Process and reagent for the determination of the luteinizing hormone and monoclonal antibodies suitable thereof

• US 5,248,593 A
• Filed: 06/29/1992
• Issued: 09/28/1993
• Est. Priority Date: 03/06/1985
• Status: Expired due to Term

• Current Assignee: Boehringer Mannheim GmbH (Roche Holding AG)
• Original Assignee: Boehringer Mannheim GmbH (Roche Holding AG)
• Inventors: Schetters, Hartmut, Erler, Klaus, Lenz, Helmut, Hubner-Parajsz, Christa
• Primary Examiner(s): Rosen, Sam
• Application Number:
  US07/908,535
• Time in Patent Office: 456 Days
• Field of Search: 530/388.1, 530/388.15, 530/388.24, 435/70.21, 435/240.27, 435/7.9, 435/7.93, 435/7.95, 436/536, 436/548
• US Class Current: 435/7.9
• CPC Class Codes: C07K 16/26 against hormones ; against hormone releasing or inhibiting factors

Abstract

The present invention provides an immunological process for the determination of the luteinizing hormone (LH), wherein at least one monoclonal antibody is used which is directed against LH and cross-reacts with other glycoprotein hormones to an extent of less than 3%.

The present invention also provides a reagent for the determination of the luteinizing hormone, wherein it contains at least one monoclonal antibody which is directed against LH and cross-reacts with other glycoprotein hormones to an extent of less than 3%.

Furthermore, the present invention provides a monoclonal antibody against the luteinizing hormone and a process and a hybridoma cell line for producing it.

Claims

1. Hybridoma cell line which produces monoclonal antibodies which specifically bind to luteinising hormone and cross react with TSH, HCG and FSH to an extent of less than 3% as determined using an enzyme linked immunosorbent assay, said cell line prepared by

(i) intraperitoneally immunizing a host animal with human luteinizing hormone;

(ii) removing the spleen of said host animal;

(iii) fusing cells of said spleen with myeloma cells;

(iv) separating fused cells from non-fused cells by culturing in selection medium;

(v) screening supernatant of fused cells for production of antibodies by mixing said supernatant with human luteinizing hormone to bind antibody in said supernatant to human luteinizing hormone and with labelled antibody against antibodies which bind to human luteinizing hormone to identify said human luteinizing hormone binding antibodies, and

(vi) repeating step (v) with a glycoprotein hormone selected from the group consisting of TSH, HCG and FSH and with a labelled antibody, to determine human luteinizing hormone specific antibodies which cross react to an extend less than 3% with said glycoprotein hormone.

2. The hybridoma cell line of claim 1, comprising a cell line deposited at the National Collection of Animal Cell Cultures under deposit number NCACC 84122001.

3. The hybridoma cell line of claim 1, comprising a cell line deposited at the National Collection of Animal Cell Cultures under deposit number NCACC 84122005.

4. Monoclonal antibody produced by the hybridoma cell line of claim 1.

5. The monoclonal antibody of claim 4, which cross reacts with TSH, HCG and FSH to an extent less than 1%.

6. The monoclonal antibody of claim 4 comprising MAB 369.

7. The monoclonal antibody of claim 4 comprising MAB 799.

8. Process for the determination of the presence of luteinizing hormone (LH) comprising contacting a sample with at least one monoclonal antibody of claim 4, under conditions favoring formation of monoclonal antibody-antigen complexes between said monoclonal antibody and LH and determining the presence of said complexes.

9. Process of claim 8 comprising monoclonal antibody MAB 369 or MAB 799.

10. Process of claim 8 comprising monoclonal antibodies MAB 369 and MAB 799.

11. Process of claim 8 wherein at least one monoclonal antibody is used which specifically binds to LH and cross reacts with TSH, HCG, and FSH to an extent of less than 1%.

12. Process of claim 8, wherein at least two monoclonal antibodies are used and at least one of said monoclonal antibodies cross reacts with TSH, HCG, and FSH to an extent of less than 1%.

13. Process of claim 8, wherein said monoclonal antibody is radioactively labelled.

14. Process of claim 8, wherein said monoclonal antibody is enzymatically labelled.

15. Process of claim 12, wherein said process comprises sandwich immunoassay.

16. Process of claim 8, wherein said monoclonal antibody comprises an Fab fragment.

17. Process of claim 13, wherein said label is ¹²⁵ I.

18. Process of claim 14, wherein said enzyme label is peroxidase or B-galactosidase.

19. Process of claim 8, wherein said antibody is fluorescently labelled.

20. Process of claim 12, comprising competitive immunoassay.

21. Process of claim 15, comprising forward sandwich immunoassay.

22. Process of claim 15 comprising simultaneous sandwich immunoassay.

23. Process of claim 15 comprising reverse sandwich immunoassay.

24. Reagent for the determination of the presence of luteinizing hormone (LH) comprising at least one monoclonal antibody of claim 4 and a buffer.

25. Reagent of claim 24, wherein said antibody cross reacts with TSH, HCG, and FSH to an extent of less than 1%.

26. Reagent of claim 24, comprising at least two monoclonal antibodies one of which cross reacts with TSH, HCG, and FSH to an extent of less than 1%.

27. Reagent of claim 24 comprising monoclonal antibodies MAB 369 or MAB 799.

28. Reagent of claim 26 comprising monoclonal antibodies MAB 369 and MAB 799.

29. A kit for determining presence of luteinzing hormone comprising separate components of reagents at least one of which is a first monoclonal antibody in accordance with claim 4 and another component which is a member selected from the group consisting of an antibody which specifically binds to the Fcγ

part of said first monoclonal antibody and a labeled antibody which specifically binds to luteinising hormone.

30. A kit as in claim 29 wherein the first antibody specifically binds to luteinizing hormone and cross reacts with TSH, HCG, and FSH to an extent of less than 1%.

31. A kit as in claim 29 wherein said separate components are both monoclonal antibodies which specifically bind to luteinizing hormone.

32. Kit of claim 29 comprising monoclonal antibody MAB 369 or monoclonal antibody MAB 799.

33. Kit of claim 31 comprising monoclonal antibodies MAB 369 and MAB 799.