Impact of ambient temperature exposure on miRNA stability in human plasma

MicroRNAs (miRNAs) are considered more stable than mRNA, but the impact of progressive thawing of biological samples after freezing as may happen during shipping delays has not been quantified. To address this, we utilized digital PCR to estimate the absolute concentrations of select miRNAs following progressive thawing of human plasma and maintenance at ambient temperature. Specifically, we quantified let-7b-3p, miR-144-5p, miR-150-5p, miR-517a-3p, miR-524-5p, and miR-1283, which have varying abundance in plasma. We observed a trend indicating a decline in miRNA concentration as plasma samples were progressively thawed. Notably, miR-150-5p and miR-517a-3p were the least stable and were degraded by 32% and 52% respectively after 24 hours of ambient temperature storage. We found that the variation in sensitivity to temperature was not due to the GC content of the miRNAs nor their initial abundance, suggesting that other factors, such as protein interactors and vesicles carrying these miRNAs, may impact sensitivity. Plasma samples from nine pregnant women from the Gen3G cohort which were stored in −80 °C for 12 years, were used to measure microRNA stability due to progressive thawing often encountered during shipping and experimental handling. We pooled samples from different participants and added two spike-in controls to reduce technical variability. A trend in microRNA (miRNA) degradation in plasma samples with progressive thawing.Degradation of miRNA was not dependent on % GC or initial abundance.We predict that factors such as protein interactors and vesicles carrying these miRNAs impact degradation rate. A trend in microRNA (miRNA) degradation in plasma samples with progressive thawing. Degradation of miRNA was not dependent on % GC or initial abundance. We predict that factors such as protein interactors and vesicles carrying these miRNAs impact degradation rate.