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HelpTest Code JAK2M JAK2 V617F Mutation Detection, Bone Marrow
Reporting Name
JAK2 V617F Mutation Detection, BMUseful For
Aiding in the distinction between a reactive blood cytosis and a chronic myeloproliferative disorder using bone marrow specimens
Testing Algorithm
For information see Myeloproliferative Neoplasm: A Diagnostic Approach to Bone Marrow Evaluation.
Method Name
Quantitative Polymerase Chain Reaction (PCR)
Performing Laboratory
Mayo Clinic Laboratories in Rochester
Specimen Type
Bone MarrowShipping Instructions
Specimen must arrive within 7 days of collection.
Specimen Required
Container/Tube:
Preferred: Lavender top (EDTA)
Acceptable: Yellow top (ACD)
Specimen Volume: 2 mL
Collection Instructions:
1. Invert several times to mix bone marrow.
2. Send bone marrow specimen in original tube. Do not aliquot.
Specimen Minimum Volume
1 mL
Specimen Stability Information
| Specimen Type | Temperature | Time | Special Container |
|---|---|---|---|
| Bone Marrow | Ambient (preferred) | 7 days | PURPLE OR PINK TOP/EDTA |
| Refrigerated | 7 days | PURPLE OR PINK TOP/EDTA |
Reject Due To
| Gross hemolysis | Reject |
| Moderately to severely clotted | Reject |
Special Instructions
Reference Values
An interpretive report will be provided.
Day(s) Performed
Monday through Friday
CPT Code Information
81270-JAK2 (Janus kinase 2) (eg, myeloproliferative disorder) gene analysis, p.Val617Phe (V617F) variant
LOINC Code Information
| Test ID | Test Order Name | Order LOINC Value |
|---|---|---|
| JAK2M | JAK2 V617F Mutation Detection, BM | 72333-8 |
| Result ID | Test Result Name | Result LOINC Value |
|---|---|---|
| 39723 | JAK2 Result | 53761-3 |
| 31157 | JAK2 V617F Mutation Detection, BM | 72333-8 |
Clinical Information
The Janus kinase 2 (JAK2) gene codes for a tyrosine kinase (JAK2) associated with the cytoplasmic portion of a variety of transmembrane cytokine and growth factor receptors important for signal transduction in hematopoietic cells. Signaling via JAK2 activation causes phosphorylation of downstream signal transducers and activators of transcription (STAT) proteins (eg, STAT5) ultimately leading to cell growth and differentiation. BCR-ABL1-negative myeloproliferative neoplasms (MPN) frequently harbor an acquired single nucleotide variant in JAK2 characterized as c.G1849T; p. Val617Phe (V617F). This variant is identified overall in approximately two-thirds of all MPN,(1-3) but the prevalence varies by MPN subtype. The JAK2 V617F variant is present in 95% to 98% of polycythemia vera patients, 50% to 60% of primary myelofibrosis (PMF) patients, and 50% to 60% of essential thrombocythemia (ET) patients. It has infrequently been described in other myeloid neoplasms, including chronic myelomonocytic leukemia and myelodysplastic syndrome.(4) This variant is not seen in chronic myelogenous leukemia or reactive conditions with elevated blood counts. Detection of the JAK2 V617F variant is useful to help establish the diagnosis of MPN. However, a negative JAK2 V617F result does not indicate absence of an MPN. Other important molecular markers in BCR-ABL1-negative MPN include CALR exon 9 variant (20%-30% of PMF and ET) and MPL exon 10 variant (5%-10% of PMF and 3%-5% of ET). Variants in JAK2, CALR, and MPL are essentially mutually exclusive.
Interpretation
The results will be reported as 1 of the 2 states:
-Negative for JAK2 V617F variant
-Positive for JAK2 V617F variant
Positive variant status is highly suggestive of a myeloid neoplasm but must be correlated with clinical and other laboratory features for definitive diagnosis.
Negative variant status does not exclude the presence of a myeloproliferative neoplasm or other neoplasm.
Results below the laboratory cutoff for positivity are of unclear clinical significance at this time.
Cautions
A positive result is not specific for a particular subtype of myeloproliferative neoplasm and clinicopathologic correlation is necessary in all cases. If this test is ordered in the setting of erythrocytosis and suspicion of polycythemia vera, interpretation requires correlation with a concurrent or recent prior bone marrow evaluation.
A negative result does not exclude the presence of a myeloproliferative neoplasm or other neoplastic process.
In rare cases, a variant other than the V617F may be present in an area that interferes with primer or probe binding and cause a false-negative result.
Method Description
Genomic DNA is extracted, and 2 polymerase chain reactions (PCR) are used for each sample. In each reaction, a short fragment of genomic DNA, including the variant site, is amplified using quantitative PCR in a real-time PCR instrument. In the first reaction, the 5' terminal base of the reverse primer matches the mutated sequence, and the PCR conditions are such that it will only bind mutated DNA. In the second reaction, the 5' terminal base of the reverse primer matches the wildtype sequence, and the PCR conditions are such that it will only bind the wild-type sequence. In both reactions, the PCR is monitored using TaqMan probe chemistry. The amount of mutated DNA and the amount of wildtype DNA is measured for each sample. In each run, the amount of mutated and wild-type DNA in a calibrator DNA sample is also measured. The calibrator is a mixture of DNA from a positive cell line (HEL) and a negative cell line (HL60) that is frozen in aliquots and expected to give an identical result in each run. Deviations in the calibrator result are assumed to be due to deviations in the run conditions and the sample results are corrected accordingly. Following each reaction, Relative Quantification Software is used to calculate the normalized mutated:wildtype ratio, which is expressed as a unitless ratio following correction with the calibrator data.
The formula for the normalized ratio is as follows:
|
Normalized ratio = |
mutated/wildtype (sample) |
|
mutated/wildtype (calibrator) |
The final result is reported as percent JAK2 V617F of total JAK2 (ie, [mutated/mutated + wildtype] x 100%), calculated from the normalized mutated:wildtype ratio.(Unpublished Mayo method)