[Study on the Sample Preservation Temperature and Period in Circulating MicroRNA Quantification Using Spike-In
Rinsho Byori. the Japanese Journal of Clinical Pathology
|November 10, 2015
Summary
Circulating microRNAs are promising biomarkers. To ensure accurate quantification using spike-in controls, avoid storing serum at 4°C for extended periods, as this degrades microRNAs like miR-223.
Area of Science:
- Biochemistry
- Molecular Biology
- Biomarker Discovery
Context:
- Circulating microRNAs (miRNAs) in body fluids show potential as non-invasive biomarkers for various diseases.
- Real-time RT-PCR quantification of circulating miRNAs often uses synthetic spike-in control RNA (e.g., cel-miR-39).
- Spike-in controls added before RNA extraction do not account for pre-extraction RNA degradation, impacting reproducibility.
Purpose:
- To assess the stability of synthetic cel-miR-39 and endogenous circulating miRNAs (miR-21, miR-223) in different sample types and conditions.
- To identify optimal pre-analytical conditions for preserving circulating miRNA integrity before RNA extraction.
- To improve the reliability of circulating miRNA quantification using spike-in controls.
Summary:
- Synthetic cel-miR-39 remained stable on ice for at least 3 hours.
- miR-21 and miR-223 showed no degradation in whole blood for 3 hours at room temperature.
- Significant degradation of miR-223 was observed in serum stored at 4°C within 24 hours, highlighting the need to avoid prolonged serum storage at this temperature.
Impact:
- Findings emphasize the critical need to prevent degradation of circulating miRNAs in samples before RNA extraction.
- Highlights the limitations of current spike-in control methods in reflecting real-world sample degradation.
- Provides crucial insights for optimizing pre-analytical workflows to enhance the accuracy and reproducibility of circulating miRNA biomarker studies.
