What tests are used to confirm polycythemia?

Tests to Confirm Polycythemia

Begin the diagnostic workup with serum erythropoietin (EPO) level measurement, followed by JAK2 mutation testing (both V617F and exon 12 mutations), and bone marrow examination with cytogenetic studies when polycythemia vera is suspected. 1, 2

Initial Laboratory Confirmation

• Confirm elevated hemoglobin/hematocrit with repeat measurements - hemoglobin >18.5 g/dL in men or >16.5 g/dL in women, hematocrit >55% in men or >49.5% in women 3
• Complete blood count (CBC) with differential to assess for accompanying thrombocytosis or leukocytosis, which suggest myeloproliferative disease 4
• Red cell indices including mean corpuscular volume (MCV) - low MCV with high red cell count suggests iron deficiency coexisting with polycythemia 3
• Reticulocyte count to evaluate bone marrow response 3
• Peripheral blood smear review to assess red cell morphology and identify abnormalities 3

Serum Erythropoietin Level

• Low EPO (<2 U/L) strongly favors polycythemia vera with >90% specificity, though sensitivity is only 64-70% 1, 2
• Normal EPO (2-12 U/L) does not exclude polycythemia vera - proceed with JAK2 testing 1, 2
• High EPO (>12 U/L) suggests secondary polycythemia, but rare cases of polycythemia vera with elevated EPO exist 2, 5, 6

Critical pitfall: In chronic hypoxic states (smoking, sleep apnea, COPD), EPO may initially be elevated but normalize once hemoglobin stabilizes at a compensatory higher level, potentially mimicking polycythemia vera 4, 2

JAK2 Mutation Testing

• Test for JAK2 V617F mutation - present in >90% of polycythemia vera cases 1, 7
• Test for JAK2 exon 12 mutations if V617F is negative - found in an additional 3-5% of polycythemia vera patients 7, 6
• JAK2 mutation testing should be performed even if EPO is elevated when clinical features suggest polycythemia vera 6

Bone Marrow Examination

• Bone marrow biopsy with histology showing hypercellularity, increased megakaryocytes with cluster formation, giant megakaryocytes with pleomorphic morphology 1, 2
• Mild reticulin fibrosis present in 12% of patients 1
• Decreased bone marrow iron stores 1
• Cytogenetic studies have limited diagnostic value (abnormalities in only 13-18% at diagnosis) but may show trisomies of chromosomes 8 or 9, or deletions of long arms of chromosomes 13 or 20 1

Additional Tests to Exclude Secondary Causes

• Serum ferritin and transferrin saturation - iron deficiency can mask erythrocytosis and coexist with polycythemia vera 4, 3
• Arterial blood gas or pulse oximetry to assess for hypoxemia 4
• Chest X-ray if chronic lung disease suspected 4
• Sleep study if nocturnal hypoxemia/obstructive sleep apnea suspected 4, 3
• Abdominal ultrasound or CT to screen for EPO-producing tumors (renal cell carcinoma, hepatocellular carcinoma) if EPO elevated 4
• Renal function tests (creatinine, BUN) and liver function tests (AST, ALT, bilirubin) 4

Specialized Testing for Equivocal Cases

• Decreased megakaryocyte expression of TPO receptor (c-mpl) on bone marrow immunohistochemistry supports polycythemia vera diagnosis 1
• Peripheral blood neutrophil assay for PRV-1 expression (high in polycythemia vera, not detectable in secondary polycythemia) 1
• Spontaneous (endogenous) erythroid colony assays when available, though limited by expertise requirements 1

Diagnostic Algorithm

When hemoglobin/hematocrit is elevated:

1. Measure serum EPO level 1, 2

   • Low EPO → Proceed to JAK2 testing
   • Normal EPO → Proceed to JAK2 testing
   • High EPO → Evaluate for secondary causes, but still consider JAK2 testing if clinical features suggest polycythemia vera

2. JAK2 mutation testing (V617F and exon 12) 7, 6

   • Positive → Polycythemia vera diagnosis likely, confirm with bone marrow biopsy
   • Negative → Consider bone marrow biopsy and specialized testing, or evaluate secondary causes

3. Bone marrow examination when polycythemia vera suspected 1, 2

   • Characteristic histology confirms diagnosis
   • Non-diagnostic → Pursue specialized testing or reevaluate in 3 months

Common pitfalls to avoid:

• Do not rely solely on red cell mass measurement - normal values do not exclude polycythemia vera 2
• Do not assume elevated EPO excludes polycythemia vera - proceed with JAK2 testing if clinical suspicion remains 5, 6
• Do not overlook iron deficiency, which can mask erythrocytosis and cause microcytic polycythemia 4, 3
• Traditional markers (splenomegaly, leukocytosis, thrombocytosis, elevated leukocyte alkaline phosphatase, increased vitamin B12) lack sensitivity and specificity for diagnosis 2