Method and assay for detection of residues

• US 20080274566A1
• Filed: 07/08/2008
• Published: 11/06/2008
• Est. Priority Date: 12/08/2003
• Status: Active Grant

• Current Assignee: Charm Sciences, Inc.
• Original Assignee: Charm Sciences, Inc.
• Inventors: Douglas, David W., Markovsky, Robert J., Saul, Steven J.
• Granted Patent: US 7,863,057 B2
• Time in Patent Office: Days
• US Class Current: 436/518
• CPC Class Codes: G01N 33/54386 Analytical elements
• CPC Class Codes: G01N 33/54388 based on lateral flow
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 436/828 Protein A

Abstract

Embodiments described herein include methods and assays for detecting an analyte in a sample using a plurality of control zone capture agents. Some embodiments include detection of multiple analytes in a sample utilizing a plurality of analyte binders and a control zone containing multiple control zone capture agents. In some embodiments, the multiple control zone capture agents capture a plurality of binders within one control zone. Test results are determined by comparison of the control zone signal to a test zone signal.

Claims

1. A lateral flow assay for detecting an analyte in a solution, the assay comprising:

(a) a binder configured for generating a detectable signal, wherein the binder can combine with an analyte from the solution to form a binder-analyte complex;

(b) a test strip configured to allow a test solution to flow, wherein the test solution contains either or both a binder and a binder-analyte complex, the test strip comprising;

(i) a test zone having immobilized thereon a test zone capture agent that, when the binder has not formed a binder-analyte complex, captures the binder,

(ii) a control zone having a plurality of control zone capture agents, each of the plurality of control zone capture agents having different affinity to the binder and each able to capture the binder, whether or not the binder has formed a binder-analyte complex, wherein capture of the binder at either the test zone or control zone results in a detectable signal and wherein a greater signal in the control zone as compared to the test zone indicates a positive result.

2. The assay of claim 1 wherein the control zone capture agents have affinity to each other.

3. The assay of claim 1 wherein at least one of the control zone capture agents comprises an antibody.

4. The assay of claim 1 wherein at least one of the control zone capture agents comprises an antibody binding protein.

5. The assay of claim 4 wherein the antibody binding protein comprises protein A.

6. The assay of claim 1 wherein at least one of the control zone capture agents comprises an antibody and another comprises protein A.

7. The assay of claim 1 wherein the binder comprises an antibody.

8. The assay of claim 1 wherein the binder comprises a multianalyte binder derived from a bacteria.

9. The assay of claim 1 wherein at least one of the control zone capture agents is immobilized on the control zone through attachment to a carrier protein.

10. The assay of claim 9 wherein the carrier protein comprises bovine serum albumin.

11. The assay of claim 1 wherein one of the plurality of control zone capture agents comprises an antibody binding protein and the other of the plurality of control zone capture agents is an antibody to which the antibody binding protein has affinity, and wherein the antibody is applied to the test strip with the binder, and flows, either with the binder or attached to the binder, to the control zone.

12. The assay of claim 11 wherein the binder comprises a beta-lactam binder and wherein one of the control zone capture agents comprises an antibody to the beta-lactam binder.

13. The assay of claim 1 further characterized in that a plurality of analytes can be detected, the assay comprising a plurality of binders and a plurality of test zones, each test zone having an immobilized test zone capture agent with affinity to one of the binders, wherein one control zone signal is compared to the plurality of test zone signals to determine the presence of analyte, and wherein a greater signal in the control zone as compared any one of the test zones indicates a positive result for a particular analyte or analyte family.

14. The assay of claim 13 wherein the one of the plurality of binders comprises a multianalyte binder with binding affinity for beta-lactam antibiotics.

15. The assay of claim 14 wherein the multianalyte binder comprises a beta-lactam binder from Geobacillus stearothermophilus.

16. The assay of claim 14 wherein one of the control zone capture agents comprises an antibody to the multianalyte beta-lactam binder.

17. The assay of claim 13 wherein one of the plurality of binders comprises an antibody.

18. The assay of claim 17 wherein the antibody comprises an antibody to cloxacillin.

19. The assay of claim 13 further comprising one or more unlabeled binders wherein the unlabeled binders reduce test sensitivity relative to the analyte to which the unlabeled binder has affinity.

20. The assay of claim 19 further comprising an assay that is positive when any one of cloxacillin, penicillin G, amoxicillin, ampicillin, amoxicillin, ceftiofur or cephapirin is present at or above the regulatory safe level.

21. The assay of claim 1 wherein at least two of the control zone capture agents are antibodies.

22. The assay of claim 1 wherein at least two control zone capture agents are antibodies and wherein each of the antibodies has affinity to a different species of animal of binder.

23. The assay of claim 1 wherein one of the control zone capture agents comprises a polyclonal antibody and the other comprises a monoclonal antibody.

24. The assay of claim 13 wherein each of the binders comprises an antibody from a different species of animal and at least one of the control zone capture agents comprises an antibody to one of those different species.

25. The assay of claim 13 wherein at least one of the control zone capture agents comprises an antibody binding protein.

26. The assay of claim 25 wherein the antibody binding protein comprises protein A.

27. A method for detecting an analyte in a test solution comprising:

a) configuring a binder for generating a detectable signal, the binder capable of combining with an analyte from the solution to form a binder-analyte complex;

b) immobilizing, within a test zone of a test strip, a test zone capture agent;

c) immobilizing, within a control zone of a strip, a plurality of control zone capture agents, each of the plurality of control zone capture agents having different affinity to the binder and each able to capture the binder;

d) applying the binder to the test strip;

e) applying the test solution to the test strip, wherein the test solution and the binder combines to allow binding of analyte to binder to form a binder-analyte complex;

f) capturing binder unbound by analyte in the test zone;

g) capturing the binder, whether or not the binder has formed a binder-analyte complex, at the control zone;

h) comparing the signal in the control zone to the signal in the test zone to determine the presence of analyte in the solution,

wherein a greater signal in the control zone, as compared to the test zone, indicates a positive result.

28. The method of claim 27 wherein the control zone capture agents have affinity to each other.

29. The method of claim 27 wherein at least one of the control zone capture agents comprises an antibody.

30. The method of claim 27 wherein at least one of the control zone capture agents comprises an antibody binding protein.

31. The method of claim 30 wherein the antibody binding protein comprises protein A.

32. The method of claim 29 wherein at least one of the antibodies has affinity to the antibody binding protein.

33. The method of claim 27 further comprising immobilizing one of the control zone capture agents on the control zone through attachment to a carrier protein.

34. The method of claim 33 wherein the carrier protein comprises bovine serum albumin.

35. The method of claim 27 wherein at least two of the plurality of binders are antibodies and each of the antibodies is raised in a different host species.

36. The method of claim 27 further comprising providing a plurality of binders and a plurality of test zones, each test zone having immobilized thereon a test zone capture agent for binding to a binder, and wherein a greater signal in the control zone, as compared to any of the test zones, indicates a positive result for an analyte or analyte family.

37. The method of claim 36 wherein one of the plurality of binders comprises a multianalyte binder with binding affinity for beta-lactam antibiotics.

38. The method of claim 37 wherein the multianalyte binder comprises a beta-lactam receptor from Geobacillus stearothermophilus.

39. The method of claim 36 wherein at least one of the plurality of binders comprises an antibody.

40. The method of claim 39 wherein the antibody comprises a polyclonal antibody.

41. The method of claim 39 wherein the antibody comprises an antibody to cloxacillin.

42. The method of claim 36 wherein at least one of the control zone capture agents comprises an antibody to a beta lactam receptor and another of the control zone capture agents comprises protein A.

43. The method of claim 36 further comprising providing one or more unlabeled binders wherein the unlabeled binders reduce test sensitivity relative to the analyte to which the unlabeled binder has affinity and wherein a positive result occurs when one or more analytes are present at or above a threshold level.

44. The method of claim 36 wherein one of the test zones capture agents comprises a beta-lactam.

45. The method of claim 44 wherein the beta-lactam comprises ceforanide.

46. The method of claim 44 wherein another of the test zone capture agents comprises cloxacillin.

47. The method of claim 46 wherein the lactam ring of the cloxacillin has been opened.

48. The method of claim 36 further comprising a test to detect amoxicillin, ampicillin, ceftiofur, cephapirin, cloxacillin and penicillin G at or below safe level.

49. The method of claim 27 wherein at least two of the control zone capture agents comprise antibodies.

50. The method of claim 27 wherein at least two of the control zone capture agents comprise antibodies and one of said antibodies comprises a polyclonal antibody and the other of said antibodies comprises a monoclonal antibody.

51. The method of claim 27 wherein at least two of the control zone capture agents comprise antibodies and each of said antibodies comprises an anti-species antibody to a different species.

52. A method for detecting a plurality of analytes present in a test solution comprising:

a) configuring a plurality of binders for generating a detectable signal, the binders capable of combining with an analyte from the solution to form a binder-analyte complex;

b) immobilizing, within a first test zone of a test strip, a first test zone capture agent;

c) immobilizing, within a second test zone of a test strip, a second test zone capture agent;

d) immobilizing, within a control zone of a strip, a plurality of control zone capture agents, each of the plurality of control zone capture agents having different affinity and each able to attach to at least one of the binders, whether or not the binder has formed a binder-analyte complex;

e) applying the plurality of binders to the test strip;

f) applying the test solution to the test strip, wherein the test solution and the plurality of binders combine to allow binding of analyte to binder to form a binder-analyte complex;

g) capturing binder unbound by analyte in either or both the first test zone and second test zone;

h) capturing the binder, whether or not the binder has formed a binder-analyte complex, at the control zone;

i) comparing the signal in the control zone to the signals in the test zones to determine the presence of analyte in the solution,

wherein a greater signal in the control zone, as compared to any one test zone, indicates a positive result.

53. The method of claim 52 wherein the control zone capture agents have affinity to each other.

54. The method of claim 52 wherein one of the control zone capture agents comprises an antibody and another of the control zone capture agents comprises an antibody binding protein.