When is testing for JAK2 (Janus Kinase 2) mutations recommended to support a diagnosis?

When to Order JAK2 Mutation Testing

Order JAK2V617F testing as the first-line molecular test in any patient with unexplained erythrocytosis (hemoglobin >18.5 g/dL in males or >16.5 g/dL in females), persistent thrombocytosis (platelets ≥450 × 10⁹/L), or suspected myelofibrosis. 1

Primary Indications for JAK2 Testing

Erythrocytosis

• Test when hemoglobin exceeds 18.5 g/dL in men or 16.5 g/dL in women, after excluding secondary causes of polycythemia 1
• JAK2V617F is present in approximately 95% of polycythemia vera cases, making it highly sensitive for this diagnosis 2, 3
• If JAK2V617F is negative in patients with persistent erythrocytosis, proceed to JAK2 exon 12 mutation testing, as these mutations account for most JAK2V617F-negative polycythemia vera cases 1, 4

Thrombocytosis

• Order testing when platelet count is persistently ≥450 × 10⁹/L without an obvious reactive cause 5
• JAK2V617F is found in 50-60% of essential thrombocythemia cases 2, 3
• If JAK2V617F is negative, test for CALR mutations first, then MPL mutations, as approximately 80% of essential thrombocythemia patients harbor one of these three driver mutations in a mutually exclusive pattern 1, 5

Suspected Myelofibrosis

• Test in patients presenting with splenomegaly, constitutional symptoms (fatigue, night sweats, weight loss), or unexplained cytopenias 6
• JAK2V617F is present in approximately 50% of primary myelofibrosis cases 2, 3

Splanchnic Vein Thrombosis

• Order JAK2V617F testing in younger patients (<60 years) with splanchnic vein thrombosis (portal, hepatic, mesenteric, or splenic vein) who lack typical thrombosis risk factors 1
• This presentation may be the first manifestation of an underlying myeloproliferative neoplasm 1

Testing Algorithm

First-Line Testing

• JAK2V617F should be the initial molecular test ordered for any suspected Philadelphia chromosome-negative myeloproliferative neoplasm 1
• The assay must be sensitive enough to detect mutant allele burden as low as 1-3% 1, 4
• Ensure your laboratory uses appropriately sensitive methods such as quantitative PCR or high-resolution melting analysis 1

Sequential Testing Based on Clinical Presentation

For erythrocytosis with negative JAK2V617F:

• Proceed to JAK2 exon 12 mutation testing 1, 4
• These mutations are found in 2-3% of polycythemia vera cases that lack JAK2V617F 7

For thrombocytosis or myelofibrosis with negative JAK2V617F:

• Test for CALR mutations first (found in ~25-30% of essential thrombocythemia and primary myelofibrosis) 1, 5
• Then test for MPL mutations if CALR is negative (found in ~3-5% of cases) 1, 4

When to Refer to Hematology

Mandatory Referral Situations

• Any positive JAK2 mutation result requires hematology referral for bone marrow biopsy and definitive diagnosis 1
• Persistent clinical suspicion despite negative molecular testing warrants hematology evaluation 1
• Bone marrow examination is necessary for definitive diagnosis in most cases, except polycythemia vera with hemoglobin >18.5 g/dL in males or >16.5 g/dL in females where clinical diagnosis may suffice 1

Critical Diagnostic Considerations

JAK2 Mutation Alone Is Not Diagnostic

• The presence of JAK2V617F confirms clonality but must be integrated with clinical findings, complete blood count, and bone marrow morphology to distinguish between polycythemia vera, essential thrombocythemia, and primary myelofibrosis 2, 8
• WHO diagnostic criteria require both molecular and morphologic findings 2

Low Allele Burden Interpretation

• JAK2 mutation values <1% require careful interpretation and may represent early disease or clonal hematopoiesis of indeterminate potential 8
• Patients with low allele burden and no hematologic abnormalities should undergo periodic monitoring with complete blood counts every 6-12 months and repeat JAK2 testing annually 8

Expanded Mutation Screening

• Consider testing beyond JAK2V617F and exon 12 mutations to include most of the pseudokinase domain (exons 12-15), as novel mutations in this region can cause myeloproliferative neoplasms 7
• RNA-based testing is more sensitive than DNA-based approaches due to relative abundance of target transcripts 7

Common Pitfalls to Avoid

• Do not order JAK2 testing for reactive thrombocytosis or polycythemia with an obvious secondary cause (hypoxia, sleep apnea, smoking, renal disease) 2
• Do not rely solely on JAK2V617F testing; if negative with high clinical suspicion, proceed with sequential mutation testing for exon 12, CALR, and MPL 1, 4
• Do not assume JAK2 negativity excludes myeloproliferative neoplasm—approximately 10-20% of essential thrombocythemia and primary myelofibrosis cases are triple-negative for JAK2, CALR, and MPL mutations 5
• Do not use insensitive assays; detection limits must reach 1-3% mutant allele burden to avoid false negatives 1, 4