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Updated: Jul 8, 2025
Detection of miRNA Targets in High-throughput Using the 3'LIFE Assay
Published on: May 25, 2015
Chemiluminescent plate assay of microRNA-155 coupled with catalytic hairpin assembly with oligonucleotide release
Oleg L Bodulev1, Maxim D Zakharov1, Ivan Yu Sakharov1
1Department of Chemistry, Lomonosov Moscow State University, Leninskie gory, bldg.1, Moscow, 119991, Russia.
Abstract:
A heterogeneous sensitive microRNA-155 assay based on a new isothermal amplification method, called catalytic hairpin assembly with oligonucleotide release (CHAOR), was developed. The principle of CHAOR was studied by non-denaturing electrophoresis. To detect the amplification product, a polyperoxidase-streptavidin conjugate (molar ratio 1:80) and an enhanced chemiluminescence reaction were used, which made it possible to increase assay sensitivity. The detection limit of microRNA-155 assay was 0.4 pM. The coefficient of variation of the chemiluminescent signal, formed upon the heterogeneous determination of miRNA-155, was less than 12 % within the working range. The efficiency of CHAOR as an amplification method was similar to that of traditional CHA, as miRNA-155 assays based on CHAOR and CHA had similar analytical parameters. In addition, the proposed assay was highly specific. Contrary to traditional CHA, CHAOR, one of whose products is a single-stranded oligonucleotide, can be used in analytical methods based on cascade amplification.
