Disease Markers

Disease Markers / 2013 / Article

Open Access

Volume 34 |Article ID 740659 | https://doi.org/10.3233/DMA-120960

Veronika Buxhofer-Ausch, Christoph Ausch, Robert Zeillinger, Christian Oberkanins, Nadia Dandachi, Angelika Reiner-Concin, Gernot Kriegshäuser, "Duplex Reverse-Hybridization Assay for The Simultaneous Detection of KRAS/BRAF Mutations in FFPE-extracted Genomic DNA from Colorectal Cancer Specimens", Disease Markers, vol. 34, Article ID 740659, 7 pages, 2013. https://doi.org/10.3233/DMA-120960

Duplex Reverse-Hybridization Assay for The Simultaneous Detection of KRAS/BRAF Mutations in FFPE-extracted Genomic DNA from Colorectal Cancer Specimens

Received14 Jan 2013
Accepted14 Jan 2013

Abstract

We report the performance evaluation of a non-quantitative reverse-hybridization assay (KRAS-BRAF StripAssay) designed for the simultaneous detection of 10 mutations in codons 12 and 13 of the KRAS gene and BRAF mutation V600E. Dilution experiments using DNA from tumor cell lines or from formalin-fixed paraffin-embedded (FFPE) colorectal cancer (CRC) tissue were performed to assess assay sensitivity. Using 50 ng of total DNA (mutant and wild-type), the KRAS-BRAF StripAssay demonstrated a detection limit of 1% mutant sequence in a background of wild-type DNA. With respect to BRAF V600E, the KRAS-BRAF StripAssay was evaluated using 60 FFPE CRC samples previously analyzed by high resolution melting (HRM). Test strip hybridization identified 2/60 (3%) samples to carry the BRAF V600E mutation, and results were in agreement with those obtained by HRM analysis. This work demonstrates the KRAS-BRAF StripAssay to be a robust and sensitive method for the detection of common KRAS/BRAF mutations in genomic DNA isolated from FFPE tissue samples.

Copyright © 2013 Hindawi Publishing Corporation. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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