Magnetic Three-Phase Single-Drop Microextraction for Rapid Amplification of the Signals of DNA and MicroRNA Analysis.

The detection of nucleic acids usually suffers from a lengthy amplification process. To obtain an enhanced signal within several seconds, a magnetic three-phase single-drop microextraction (MTP-SDME) approach was developed for the quantification of nucleic acids. First, a target-triggered recycling amplification strategy was used to constitute magnetic branched DNA/Fe3O4 networks, which displayed peroxidase-like catalytic activity toward the 3,3',5,5'-tetramethylbenzidine colorimetric reaction. The networks were separated and enriched by rapid (6 s) MTP-SDME (with only 6 μL of solvent required), thereby producing highly sensitive signals for the quantification of nucleic acids. The signals were significantly amplified by the triple strategy (network formation, MTP-SDME, and catalytic reaction). The application of magnetic extraction minimized the background signal, avoided sample matrix effects, and enhanced the analyte signals. This assay achieved linear calibration curves of between 0.5 aM and 1 pM for microRNA-122 (miRNA-122) and between 1 aM and 1 pM for HBV-T (a DNA fragment from hepatitis B virus). Limits of detection of 0.15 aM for miRNA-122 and 0.34 aM for HBV-T were attained, with relative standard deviations of <5.0% (n = 3). Furthermore, the procedure was applied to determine miRNA-122 and HBV-T in genuine serum samples from hepatocellular carcinoma patients.