Immunometric assay for the detection of human chorionic gonadotropin

• US 4,804,626 A
• Filed: 10/22/1986
• Issued: 02/14/1989
• Est. Priority Date: 10/22/1986
• Status: Expired due to Term

• Current Assignee: The General Hospital Corporation (Mass General Brigham, Inc.)
• Original Assignee: The General Hospital Corporation (Mass General Brigham, Inc.)
• Inventors: Wands, Jack R., Bellet, Dominique, Ozturk, Mehmet
• Primary Examiner(s): Nucker, Christine M.
• Application Number:
  US06/921,508
• Time in Patent Office: 846 Days
• Field of Search: 435/7, 435/240, 435/27, 435/810, 436/510, 436/531, 436/534, 436/548, 436/800, 436/804, 436/808, 436/813, 436/814, 436/818, 436/819, 530/387, 530/808, 530/809
• US Class Current: 435/7.94
• CPC Class Codes: C07K 16/26 against hormones ; against hormone releasing or inhibiting factors
• CPC Class Codes: C07K 2317/34 Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
• CPC Class Codes: Y10S 435/81 Packaged device or kit
• CPC Class Codes: Y10S 436/80 Fluorescent dyes, e.g. rhodamine
• CPC Class Codes: Y10S 436/804 Radioisotope, e.g. radioimmunoassay
• CPC Class Codes: Y10S 436/808 Automated or kit
• CPC Class Codes: Y10S 436/813 Cancer
• CPC Class Codes: Y10S 436/814 Pregnancy
• CPC Class Codes: Y10S 436/818 Human chorionic gonadotropin
• CPC Class Codes: Y10S 436/819 Multifunctional antigen or antibody
• CPC Class Codes: Y10S 530/808 Materials and products related to genetic engineering or hybrid or fused cell technology, e.g. hybridoma, monoclonal products
• CPC Class Codes: Y10S 530/809 Fused cells, e.g. hybridoma

Abstract

A highly sensitive and specific monoclonal-immuno-radiometric assay (M-IRMA) for hCG, using monoclonal antibodies (Mabs) directed against a 37-amino acid synthetic polypeptide analogous to the carboxyl terminus (CTP) of beta-hCG. Accordingly, in one embodiment, a method is described for the determination of human chorionic gonadotr

The present invention was made utilizing funds of the United States Government. The U.S. government is therefore granted a royalty-free, non-exclusive, world wide, paid-up license in this invention.

Claims

1. A method for the determination of human chorionic gonadotropin in a sample, which comprises:

(a) contacting said sample with a first capture monoclonal antibody and a second capture monoclonal antibody which are bound on a carrier, wherein said first and second capture monoclonal antibodies are epitopically specific and non cross-reactive for distinct epitopes of the carboxy terminus region of the beta-subunit of human chorionic gonadotropin;

(b) incubating the components of step (a) for a period of time and under conditions sufficient to form an immune complex between said human chorionic gonadotropin, said first capture monoclonal antibody, said second capture monoclonal antibody and said carrier,

(c) adding to said complex of step (b) a detectably labeled indicator monoclonal antibody, wherein said indicator monoclonal antibody is epitopically specific for the alpha-subunit of human chorionic gonadotropin; and

(d) determining the detectably labeled indicator monoclonal antibody in said carrier or in liquid phase, which is indicative of the amount of human chorionic gonadotropin in said sample.

2. The method of claim 1 wherein at least one of said first and second capture antibodies has a Kasn for hCG which is equal to or greater than about 10⁸ /M but less than about 10⁹ /M.

3. The method of claim 1 wherein at least one of said first and second capture antibodies has a Kasn for hCG which is equal to or greater than about 10⁹ /M.

4. The method of claim 1 wherein said first capture monoclonal antibody is secreted by cell line FB12 and said second capture monoclonal antibody is secreted by a cell line selected from the group consisting of FB08 and FB09.

5. The method of claim 1 wherein said first capture monoclonal antibody is secreted by cell line FB12 and said second capture monoclonal antibody is derived from cell line FB09.

6. The method of claim 1 wherein said indicator monoclonal antibody is secreted by cell line HT 13.

7. The method of any of claims 2, 3 or 4 wherein said indicator monoclonal antibody is secreted by cell line HT 13.

8. The method of claim 5 wherein said indicator monoclonal antibody is secreted by cell line HT 13.

9. A kit useful for the detection of human chorionic gonadotropin comprising carrier means being compartmentalized to receive in close confinement therein one or more containers wherein:

(a) a first container contains a first capture monoclonal antibody and a second capture monoclonal antibody which are bound on a carrier, wherein said first and second capture monoclonal antibodies are epitopically specific and noncross reactive for distinct epitopes of the carboxy terminus region of the beta-subunit of human chorionic gonadotropin; and

(b) a second container contains a detectably labeled indicator monoclonal antibody, wherein said indicator monoclonal antibody is epitopically specific for the alpha-subunit of human chorionic gonadotropin.

10. The kit of claim 9 wherein at least one of said first and second capture antibodies has a Kasn for hCG which is equal to or greater than about 10⁸ /M but less than about 10⁹ /M.

11. The kit of claim 9 wherein at least one of said first and second capture antibodies has a Kasn for hCG which is equal to or greater than about 10⁹ /M.

12. The kit of claim 9 wherein said first capture monoclonal antibody is secreted by cell line FB12 and said second capture monoclonal antibody is secreted by a cell line selected from the group consisting of FB08 and FB09.

13. The kit of claim 9 wherein said first capture monoclonal antibody is secreted by cell line FB12 and said second capture monoclonal antibody is secreted by cell line FB09.

14. The kit of claim 9 wherein said indicator monoclonal antibody is secreted by cell line HT 13.

15. The kit of any of claims 10, 11 or 12 wherein said indicator monoclonal antibody is secreted by cell line HT 13.

16. The kit of claim 13 wherein said indicator monoclonal antibody is secreted by cell line HT 13.

17. An insoluble immunological reagent support useful in the determination of human chorionic gonadotropin in a sample, which comprises an insoluble support comprising two immobilized capture monoclonal antibodies epitopically specific and non cross-reactive for distinct epitopes of the carboxy terminal region of the beta-subunit of human chorionic gonadotropin.

18. The reagent of claim 17 wherein at least one of said immobilized capture monoclonal antibodies has a Kasn for hCG which is equal to or greater than about 10⁸ /M but less than about 10⁹ /M.

19. The reagent of claim 17 wherein at least one of said immobilized capture monoclonal antibodies has a Kasn for hCG which is equal to or greater than about 10⁹ /M.

20. The reagent of claim 17 wherein the first of said immobilized capture monoclonal antibodies is secreted by cell line FB12, and the second of said immobilized capture monoclonal antibodies is secreted by a cell line selected from the group consisting of FB08 and FB09.

21. The reagent of claim 17 wherein the first of said immobilized capture monoclonal antibodies is secreted by cell line FB12 and the second of said immobilized capture monoclonal antibodies is secreted by cell line FB09.

22. The cell line FB08.

23. The cell line FB09.

24. The cell line FB12.

25. The cell line HT 13.

26. The monoclonal antibody secreted by cell line FB08.

27. The monoclonal antibody secreted by cell line FB09.

28. The monoclonal antibody secreted by cell line FB12.

29. The monoclonal antibody secreted by cell line HT 13.