NOT exclude PV; JAK2 testing becomes essential 1, 2
• Elevated EPO (>12 U/L) → Suggests secondary polycythemia; evaluate for hypoxia-driven or tumor-related causes 1, 2
• Low mean corpuscular volume (MCV) → Indicates iron deficiency that may be masking higher hemoglobin; check iron studies and look for other PV features (thrombocytosis, leukocytosis, splenomegaly) 1

Secondary Testing Based on Initial Results

If JAK2 V617F is Negative

• JAK2 exon 12 mutation testing: Accounts for 2-4% of PV cases that lack V617F mutation 1, 4
• Combined JAK2 testing (V617F + exon 12) achieves 97-99% sensitivity for PV 1

If EPO is Elevated (Suggesting Secondary Polycythemia)

Systematically exclude secondary causes in this order:

1. Smoking history and carboxyhemoglobin level: Smoker's polycythemia is the most common secondary cause and resolves with cessation 1, 6
2. Arterial oxygen saturation and chest X-ray: Rule out chronic lung disease (COPD, pulmonary fibrosis) or right-to-left cardiopulmonary shunts 1
3. Sleep study: Evaluate for obstructive sleep apnea causing nocturnal hypoxemia 1
4. Abdominal ultrasound or CT: Screen for EPO-producing tumors (renal cell carcinoma, hepatocellular carcinoma, uterine leiomyoma, pheochromocytoma) 1, 6
5. Medication review: Check for exogenous testosterone, anabolic steroids, or administered erythropoietin 1
6. Renal function tests: Evaluate for post-renal-transplant erythrocytosis 1

Bone Marrow Examination

Perform bone marrow biopsy with cytogenetic studies when PV is suspected based on low/normal EPO and/or positive JAK2 mutation, particularly before initiating cytoreductive therapy. 2, 4

Diagnostic Findings Supporting PV

• Hypercellularity for age with trilineage growth (panmyelosis) 2, 4, 3
• Increased megakaryocytes with cluster formation and pleomorphic morphology 2
• Cytogenetic abnormalities present in 13-18% but have limited diagnostic value 2

WHO 2016 Diagnostic Criteria Application

Diagnosis requires EITHER:

• All 3 major criteria, OR
• First 2 major criteria + 1 minor criterion 4

Major Criteria

1. Hemoglobin ≥18.5 g/dL (men) or ≥16.5 g/dL (women), OR hematocrit ≥49% (men) or ≥48% (women) 4
2. JAK2 V617F or JAK2 exon 12 mutation 4
3. Bone marrow biopsy showing hypercellularity with trilineage growth 4

Minor Criteria

1. Serum EPO below normal reference range 4
2. Endogenous erythroid colony formation in vitro (rarely available) 2, 4

Clinical Features That Trigger Immediate Workup

• Sustained hemoglobin increase ≥2 g/dL from baseline, even if within normal reference range 1, 4
• Borderline-high hematocrit with PV-related features: thrombocytosis, leukocytosis, microcytosis (from iron deficiency), splenomegaly (36% of cases), aquagenic pruritus (33%), erythromelalgia (5.3%), or unusual thrombosis (Budd-Chiari syndrome) 1, 2, 3
• Values above 99th percentile for age, sex, and altitude-adjusted reference ranges 4

Critical Diagnostic Pitfalls to Avoid

• Do not assume normal EPO excludes PV: EPO sensitivity for PV is only 64-70%; normal EPO with elevated hemoglobin still requires JAK2 testing 1, 6
• Do not miss iron deficiency: Low MCV or MCHC <32% indicates iron deficiency that can mask erythrocytosis by suppressing hemoglobin into the normal range 1, 2, 4
• Do not rely on traditional markers alone: Splenomegaly, leukocytosis, thrombocytosis, elevated leukocyte alkaline phosphatase, and increased vitamin B12 levels lack both sensitivity and specificity for PV 2
• Do not perform unnecessary red cell mass measurements: Modern JAK2 testing and EPO levels have largely replaced the need for these measurements except in truly equivocal cases 1
• Verify hydration status first: Dehydration is the most common cause of falsely elevated hematocrit 4

Specialized Testing for Equivocal Cases

When diagnosis remains uncertain after standard workup:

• Bone marrow immunohistochemistry for c-mpl (thrombopoietin receptor): Decreased expression supports PV diagnosis 1, 2
• Neutrophil PRV-1 expression assay: Can distinguish PV from secondary polycythemia but has limited availability 1, 2
• Spontaneous erythroid colony assay: Distinguishes PV from secondary polycythemia but requires considerable expertise and is rarely used 1, 2