METHOD

• US 20100297683A1
• Filed: 10/16/2007
• Published: 11/25/2010
• Est. Priority Date: 10/16/2006
• Status: Active Grant

• Current Assignee: Queen Mary And Westfield College
• Original Assignee: Queen Mary And Westfield College
• Inventors: Kamarun, Dzaraini, Watkinson, Michael, Stair, Jadqueline, Krause, Steffi
• Granted Patent: US 8,278,064 B2
• Time in Patent Office: Days
• US Class Current: 435/23
• CPC Class Codes: C12Q 1/001 Enzyme electrodes
• CPC Class Codes: C12Q 1/37 involving peptidase or proteinase
• CPC Class Codes: G01N 33/545 Synthetic resin

Abstract

The present invention provides a method for the detection of a protease comprising the use of a substrate partially coated with a film of a synthetic polymeric matrix comprising a peptide of up to 20 amino acids. Apparatus for performing such methods is also disclosed.

Claims

1. A method for the detection of a protease, comprising:

(i) contacting a sample to be assayed with a detection means comprising a substrate partially coated with a film of a synthetic polymeric matrix; and

(ii) measuring a signal output of said detection means;

wherein the synthetic polymeric matrix comprises a peptide comprising up to 20 amino acids.

2. A method as claimed in claim 1, in which the synthetic polymeric matrix is cross-linked with a peptide comprising up to 20 amino acids.

3. A method as claimed in claim 1, in which the peptide is integrated into the backbone of the synthetic polymeric matrix.

4. A method as claimed in claim 1, in which the peptide is used as a polymerisable monomer to synthesise the polymeric matrix.

5. A method as claimed in claim 1, in which the synthetic polymeric matrix is a hydrogel.

6. A method as claimed in claim 1, in which the substrate is a metal coated piezoelectric material, metallised glass, a plastic material printed with at least one carbon electrode, an electrode, a capacitor, or a transducer.

7. A method as claimed in claim 1, in which the detection means is a piezoelectric quartz crystal microbalance.

8. A method as claimed in claim 1, in which the detection means is a metallised glass slide.

9. A method as claimed in claim 1, in which the detection means is an electrode.

10. A method as claimed in claim 1, in which the synthetic polymeric matrix comprises a polymer selected from:

polyvinylpyrrolidone, acrylamide, polymers formed from acrylic and methacrylic monomers, polyethylene glycol, polyethylene glycol acrylate, ethylene glycol methyl ether acrylate, dendrimers, poly(ethylene oxide) (PEO) and polyvinyl alcohol.

11. A method as claimed in claim 1, in which the synthetic polymeric matrix comprises a polymer which is oxidised dextran.

12. A method as claimed in claim 1 in which the protease is a serine protease, a matrix metalloproteinase or a gingipain protease.

13. A method as claimed in claim 12, in which the serine protease is elastase, trypsin, chymotrypsin, subtilisin, cathepsin G or MMP8.

14. A method as claimed in claim 13, in which the serine protease is elastase.

15. A method as claimed in claim 13, in which the serine protease is cathepsin G.

16. A method as claimed in claim 12, in which the peptide comprises the sequence:

17. A method as claimed in claim 14, in which the peptide comprises the sequence:

18. A method as claimed in claim 15, in which the peptide comprises the sequence:

Ala-Ala-Pro-Phe (AAPF) (SEQ ID NO;

6), or Ala-Ala-Pro-Phe-Phe-Lys (AAPFFK) (SEQ ID NO;

7).

19-23. -23. (canceled)

24. Apparatus for detection of a protease comprising a detection means comprising a substrate partially coated with a film of a synthetic polymeric matrix, wherein said synthetic polymeric matrix comprises a peptide comprising up to 20 amino acids.

25-29. -29. (canceled)