title: The focus on sample quality: Influence of colon tissue collection on reliability of qPCR data.
abstract: Successful molecular analyses of human solid tissues require intact biological material with well-preserved nucleic acids, proteins, and other cell structures. Pre-analytical handling, comprising of the collection of material at the operating theatre, is among the first critical steps that influence sample quality. The aim of this study was to compare the experimental outcomes obtained from samples collected and stored by the conventional means of snap freezing and by PAXgene Tissue System (Qiagen). These approaches were evaluated by measuring rRNA and mRNA integrity of the samples (RNA Quality Indicator and Differential Amplification Method) and by gene expression profiling. The collection procedures of the biological material were implemented in two hospitals during colon cancer surgery in order to identify the impact of the collection method on the experimental outcome. Our study shows that the pre-analytical sample handling has a significant effect on the quality of RNA and on the variability of qPCR data. PAXgene collection mode proved to be more easily implemented in the operating room and moreover the quality of RNA obtained from human colon tissues by this method is superior to the one obtained by snap freezing.
authors.affiliation: Institute of Biotechnology, BIOCEV Centre, Czech Academy of Sciences, Průmyslová 595, 252 42, Vestec u Prahy, Czech Republic.
authors.forename: Vlasta
authors.initials: V
authors.lastname: Korenkova
authors.affiliation: Institute of Experimental Medicine, Czech Academy of Sciences, Prague, Czech Republic.
authors.forename: Jana
authors.initials: J
authors.lastname: Slyskova
authors.affiliation: Institute of Biotechnology, BIOCEV Centre, Czech Academy of Sciences, Průmyslová 595, 252 42, Vestec u Prahy, Czech Republic.
authors.forename: Vendula
authors.initials: V
authors.lastname: Novosadova
authors.affiliation: Unit of Medical Statistics, Biometry and Bioinformatics, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Istituto Nazionale dei Tumori, Milan, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Pizzamiglio
authors.affiliation: Institute of Biotechnology, BIOCEV Centre, Czech Academy of Sciences, Průmyslová 595, 252 42, Vestec u Prahy, Czech Republic.
authors.forename: Lucie
authors.initials: L
authors.lastname: Langerova
authors.affiliation: TATAA Biocenter AB, Göteborg, Sweden.
authors.forename: Jens
authors.initials: J
authors.lastname: Bjorkman
authors.affiliation: Deparment of Surgery, Teaching Hospital and Medical School Pilsen, Charles University in Prague, Pilsen, Czech Republic.|Biomedical Centre, Medical School Pilsen, Charles University in Prague, Pilsen, Czech Republic.
authors.forename: Ondrej
authors.initials: O
authors.lastname: Vycital
authors.affiliation: Deparment of Surgery, Teaching Hospital and Medical School Pilsen, Charles University in Prague, Pilsen, Czech Republic.|Biomedical Centre, Medical School Pilsen, Charles University in Prague, Pilsen, Czech Republic.
authors.forename: Vaclav
authors.initials: V
authors.lastname: Liska
authors.affiliation: Surgical Department, Thomayer Hospital, First Faculty of Medicine, Charles University in Prague, Prague, Czech Republic.
authors.forename: Miroslav
authors.initials: M
authors.lastname: Levy
authors.affiliation: Surgical Department, Thomayer Hospital, First Faculty of Medicine, Charles University in Prague, Prague, Czech Republic.
authors.forename: Karel
authors.initials: K
authors.lastname: Veskrna
authors.affiliation: Institute of Experimental Medicine, Czech Academy of Sciences, Prague, Czech Republic.|Biomedical Centre, Medical School Pilsen, Charles University in Prague, Pilsen, Czech Republic.|Institute of Biology and Medical Genetics, First Faculty of Medicine, Charles University in Prague, Prague, Czech Republic.
authors.forename: Pavel
authors.initials: P
authors.lastname: Vodicka
authors.affiliation: Institute of Experimental Medicine, Czech Academy of Sciences, Prague, Czech Republic.|Biomedical Centre, Medical School Pilsen, Charles University in Prague, Pilsen, Czech Republic.|Institute of Biology and Medical Genetics, First Faculty of Medicine, Charles University in Prague, Prague, Czech Republic.
authors.forename: Ludmila
authors.initials: L
authors.lastname: Vodickova
authors.affiliation: Institute of Biotechnology, BIOCEV Centre, Czech Academy of Sciences, Průmyslová 595, 252 42, Vestec u Prahy, Czech Republic.|TATAA Biocenter AB, Göteborg, Sweden.
authors.forename: Mikael
authors.initials: M
authors.lastname: Kubista
authors.affiliation: Unit of Medical Statistics, Biometry and Bioinformatics, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Istituto Nazionale dei Tumori, Milan, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Verderio
mesh_terms: D002277:Carcinoma / Q000737:chemistry* / Q000601:surgery; D003106:Colon / Q000737:chemistry*; D003110:Colonic Neoplasms / Q000737:chemistry* / Q000601:surgery; D015925:Cryopreservation / Q000295:instrumentation / Q000379:methods*; D004264:DNA Topoisomerases, Type I / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D020869:Gene Expression Profiling / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods; D006801:Humans; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics*; D009584:Nitrogen; D019188:Organ Preservation Solutions; D011309:Preservation, Biological / Q000295:instrumentation / Q000379:methods*; D011786:Quality Control; D012334:RNA, Neoplasm / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification*; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D013048:Specimen Handling / Q000295:instrumentation / Q000379:methods*; D016707:Tissue Fixation / Q000379:methods
chemical_list: D004273:DNA, Neoplasm; D004275:DNA, Ribosomal; D009363:Neoplasm Proteins; D019188:Organ Preservation Solutions; D012334:RNA, Neoplasm; D012337:RNA, Ribosomal, 18S; D004264:DNA Topoisomerases, Type I; C528909:TOP1 protein, human; D009584:Nitrogen