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Assessment of cellularity, genomic DNA yields, and technical platforms for BRAF mutational testing in thyroid fine‐needle aspirate samples
Author(s) -
Dyhdalo Kathryn,
MacNamara Stephen,
Brainard Jennifer,
Underwood Dawn,
Tubbs Raymond,
Yang Bin
Publication year - 2014
Publication title -
cancer cytopathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.29
H-Index - 57
eISSN - 1934-6638
pISSN - 1934-662X
DOI - 10.1002/cncy.21356
Subject(s) - medicine , sanger sequencing , genomic dna , pathology , mutation , polymerase chain reaction , microbiology and biotechnology , thyroid carcinoma , fine needle aspiration , thyroid , cytology , dna , biology , gene , biopsy , genetics
BACKGROUND BRAF mutation V600E (substitution Val600Glu) is a molecular signature for papillary thyroid carcinoma (PTC). Testing for BRAF mutation is clinically useful in providing prognostic prediction and facilitating accurate diagnosis of PTC in thyroid fine‐needle aspirate (FNA) samples. METHODS This study assessed the correlation of cellularity with DNA yield and compared 2 technical platforms with different sensitivities in detection of BRAF mutation in cytologic specimens. Cellularity was evaluated based on groups of 10+ cells on a ThinPrep slide: 1+ (1‐5 groups), 2+ (6‐10 groups), 3+ (11‐20 groups), and 4+ (> 20 groups). Genomic DNA was extracted from residual materials of thyroid FNAs after cytologic diagnosis. RESULTS Approximately 49% of thyroid FNA samples had low cellularity (1‐2+). DNA yield is proportionate with increased cellularity and increased nearly 4‐fold from 1+ to 4+ cellularity in cytologic samples. When applied to BRAF mutational assay, using a cutoff of 6 groups of follicular cells with 10+ cells per group, 96.7% of cases yielded enough DNA for at least one testing for BRAF mutation. Five specimens (11.6%) with lower cellularity did not yield sufficient DNA for duplicate testing. Comparison of Sanger sequencing to allele‐specific polymerase chain reaction methods shows the latter confers better sensitivity in detection of BRAF mutation, especially in limited cytologic specimens with a lower percentage of malignant cells. CONCLUSIONS This study demonstrates that by using 6 groups of 10+ follicular cells as a cutoff, nearly 97% of thyroid FNA samples contain enough DNA for BRAF mutational assay. Careful selection of a molecular testing system with high sensitivity facilitates the successful conduction of molecular testing in limited cytologic specimens. Cancer (Cancer Cytopathol) 2014;122:114–22 © 2013 American Cancer Society .