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. 2023 Jan 4;28:101613. doi: 10.1016/j.tranon.2022.101613

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.