Abstract

Breakfast Briefing: From Discovery to Diagnostic: Building miRNA-based Tools for Pediatric Heart Failure Patients

Pete Mariner, Founder, CoramiR Biomedical

Circulating miRNAs are important biomarkers of disease, and several methodologies are available that identify circulating miRNAs with varied specificity and reproducibility. We have previously shown that circulating miRNAs can accurately predict recovery from heart failure in pediatric dilated cardiomyopathy patients, and to determine cardiac allograft vasculopathy. Our previous work was performed using serum-extracted RNA and Life Technologies miRNA array cards. Here we describe an improvement to the original methodology that consists of performing miRNA arrays directly from serum, without the need of RNA extraction. Importantly, although 250 µl of serum were necessary to perform RNA extraction, only 3 µl are necessary to perform serum-miRNA-arrays. Furthermore, a thorough description of the methodology used to improve release of miRNAs from microparticles is provided. Moreover, whereas circulating miRNA studies are often based on relative comparison values, an absolute quantification of circulating miRNAs is necessary of tests will be standardize across laboratories. We have successfully generated an absolute quantification platform that can be easily implemented in RT-PCR confirmatory studies. In summary, the work described here provides specific guidelines to detect and quantify circulating miRNAs.