SPIDIA-RNA: First external quality assessment for the pre-analytical phase of blood samples used for RNA based analyses

► Study of pre-analytical variables involved in RNA extraction from blood samples. ► Implementation of a Pan European EQA for evaluation of RNA quality. ► RNA quality parameters: purity, quantity, integrity, gene expression stability. The diagnostic use of in vitro molecular assays can be limited by...

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Published in:Methods (San Diego, Calif.) Calif.), 2013-01, Vol.59 (1), p.20-31
Main Authors: Pazzagli, M., Malentacchi, F., Simi, L., Orlando, C., Wyrich, R., Günther, K., Hartmann, C.C., Verderio, P., Pizzamiglio, S., Ciniselli, C.M., Tichopad, A., Kubista, M., Gelmini, S.
Format: Article
Language:English
Subjects:
Blood Chemical Analysis - standards
Blood samples
Blood Specimen Collection - standards
Clinical trials
Edetic acid
Europe
External quality assessment
Gene Expression Profiling - standards
Guidelines as Topic
Humans
Laboratory Proficiency Testing
Pre-analytical phase
Quality control
RNA
RNA - blood
RNA - isolation & purification
RNA quality
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Summary:► Study of pre-analytical variables involved in RNA extraction from blood samples. ► Implementation of a Pan European EQA for evaluation of RNA quality. ► RNA quality parameters: purity, quantity, integrity, gene expression stability. The diagnostic use of in vitro molecular assays can be limited by the lack of guidelines for collection, handling, stabilization and storage of patient specimens. One of the major goals of the EC funded project SPIDIA (www.spidia.eu) is to develop evidence-based quality guidelines for the pre-analytical phase of blood samples used for molecular testing which requires intracellular RNA analytes. To this end, a survey and a pan-European external quality assessment (EQA) were implemented. This report is the summary of the results of that trial. With the European Federation of Laboratory Medicine (EFLM) support, 124 applications for participation in the trial were received from 27 different European countries, and 102 laboratories actually participated in the trial. Each participating laboratory described their respective laboratory policies and practices as well as blood collection tubes typically used in performing this type of testing. The participating laboratories received two identical blood specimens: in an EDTA tubes (unstabilized blood; n=67) or in tubes designed specifically for the stabilization of intracellular RNA in blood (PAXgene® Blood RNA tubes; n=35). Laboratories were requested to perform RNA extraction according to the laboratory’s own procedure as soon as possible upon receipt of the tubes for one tube and 24h after the first extraction for the second tube. Participants (n=93) returned the two extracted RNAs to SPIDIA facility for analysis, and provided details about the reagents and protocols they used for the extraction. At the SPIDIA facility responsible for coordinating the study, the survey data were classified, and the extracted RNA samples were evaluated for purity, yield, integrity, stability, and the presence of interfering substances affecting RT-qPCR assays. All participants received a report comparing the performance of the RNA they submitted to that of the other participants. All the results obtained by participants for each RNA quality parameter were classified as “in control”, “warning”, “out of control” and “missing” by consensus mean analysis. From the survey data, the most variable parameters were the volume of blood collected and the time and storage temperature between blood collection and
ISSN:1046-2023
1095-9130
DOI:10.1016/j.ymeth.2012.10.007