title: KRAS detection in colonic tumors by DNA extraction from FTA paper: the molecular touch-prep.
abstract: DNA isolated from formalin-fixed paraffin-embedded (FFPE) tissue is usually more degraded and contains more polymerase chain reaction (PCR) inhibitors than DNA isolated from nonfixed tissue. In addition, the tumor size and cellular heterogeneity found in tissue sections can often impact testing for molecular biomarkers. As a potential remedy to this situation, we evaluated the use of Whatman FTA paper cards for collection of colorectal tumor samples before tissue fixation and for isolation of DNA for use in a real-time PCR-based KRAS mutation assay. Eleven colon tumor samples were collected by making a cut into the fresh tumor and applying the Whatman FTA paper to the cut surface. Matched FFPE tissue blocks from these tumors were also collected for comparison. KRAS mutation analysis was carried out using the Applied Biosystems 7500 Fast Real-time PCR System using 7 independent custom TaqMan PCR assays. Of the 11 colon tumors sampled, 6 were positive for KRAS mutations in both the Whatman FTA paper preparations and corresponding FFPE samples. Whatman FTA paper cards for collection of colorectal tumor samples before tissue fixation and for isolation of DNA have many advantages including ease of use, intrinsic antimicrobial properties, long storage potential (stability of DNA over time), and a faster turnaround time for results. Extracted DNA should be suitable for most molecular diagnostic assays that use PCR techniques. This novel means of DNA preservation from surgical specimens would benefit from additional study and validation as a dependable and practical technique to preserve specimens for molecular testing.
authors.affiliation: Department of Pathology, Dartmouth-Hitchcock Medical Center, Dartmouth Medical School, Hanover, NH, USA.
authors.forename: Melissa L
authors.initials: ML
authors.lastname: Petras
authors.forename: Joel A
authors.initials: JA
authors.lastname: Lefferts
authors.forename: Brian P
authors.initials: BP
authors.lastname: Ward
authors.forename: Arief A
authors.initials: AA
authors.lastname: Suriawinata
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Tsongalis
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D001706:Biopsy; D003110:Colonic Neoplasms / Q000235:genetics*; D004273:DNA, Neoplasm / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification*; D005260:Female; D011905:Genes, ras; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques; D009154:Mutation; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction / Q000379:methods; D011309:Preservation, Biological; D011518:Proto-Oncogene Proteins / Q000032:analysis* / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D013048:Specimen Handling / Q000379:methods; D016707:Tissue Fixation; D018631:ras Proteins / Q000032:analysis* / Q000235:genetics*
chemical_list: D004273:DNA, Neoplasm; C117307:KRAS protein, human; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins