COMPETITIVE BIOSENSOR HAVING ELEVATED SENSITIVITY

• US 20130337468A1
• Filed: 12/13/2011
• Published: 12/19/2013
• Est. Priority Date: 12/17/2010
• Status: Active Grant

• Current Assignee: EyeSense AG
• Original Assignee: EyeSense AG
• Inventors: Mller, Achim, Nikolaus, Katharina, Herberchtsmeier, Peter, Knuth, Monika
• Granted Patent: US 10,429,380 B2
• Time in Patent Office: Days
• US Class Current: 435/7.1
• CPC Class Codes: G01N 33/542 with steric inhibition or signal modification, e.g. fluorescent quenching
• CPC Class Codes: G01N 33/5436 with ligand physically entrapped within the solid phase liposomes G01N33/5432; immunological test elements G01N33/54386
• CPC Class Codes: G01N 33/66 involving blood sugars, e.g. galactose

Abstract

The present invention relates to measures for determining glucose and for diagnosing diseases based on impaired glucose metabolism. In particular the present invention relates to a device comprising a hydrogel having a glucose-binding protein and a ligand of the glucose-binding protein incorporated therein, wherein the hydrogel comprises a first hydrogel matrix made of alginate and a second hydrogel matrix which forms an interpenetrating network within the first hydrogel matrix. The invention further relates to the use of such a device for determining the glucose content in a sample and to the use of the device for diagnosing impaired glucose metabolism in a test subject.

Claims

1-17. -17. (canceled)

18. A device comprising a hydrogel having a glucose-binding protein incorporated therein and a ligand of the glucose-binding protein, wherein the hydrogel comprises:

(a) a first hydrogel matrix made of alginate; and

(b) a second hydrogel matrix which forms an interpenetrating network within the first hydrogel matrix.

19. The device of claim 18, wherein the second hydrogel matrix comprises a water-soluble polymer with at least one cross-linkable group per molecule and a molecular weight of at most 500,000.

20. The device of claim 18, wherein the second hydrogel matrix is selected from the group consisting of polyvinyl alcohols (PVAs;

including copolymers with vinyl acetates and/or ethylene), polyethylene glycols (PEGs), and polyhydroxyalkanoates (PHAs), poly (2-methyl-2 oxazolines), poly (2-ethyl-2 oxazolines), poly (2-hydroxyethyl-2 oxazolines), poly (2-(1-(hydroxymethyl)-ethyl)-2 oxazolines), poly-(hydroxyethyl methacrylate) (PHEMA), poly-(hydroxyethyl acrylate) (PHEA), poly-vinylpyrolidones, poly-(dimethyl)acrylamide, poly-(hydroxyethyl)acrylamide, poly-(ethylene-co-vinyl alcohol), poly(vinyl acetate-co-vinyl alcohol), poly(ethylene-co-vinyl acetate-co-vinyl alcohol), and poly(ethylene glycol-co-propylene glycol).

21. The device of claim 18, wherein the first hydrogen matrix, the second hydrogel matrix, or a combination thereof is capable of interacting with the glucose-binding protein.

22. The device of claim 18, wherein the glucose-binding protein is selected from the group consisting of lectins, enzymes which bind glucose as substrate, antibodies which specifically recognize glucose, and aptamers which specifically recognize glucose.

23. The device of claim 18, wherein the ligand of the glucose-binding protein is an oligosaccharide, a glycosylated macromolecule, or a glycosylated nanoparticle.

24. The device of claim 18, wherein the glucose-binding protein is concanavalin A.

25. The device of claim 24, wherein the concanavalin A concentration is greater than about 0.5 mg/(g matrix).

26. The device of claim 25, wherein the concanavalin A concentration lies between about 2 and about 60 mg/(g matrix).

27. The device of claim 24, wherein the concanavalin A exhibits increased water solubility as compared to unmodified concanavalin A owing to chemical modification.

28. The device of claim 27, wherein the modification is selected from the group consisting of pegylation, acetylation, succinylation, and polyoxazolinylation.

29. The device of claim 18, wherein the glucose-binding protein is linked to an oxazine dye and the ligand of the glucose-binding protein to a rhodamine dye.

30. A method for determining the quantity of glucose in a sample comprising:

(a) contacting the device of claim 18 with a sample for a period and under conditions which enables the binding of the glucose present in the sample to the glucose-binding protein in the device; and

(b) determining the quantity of ligand displaced from the glucose in the device, whereby the quantity of glucose is determined in the sample.

31. The method of claim 30, wherein the sample has been obtained from a test subject which exhibits impaired glucose metabolism or which presumably exhibits impaired glucose metabolism.

32. The method of claim 31, wherein the impaired glucose metabolism is caused by diabetes mellitus or metabolic syndrome.

33. A method for diagnosing impaired glucose metabolism in a test subject, comprising:

(a) contacting the device of claim 18 with a sample from a test subject;

(b) determining the quantity of glucose in the sample of step (a); and

(c) comparing the quantity of glucose obtained in step (b) to reference quantities of glucose which are indicative for impaired glucose metabolism, thereby diagnosing impaired glucose metabolism in the test subject.

34. The method of claim 33, wherein the impaired glucose metabolism is caused by diabetes mellitus or metabolic syndrome.

35. A method for determining the need for a therapeutic measure in a test subject with impaired glucose metabolism, comprising:

(a) method steps (a), (b), and (c) of claim 33; and

(b) determining the need for a therapeutic measure in the test subject with impaired glucose metabolism.