Journal
CLINICAL CHEMISTRY AND LABORATORY MEDICINE
Volume 55, Issue 5, Pages 608-621Publisher
WALTER DE GRUYTER GMBH
DOI: 10.1515/cclm-2016-0471
Keywords
biomarker; circulating; FFPE; miRNA; pre-analytical; standardization; tissue; variables
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microRNAs (miRNAs) hold promise as biomarkers for a variety of disease processes and for determining cell differentiation. These short RNA species are robust, survive harsh treatment and storage conditions and may be extracted from blood and tissue. Pre-analytical variables are critical confounders in the analysis of miRNAs: we elucidate these and identify best practices for minimizing sample variation in blood and tissue specimens. Preanalytical variables addressed include patient-intrinsic variation, time and temperature from sample collection to storage or processing, processing methods, contamination by cells and blood components, RNA extraction method, normalization, and storage time/conditions. For circulating miRNAs, hemolysis and blood cell contamination significantly affect profiles; samples should be processed within 2 h of collection; ethylene diamine tetraacetic acid (EDTA) is preferred while heparin should be avoided; samples should be double spun or filtered; room temperature or 4 degrees C storage for up to 24 h is preferred; miRNAs are stable for at least 1 year at -20 degrees C or -80 degrees C. For tissuebased analysis, warm ischemic time should be < 1 h; cold ischemic time (4 degrees C) < 24 h; common fixative used for all specimens; formalin fix up to 72 h prior to processing; enrich for cells of interest; validate candidate biomarkers with in situ visualization. Most importantly, all specimen types should have standard and common workflows with careful documentation of relevant pre-analytical variables.
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