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PLoS One. 2014 Nov 10;9(11):e112293. doi: 10.1371/journal.pone.0112293. eCollection 2014.

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

Author information

1
Department of Biomedical Experimental and Clinical Sciences, University of Florence, Florence, Italy.
2
Qiagen GmbH, Hilden, Germany.
3
Unit of Medical Statistics, Biometry and Bioinformatics, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Istituto Nazionale dei Tumori, Milan, Italy.
4
DiaGenic ASA, Oslo, Norway.
5
Institute of Biotechnology, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
6
Becton Dickinson (BD), Franklin Lakes, New Jersey, United States of America.
7
Labonnet Ltd, St. Ramat-Hasharon, Israel.
8
Institute of Biotechnology, Academy of Sciences of the Czech Republic, Prague, Czech Republic; TATAA Biocenter AB, Gothenburg, Sweden.

Abstract

One purpose of the EC funded project, SPIDIA, is to develop evidence-based quality guidelines for the pre-analytical handling of blood samples for RNA molecular testing. To this end, two pan-European External Quality Assessments (EQAs) were implemented. Here we report the results of the second SPIDIA-RNA EQA. This second study included modifications in the protocol related to the blood collection process, the shipping conditions and pre-analytical specimen handling for participants. Participating laboratories received two identical proficiency blood specimens collected in tubes with or without an RNA stabilizer. For pre-defined specimen storage times and temperatures, laboratories were asked to perform RNA extraction from whole blood according to their usual procedure and to return extracted RNA to the SPIDIA facility for further analysis. These RNA samples were evaluated for purity, yield, integrity, stability, presence of interfering substances, and gene expression levels for the validated markers of RNA stability: FOS, IL1B, IL8, GAPDH, FOSB and TNFRSF10c. Analysis of the gene expression results of FOS, IL8, FOSB, and TNFRSF10c, however, indicated that the levels of these transcripts were significantly affected by blood collection tube type and storage temperature. These results demonstrated that only blood collection tubes containing a cellular RNA stabilizer allowed reliable gene expression analysis within 48 h from blood collection for all the genes investigated. The results of these two EQAs have been proposed for use in the development of a Technical Specification by the European Committee for Standardization.

PMID:
25384019
PMCID:
PMC4226503
DOI:
10.1371/journal.pone.0112293
[Indexed for MEDLINE]
Free PMC Article

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