ELECTROCHEMICAL AFFINITY ASSAY

• US 20020137193A1
• Filed: 06/18/2001
• Published: 09/26/2002
• Est. Priority Date: 08/24/1998
• Status: Active Grant

• Current Assignee: Abbott Diabetes Care Incorporated (Abbott Laboratories Incorporated), Marconi Communications Limited (Marconi Holdings Company Limited)
• Original Assignee: Marconi Communications Limited (Marconi Holdings Company Limited)
• Inventors: Heller, Adam, Campbell, Charles N.
• Granted Patent: US 6,576,461 B2
• Time in Patent Office: Days
• US Class Current: 435/287.1
• CPC Class Codes: C12Q 1/002 Electrode membranes
• CPC Class Codes: C12Q 1/005 involving specific analytes or enzymes including groups of enzymes, e.g. oxydases; C12Q1/004 takes precedence
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: Y10S 435/817 Enzyme or microbe electrode
• CPC Class Codes: Y10S 436/806 Electrical property or magnetic property

Abstract

Abstract of Disclosure

An electrochemical affinity assay system for detection of ligand-ligand receptor binding.

Claims

1. 18.A method for detecting a second member of a ligand-ligand receptor pair in a sample;

the method comprising;

contacting the sample with an electrode having immobilized thereon a first member of the ligand-ligand receptor pair and with a third member that binds the second member to form a first member-second member-third member complex, the third member labeled with a detection marker;

wherein the detection marker-labeled first member-second member-third member complex generates an electrical signal when a potential is applied to the electrode; and

correlating the generated electrical signal with the presence or amount of the second member in the sample.

18. 35.The method according to claim 18, wherein contacting further comprises the step of rotating the electrode within the sample.

2. 19.The method according to claim 18, wherein the first member is immobilized in a redox polymer on the electrode.

3. 20.The method according to claim 18, wherein the electrode further comprises an immobilized substrate generating enzyme, the substrate generating enzyme generating a detection compound.

9. 26.The method according to claim 18, wherein the first member is immobilized in a film coating the electrode.

11. 28.The method according to claim 18, wherein the first member is non-covalently bound to a film coating the electrode.

15. 32.The method according to claim 18, wherein the detection marker comprises peroxidase.

17. 34.The method according to claim 18, wherein the potential applied to the electrode is not more reducing than -0.0V versus the saturated calomel electrode (SCE) and not more oxidizing than +0.4V versus the saturated calomel electrode (SCE).

19. 36.The method according to claim 18, wherein the sample comprises blood.

20. 37.A method for detecting a second member of a ligand-ligand receptor pair in a sample;

the method comprising;

contacting the sample with a sensor wherein the sensor comprises;

an electrode;

a redox polymer film on the electrode;

a substrate generating enzyme on the electrode that generates a detection compound;

a first member of the ligand-ligand receptor pair that binds the second member, the first member on the electrode;

and with a third member that binds the second member, the third member labeled with a detection marker that reacts with the detection compound to form a first member-second member-third member complex;

wherein the detection marker-labeled first member-second member-third member complex generates an electrical signal in the presence of the detection compound when a potential is applied to the electrode; and

correlating the generated electrical signal with the presence or amount of the second member in the sample.

4. 21. The method according to claim 20, wherein the substrate generating enzyme is immobilized in the redox polymer.

5. 22.The method according to claim 20, wherein the substrate generating enzyme comprises one or more reaction centers, and wherein the reaction centers do not exchange electrons with the electrode.

7. 24.The method according to claim 20, wherein the substrate generating enzyme comprises choline oxidase.

14. 31.The method according to claim 20, wherein the detection compound comprises hydrogen peroxide.

21. 38.The method according to claim 37, wherein the substrate generating enzyme comprises one or more reaction centers, and wherein the reaction centers do not exchange electrons with the redox polymer.

6. 23.The method according to claim 21, wherein the substrate generating enzyme comprises one or more reaction centers, and wherein the reaction centers do not exchange electrons with the redox polymer.

8. 25.The method according to claim 21, wherein the substrate generating enzyme comprises choline oxidase.

22. 39.The method according to claim 37, wherein the substrate generating enzyme is choline oxidase.

23. 40.The method according to claim 37, wherein the redox polymer film on the electrode further comprises a binding agent that binds the first member.

24. 41.The method according to claim 40, wherein the binding agent covalently binds the first member.

25. 42.The method according to claim 40, wherein the binding agent non-covalently binds the first member.

26. 43.The method according to claim 40, wherein the binding agent is avidin or streptavidin, and the first member is biotinylated.

10. 27. The method according to claim 26, wherein the film comprises a binding agent that binds the first member.

27. 44.The method according to claim 37, wherein the detection compound is hydrogen peroxide.

12. 29. The method according to claim 27, wherein the binding agent covalently binds the first member.

13. 30.The method according to claim 27, wherein the binding agent is avidin or streptavidin, and the first member is biotinylated.

28. 45.The method according to claim 44, wherein the detection marker is peroxidase

46.The method according to claim 45, wherein the peroxidase is horseradish peroxidase or soybean peroxidase.

29. 47.The method according to claim 37, wherein potential applied to the electrode is not more reducing than -0.0V versus the saturated calomel electrode (SCE) and not more oxidizing than +0.4V versus the saturated calomel electrode (SCE).

30. 48.The method according to claim 37, wherein the contacting further comprises rotating the electrode within the sample.

31. 49.The method according to claim 37, wherein the sample comprises blood.

32. 50.The method according to claim 37, wherein the first member of the ligand-ligand receptor pair is an antigen and the second member of the ligand-ligand receptor pair is an antibody.

16. 33.The method according to claim 32, wherein the peroxidase is horseradish peroxidase or soybean peroxidase.

33. 51.The method according to claim 37 wherein the first member of the ligand-ligand receptor pair is a growth factor receptor and the second member of the ligand-ligand receptor pair is a growth factor.

34. 52.The method according to claim 37 wherein the first member of the ligand-ligand receptor pair and the second member of the ligand-ligand receptor pair are complementary strands of nucleic acid.