Device and methods of using device for detection of hyperammonemia
Inventors
Ayyub, Omar Bilal • Behrens, Adam Michael • Kofinas, Peter • Summar, Marshall Lynn • Cabrera-Luque, Juan Manuel • Cunningham, Gary • Simeonov, Anton • Marugan, Juan
Assignees
Childrens National Medical Center Inc • University of Maryland College Park • US Department of Health and Human Services
Publication Number
US-10620187-B2
Publication Date
2020-04-14
Expiration Date
2034-09-02
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Abstract
The present disclosure relates to a biosensor capable of measuring the total concentration of one or a plurality of ammonia or ammonium ions with the use of indophenol reagents in the presence of an ionomer. In some embodiments, the biosensor comprises a perfluorinated membrane that comprises an ionomer in contact with an alkali buffer in a vessel configured to receive a sample, such as whole blood. The disclosure also relates to a method of detecting or quantifying the ammonia or ammonium ion concentration in whole blood in a point of care bio sensor without reliance on gas chromatography or any measurement that takes more than about twenty minutes.
Core Innovation
The invention relates to a biosensor and methods for detecting or quantifying ammonia or ammonium ion concentration in a sample of bodily fluid, such as whole blood, using reagents for the indophenol or Berthelot reaction. The biosensor comprises at least a first vessel containing an alkali buffer and a second vessel containing phenolic reagents configured for the indophenol reaction, separated by a membrane comprising an ionomer that allows ammonia to pass between vessels. The reaction produces a color change detectable by photodiodes or spectrophotometers to determine ammonia levels without reliance on gas chromatography or extended measurement times.
The problem addressed is the lack of rapid, point-of-care testing devices for hyperammonemia, a potentially fatal condition associated with diseases like liver cirrhosis and urea cycle disorders. Conventional tests rely on centralized laboratory methods such as fluorometry and tandem mass spectroscopy, which require cumbersome and expensive equipment and can take multiple days. This delay impedes timely diagnosis and treatment, particularly critical in neonatal infants and in managing liver conditions.
The biosensor system innovatively utilizes a cation exchange membrane, such as Nafion, to separate the sample and reagents while allowing selective passage of ammonia. This enables rapid diffusion and isolation of ammonia from complex matrices like whole blood, minimizing interference from proteins and amino acids. The system includes methods for quantitative colorimetric analysis of the indophenol reaction product, allowing determination of ammonia concentration within about twenty minutes. Further embodiments incorporate electrodes and metabolic enzymes immobilized in hydrogels to measure amino acid concentrations electrochemically, broadening diagnostic capabilities to aminoacidopathies.
Claims Coverage
The patent includes multiple independent claims defining methods and devices for quantifying ammonia or ammonium ion concentration in bodily fluid samples using a biosensor with specific structural and functional features.
Biosensor design featuring a cationically selective membrane for ammonia detection
A biosensor comprising at least first and second vessels separated by a fluid exchange opening, with a membrane comprising an ionomer positioned at the opening, allowing ammonia transport. The vessels include reagents such as hypohalite, alkali buffer, and phenolic reagents for the indophenol reaction. The biosensor accepts fluid via a conduit from an external sample source and holds sample volumes from about 5 μL to about 100 μL.
Ammonia quantification method without pre-treatment of bodily fluid samples
A method of quantifying ammonia concentration in bodily fluid by contacting the fluid with the biosensor described above without exposing the fluid to any external stimuli or reagent prior to contact, enabling direct analysis of samples such as whole blood.
Comparative analysis of ammonia concentration to metabolic disease thresholds
Methods including comparing measured ammonia concentration values obtained from the biosensor to predetermined threshold values associated with metabolic diseases such as hyperammonemia to facilitate diagnosis.
The independent claims cover a biosensor system with vessels separated by an ionomer membrane for passage of ammonia, methods for direct sample contact without pre-treatment, and diagnostic methods comparing ammonia concentrations to threshold values indicative of metabolic diseases.
Stated Advantages
Enables rapid detection of ammonia or ammonium ion concentrations in whole blood or bodily fluids at the point of care, with results within approximately twenty minutes.
Does not require complex, cumbersome, or expensive laboratory equipment such as gas chromatography, fluorometry, or tandem mass spectroscopy.
Provides stable reagent formulations that avoid reliance on enzymatic components prone to degradation, thereby allowing extended shelf life and ease of storage.
Incorporates membrane technology to effectively isolate ammonia from interfering biological molecules like proteins and amino acids, enhancing assay specificity and accuracy.
Can be modified inexpensively to detect amino acid concentrations for diagnosing and monitoring aminoacidopathies and related metabolic diseases.
Documented Applications
Point-of-care diagnosis and monitoring of hyperammonemia in subjects by detecting ammonia levels in whole blood.
Detection and quantification of amino acids in bodily fluids for diagnosing aminoacidopathies.
Environmental and civil engineering applications, such as monitoring ammonia levels in wastewater treatment plants.
Home-based testing for ammonia concentrations in aquariums, pipes, or similar environments.
Real-time measurement of ammonia or ammonium ion and metabolites in blood of patients to guide clinical management.
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