Long-term Storage of Urine and Urinary EVs: A Systematic Evaluation of microRNA Stability

However, their stability during long-term storage and freeze–thaw cycles remain insufficiently characterized, limiting both research throughput and clinical application. In this study, we systematically evaluated the temporal stability of urine for up to 12 months.

In addition, as a point of comparison, we examined the stability of uEVs, miRNAs purified from uEVs, and cDNA libraries synthesized from miRNAs for up to 3 months. Next generation sequencing-based profiling demonstrated that the abundance of miRNAs and subject-specific expression profiles were largely preserved under all storage conditions during the observation periods. At the individual miRNA level, expression patterns were generally maintained in urine for 12 months and in uEVs and cDNA libraries for 3 months, whereas partial degradation was observed when stored as RNA.