POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices

Abstract Oligonucleotide therapeutics use short interfering RNA (siRNA) or antisense oligonucleotide (ASO) molecules to exploit endogenous systems—neutralizing target RNA to prevent subsequent protein translation. While the potential clinical application is vast, delivery efficiency and extrahepatic...

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Autores principales:	Mai B. Thayer, Sara C. Humphreys, Kyu S. Chung, Julie M. Lade, Kevin D. Cook, Brooke M. Rock
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2020
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/22cff2a3388640828a7f309b40b4cccd
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spelling	oai:doaj.org-article:22cff2a3388640828a7f309b40b4cccd2021-12-02T17:45:12ZPOE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices10.1038/s41598-020-66829-62045-2322https://doaj.org/article/22cff2a3388640828a7f309b40b4cccd2020-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-020-66829-6https://doaj.org/toc/2045-2322Abstract Oligonucleotide therapeutics use short interfering RNA (siRNA) or antisense oligonucleotide (ASO) molecules to exploit endogenous systems—neutralizing target RNA to prevent subsequent protein translation. While the potential clinical application is vast, delivery efficiency and extrahepatic targeting is challenging. Bioanalytical assays are important in building understanding of these complex relationships. The literature currently lacks description of robust and sensitive methods to measure siRNA and ASOs in complex biological matrices. Described herein is a non-enzymatic hybridization-based immunoassay that enables quantification of individual siRNA strands (antisense or sense) in serum, urine, bile, and liver and kidney homogenates. Assay utility is also demonstrated in ASOs. The assay improves upon previous works by abolishing enzymatic steps and further incorporating Locked Nucleic Acid (LNA) nucleotide modifications to increase analyte hybridization affinity and improve sensitivity, specificity, and robustness. We report an assay with an ultrasensitive dynamic range of 0.3 to 16,700 pM for siRNA in serum. The assay was submitted to full qualification for accuracy and precision in both serum and tissue matrices and assay performance was assessed with single and mixed analytes. The reliable LNA-hybridization-based approach removes the need for matrix sample extraction, enrichment or amplification steps which may be impeded by more advanced chemical modifications.Mai B. ThayerSara C. HumphreysKyu S. ChungJulie M. LadeKevin D. CookBrooke M. RockNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 10, Iss 1, Pp 1-9 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Mai B. Thayer Sara C. Humphreys Kyu S. Chung Julie M. Lade Kevin D. Cook Brooke M. Rock POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
description	Abstract Oligonucleotide therapeutics use short interfering RNA (siRNA) or antisense oligonucleotide (ASO) molecules to exploit endogenous systems—neutralizing target RNA to prevent subsequent protein translation. While the potential clinical application is vast, delivery efficiency and extrahepatic targeting is challenging. Bioanalytical assays are important in building understanding of these complex relationships. The literature currently lacks description of robust and sensitive methods to measure siRNA and ASOs in complex biological matrices. Described herein is a non-enzymatic hybridization-based immunoassay that enables quantification of individual siRNA strands (antisense or sense) in serum, urine, bile, and liver and kidney homogenates. Assay utility is also demonstrated in ASOs. The assay improves upon previous works by abolishing enzymatic steps and further incorporating Locked Nucleic Acid (LNA) nucleotide modifications to increase analyte hybridization affinity and improve sensitivity, specificity, and robustness. We report an assay with an ultrasensitive dynamic range of 0.3 to 16,700 pM for siRNA in serum. The assay was submitted to full qualification for accuracy and precision in both serum and tissue matrices and assay performance was assessed with single and mixed analytes. The reliable LNA-hybridization-based approach removes the need for matrix sample extraction, enrichment or amplification steps which may be impeded by more advanced chemical modifications.
format	article
author	Mai B. Thayer Sara C. Humphreys Kyu S. Chung Julie M. Lade Kevin D. Cook Brooke M. Rock
author_facet	Mai B. Thayer Sara C. Humphreys Kyu S. Chung Julie M. Lade Kevin D. Cook Brooke M. Rock
author_sort	Mai B. Thayer
title	POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
title_short	POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
title_full	POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
title_fullStr	POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
title_full_unstemmed	POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
title_sort	poe immunoassay: plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices
publisher	Nature Portfolio
publishDate	2020
url	https://doaj.org/article/22cff2a3388640828a7f309b40b4cccd
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POE Immunoassay: Plate-based oligonucleotide electro-chemiluminescent immunoassay for the quantification of nucleic acids in biological matrices

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