Background
MicroRNAs (miRNAs) can be used for early diagnosis of myocardial infarction. However, due to a lack of standardized operating procedures, their value for clinical application is low.
Conclusion
Preliminary standardization for miRNA detection provides a reference for clinical blood testing of miRNAs.
Methods
Detection of plasma miRNAs was optimized by analyzing factors influencing miRNA variance and myocardial infarction risk scores during analysis (extraction, reverse transcription, and real-time PCR) and pre-analysis (dietary status, anticoagulants, storage conditions, and hemolysis).
Results
Regarding variable factors during analysis, the centrifugal column method was superior to Trizol LS reagent when extracting miRNA from plasma. Recovery rate was highest with plasma volumes of 200 and 300 µL. During analysis, the main source of miRNA detection inaccuracy was derived from RNA extraction (mainly organic extraction), and not reverse transcription or PCR. MiRNA variance could be reduced by use of an internal reference. During analysis, 95% of risk score variation fluctuated within a range of 6.267. The variable factors pre-analysis mainly involved dietary status, anticoagulant selection, and storage conditions. Hemolysis positively correlated with miRNA levels, but there was no significant change in risk score after internal reference calibration.