Table 1.

Main high-throughput approaches for miRNAs profiling, comparing their features and limitations in detecting these molecules.

Detection methods	Principle/Main functions	Advantages	Limitations
qRT-PCR	-Amplification-based -Best used for validation of targets obtained by large-scale approaches	-Convenient/simple -Relatively fast -Relatively inexpensive -Relatively easy data analysis -High sensitivity and specificity -Absolute and relative quantification	-Challenges in primers design -Controversial reference genes -Difficulty to discern subfamily targets -Sensitive to gDNA contaminants
Microarrays	-Hybridization-based -Best used for preliminary screening	-Relatively fast -Easily standardized -Biochips readily available	-Low sensitivity -Low specificity for targets in the same subfamily -Cross-hybridization and background noise issues -Restricted to array content -Relatively expensive custom-made chips
Deep sequencing	-Sequencing-based -Best used for large-scale novel biomarkers identification	-Detection of novel targets -High dynamic range, sensitivity, and, specificity -No prior sequence knowledge required -Absolute quantification	-Relatively high cost -Complex data processing and interpretation -Requirement of specialized personnel and equipment -Sequencing bias -Time-consuming
Nanostring's nCounter	-Hybridization-based	-No amplification steps -Relatively fast -Absolute quantification -High accuracy	-Only available in specialized facilities with limited accessibility -Reduced sensitivity

Abbreviations: qRT-PCR; quantitative real-time reverse-transcription-polymerase chain reaction, gDNA; genomic DNA.