Pre-Analytical Conditions Influence Cell-Free MicroRNA Stability in Blood Plasma Samples

Scientists worldwide are working to enhance the quality of diagnostics and prognosis for various diseases, including cancer, by analyzing different body fluids such as blood, urine, and saliva. MicroRNAs, which range from 18 to 25 nucleotides in length, are well-established post-transcriptional regulators of gene expression and play a crucial role in intracellular communication. These molecules, present in human biological fluids, are released from both normal and tumor cells, with about 3,000 individual microRNAs identified. These microRNAs circulate within biological fluids, often bound to biopolymers or packed into microvesicles, which makes them relatively stable. As a result, they are considered an ideal source of diagnostic material for liquid biopsies to detect tumors or other pathological conditions. For microRNAs to be utilized effectively as biomarkers, systematic and universally accepted guidelines for biosample processing must be developed, particularly concerning the stability of microRNAs during storage. Now, new research has shown that pre-analytical factors, such as the storage conditions of blood plasma or extracellular vesicles, significantly impact microRNA stability, thereby influencing the detected concentrations of specific microRNAs.

In a study conducted by researchers at the Novosibirsk Institute of Chemical Biology and Fundamental Medicine (ICBFM, Novosibirsk, Russia), the team investigated how different plasma storage conditions affected the stability of endogenous microRNAs in human blood plasma. The study focused on four endogenous microRNAs (miR-16, miR-19b, miR-23a, miR-451a) and the exogenous microRNA cel-miR-39, evaluating their stability under short- and long-term incubation at various temperatures. Additionally, the team examined how long-term storage affected the stability of microRNAs within extracellular vesicles (EVs). They also compared microRNA yields from fresh and archived plasma samples and assessed the impact of variations in miRNA extraction protocols and the use of RNA stabilizing agents on the efficiency of isolation.

The researchers employed a single-phase miRNA isolation method, which they had successfully used in previous studies to identify miRNA biomarkers for lung cancer in blood. Published in the ExRNA journal, the study confirmed that the degradation rate of microRNAs is influenced by their structure and packaging. They also found that adding various stabilization solutions to biofluids can impact the efficiency of miRNA extraction. The long-term findings from this study emphasize the importance of analyzing cell-free nucleic acids, including microRNAs, within 2 to 4 weeks after biological samples are collected to ensure the accuracy and reliability of diagnostic results.