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Stability of Circulating Blood-Based MicroRNAs - Pre-Analytic Methodological Considerations
Charlotte Glinge1, Sebastian Clauss2,3,4, Kim Boddum5
1Department of Cardiology, the Heart Centre, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark.
Plos One
|February 3, 2017
Summary
Proper blood sample handling is crucial for accurate microRNA (miRNA) biomarker analysis. Standardizing collection tubes, storage, and processing conditions ensures reliable miRNA quantification in research and clinical settings.
Area of Science:
- Biochemistry
- Molecular Biology
- Biomarker Discovery
Background:
- MicroRNAs (miRNAs) show promise as non-invasive biomarkers for diagnosis, prognosis, and therapy.
- Standardized methodologies for miRNA analysis in blood are lacking.
- This study systematically evaluates miRNA stability under various pre-analytical conditions.
Purpose of the Study:
- To investigate the stability of circulating miRNAs in blood.
- To assess the impact of different collection tubes, storage temperatures, physical disturbance, and freeze-thaw cycles on miRNA levels.
- To provide recommendations for standardized miRNA sample handling.
Main Methods:
- Blood samples from 12 healthy donors were collected in EDTA, citrate, lithium-heparin, and serum tubes.
- MiRNA expression (miR-1, miR-21, miR-29b) was measured after varying processing times, long-term storage (-80°C), physical disturbance, and freeze-thaw cycles.
Main Results:
- Lithium-heparin tubes were unsuitable for miRNA quantification.
- MiRNAs remained stable in whole blood at room temperature for 24h, but alterations occurred in separated fractions.
- Significant changes in miR-21 and miR-29b were observed after 72h at room temperature.
- Physical disturbance affected miR-1 and miR-21 levels in plasma/serum.
- Long-term storage (-80°C) showed stability in plasma but not whole blood.
- Multiple freeze-thaw cycles reduced miRNA concentrations in plasma and serum.
Conclusions:
- Proper sample collection and preparation are critical for reliable circulating miRNA measurements.
- Standardization of collection tubes, preparation, handling, and storage is essential to avoid confounding variables.
- These findings are vital for the consistent application of miRNA biomarkers in clinical trials and research.
