Bovine pregnancy test

• US 20030073248A1
• Filed: 09/24/2002
• Published: 04/17/2003
• Est. Priority Date: 09/28/2001
• Status: Active Grant

• Current Assignee: Aspenbio Pharma, Inc.
• Original Assignee: Aspenbio Pharma, Inc.
• Inventors: Landmann, Cathy, Colgin, Mark, Hurst, Roger, Newman, Diane, Roth, J.W.
• Granted Patent: US 7,393,696 B2
• Time in Patent Office: Days
• US Class Current: 436/510
• CPC Class Codes: A61D 17/006 for detecting pregnancy of animals
• CPC Class Codes: G01N 2800/368 Pregnancy complicated by disease or abnormalities of pregnancy, e.g. preeclampsia, preterm labour

Abstract

This invention provides bovine pregnancy test methods and devices. The test is also suitable for other ruminant and/or ungulate animals. Antigens from Group A (early pregnancy antigens), and/or Group B (mid-pregnancy antigens), and Group C (early, mid- and late pregnancy antigens) are detected in a fluid from the animal, and pregnancy is reliably determined. The pregnancy assays of this invention are preferably carried out using immunoassay devices which provide immediate results in the field.

Claims

1. A method for determining pregnancy of a ruminant and/or ungulate animal, the method comprising:

testing a fluid or cell derived from said animal for the presence of at least one Group A or Group B antigen;

also testing said fluid for the presence of at least one Group C antigen;

determining that the animal is pregnant if the test is positive for the presence of at least one Group A or Group B antigen and at least one Group C antigen; and

determining that the animal is not pregnant if the test is negative for the presence of at least one Group C antigen.

2. The method of claim 1 wherein the animal is selected from the group consisting cattle, sheep, goats, yak, water buffalo, bison, antelopes, gazelles, elk, reindeer, moose, bighorn sheep, swine, horses, giraffes, bactrian and dromedary camels, llamas, alpacas, and vicunas.

3. The method of claim 1 wherein the animal is a cow.

4. The method of claim 1 comprising testing said fluid for the presence of at least one Group A antigen and at least one Group B antigen.

5. The method of claim 1 wherein testing for the presence of said Group A or Group B antigen is done by means of contacting said fluid with an antibody for said antigen.

6. The method of claim 5 wherein said antibody is a polyclonal antibody.

7. The method of claim 5 wherein said antibody is a monoclonal antibody.

8. The method of claim 5 wherein said antibody is present on a test strip.

9. The method of claim 5 wherein antibodies for both Group A and Group B antigens are on the same test strip.

10. The method of claim 1 wherein testing for the presence of said Group C antigen is done by means of contacting said fluid with an antibody for said antigen.

11. The method of claim 10 wherein said antibody is a polyclonal antibody.

12. The method of claim 10 wherein said antibody is a monoclonal antibody.

13. The method of claim 10 wherein said antibody is present on a test strip.

14. The method of claim 10 wherein antibodies for Group A or Group B antigens are on the same test strip with antibodies for a Group C antigen.

15. The method of claim 1 wherein said Group A antigen is selected from the group consisting of ISG17, Mx, GCP-2, 2′

,5′

oligoadenylate synthetase, β

2-microglobulin, IRF-1, IRF-2, I-8U, 1-8D and Leu-13/9-27, COX-2, BAG, EPF, PSPB, PAG-1, PAG-4, PAG-5, and PAG-9.

16. The method of claim 1 wherein said Group B antigen is selected from the group consisting of PSP-60, PAG-6, and PAG-7.

17. The method of claim 1 wherein said Group C antigen is progesterone.

18. The method of claim 1 wherein said fluid is selected from the group consisting of whole blood, plasma, serum, urine, milk, nasal secretions, ocular secretions, vaginal secretions, saliva and cell lysates.

19. A method for determining readiness for breeding of a female animal of a ruminant and/or ungulate species, said method comprising:

a) determining that the animal is not pregnant using the method of claim 1;

b) monitoring for and detecting behavioral estrus;

thereby determining readiness for breeding.

20. The method of claim 18 wherein method step a) comprises obtaining a sample from said animal on day 18-21 of said animal'"'"'s estrus cycle.

21. A method for readying a female animal of a ruminant and/or ungulate species for breeding, said method comprising:

a) determining that the animal is not pregnant using the method of claim 1;

b) forcing said animal into heat;

thereby readying said animal for breeding.

22. The method of claim 21 wherein method step a) comprises obtaining a sample from said animal on day 18-21 of said animal'"'"'s estrus cycle.

23. The method of claim 21 also comprising breeding said animal.

24. A pregnancy assay for determining pregnancy or lack of pregnancy of a ruminant and/or ungulate animal, said assay comprising:

(a) a cassette; and

(b) at least one test strip housed within said cassette;

(c) first antibodies specific to Group A or specific to Group B antigen, and first antibodies specific to Group C antigen on said at least one test strip, said first antibodies being capable of binding to the corresponding antigen to form first antibody-antigen complexes;

(d) binding partners specific to each such complex on said test strip; and

(e) labels attached to said first antibodies or said binding partners.

25. The pregnancy assay of claim 24 comprising a plurality of test strips, each having at least one said first antibody and at least one binding partner for said at least one first antibody thereon.

26. The pregnancy test of claim 25 comprising two test strips, a first test strip having thereon a first antibody specific to Group A or Group B antigen and a binding partner specific to a Group A or Group B antibody-antigen complex;

and a second test strip having thereon a second antibody specific to Group C antigen and a binding partner specific to a Group C antibody-antigen complex.

27. The pregnancy assay of claim 25 wherein said first test strip has thereon a first antibody specific to Group A antigen and a binding partner specific to a Group A antigen-antibody complex and a first antibody specific to Group B antigen and a binding partner specific to a Group B antigen-antibody complex.

28. The pregnancy assay of claim 25 comprising a first test strip having thereon a first antibody specific to Group A antigen and a binding partner specific to a Group A antigen-antibody complex;

a second test strip having thereon a first antibody specific to Group B antigen and a binding partner specific to a Group B antigen-antibody complex; and

a third test strip having thereon a first antibody specific to Group C antigen and a binding partner specific to Group C antigen-antibody complex.

29. The pregnancy assay of claim 24 wherein said binding partners are second antibodies specific to said Group A or Group B antigen, and to said Group C antigen.

30. The pregnancy assay of claim 24 wherein said binding partners are second antibodies specific to each of said first antibodies.

31. The pregnancy test of claim 24 wherein said binding partners are detection antigens capable of binding specifically to each of said first antibodies.

32. The pregnancy test of claim 24 wherein said binding partners are immobilized on said test strip.

33. The bovine pregnancy test of claim 24 wherein said binding partners are not immobilized on said test strip.

34. The bovine pregnancy test of claim 24 wherein said first antibodies are immobilized on said test strip.

35. The bovine pregnancy test of claim 24 wherein said first antibodies are not immobilized on said test strip.

36. The bovine pregnancy test of claim 24 wherein said first antibodies are labeled.

37. The assay of claim 36 wherein labels attached to said first antibodies specific to Antigen A or Antigen B are detectably different from said labels attached to said first antibodies specific to Antigen C.

38. The assay of claim 37 comprising first labeled antibodies specific to Antigen A and first labeled antibodies specific to Antigen B, wherein the labels attached to each antigen are detectably different from each other.

39. The bovine pregnancy test of claim 24 wherein said first antibodies are not labeled.

40. The bovine pregnancy test of claim 24 wherein said binding partners are labeled.

41. The bovine pregnancy test of claim 24 wherein said binding partners are not labeled.

42. The assay of claim 24 wherein said labeled first antibodies or said labeled binding partners immobilized on said test strip are immobilized in different patterns corresponding to whether they bind to Group A antigens or antibodies, Group B antigens or antibodies, or Group C antigens or antibodies.

43. The assay of claim 24 wherein said cassette comprises a sample aperture for introducing sample fluid into said assay.

44. The assay device of claim 43 also comprising a sample pad positioned beneath said sample aperture.

45. The assay device of claim 43 also comprising a filter positioned downstream from said sample aperture.

46. The assay of claim 24 wherein said cassette comprises at least one test window positioned above the point(s) on said test strip(s) wherein said labeled first antibodies, said labeled second antibodies, or said labeled detection antigens are immobilized.

47. The assay of claim 24 also comprising control Group A and/or Group B antigens, and control Group C antigens on said test strip(s).

48. The assay device of claim 47 wherein said cassette also comprises at least one control window positioned above said test strip(s) at the point(s) where said control antigens or said binding partners are immobilized.

49. A bovine pregnancy assay device comprising:

(1) a test cassette having an upstream end and a downstream end comprising;

(a) a sample aperture near the upstream end;

(b) a test window downstream from the sample aperture;

(c) a control window downstream from the test window;

(2) a test strip housed within said test cassette comprising;

(a) a test region which comprises;

(i) a sample pad positioned beneath said sample aperture;

(ii) a filter positioned adjacent to and downstream from said sample pad;

(iii) a detection antibody area positioned downstream from said filter comprising removable labeled first antibodies to Antigen A or Antigen B, and removable labeled first antibodies to Antigen C, wherein said first antibodies to Antigen A or Antigen B are labeled with detectably different labels from the labels of said first antibodies to Antigen C;

(iv) a capture antibody area positioned downstream from said detection antibody area comprising second antibodies to Antigen A or Antigen B, and second antibodies to Antigen C, bound to said test strip, wherein said second antibodies to Antigen A or Antigen B are bound to known a known location in said capture antibody area and said second antibodies to Antigens C are bound to a known different locations in said capture antibody area; and

(b) a control region downstream from said test region comprising;

(i) a control antigen area comprising removable Antigen A or Antigen B and Antigen C;

(ii) a second capture antibody region downstream from said control antigen area.

50. The assay device of claim 49 comprising first and second antibodies to Antigens A and B comprising control antigens A and B.

51. A method of making a test device for determining pregnancy status of a cow comprising:

providing a test strip;

providing a housing for a test strip;

removably or fixedly placing on said test strip labeled or unlabeled first antibodies specific to Group A or specific to Group B antigen, and labeled or unlabeled first antibodies specific to Group C antigen;

removably or fixedly placing on said test strip;

labeled or unlabeled second antibodies specific to Group A or Group B antigen; and

labeled or unlabeled second antibodies specific to Group C antigen;

or

labeled or unlabeled detection antigens capable of specifically binding to said first or second antibodies to Group A antigen, or labeled or unlabeled detection antigens capable of specifically binding to said first or second antibodies to Group B antigen; and

labeled or unlabeled detection antigens capable of specifically binding to said first or second antibodies to Group C antigen; and

placing said test strip within said housing.

52. The method of claim 51 also comprising:

removably or fixedly placing on said test strip control Group A or control Group B antigen and control Group C antigen; and

removably or fixedly placing on said test strip binding partners specific to said control Group A or control Group B antigen, and specific to said control Group C antigen.

53. The method of claim 51 comprising placing a sample pad within said housing.

54. The method of claim 51 comprising placing a filter within said housing.

55. An assay kit for determining pregnancy in a ruminant and/or ungulate animal, said kit comprising, in close association:

a) a test strip;

b) first antibodies that bind immunologically to Antigens A and/or B and Antigen C;

c) second antibodies that bind to said first antibody(ies);

d) control antigens of Group A and/or Group B;

e) instructions for use of said kit.