Quantification of MicroRNAs in Urine-Derived Specimens

MicroRNAs are small noncoding RNAs which regulate the expression of genes involved in a multitude of cellular processes. Dysregulation of microRNAs and—in consequence—of the affected pathways is frequently observed in numerous pathologies including cancers. Therefore, tumor-related alterations in microRNA expression and function can reflect molecular processes of tumor onset and progression qualifying microRNAs as potential diagnostic and prognostic biomarkers. In particular, microRNAs with differential expression in bladder cancer (BCa) might represent promising tools for noninvasive tumor detection in urine. This would be helpful not only for diagnostic and monitoring purposes but also for therapeutic decisions. Detection and quantification of BCa-associated microRNAs in urine can be performed using the cellular sediment, which also contains BCa cells, or in exosomes originating from those cells. Methods for isolation of exosomes from urine, extraction of total RNA from cells and exosomes as well as techniques for RNA quantification, reverse transcription, and qPCR-based quantification of microRNA expression levels are described herein.