Aiding in the distinction between a reactive blood cytosis and a chronic myeloproliferative disorder in extracted DNA specimens
Point Mutation Detection in DNA Using Quantitative Polymerase
Janus kinase 2 gene Tyrosine kinase mutation
Tyrosine Kinase Mutation
Varies
Specimen Type: Extracted DNA from blood or bone marrow
Container/Tube: 1.5- to 2-mL tube with indication of volume and concentration of the DNA
Specimen Volume: Entire specimen
Collection Instructions: Label specimen as extracted DNA from blood or bone marrow
Specimen Stability Information: Refrigerated/Ambient
1. Hematopathology Patient Information (T676) in Special Instructions
2. If not ordering electronically, complete, print, and send a Hematopathology/Cytogenetics Test Request (T726) with the specimen.
Extracted DNA from blood or bone marrow: 50 microliter at 20 ng/microliter
Other | Bone marrow biopsies, slides, paraffin shavings Frozen tissues and paraffin-embedded tissues Paraffin-embedded bone marrow aspirates Moderately to severely clotted |
Specimen Type | Temperature | Time | Special Container |
---|---|---|---|
Varies | Varies |
Aiding in the distinction between a reactive blood cytosis and a chronic myeloproliferative disorder in extracted DNA specimens
An interpretive report will be provided.
The results will be reported as 1 of the 2 states:
-Negative for JAK2 V617F mutation
-Positive for JAK2 V617F mutation
Positive mutation status is highly suggestive of a myeloid neoplasm, but must be correlated with clinical and other laboratory features for a definitive diagnosis.
Negative mutation 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.
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 mutation other than the V617F may be present in an area that interferes with primer or probe binding and cause a false-negative result.
Analytical sensitivity is determined at 0.06% (by dilution of a JAK2 V617F-positive cell line DNA into a negative cell line DNA).
1. Baxter EJ, Scott LM, Campbell PJ, et al: Acquired mutation of
2. James C, Ugo V, Le Couedic JP, et al: A unique clonal JAK2
3. Kralovics R, Passamonti F, Buser AS, et al: A gain-of-function
4. Steensma DP, Dewald GW, Lasho TL, et al: The JAK2 V617F
Genomic DNA is extracted and 2 PCR reactions are used for each sample. In each reaction, a short fragment of genomic DNA, including the mutation site, is amplified using quantitative PCR in a real-time PCR instrument (LightCycler 480, Roche). 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 wild-type 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 wild-type 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, LightCycler 480 Relative Quantification Software is used to calculate the normalized mutated:wild-type 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/wild-type (sample) |
mutated/wild-type (calibrator) | |
| |
The final result is reported as percent JAK2 V617F of total JAK2, ie [mutated/mutated + wild-type] x 100%, calculated from the normalized mutated:wild-type ratio.(Instruction manual: Roche Applied Science Technical Note No. LC 13/2001. Relative Quantification; LightCycler 480, 2006)
Monday through Friday
This test was developed, and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the US Food and Drug Administration.
81270-JAK2 (Janus kinase 2) (eg, myeloproliferative disorder) gene analysis, p.Val617Phe (V617F) variant
Test Id | Test Order Name | Order LOINC Value |
---|---|---|
JAK2V | JAK2 V617F Mutation Detection, V | 43399-5 |
Result Id | Test Result Name |
Result LOINC Value
Applies only to results expressed in units of measure originally reported by the performing laboratory. These values do not apply to results that are converted to other units of measure.
|
---|---|---|
31160 | JAK2 V617F Mutation Detection, V | 43399-5 |
39724 | JAK2 Result | 53761-3 |