title: Duplex reverse-hybridization assay for the simultaneous detection of KRAS/BRAF mutations in FFPE-extracted genomic DNA from colorectal cancer specimens.
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.
authors: Second Department of Medicine, Donauspital, Vienna, Austria.
authors: Veronika
authors: V
authors: Buxhofer-Ausch
authors: Christoph
authors: C
authors: Ausch
authors: Robert
authors: R
authors: Zeillinger
authors: Christian
authors: C
authors: Oberkanins
authors: Nadia
authors: N
authors: Dandachi
authors: Angelika
authors: A
authors: Reiner-Concin
authors: Gernot
authors: G
authors: Kriegshäuser
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D015179:Colorectal Neoplasms / Q000235:genetics*; D004273:DNA, Neoplasm / Q000032:analysis* / Q000235:genetics; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D008875:Middle Aged; D009154:Mutation / Q000235:genetics*; D009367:Neoplasm Staging; D009693:Nucleic Acid Hybridization; D016612:Paraffin Embedding; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins / Q000235:genetics*
chemical_list: D004273:DNA, Neoplasm; C117307:KRAS protein, human; D011518:Proto-Oncogene Proteins; D005557:Formaldehyde; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins