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Updated: Nov 4, 2025
Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs
Published on: April 14, 2015
Methodological considerations for measuring biofluid-based microRNA biomarkers
Brian N Chorley1, Elnaz Atabakhsh2, Graeme Doran2
1U.S. Environmental Protection Agency, Research Triangle Park, NC, USA.
Abstract:
MicroRNAs (miRNAs) are small non-coding RNA that regulate the expression of messenger RNA and are implicated in almost all cellular processes. Importantly, miRNAs can be released extracellularly and are stable in these matrices where they may serve as indicators of organ or cell-specific toxicity, disease, and biological status. There has thus been great enthusiasm for developing miRNAs as biomarkers of adverse outcomes for scientific, regulatory, and clinical purposes. Despite advances in measurement capabilities for miRNAs, miRNAs are still not routinely employed as noninvasive biomarkers. This is in part due to the lack of standard approaches for sample preparation and miRNA measurement and uncertainty in their biological interpretation. Members of the microRNA Biomarkers Workgroup within the Health and Environmental Sciences Institute's (HESI) Committee on Emerging Systems Toxicology for the Assessment of Risk (eSTAR) are a consortium of private- and public-sector scientists dedicated to developing miRNAs as applied biomarkers. Here, we explore major impediments to routine acceptance and use of miRNA biomarkers and case examples of successes and deficiencies in development. Finally, we provide insight on miRNA measurement, collection, and analysis tools to provide solid footing for addressing knowledge gaps toward routine biomarker use.
Insights
MicroRNAs (miRNAs), stable extracellular molecules, show promise as noninvasive biomarkers for disease and toxicity. Standardization in their measurement and interpretation is crucial for routine clinical and regulatory use.
Area of Science:
- Molecular Biology
- Biomarker Discovery
- Toxicology
Background:
- MicroRNAs (miRNAs) are small non-coding RNAs regulating gene expression, involved in numerous cellular processes.
- Extracellular miRNAs are stable and can indicate organ/cell-specific toxicity, disease, and biological status.
- Significant interest exists in utilizing miRNAs as noninvasive biomarkers for adverse outcomes in scientific, regulatory, and clinical settings.
Purpose of the Study:
- To explore major impediments hindering the routine acceptance and application of miRNA biomarkers.
- To examine case examples illustrating both successes and deficiencies in miRNA biomarker development.
- To provide insights into miRNA measurement, collection, and analysis to address knowledge gaps.
Main Methods:
- Review of current literature and expert group discussions on miRNA biomarker development.
- Analysis of challenges in standardization of sample preparation and miRNA measurement techniques.
- Exploration of biological interpretation uncertainties for miRNA biomarkers.
Main Results:
- Lack of standardized protocols for sample handling and miRNA quantification impedes routine use.
- Uncertainty in the biological interpretation of miRNA levels contributes to biomarker development challenges.
- Despite challenges, progress has been made, with specific examples highlighting both successful and deficient development pathways.
Conclusions:
- Standardized approaches for sample preparation, measurement, and interpretation are essential for the widespread adoption of miRNA biomarkers.
- Addressing current knowledge gaps through robust tools and methodologies will facilitate the routine application of miRNAs in diagnostics and risk assessment.
- Collaborative efforts are key to overcoming hurdles and realizing the full potential of miRNA biomarkers.
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