Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes

Abstract Reliable ammonia quantification assays are essential for monitoring ammonemia in patients with liver diseases. In this study, we describe the development process of a microplate-based assay for accurate, precise, and robust ammonia quantification in biological fluids, following regulatory g...

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Autores principales:	Anastasia Spyrogianni, Charlotte Gourmel, Leopold Hofmann, Jessica Marbach, Jean-Christophe Leroux
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/b78edb55964b4e9caae71087417608c5
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spelling	oai:doaj.org-article:b78edb55964b4e9caae71087417608c52021-11-14T12:20:23ZOptimization of an ammonia assay based on transmembrane pH-gradient polymersomes10.1038/s41598-021-01137-12045-2322https://doaj.org/article/b78edb55964b4e9caae71087417608c52021-11-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-01137-1https://doaj.org/toc/2045-2322Abstract Reliable ammonia quantification assays are essential for monitoring ammonemia in patients with liver diseases. In this study, we describe the development process of a microplate-based assay for accurate, precise, and robust ammonia quantification in biological fluids, following regulatory guidelines on bioanalytical method validation. The assay is based on transmembrane pH-gradient polymersomes that encapsulate a pH-sensitive ratiometric fluorophore, the fluorescence signal of which correlates with the ammonia concentration in the sample. Using a four-parameter logistic regression, the assay had a large quantification range (30–800 μM ammonia). As for selectivity, the presence of amino acids or pyruvate (up to clinically relevant concentrations) showed no assay interference. In samples with low bilirubin levels, polymersomes containing the fluorophore pyranine provided accurate ammonia quantification. In samples with high bilirubin concentrations, billirubin’s optical interference was alleviated when replacing pyranine with a close to near-infrared hemicyanine fluorophore. Finally, the assay could correctly retrieve the ammonia concentration in ammonia-spiked human plasma samples, which was confirmed by comparing our measurements with the data obtained using a commercially available point-of-care device for ammonia.Anastasia SpyrogianniCharlotte GourmelLeopold HofmannJessica MarbachJean-Christophe LerouxNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-15 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Anastasia Spyrogianni Charlotte Gourmel Leopold Hofmann Jessica Marbach Jean-Christophe Leroux Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes
description	Abstract Reliable ammonia quantification assays are essential for monitoring ammonemia in patients with liver diseases. In this study, we describe the development process of a microplate-based assay for accurate, precise, and robust ammonia quantification in biological fluids, following regulatory guidelines on bioanalytical method validation. The assay is based on transmembrane pH-gradient polymersomes that encapsulate a pH-sensitive ratiometric fluorophore, the fluorescence signal of which correlates with the ammonia concentration in the sample. Using a four-parameter logistic regression, the assay had a large quantification range (30–800 μM ammonia). As for selectivity, the presence of amino acids or pyruvate (up to clinically relevant concentrations) showed no assay interference. In samples with low bilirubin levels, polymersomes containing the fluorophore pyranine provided accurate ammonia quantification. In samples with high bilirubin concentrations, billirubin’s optical interference was alleviated when replacing pyranine with a close to near-infrared hemicyanine fluorophore. Finally, the assay could correctly retrieve the ammonia concentration in ammonia-spiked human plasma samples, which was confirmed by comparing our measurements with the data obtained using a commercially available point-of-care device for ammonia.
format	article
author	Anastasia Spyrogianni Charlotte Gourmel Leopold Hofmann Jessica Marbach Jean-Christophe Leroux
author_facet	Anastasia Spyrogianni Charlotte Gourmel Leopold Hofmann Jessica Marbach Jean-Christophe Leroux
author_sort	Anastasia Spyrogianni
title	Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes
title_short	Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes
title_full	Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes
title_fullStr	Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes
title_full_unstemmed	Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes
title_sort	optimization of an ammonia assay based on transmembrane ph-gradient polymersomes
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/b78edb55964b4e9caae71087417608c5
work_keys_str_mv	AT anastasiaspyrogianni optimizationofanammoniaassaybasedontransmembranephgradientpolymersomes AT charlottegourmel optimizationofanammoniaassaybasedontransmembranephgradientpolymersomes AT leopoldhofmann optimizationofanammoniaassaybasedontransmembranephgradientpolymersomes AT jessicamarbach optimizationofanammoniaassaybasedontransmembranephgradientpolymersomes AT jeanchristopheleroux optimizationofanammoniaassaybasedontransmembranephgradientpolymersomes
_version_	1718429309396320256

Optimization of an ammonia assay based on transmembrane pH-gradient polymersomes

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