Isothermal and enzyme-free microRNA assay based on catalytic hairpin assembly and lanthanide-labeled probes

Session: Analytical Division Poster:  

MicroRNAs (miRNAs) are a category of short non-coding RNA molecules which play an essential role in regulating gene expression. Correspondingly, the aberrant expression of miRNAs is commonly related to the development of various diseases including cancers and cardiovascular diseases. Hence, miRNA is considered as an ideal biomarker for disease diagnosis and its quantification assays are well developed. Herein, an isothermal and enzyme-free miRNA assay based on catalytic hairpin assembly (CHA) amplification and inductively coupled plasma mass spectrometry (ICP-MS) detection is proposed. Two hairpin probes are employed in CHA reaction with lanthanide tag on HP1 for ICP-MS detection and biotin on HP2 for immobilization on magnetic beads. In the presence of target miRNA, the two independent probes open up their stem-loop structure and hybridized. Significantly, target miRNA will be released after the hybridization which can be utilized to trigger another CHA reaction. Ultimately, one target miRNA molecule promotes a number of CHA cycles and produces several copies of hybridized complex leading to the enhanced ICP-MS signal. Based on this approach, miRNA-141 was successfully analyzed with high sensitivity and specificity. The limit of detection (LOD) was 20 fmol of miRNA-141 with the linear range from 20 fmol to 1 pmol. Meanwhile, miRNA-429, miRNA-200a, miRNA-200b, miRNA-200c can be specifically discriminated from miRNA-141. The developed method was also applied to analyse miRNA-141 in tumor cell lysate to demonstrate its practical applicability. Furthermore, this miRNA assay has the potential to become a high-throughput method by employing distinct lanthanide elements.