Accurate quantitation of JAK2 V617F allele burden by array-based digital PCR

Area of Science:

• Molecular Diagnostics
• Genetics
• Oncology

Background:

• The JAK2 V617F mutation is a key driver in myeloproliferative neoplasms (MPN).
• Allele burden of JAK2 V617F correlates with MPN patient characteristics and outcomes.
• Allele-specific quantitative PCR (RQ-PCR) is the standard for JAK2 V617F detection, while digital PCR offers precise DNA quantitation.

Purpose of the Study:

• To compare the performance of array-based digital PCR with RQ-PCR for JAK2 V617F allele quantitation.
• To evaluate the suitability of digital PCR as a diagnostic tool for MPN.

Main Methods:

• A comparison was conducted between array-based digital PCR (QuantStudio™ 3D) and a CE-registered RQ-PCR assay.
• Thirty JAK2 V617F-positive samples and thirteen negative samples were analyzed.

Main Results:

• JAK2 V617F allele burden in tested samples varied widely (below 1% to over 90%).
• A strong linear correlation (r² = 0.9983) was observed between digital PCR and RQ-PCR results.
• Digital PCR confirmed the negative results for the 13 samples that were negative by RQ-PCR.

Conclusions:

• Array-based digital PCR is a reliable method for quantifying JAK2 V617F allele burden.
• Digital PCR demonstrates excellent agreement with allele-specific RQ-PCR, supporting its use in clinical settings for MPN diagnosis and monitoring.