Salivary protein glycosylation test for diagnosis and monitoring of diabetes

• US 9,383,352 B2
• Filed: 03/26/2014
• Issued: 07/05/2016
• Est. Priority Date: 03/26/2014
• Status: Active Grant

• Current Assignee: DiabetOmics, Inc.
• Original Assignee: DiabetOmics, Inc.
• Inventors: Nagalla, Srinivasa R., Bean, Eric S.
• Primary Examiner(s): Wallenhorst, Maureen
• Application Number:
  US14/226,131
• Publication Number:
  US 20150276723A1
• Time in Patent Office: 832 Days
• Field of Search: 436/63, 436/87, 436/94, 436/95, 436/124, 436/125, 436/127, 436/128, 436/164, 436/166, 436/169, 436/170, 435/14, 435/287.7, 435/287.8, 435/288.7, 422/400, 422/420, 422/421, 422/82.05, 422/430
• US Class Current: 1/1
• CPC Class Codes: G01N 21/78 producing a change of colour
• CPC Class Codes: G01N 2800/042 Disorders of carbohydrate metabolism, e.g. diabetes, glucose metabolism
• CPC Class Codes: G01N 2800/56 Staging of a disease; Further complications associated with the disease
• CPC Class Codes: G01N 33/525 Multi-layer analytical elements
• CPC Class Codes: G01N 33/66 involving blood sugars, e.g. galactose
• CPC Class Codes: G01N 33/6872 Intracellular protein regulatory factors and their receptors, e.g. including ion channels
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]
• CPC Class Codes: Y10T 436/144444 Glucose
• CPC Class Codes: Y10T 436/200833 Carbonyl, ether, aldehyde or ketone containing

Abstract

Disclosed herein are methods and tests for diagnosing and/or monitoring a metabolic condition such as diabetes in a subject, wherein the methods and tests measure salivary glycoproteins. Some of the methods are based on the oxidation of glycoproteins in a sample from the subject, such as saliva or urine, for example using sodium metaperiodate, and then detecting the aldehydes generated during oxidation using a chemical detection method. Also disclosed are kits and lateral flow devices for detecting glycoproteins in a saliva sample.

Claims

1. A method of determining a glycemic state in a subject, the method comprising:

chemically oxidizing a saliva sample from the subject, thereby oxidizing one or more glycoproteins in the saliva sample to generate one or more oxidation products, wherein the one or more oxidation products comprise one or more aldehydes;

quantifying the one or more aldehydes in the oxidized saliva sample; and

comparing a level of the one or more aldehydes to a normal control value, wherein an increase in the level of the one or more aldehydes indicates that the subject has an elevated glycemic state.

2. The method of claim 1, wherein determining the glycemic state in the subject comprises determining an average blood glucose value for the subject.

3. The method of claim 2, wherein the average blood glucose value is a three-week average blood glucose value.

4. The method of claim 1, wherein the method is a method of diagnosing diabetes in the subject, and wherein the subject is determined to have diabetes if the level of the one or more aldehydes exceeds the normal control value.

5. The method of claim 2, wherein the method is a method of monitoring glycemic control in the subject.

6. The method of claim 1, wherein oxidizing the saliva sample comprises reacting the saliva sample with sodium metaperiodate.

7. The method of claim 1, wherein oxidizing the saliva sample comprises applying the saliva sample to a sodium metaperiodate membrane.

8. The method of claim 1, wherein quantifying aldehydes in the oxidized saliva sample comprises using a chemical detection method.

9. The method of claim 1, wherein the one or more glycoproteins comprise sialic acid and fucose.

10. A lateral flow device for detecting glycoproteins in a saliva sample, the lateral flow device comprising:

a first membrane comprising sodium metaperiodate;

a second membrane in operable contact with the first membrane, the second membrane comprising NaOH;

a third membrane in operable contact with the second membrane, the third membrane comprising immobilized 4-amino-3-hydrazino-5-mercapto-1,2,4-triazole (AHMT); and

a mesh layer disposed between the second membrane and the third membrane.

11. The lateral flow device of claim 10, wherein the first membrane oxidizes sialic acid and fucose in the saliva sample.

12. The lateral flow device of claim 11, wherein the third membrane detects aldehydes in the oxidized saliva sample.

13. The lateral flow device of claim 12, wherein the third membrane produces a colorimetric result corresponding to a quantity of aldehydes in the oxidized saliva sample.

14. A kit for detecting glycoproteins in a saliva sample, the kit comprising:

a reagent for oxidizing glycoproteins in a saliva sample;

a test strip for detecting aldehydes in an oxidized saliva sample, the test strip comprising;

a first membrane comprising NaOH;

a second membrane in operable contact with the first membrane, the second membrane comprising immobilized 4-amino-3-hydrazino-5-mercapto-1,2,4-triazole (AHMT);

a mesh layer disposed between the first membrane and the second membrane, and

instructions for using the kit.

15. The kit of claim 14, wherein the reagent for oxidizing glycoproteins in the saliva sample comprises sodium metaperiodate.

16. The kit of claim 14, wherein the reagent for oxidizing glycoproteins in the saliva sample comprises a membrane that comprises sodium metaperiodate, wherein the membrane that comprises sodium metaperiodate is in operable contact with the first membrane.

17. The kit of claim 14, further comprising a colorimeter for reading the test strip.

18. The kit of claim 17, wherein the colorimeter generates a number corresponding to an amount of aldehydes detected by the test strip.

19. The kit of claim 18, wherein the number corresponding to the amount of aldehydes detected by the test strip ranges from 4.0-12.0, and wherein a number higher than 6.0 reflects an elevated average blood glucose value compared to a normal average blood glucose value.