title: Risk for molecular contamination of tissue samples evaluated for targeted anti-cancer therapy.
abstract: With the increasing usage of sensitive PCR technology for pharmacogenetics, cross contamination becomes a significant concern. Researchers employed techniques which basically include replacing laboratory equipment after each sample preparation; however, there are no recommended guidelines. In the present work we wanted to evaluate the risk of cross contamination during tissue processing using the routine precaution measures. Twenty-one surgical samples of lung adenocarcinoma were used, of which 7 contained EGFR exon 19 mutation, 7 contained EGFR exon 21 mutation (p.L858R) and 7 were EGFR wild-type. The samples were ordered by alternating the mutation group to maximize the potential for cross contamination and underwent tissue sectioning and de-paraffinization. The entire process was performed using the same tools. Following DNA extraction all samples underwent PCR amplification and were scrutinized for small fractions of EGFR mutation using deep sequencing with the Ion torrent PGM technology. Twenty samples yielded results. The fraction of mutated copies was 41 ± 23% (range 11-66) for the cases with known exon 19 mutation and 48±24% (range 0-65) for the cases with known exon 21 mutations. No in-frame exon 19 deletion mutations were identified in the wild-type (WT) and exon 21 groups. The fraction of EGFR exon 21 (codon 858) mutations was 0.018±0.014% (range 0-0.05%) in the WT and exon 19 groups, which was not statistically different than the background sequencing artifact noise for the same base-pair alteration (p = 0.21). Our results suggest that standard precautions are sufficient for molecular pathology diagnosis of surgical samples and are not associated with increased risk of cross contamination.
authors.affiliation: B. Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.|Institute of Pathology, Rambam Health Care Campus, Haifa, Israel.
authors.forename: Eyal
authors.initials: E
authors.lastname: Asor
authors.affiliation: Institute of Pathology, Rambam Health Care Campus, Haifa, Israel.
authors.forename: Michael Y
authors.initials: MY
authors.lastname: Stav
authors.affiliation: B. Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.
authors.forename: Einav
authors.initials: E
authors.lastname: Simon
authors.affiliation: Institute of Pathology, Tel-Aviv Sourasky Medical Center, Tel-Aviv, Israel.
authors.forename: Ibrahim
authors.initials: I
authors.lastname: Fahoum
authors.affiliation: B. Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.|Institute of Pathology, Rambam Health Care Campus, Haifa, Israel.
authors.forename: Edmond
authors.initials: E
authors.lastname: Sabo
authors.affiliation: B. Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.|Institute of Pathology, Rambam Health Care Campus, Haifa, Israel.
authors.forename: Ofer
authors.initials: O
authors.lastname: Ben-Izhak
authors.affiliation: Institute of Pathology, Tel-Aviv Sourasky Medical Center, Tel-Aviv, Israel.|Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv, Israel.
authors.forename: Dov
authors.initials: D
authors.lastname: Hershkovitz
mesh_terms: D000230:Adenocarcinoma / Q000235:genetics* / Q000473:pathology; D000077192:Adenocarcinoma of Lung; D066246:ErbB Receptors / Q000235:genetics; D005091:Exons; D006801:Humans; D057230:Limit of Detection; D008175:Lung Neoplasms / Q000235:genetics* / Q000473:pathology; D058990:Molecular Targeted Therapy; D009154:Mutation; D059645:Mutation Rate; D016612:Paraffin Embedding / Q000379:methods / Q000592:standards; D057089:Pathology, Molecular / Q000379:methods* / Q000592:standards; D016133:Polymerase Chain Reaction / Q000379:methods; D016707:Tissue Fixation / Q000379:methods / Q000592:standards
chemical_list: C512478:EGFR protein, human; D066246:ErbB Receptors