Highly Sensitive Detection of Multiple MicroRNAs by High-Performance Liquid Chromatography Coupled with Long and Short Probe-Based Recycling Amplification.
2020
This
report demonstrated the utility of high-performance liquid
chromatography (HPLC)-fluorescence detection for selective separation
and sensitive quantification of multiple microRNAs (miRNAs). A duplex
specific nuclease (DSN)-assisted target recycling amplification strategy
was developed to enhance the signals of miRNAs, which alleviates the
low sensitivity of conventional HPLC to nucleic acids. To separate
the signals of different miRNAs, DNA probes with different lengths
and base sequences were immobilized on magnetic beads. The application
of an effective magnetic separation minimized the background signal
and extended the dynamic range. This assay achieved a limit of detection
of 0.39 fM for miRNA-122, 0.30 fM for miRNA-155, and 0.26 fM for miRNA-21,
respectively. The proposed assay was successfully applied to detect
simultaneously miRNA-122, miRNA-155, and miRNA-21 in serum samples
from healthy persons and cervical cancer patients, and the results
were then compared with those of quantitative real-time-polymerase
chain reaction amplification.
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