Diagnostic assay for fructosamines

• US 5,370,990 A
• Filed: 07/27/1992
• Issued: 12/06/1994
• Est. Priority Date: 07/29/1991
• Status: Expired due to Term

• Current Assignee: Genzyme Ltd (Sanofi )
• Original Assignee: Genzyme Corporation (Sanofi )
• Inventors: Power, John A., Lovelady, John A., Staniford, Julie M.
• Primary Examiner(s): Wityshyn, Michael G.
• Assistant Examiner(s): Leary, Louise N.
• Application Number:
  US07/919,434
• Time in Patent Office: 862 Days
• Field of Search: 435/4, 435/240.27, 435/249, 435/255, 435/252, 436/88, 436/87
• US Class Current: 435/4
• CPC Class Codes: C12Q 1/26 involving oxidoreductase
• CPC Class Codes: C12Q 1/37 involving peptidase or proteinase

Abstract

A method for the determination of glycated protein in a sample characterised in that it comprises treating the sample with a protease and treating the protease-treated sample with a ketoamine oxidase, a product of this reaction being measured is disclosed.

Claims

1. A method for the determination of a glycated protein in a sample comprising the steps of:

a) treating the sample with a protease;

b) treating the protease-treated sample with a ketoamine oxidase obtained from a microorganism selected from the group consisting of Klebsiella, Fusarium, Acremonium, and Debraryomyces; and

c) measuring the hydrogen peroxide product of the reaction in step (b).

2. The method of claim 1 wherein the ketoamine oxidase is obtained from Debaryomyces vanrijiae var. vanrijiae.

3. The method of claim 1 wherein the protease is selected from the group consisting of proteinase K, pronase E, ananain, thermolysin, subtilisin and bovine pancreatic proteases.

4. A method of claim 1 wherein the protease treatment is performed in the presence of a detergent.

5. The method of claim 4 wherein the detergent is selected from the group consisting of SDS, Brij 35 and Tween 20.

6. A kit for the determination of glycated protein which comprises a protease and a ketoamine oxidase, said ketoamine oxidase being obtained from a microorganism selected from the group consisting of Klebsiella, Fusarium, Acremonium, and Debraryomyces.

7. A ketoamine oxidase which catalyses the oxidation of the carbon atom in position 1 of a sugar moiety of a glycated protein with consequent hydrolytic disruption of an amine bond to release a sugar osone and hydrogen peroxide from an amino acid, said ketoamine oxidase being obtained from a microorganism selected from the group consisting of Klebsiella, Fusarium, Acremonium, and Debraryomyces.

8. The ketoamine oxidase of claim 7 which is obtained by culturing the microorganism in the presence of a model substrate.

9. A process for producing a ketoamine oxidase comprising the steps of:

a) preparing a cell culture medium containing a microorganism selected from the group consisting of Klebsiella, Fusarium, Acremonium, and Debraryomyces, said microorganism being capable of producing ketoamine oxidase;

b) adding fructosyl valine or butylamino deoxy fructose to the culture medium as the sole nitrogen source for said microorganism, and;

c) obtaining ketoamine oxidase from said culture medium.