OBJECTIVE: The impact of single-nucleotide polymorphisms (SNPs) on tumour susceptibility and pathogenesis has gained enormous attention. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based genotyping facilitates the analysis of short DNA amplicons and is, therefore, a promising tool for the investigation of formalin-fixed paraffin-embedded (FFPE) tissue samples, particularly in targeted genotyping analysis. METHODS: To examine the applicability of genotyping FFPE DNA with MALDI-TOF MS in multiplex reactions, we investigated five DNA samples extracted from FFPE tumour specimens from follicular lymphoma patients using different extraction methods (phenol-chloroform, commercial kit). Thirty-one SNPs from 25 genes, integrated in different-sized multiplex assays (7-plex, 10-plex, 14-plex, 24-plex), were analyzed. To investigate the reliability of genotyping tumour-derived DNA extracted from FFPE tissue, we examined 64 FFPE tumour specimens in comparison with matched germline DNA samples. RESULTS: Call rates of 99.6 (274/275) and 93.5% (257/275) were observed for the DNA extracted with the phenol-chloroform approach or the commercial extraction kit, respectively. Increasing the number of SNPs per assay resulted in reduced genotyping call rates and genotyping quality, especially in the DNA samples isolated with the commercial extraction kit. When comparing the genotypes of DNA derived from germline and tumour (FFPE) specimens, a perfect concordance rate of 100% was detected. CONCLUSION: Our data delineate that MALDI-TOF-based genotyping of FFPE DNA is reliable and reproducible even in multiplex reactions, enabling the retrospective investigation of FFPE study cohorts in future experiments.
OBJECTIVE: The impact of single-nucleotide polymorphisms (SNPs) on tumour susceptibility and pathogenesis has gained enormous attention. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based genotyping facilitates the analysis of short DNA amplicons and is, therefore, a promising tool for the investigation of formalin-fixed paraffin-embedded (FFPE) tissue samples, particularly in targeted genotyping analysis. METHODS: To examine the applicability of genotyping FFPE DNA with MALDI-TOF MS in multiplex reactions, we investigated five DNA samples extracted from FFPE tumour specimens from follicular lymphomapatients using different extraction methods (phenol-chloroform, commercial kit). Thirty-one SNPs from 25 genes, integrated in different-sized multiplex assays (7-plex, 10-plex, 14-plex, 24-plex), were analyzed. To investigate the reliability of genotyping tumour-derived DNA extracted from FFPE tissue, we examined 64 FFPE tumour specimens in comparison with matched germline DNA samples. RESULTS: Call rates of 99.6 (274/275) and 93.5% (257/275) were observed for the DNA extracted with the phenol-chloroform approach or the commercial extraction kit, respectively. Increasing the number of SNPs per assay resulted in reduced genotyping call rates and genotyping quality, especially in the DNA samples isolated with the commercial extraction kit. When comparing the genotypes of DNA derived from germline and tumour (FFPE) specimens, a perfect concordance rate of 100% was detected. CONCLUSION: Our data delineate that MALDI-TOF-based genotyping of FFPE DNA is reliable and reproducible even in multiplex reactions, enabling the retrospective investigation of FFPE study cohorts in future experiments.
Authors: Daniel L Hertz; Kelley M Kidwell; Jacklyn N Thibert; Christina Gersch; Meredith M Regan; Todd C Skaar; N Lynn Henry; Daniel F Hayes; Catherine H Van Poznak; James M Rae Journal: Mol Oncol Date: 2015-07-29 Impact factor: 6.603
Authors: Shenli Zhang; Iain B Tan; Nur S Sapari; Heike I Grabsch; Alicia Okines; Elizabeth C Smyth; Toru Aoyama; Lindsay C Hewitt; Imran Inam; Dan Bottomley; Matthew Nankivell; Sally P Stenning; David Cunningham; Andrew Wotherspoon; Akira Tsuburaya; Takaki Yoshikawa; Richie Soong; Patrick Tan Journal: J Mol Diagn Date: 2015-03-04 Impact factor: 5.568
Authors: Heather L Gustafson; Song Yao; Bryan H Goldman; Kristy Lee; Catherine M Spier; Michael L LeBlanc; Lisa M Rimsza; James R Cerhan; Thomas M Habermann; Brian K Link; Matthew J Maurer; Susan L Slager; Daniel O Persky; Thomas P Miller; Richard I Fisher; Christine B Ambrosone; Margaret M Briehl Journal: Am J Hematol Date: 2014-04-12 Impact factor: 10.047