Device and methods of using device for detection of hyperammonemia

• US 10,591,495 B2
• Filed: 04/27/2016
• Issued: 03/17/2020
• Est. Priority Date: 04/27/2015
• Status: Active Grant

• Current Assignee: Children's National Medical Center, University of Maryland, College Park
• Original Assignee: University of Maryland, College Park
• Inventors: Ayyub, Omar Bilal, Behrens, Adam Michael, Kofinas, Peter, Summar, II, Marshall Lynn, Cabrera-Luque, Juan Manuel, Cunningham, Gary
• Primary Examiner(s): White, Dennis
• Application Number:
  US15/570,269
• Publication Number:
  US 20180143210A1
• Time in Patent Office: 1,420 Days
• CPC Class Codes: A61K 31/216 of acids having aromatic rings, e.g. benactizyne, clofibrate
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: B01L 2200/16 Reagents, handling or storing thereof
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: G01N 2021/775 Indicator and selective membrane
• CPC Class Codes: G01N 2021/7759 Dipstick; Test strip
• CPC Class Codes: G01N 21/78 producing a change of colour
• CPC Class Codes: G01N 2800/04 Endocrine or metabolic disorders
• CPC Class Codes: G01N 33/6806 Determination of free amino acids
• CPC Class Codes: G01N 33/84 involving inorganic compounds or pH

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 perflurinated 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.

Claims

1. A biosensor comprising:

at least a first and second vessel;

a fluid exchange opening positioned between the first and the second vessel;

a membrane positioned across or over the fluid exchange opening; and

a catalyst in solid phase within the second vessel or within a first conduit in fluid communication with the second vessel, wherein the catalyst is nitroprusside or salt thereof with a mass of from about 5.5 micrograms to about 7.8 micrograms;

wherein the membrane comprises an ionomer.

2. The biosensor of claim 1 further comprising a compound comprising a phenyl group in solid phase within the second vessel or within a conduit in fluid communication with the second vessel.

3. The biosensor of claim 2, wherein the compound comprising a phenyl group has a mass of from about 75 micrograms to about 85 micrograms.

4. The biosensor of claim 1, wherein at least the first conduit is in fluid communication with at least the first vessel, the first conduit configured to receive a fluid from a point external to the biosensor.

5. The biosensor of claim 1, wherein the first vessel or the second vessel:

(i) comprises one or a combination of a hypohalite, an alkali buffer, and at least one;

phenolic reagent or indophenol related compound;

or (ii) are in fluid communication with a second conduit connected to a storage vessel that comprises a fluid comprising individually or in combination;

a hypohalite, an alkali buffer, and at least one;

phenolic reagent or indophenol related compound.

6. The biosensor of claim 5, wherein the first vessel or the second vessel are in fluid communication with at least one reagent conduit configured to receive a fluid from a reagent storage vessel at a point distal from the first or second vessel.

7. The biosensor of claim 6, wherein the first vessel or the second vessel is in fluid communication with:

(i) a first reagent conduit configured to receive a fluid from a first reagent storage vessel comprising a compound comprising a phenyl group in liquid phase;

(ii) a second reagent conduit configured to receive a fluid from a second reagent storage vessel comprising a hypohalite in liquid phase; and

(iii) a third reagent conduit configured to receive a fluid from a third reagent storage vessel comprising an alkali buffer in liquid phase.

8. The biosensor of claim 6, wherein at least one reagent storage vessel is a blister pack comprising a seal covering the blister pack on a portion of that blister pack adjacent to at least one reagent conduit, wherein the seal is configured for mechanical rupture, such that, upon rupture, a fluid circuit is formed between the first vessel or the second vessel and the reagent storage vessel and fluid contents from the blister pack are capable of fluid flow to the first vessel or the second vessel.

9. The biosensor of claim 1, wherein either the first or the second vessels individually comprise a hypohalite, an alkali buffer, and the catalyst;

wherein the first and second vessel are in fluid communication with a detection vessel by at least one reaction conduit, wherein the reaction conduit comprises a first portion positioned proximate to the second vessel, said first portion comprising the catalyst in solid phase, and wherein said first portion comprises the compound comprising a phenyl group in solid phase.

10. The biosensor of claim 9, wherein the alkali buffer comprises from about 0.1 M to about 5 M sodium acetate or sodium chloride.

11. The biosensor of claim 1, wherein the biosensor is free of one or more of the following:

(i) uricase or a functional fragment thereof;

(ii) a hydrogel comprising dextran or a derivative thereof;

(iii) a bacterial cell;

(iv) an electronic dipole configured for electrophoresis;

(v) 3, 4-DHB, (vi) a vaporizer, gas chromatograph, or a heating element configured for converting liquid ammonia into a gaseous state; and

(vii) a membrane comprising cellulose or a derivative thereof.

12. The biosensor of claim 1, wherein the first vessel has a volume from about 5 μ

l to about 100 μ

l, and, optionally, the second vessel has a volume from about 5 μ

l to about 100 μ

l.

13. The biosensor of claim 1 further comprising:

(i) a circuit comprising the at least one wire, and a digital display operably connected to a processor configured to receive digital information from the spectrophotometer and to send digital information to the digital display; and

(ii) a test strip comprising the first conduit configured for receiving a volume of bodily fluid.

14. The biosensor of claim 1, further comprising at least one indophenol reagent or indophenol related compound selected from:

phenol, 2-phenylphenol, or napthol;

wherein the alkali buffer is about 1.0 M sodium chloride, calcium chloride, zinc chloride, sodium acetate, calcium acetate, or zinc acetate; and

wherein the membrane comprises Nafion.

15. The biosensor of claim 1, capable of being stored at about room temperature or frozen for no less than 50, 100, 150, 200, or 245 days.

16. The biosensor of claim 1, wherein the membrane comprises Nafion which has not been pre-treated with an acid solution and/or a hydrogen peroxide solution.

17. A system comprising:

(i) the biosensor of claim 1 in operable connection to at least one computer storage memory;

(ii) a computer processor in operable connection with the at least one computer storage memory;

wherein the computer processor is in operable connection with at least one light emitting diode (LED), amplification circuit, battery, and stepper motor.

18. A method of diagnosing a metabolic disease in a subject comprising:

(a) contacting a sample of bodily fluid to the biosensor of claim 1;

(b) quantifying one or more concentration values of ammonia in the sample;

(c) comparing the one or more concentration values of ammonia in the sample to a threshold value of ammonia concentration identified as being in a healthy range; and

(d) identifying the subject as having a metabolic disease if the one or more concentration values of ammonia in the sample exceed or fall below the threshold value.

19. The method of claim 18, wherein the metabolic disease is hyperammonemia.

20. A method of treating a metabolic disease comprising:

(a) contacting a sample of bodily fluid to the biosensor of claim 1;

(b) quantifying one or more concentration values of ammonia in the sample;

(c) comparing the one or more concentration values of ammonia in the sample to a threshold value of ammonia concentration identified as being in a healthy range; and

(d) identifying the subject as having a metabolic disease if the one or more concentration values of ammonia in the sample exceed or fall below the threshold value;

(e) administering a therapeutically effective amount of a therapeutic agent to treat the metabolic disease.

21. The method of claim 20, wherein the metabolic disease is hyperammonemia.

28. A method of quantifying a concentration of ammonia or ammonium ion in a sample of bodily fluid comprising contacting the sample to the biosensor of claim 1.

29. The method of claim 28, wherein the step of contacting comprises exposing the sample to the biosensor of claim 1 for a period of time sufficient to allow modification of an indophenol reagent or indophenol related compound to an indophenol or indophenol reaction product.

30. The method of claim 28, wherein the method does not comprise exposing the sample of bodily fluid to any external stimuli or reagent prior to contacting the sample to the biosensor.

31. The method of claim 28, wherein the sample of bodily fluid is whole blood from a subject.

22. A biosensor comprising:

at least a first and second vessel configured for receiving a sample from a point exterior to the biosensor;

a fluid exchange opening positioned between the first and the second vessel;

a membrane positioned across or over the fluid exchange opening; and

a catalyst in liquid phase or solid phase a hypohalite in liquid phase;

an alkali buffer in liquid phase;

a phenolic reagent in liquid phase or solid phase; and

a fluid circuit comprising, in fluid communication;

the first and second vessel;

a reagent conduit; and

a detection vessel positioned distal from the first and second vessel;

wherein the catalyst is sodium nitroprusside or a salt thereof, and, if the catalyst is in solid phase, the biosensor comprises from about 5.8 to about 7.3 micrograms of sodium nitroprusside or a salt thereof in solid phase; and

, if the catalyst is in liquid phase, the sodium nitroprusside or a salt thereof is at a concentration greater than 8.4 μ

M.

23. The biosensor of claim 22, wherein the catalyst is sodium nitroprusside in liquid phase at a concentration from about 1.0 mM to about 2.0 mM.

24. The biosensor of claim 22 comprising:

a first operable condition, in which the first vessel comprises a sample;

a second operable condition, in which the first vessel comprises a sample and an aqueous buffer;

a third operable condition, in which the reagent conduit comprises a mixture of the aqueous buffer, the alkali buffer, the hypohalite, the catalyst and the phenolic reagent and the mixture moves toward the detection vessel and the catalyst mixes with the sample for a time period sufficient for the catalyst to catalyze an indophenol reaction; and

a fourth operable condition, in which the detection vessel comprises the mixture and a light source positioned proximate to the detection vessel capable of emitting light through the detection vessel sufficient to quantify the amount of ammonia in the sample;

wherein, in the fourth operable condition, the light source is a photodiode capable of emitting light in the detection vessel and producing an amount of light sufficient to enable absorbance readings detected at a wavelength of light from about 600 to about 650 nm.

25. The biosensor of claim 22, wherein, if the sodium nitroprusside or salt thereof is in liquid phase, the sodium nitroprusside or salt thereof is at a concentration greater than 16.6 μ

M.

26. The biosensor of claim 22, wherein, if the sodium nitroprusside or salt thereof is in liquid phase, the sodium nitroprusside or salt thereof is at a concentration from 665 to 1996 μ

M.

27. A kit comprising a solid support that comprises:

at least a first and second vessel;

a fluid exchange opening positioned between the first and the second vessel;

at least a first conduit in fluid communication with at least the first vessel, the first conduit configured to receive a fluid from a point external to the biosensor; and

a membrane positioned at the fluid exchange opening;

a catalyst in liquid phase or solid phase;

a hypohalite in liquid phase;

an alkali buffer in liquid phase;

a phenolic reagent in liquid phase or solid phase; and

a fluid circuit comprising, in fluid communication;

the first and second vessel;

a reagent conduit; and

a detection vessel positioned distal from the first and second vessel;

wherein the catalyst is sodium nitroprusside, and, if the catalyst is in solid phase, the biosensor comprises from about 5.8 to about 7.3 micrograms of sodium nitroprusside or a salt thereof in solid phase; and

, if the catalyst is in liquid phase, the sodium nitroprusside or a salt thereof is at a concentration greater than 8.4 μ

M; and

wherein the membrane comprises an ionomer.