Detection of autoantibodies reactive with pancreatic islet cell antigenic molecules and/or insulin

1. A method of screening an undiluted sample of serum or plasma obtained from an animal subject for analyte autoantibodies reactive with the antigenic molecule protein tyrosine phosphatase-like islet cell antigen (IA-2), said method comprising:

(a) immobilizing antigenic molecules from one or more first sources to a solid support in an amount so as to form a monovalent intermediate complex comprising [antigenic molecule of said first source]-[analyte autoantibody] capable of forming a divalent complex with antigenic molecules of a second source, wherein said antigenic molecules of the one or more first sources are capable of interacting with analyte autoantibodies in a sample of serum or plasma from said subject or in a calibrator and comprises molecules of IA-2;

(b) contacting said immobilized antigenic molecules of the one or more first sources with:

(1) an undiluted sample of serum or plasma from said subject, or said calibrator; and

(2) antigenic molecules of a one or more second sources comprising a direct or indirect label, wherein said antigenic molecules of one or more second sources are capable of interacting with analyte autoantibodies in a sample of serum or plasma from said subject or in said calibrator, and comprises molecules of IA-2,

whereby analyte autoantibodies when present in said sample of serum or plasma interact with said antigenic molecules of the first and second sources to form one or more divalent complexes comprising [antigenic molecule of said first source]-[analyte autoantibody]-[antigenic molecule of said second source]; and

(c) detecting the presence of complexes formed in (b) to provide an indication of analyte autoantibodies present in said sample.

2. The method according to claim 1 , wherein the antigenic molecules of said one or more first or second sources consist essentially of IA-2.

3. The method according to claim 1 , wherein the antigenic molecules of said one or more first and second sources consist essentially of IA-2.

4. The method according to claim 1 wherein the antigenic molecules of said one or more first or second sources when present in one or more complexes comprise IA-2.

5. The method according to claim 1 , wherein the antigenic molecules of said one or more first and/or second sources consist essentially of fusion molecules.

6. The method according to claim 1 , wherein the antigenic molecules of said one or more first and/or second sources comprises fusion molecules.

7. The method according to claim 1 , wherein both antigenic molecules of said one or more first and second sources when present in the one or more complexes comprise IA-2, with which analyte autoantibodies when present in said sample can interact.

8. The method according to claim 1 , wherein one antigenic molecules of said one or more first or second source when present in the one or more complexes comprise IA-2, and the other antigenic molecule when present in the one or more complexes comprises one IA-2, with which analyte autoantibodies when present in said sample can interact.

9. The method according to claim 1 , wherein said immobilised antigenic molecules of said one or more first sources is contacted with the sample obtained from the subject so as to form the monovalent intermediate complex comprising [antigenic molecule of first source]-[analyte autoantibody] and the formed immobilised monovalent intermediate complex is subsequently contacted with said antigenic molecules of said one or more second sources, present in solution phase, so as to form said divalent complexes immobilised to the solid support via the antigenic molecule of the first source.

10. The method according to claim 1 , wherein the said sample of serum or plasma is obtained from an animal subject suspected of suffering from, susceptible to or having one or more of the following: type 1 diabetes mellitus and/or stiff man syndrome, type 2 diabetes mellitus, one or more autoimmune thyroid diseases, celiac disease, one or more connective tissue diseases, adrenal autoimmunity, or a combination of two or more different autoimmune diseases.

11. The method according to claim 1 , wherein the calibrator is diluted 32 fold or less.

12. The method according to claim 1 , wherein the calibrator is diluted 16 fold or less.

13. The method according to claim 1 , wherein the calibrator is diluted 8-fold or less.

14. A method of screening a diluted sample of serum or plasma obtained from an animal subject for analyte autoantibodies reactive with the antigenic molecule protein tyrosine phosphatase-like islet cell antigen (IA-2), said method comprising:

(a) immobilizing antigenic molecules from one or more first sources to a solid support in an amount so as to form a monovalent intermediate complex comprising [antigenic molecule of said first source]-[analyte autoantibody] capable of forming a divalent complex with antigenic molecules of the second source, wherein said first source of antigenic molecule is capable of interacting with analyte autoantibodies in a sample of serum or plasma from said subject, or a calibrator and comprises molecules of IA-2;

(b) contacting said immobilized one or more first sources of antigenic molecules with:

(1) a diluted sample of serum or plasma from said subject, wherein the sample is diluted 32-fold or less; or said calibrator; and

(2) antigenic molecules of one or more second sources comprising a direct or indirect label, wherein said antigenic molecules of the one of more second sources are capable of interacting with analyte autoantibodies in a sample of serum or plasma from said subject or said calibrator and comprises molecules of IA-2,

whereby analyte autoantibodies when present in said sample of serum or plasma interact with said antigenic molecules of the first and second sources to form one or more divalent complexes comprising [antigenic molecule of said first source]-[analyte autoantibody]-[antigenic molecule of said second source]; and

(c) detecting the presence of complexes formed in (b) to provide an indication of analyte autoantibodies present in said sample.

15. The method according to claim 14 , wherein the antigenic molecules of said one or more first or second sources consist essentially of IA-2.

16. The method according to claim 14 , wherein the antigenic molecules of said one or more first and second sources consist essentially of IA-2.

17. The method according to claim 14 , wherein the antigenic molecules of said one or more first or second sources when present in one or more complexes comprise IA-2.

18. The method according to claim 14 , wherein the antigenic molecules of said one or more first and/or second sources consist essentially of fusion molecules.

19. The method according to claim 14 , wherein the antigenic molecules of said one or more first and/or second sources comprises fusion molecules.

20. The method according to claim 14 , wherein both antigenic molecules of said one or more first and second sources when present in the one or more complexes comprise IA-2, with which analyte autoantibodies when present in said sample can interact.

21. The method according to claim 14 , wherein one antigenic molecules of said one or more first or second source when present in the one or more complexes comprise IA-2, the other antigenic molecule when present in the IA-2, with which analyte autoantibodies when present in said sample can interact.

22. The method according to claim 14 , wherein said immobilised antigenic molecules of said one or more first sources is contacted with the sample obtained from the subject so as to form the monovalent intermediate complex comprising [antigenic molecule of first source]-[analyte autoantibody] and the formed immobilised monovalent intermediate complex is subsequently contacted with said antigenic molecules of said one or more second sources, present in solution phase, so as to form said divalent complexes immobilised to the solid support via the antigenic molecule of the first source.

23. The method according to claim 14 , wherein the said sample of serum or plasma is obtained from an animal subject suspected of suffering from, susceptible to or having one or more of the following: type 1 diabetes mellitus and/or stiff man syndrome, type 2 diabetes mellitus, one or more autoimmune thyroid diseases, celiac disease, one or more connective tissue diseases, adrenal autoimmunity, or a combination of two or more different autoimmune diseases.

24. The method according to claim 14 , wherein the calibrator is diluted 32 fold or less.

25. The method according to claim 14 , wherein the calibrator is diluted 16 fold or less.

26. The method according to claim 14 , wherein the calibrator is diluted 8-fold or less.

27. The method according to claim 14 , wherein the sample is diluted 16-fold or less.

28. The method according to claim 14 , wherein the label comprises an enzymatic label.

29. The method according to claim 14 , wherein the label comprises alkaline phosphatases, horse radish peroxidase or biotin.