Multiplex Digital Quantification of β-Lactamase Genes in Antibiotic-Resistant Bacteria by Counting Gold Nanoparticle Labels on Silicon Microchips

Presnova, G.V.; Presnov, D.E.; Filippova, A.A.; Tsiniaikin, I.I.; Ulyashova, M.M.; Rubtsova, M.Y.

Статья опубликована в высокорейтинговом журнале

Информация о цитировании статьи получена из Scopus
Статья опубликована в журнале из списка Web of Science и/или Scopus
Дата последнего поиска статьи во внешних источниках: 11 мая 2022 г.

Авторы: Presnova G.V., Presnov D.E., Filippova A.A., Tsiniaikin I.I., Ulyashova M.M., Rubtsova M.Y.
Журнал: Biosensors
Том: 12
Год издания: 2022
Издательство: MDPI
Местоположение издательства: Basel, Switzerland
Номер статьи: 226
DOI: 10.3390/bios12040226
Аннотация: Digital quantification based on counting of individual molecules is a promising approach for different biomedical applications due to its enhanced sensitivity. Here, we present a method for the digital detection of nucleic acids (DNA and RNA) on silicon microchips based on the counting of gold nanoparticles (GNPs) in DNA duplexes by scanning electron microscopy (SEM). Bio-tin-labeled DNA is hybridized with capture oligonucleotide probes immobilized on the micro-chips. Then biotin is revealed by a streptavidin–GNP conjugate followed by the detection of GNPs. Sharp images of each nanoparticle allow the visualization of hybridization results on a sin-gle-molecule level. The technique was shown to provide highly sensitive quantification of both short oligonucleotide and long double-strand DNA sequences up to 800 bp. The lowest limit of detection of 0.04 pM was determined for short 19-mer oligonucleotide. The method’s applicability was demonstrated for the multiplex quantification of several β-lactamase genes responsible for the development of bacterial resistance against β-lactam antibiotics. Determination of nucleic acids is effective for both specific DNA in lysates and mRNA in transcripts. The method is also character-ized by high selectivity for single-nucleotide polymorphism discrimination. The proposed principle of digital quantification is a perspective for studying the mechanisms of bacterial antibiotic resistance and bacterial response to drugs.
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Multiplex Digital Quantification of β-Lactamase Genes in Antibiotic-Resistant Bacteria by Counting Gold Nanoparticle Labels on Silicon Microchipsстатья

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