Preanalytic Methodological Considerations and Sample Quality Control of Circulating miRNAs

Abstract

As miRNAs emerge as potential circulating biomarkers for the diagnosis or prognosis of a wide variety of diseases, the quantification of miRNA necessitates careful preanalytic considerations and sample quality control becomes crucial. This study comprehensively analyzed the profiles of 356 miRNAs by quantitative RT-PCR in various blood sample types, with various processing protocols. The comprehensive analysis investigated the correlations of individual miRNAs with certain confounding factors. On the basis of these profiles, a panel of 7 miRNAs was established for the quality control of samples corresponding to hemolysis and platelet contamination. The panel was used to investigate the confounding impacts based on the size of the blood collection tube, the centrifugation protocol, post-freeze-thaw spinning, and whole blood storage. A standard dual-spin workflow for the processing of blood had been established for optimal sample quality. The real-time stability of 356 miRNAs was also investigated with demonstration of the temperature and time-induced miRNA degradation profile. Stability-related miRNAs were identified from real-time stability study and further incorporated into the quality control panel. This quality control panel enables the assessment of sample quality for more robust and reliable detection of circulating miRNAs.