Method for diagnosing diseases based on levels of anti-glycan antibodies

1. A method of diagnosing Crohn's disease in a subject, the method comprising

providing a test sample from a subject with digestive symptoms of Crohn's disease;

detecting a level of an IgA anti-GlcNAc (β 1-4) GlcNAc (β) antibody (ACCA) in said sample by binding of said IgA ACCA in said sample to a carbohydrate reagent comprising an isolated GlcNAc (β 1-4) GlcNAc (β) glycan; and

diagnosing Crohn's Disease in said subject by detection of an elevated level of said IgA ACCA in said test sample relative to a control reference level.

2. The method of claim 1 , wherein said control reference level is a level from one or more individuals without Crohn's disease.

3. The method of claim 1 , wherein said control reference level is a level from one or more individuals with a gastrointestinal disorder that is irritable bowel syndrome or ulcerative colitis.

4. The method of claim 1 , wherein said control reference level is a level from one or more individuals that do not have a gastrointestinal disorder.

5. The method of claim 1 , wherein said ACCA is detected with a fluorescent antibody.

6. The method of claim 1 , wherein said ACCA is detected with an enzyme-linked immunoabsorbent assay (ELISA).

7. The method of claim 1 , wherein said method further comprises detecting a level of an anti-Glc (β 1-3) Glc (β) antibody (ALCA) or an anti-polysaccharide β-D (1-3) Glucan antibody in said sample, wherein said subject is assessed as having Crohn's disease if said ALCA or said polysaccharide β-D (1-3) Glucan antibody levels are elevated in said sample relative to a control reference level.

8. The method of claim 7 , further comprising determining an isotype of said anti-Glc (β 1-3) Glc (β) antibody or said anti-polysaccharide β-D (1-3) Glucan antibody in said sample.

9. The method of claim 8 , wherein said anti-Glc (β 1-3) Glc (β) antibody or said anti-polysaccharide β-D (1-3) Glucan antibody in said sample is an IgA isotype antibody.

10. The method of claim 8 , wherein said anti-Glc (β 1-3) Glc (β) antibody or said anti-polysaccharide β-D (1-3) Glucan antibody in said sample is an IgC isotype antibody.

11. The method of claim 10 , wherein said method further comprises detecting a level of an anti-Glc (β) IgG antibody, an anti-Glc (β 1-3) Glc (β) IgG antibody, an anti-Glc (β 1-4) Glc (β) IgG antibody, an anti-GlcNAc (β) 6-sulfate IgG antibody, or an anti-Xylan IgG antibody in said sample, wherein said subject is assessed as having Crohn's disease if the levels of each of said detected antibodies are elevated in said sample relative to a control reference level.

12. The method of claim 1 , wherein said method further comprises detecting a level of an anti-Glc (β 1-3) Glc (β) antibody (ALCA) and an anti-polysaccharide β-D (1-3) Glucan antibody in said sample, wherein said subject is assessed as having Crohn's disease if said ALCA and said polvsaccharide (β 1-3) Glucan antibody levels are elevated in said sample relative to a control reference level.

13. The method of claim 1 , further comprising

detecting a level of an anti-Glc (β 1-3) Glc (β) antibody (ALCA) in said sample by binding of said ALCA in said sample to a carbohydrate reagent comprising an isolated Glc (β 1-3) Glc (β) glycan and

diagnosing Crohn's Disease in said subject by detection of an elevated level of said anti-Glc (β 1-3) Glc (β) antibody in said test sample relative to a control reference level.

14. The method of claim 1 , further comprising determining whether said test sample has anti-neutrophil cytoplasmic antibodies (ANCA), wherein the subject is assessed as having Crohn's Disease if said ANCA are absent from said sample.

15. The method of claim 1 , further comprising detecting in said test sample a level of an anti-Mannan antibody (ASCA), wherein the subject is assessed as having Crohn's disease if said ASCA levels are elevated in said sample relative to a control reference level.

16. The method of claim 15 , further comprising determining whether said test sample has anti-neutrophil cytoplasmic antibodies (ANCA), wherein the subject is assessed as having Crohn's Disease if said ANCA are absent from said sample.

17. The method of claim 1 , further comprising

detecting a level of an anti-mannan antibody (ASCA) in said sample by binding of said ASCA in said sample to a carbohydrate reagent comprising an isolated mannan and

diagnosing Crohn's Disease in said subject by detection of an elevated level of said anti-mannan antibody in said test sample relative to a control reference level.

18. The method of claim 1 , wherein said method further comprises detecting a level of one, two, or three of anti-Man (α 1-3)[Man (α 1-6)] Man (α) antibody, anti-Man (α 1-2) Man (α) antibody, anti-Man (α 1-6) Man (α) antibody (AMCA) or an anti-Mannan antibody (ASCA) in said sample, wherein said subject is assessed as having Crohn's disease if the levels of each of said detected antibodies are elevated in said sample relative to a control reference level.

19. The method of claim 1 , further comprising

detecting a level of an anti-Man (α 1-3) Man (α) antibody (AMCA) in said sample by binding of said AMCA in said sample to a carbohydrate reagent comprising an isolated Man (α 1-3) Man (α) glycan and

diagnosing Crohn's Disease in said subject by detection of an elevated level of said anti-Man (α 1-3) Man (α) antibody in said test sample relative to a control reference level.

20. The method of claim 1 , wherein said test sample is a biological fluid.

21. The method of claim 20 , wherein said biological fluid is whole blood, serum, plasma, urine, or saliva.

22. The method of claim 20 , wherein said biological fluid is serum.

23. A method of diagnosing Crohn's disease in a subject, the method comprising

providing a test sample from a subject with digestive symptoms of Crohn's disease;

detecting a level of an anti-Glc (β 1-3) Glc (β) antibody (ALCA) in said sample by binding of said ALCA in said sample to a carbohydrate reagent comprising an isolated Glc (β 1-3) Glc (β) glycan; and

diagnosing Crohn's Disease in said subject by detection of an elevated level of said antibody in said test sample relative to a control reference level.

24. The method of claim 23 , wherein said method comprises detecting a level of an IgG ALCA in said sample.

25. The method of claim 24 , wherein said method further comprises detecting a level of an IgG anti-Man (α 1-3) Man (α) antibody in said sample, wherein said subject is assessed as having Crohn's disease if said IgG anti-Man (α 1-3) Man (α) antibody level is elevated in said sample relative to a control reference level.

26. The method of claim 23 , wherein said method comprises detecting a level of an IgG anti-Glc (β 1-3) Glc (β) antibody (ALCA) and an IgG anti-Man (α 1-3) Man (α) antibody (AMCA) in said sample, wherein said subject is assessed as having Crohn's disease if said IgG ALCA and said IgG AMCA levels are elevated in said sample relative to a control reference level.

27. The method of claim 23 , wherein said method further comprises detecting a level of an IgG anti-Mannan antibody (ASCA) or an IgA anti-Mannan antibody (ASCA) in said sample, wherein said subject is assessed as having Crohn's Disease if said IgG ASCA or said IgA ASCA is elevated in said sample relative to a control reference level.

28. The method of claim 27 , wherein said method comprises detecting the level of IgG ASCA in said sample.

29. The method of claim 27 , wherein said method comprises detecting the level of IgA ASCA in said sample.

30. The method of claim 27 , wherein said method further comprises determining whether said sample has anti-neutrophil cytoplasmic antibodies (ANCA), wherein said subject is assessed as having Crohn's Disease if said ANCA are absent from said sample.

31. The method of claim 23 , wherein said control reference level is a level from one or more individuals without Crohn's disease.

32. The method of claim 23 , wherein said control reference level is a level from one or more individuals with a gastrointestinal disorder that is irritable bowel syndrome or ulcerative colitis.

33. The method of claim 23 , wherein said control reference level is a level from one or more individuals that do not have a gastrointestinal disorder.

34. The method of claim 23 , wherein said ALCA is detected with a fluorescent antibody.

35. The method of claim 23 , wherein said ALCA is detected with an enzyme-linked inununoabsorbent assay (ELISA).

36. The method of claim 23 , further comprising determining an isotype of said antibody, wherein said antibody is an IgA isotype antibody.

37. The method of claim 23 , wherein said method further comprises detecting a level of an anti-GlcNAc (β 1-4) GleNAc (β) antibody (ACCA) or an anti-polysaceharide β-D (1-3) Glucan antibody in said sample, wherein said subject is assessed as having Crohn's disease if said ACCA or said anti-polysaceharide β-D (1-3) Glucan antibody levels are elevated in said sample relative to a control reference level.

38. The method of claim 37 , further comprising determining an isotype of said ALCA and said detected ACCA or said detected anti-polysaccharide β-D (1-3) Glucan antibody in said sample.

39. The method of claim 38 , wherein said detected ACCA or said detected anti-polysaccharide β-D (1-3) Glucan antibody in said sample is an IgA isotype antibody.

40. The method of claim 38 , wherein said ALCA and said detected ACCA or said detected anti-polysaccharide β-D (1-3) Glucan antibody in said sample is an IgG isotype antibody.

41. The method of claim 40 , wherein said method further comprises detecting a level of an anti-Glc (β) IgG antibody, an anti-Glc (β 1-3) Glc (β) IgG antibody, an anti-Glc (β 1-4) Glc (β) IgG antibody, an anti-GlcNAc (β) 6-sulfate IgG antibody, or an anti-Xylan IgG antibody in said sample, wherein said subject is assessed as having Crohn's disease if the levels of each of said detected antibodies are elevated in said sample relative to a control reference level.

42. The method of claim 23 , wherein said method further comprises detecting a level of an anti-GleNAc (β 1-4) GleNAc (β) antibody (ACCA) and an anti-polysaceharide β-D (1-3) Glucan antibody in said sample, wherein said subject is assessed as having Crohn's disease if said ACCA and said anti-polysaccharide β-D (1-3) Glucan antibody levels are elevated in said sample relative to a control reference level.

43. The method of claim 23 , further comprising determining whether said test sample has anti-neutrophil cytoplasmic antibodies (ANCA), wherein the subject is assessed as having Crohn's Disease if said ANCA are absent from said sample.

44. The method of claim 23 , further comprising detecting a level of anti-Mannan antibody (ASCA) in said sample, wherein the subject is assessed as having Crohn's disease if said ASCA is elevated in said sample relative to a control reference level.

45. The method of claim 44 , further comprising determining whether said test sample has anti-neutrophil cytoplasmic antibodies (ANCA), wherein the subject is assessed as having Crohn's Disease if said ANCA are absent from said sample.

46. The method of claim 23 , wherein said test sample is a biological fluid.

47. The method of claim 46 , wherein said biological fluid is whole blood, serum, plasma, urine, or saliva.

48. The method of claim 46 , wherein said biological fluid is serum.

49. The method of claim 23 , wherein said method further comprises detecting a level of said anti-Glc (β 1-3) Glc (β) antibody (ALCA), and one, two, or three of anti-Man (α 1-3) Man (α) antibody, anti-Man (α 1-3)[Man (α 1-6)] Man (α) antibody, anti-Man (α 1-2) Man (α) antibody, anti-Man (α 1-6) Man (α) antibody (AMCA) or an anti-Mannan antibody (ASCA ) in said sample, wherein said subject is assessed as having Crohn's disease if the levels of each of said detected antibodies are elevated in said sample relative to a control reference level.

50. A method of differentially diagnosing Crohn's disease or inflammatory bowel disease in a subject with digestive symptoms of Crohn's disease or inflammatory bowel disease, the method comprising

providing a test sample from the subject;

detecting a level of an anti-neutrophil cytoplasmic antibody (ANCA) in said sample;

detecting a level of an IgG anti-Glc (β 1-3) Glc (β) antibody (ALCA) in said sample by binding of said ALCA in said sample to a carbohydrate reagent comprising an isolated Glc (β 1-3) Glc (β) glycan;

the method further comprising identifying a specific anti-Mannan antibody (ASCA) selected from the group consisting of

IgG ASCA and

IgA ASCA,

wherein absence of ANCA and presence of at least one of said IgG ALCA IgG ASCA, or IgA ASCA antibodies in said test sample indicates the subject has Crohn's disease, and

wherein the subject is assessed as having inflammatory bowel disease if ANCA is present and at least one of said IgG ALCA, IgC ASCA, IgA ASCA antibodies are present in said test sample.

51. A method of diagnosing Crohn's disease in a subject, the method comprising providing a test sample from a subject with digestive symptoms of Crohn's disease;

detecting a level of an anti-GlcNAc (β 1-4) GlcNAc (β) antibody (ACCA) in said sample by binding of said ACCA in said sample to an isolated GlcNAc (β 1-4) GlcNAc (β) on a solid phase;

detecting a level of an anti-Glc (β 1-3) Glc (β) antibody (ALCA) in said sample by binding of said ALCA in said sample to an isolated Glc (β 1-3) Glc (β) on a solid phase;

detecting a level of an anti-mannan antibody (ASCA) in said sample by binding of said ASCA in said sample to an isolated mannan on a solid phase; and

detecting a level of an anti-Man(α1,3)Man(α) antibody (AMCA) in said sample by binding of said AMCA in said sample to an isolated Man(α1,3)Man(α) on a solid phase;

wherein elevated levels of at least two of said antibodies in said test sample relative to control reference levels indicate the subject has Crohn's disease.

52. The method of claim 51 , wherein elevated levels of at least three of said antibodies in said test sample relative to control reference levels indicate the subject has Crohn's disease.

53. The method of claim 51 , wherein said GlcNAc (β 1-4) GlcNAc (β) glyean, said Glc(β 1-3) Glc (β), said mannan, and said Man(α 1-3)Man (α)are attached via a linker to said solid phase.

54. The method of claim 51 , wherein said GlcNAc (β 1-4) GlcNAc (β), said Glc(β 1-3) Glc (β), said mannan, and said Man(α1,3)Man(α) are covalently attached to said solid phase.