title: Impact of preanalytical handling and timing for peripheral blood mononuclear cells isolation and RNA studies: the experience of the Interinstitutional Multidisciplinary BioBank (BioBIM).
abstract: Multicenter studies and biobanking projects require blood transportation from the participating center to a central collection or diagnostic laboratory. The impact of time delays between venous blood collection and peripheral blood mononuclear cells (PBMC) isolation prior to RNA extraction may affect the quality and quantity of isolated nucleic acids for genomic applications. Thus, standard operating procedure (SOP) optimization for the treatment of biological samples before RNA extraction is crucial in a biological repository. In order to define SOPs for whole blood preservation prior to RNA extraction, we sought to determine whether different blood storage times (0, 3, 6, 10, 24, and 30 hours) prior to PBMCs isolation and storage at -80°C, could affect the quality and quantity of extracted RNA. After spectrophotometric quantification, the quality and integrity of RNA were assessed by agarose gel electrophoresis, RNA integrity number and real time-PCR (RT-PCR). Across the different time points we did not observe significant differences within the first 24 hours of blood storage at room temperature, while a significant loss in RNA yield and integrity was detected between 24 and 30 hours. We conclude that time delays before PBMCs isolation prior to RNA extraction may have a significant impact on downstream molecular biological applications.
authors.affiliation: Laboratory of Molecular Diagnostics, Department of Laboratory Medicine & Advanced Biotechnologies, IRCCS San Raffaele Pisana, Rome - Italy. raffaele.palmirotta@sanraffaele.it
authors.forename: Raffaele
authors.initials: R
authors.lastname: Palmirotta
authors.forename: Maria Laura
authors.initials: ML
authors.lastname: De Marchis
authors.forename: Giorgia
authors.initials: G
authors.lastname: Ludovici
authors.forename: Barbara
authors.initials: B
authors.lastname: Leone
authors.forename: Annalisa
authors.initials: A
authors.lastname: Savonarola
authors.forename: Cristiano
authors.initials: C
authors.lastname: Ialongo
authors.forename: Antonella
authors.initials: A
authors.lastname: Spila
authors.forename: Francesco
authors.initials: F
authors.lastname: De Angelis
authors.forename: Patrizia
authors.initials: P
authors.lastname: Ferroni
authors.forename: David
authors.initials: D
authors.lastname: Della-Morte
authors.forename: Fiorella
authors.initials: F
authors.lastname: Guadagni
mesh_terms: D001793:Blood Preservation / Q000379:methods*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000737:chemistry*; D009000:Monocytes / Q000737:chemistry*; D012313:RNA / Q000032:analysis / Q000097:blood*; D060888:Real-Time Polymerase Chain Reaction; D013053:Spectrophotometry
chemical_list: D012313:RNA