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Updated: Oct 15, 2025
Employing Digital Droplet PCR to Detect BRAF V600E Mutations in Formalin-fixed Paraffin-embedded Reference Standard Cell Lines
Published on: October 8, 2015
Utility of droplet digital polymerase chain reaction for TERT and BRAF mutational profiling of thyroid nodules
Brandon R Rosvall1, Morris Kostiuk1, Jordana Williams1
1Division of Otolaryngology - Head and Neck Surgery, Department of Surgery, University of Alberta, 1E4 Walter Mackenzie Center, 8440 112 St, Edmonton, Alberta, T6G 2B7, Canada.
Background:
Mutations involving BRAF and TERT are important predictors of disease severity in thyroid cancer, but molecular testing is limited by cost and lack of adequate tissue sample. This study aimed to assess the utility of BRAFV600E and TERT testing using droplet digital PCR (ddPCR) as a diagnostic and prognostic tool for thyroid fine needle aspirate biopsy (FNAB).
Methods:
Patients with thyroid nodules were prospectively enrolled from March 2015 to September 2018. Pre-operative FNAB was collected for standard cytology and molecular testing. BRAFV600E and TERT levels were analyzed by ddPCR. Cytology (Bethesda system) and ddPCR results were correlated to surgical pathology.
Results:
A total of 222 patients were enrolled, of which 124 received thyroid surgery. Pre-operative cytology alone with Bethesda ≥5 was 100% specific and 70% sensitive for malignancy on final surgical pathology. BRAFV600E positivity or TERT overexpression was 100% specific and 60.0% sensitive. Combining cytology (Bethesda ≥5) with BRAFV600E and TERT testing increased the sensitivity of a malignant diagnosis to 80.0%. High TERT levels and/or BRAFV600E was associated with aggressive or advanced stage pathology.
Conclusions:
Combining cytology with ddPCR analysis of BRAFV600E and TERT can improve the diagnostic accuracy of thyroid FNAB, and help predict aggressive pathology.
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