Quantitation of MicroRNA-155 in Human Cells by Heterogeneous Enzyme-Linked Oligonucleotide Assay Coupled with Mismatched Catalytic Hairpin Assembly Reaction

Авторы: Bodulev Oleg L., Galkin Ivan I., Zhao Shulin, Pletyushkina Olga Y., Sakharov Ivan Y.
Журнал: Biosensors
Том: 12
Номер: 8
Год издания: 2022
Издательство: MDPI
Местоположение издательства: Basel, Switzerland
Номер статьи: 570
DOI: 10.3390/bios12080570
Аннотация: In the present work, we describe the development of a chemiluminescent enzyme-linkedoligonucleotide assay coupled with mismatched catalytic hairpin assembly (mCHA) amplification forthe quantitative determination of microRNA-155. To improve its sensitivity, a polymerase-free mCHAreaction was applied as an isothermal amplification method. The detection limit of the proposedassay was 400 fM. In addition, the high specificity of the assay was demonstrated. The proposed assayallowed assessment of the content of microRNA-155 in human cancer lines such as HepG2, Caco2,MCF7, and HeLa. The quantitation of microRNA-155 was performed after purification of short RNAs(less than 200 nt) from cell lysates since a high matrix effect was observed without this pre-treatment.The results of the quantitative determination of the microRNA content in cells were normalized usingnematode microRNA-39, the concentration of which was determined using a heterogeneous assaydeveloped by us using a strategy identical to that of the microRNA-155 assay.
Добавил в систему: Сахаров Иван Юрьевич

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