Claims for Patent: 4,469,787

Summary for Patent: 4,469,787

Title:	Immunoassay involving soluble complex of second antibody and labeled binding protein
Abstract:	A method and kit for determining the presence of a polyvalent ligand in an aqueous fluid. The method involves incubating the fluid with an immobilized antibody to the ligand to form an immobilized ligand-antibody complex, then incubating the immobilized ligand-antibody complex with a solution of a soluble complex of a second antibody (to the ligand) and a labeled binding protein, such as protein A. The labeled binding protein is specifically bound to the Fc portion of the second antibody. The unbound second antibody and labeled binding protein are washed from the immobilized complex, and the presence of bound labeled binding protein is determined as a measure of the concentration of the ligand in the aqueous fluid.
Inventor(s):	Woods; James W. (Creve Coeur, MO), St. Denis; John N. (Maryland Hts., MO), Chapman; Dana A. (Overland, MO)
Assignee:	Mallinckrodt Inc. (St. Louis, MO)
Application Number:	06/378,126
Patent Claims:	1. A method for quantitatively determining the presence of polyvalent ligand in an aqueous fluid which comprises: (a) incubating said fluid with a solid support having a first antibody to said ligand bound thereto to form an immobilized antibody-ligand complex; (b) incubating said immobilized antibody-ligand complex with a solution containing a soluble complex of a second antibody to said ligand and a labeled binding protein specifically bound to the F.sub.c portion of said second antibody; (c) washing unbound antibody and labeled binding protein from the solid support; and (d) determining the presence of labeled binding protein bound to said solid support or remaining in solution, as a measure of the concentration of the ligand in the aqueous fluid; wherein said method is further characterized in that the solution of second antibody-labeled binding protein complex contains an excess concentration of second antibody to provide a substantially linear relationship between the concentration of ligand and the amount of labeled binding protein bound to the solid support, while maintaining analytically acceptable low levels of background interference. 2. The method of claim 1, wherein the molar ratio of soluble second antibody to labeled binding protein is from about 250:1 to about 5,000:1. 3. The method of claim 1, wherein the molar ratio of soluble second antibody to labeled binding protein is from about 50:1 to about 24,000:1. 4. The method of claim 3, wherein the labeled binding protein is protein A obtained from Staphylococcus aureus. 5. The method of claim 4, wherein the ligand is ferritin. 6. The method of claim 4, wherein the ligand is selected from the group consisting of ferritin, thyroid stimulating hormone, creatinine phosphokinase, prostatic acid-phosphatase, and human chorionic gonadotroin, alpha fetoprotein, parathyroid hormone, insulin and C-peptide. 7. The method of claim 6, wherein the first antibody is an intact antibody. 8. The method of claim 5 or 6, wherein the labeled binding protein is selected from the group consisting of .sup.125 I-Protein A, .sup.3 H-Protein A and .sup.14 C-Protein A. 9. The method of claim 5 or 6, wherein the labeled binding protein is enzyme-labeled Protein A. 10. The method of claim 5 or 6, wherein the labeled binding protein is protein A labeled with a fluorescent substance. 11. An immunoassay test kit for determining the presence of a ligand in an aqueous fluid comprising a solid support coated with an antibody to said ligand and, in a separate container, a solution containing a soluble complex of a second antibody to said ligand and a labeled binding protein, wherein the concentrations of second antibody and labeled binding protein in said solution are in a molar ratio of from about 50:1 to about 24,000:1 respectively. 12. The test kit of claim 11, wherein the molar ratio of second antibody to labeled binding protein is from about 250:1 to about 5,000:1. 13. The test kit of claim 12, wherein the labeled binding protein is radiolabeled Protein A. 14. The test kit of claim 12, wherein the labeled binding protein is enzyme-labeled Protein A, and said test kit further comprises a solution of a chromogenic substrate for said enzyme. 15. A method for quantitatively determining the presence of polyvalent ligand in an aqueous fluid which comprises: (a) incubating said fluid with a solid support having a first antibody to said ligand bound therto to form an immobilized antibody-ligand complex; (b) incubating said immobilized antibody-ligand complex with a solution containing a soluble complex of a second antibody to said ligand and a labeled binding protein specifically bound to the Fc portion of said second antibody; (c) washing unbound antibody and labeled binding protein from the solid support; and (d) determining the presence of labeled binding protein bound to said solid support or remaining in solution, as a measure of the concentration of the ligand in the aqueous fluid; wherein said method is characterized in that the first antibody is an IgG immunoglobin which has been enzymatically digested to remove the Fc portion. 16. The method of claim 15, wherein the molar ratio of soluble second antibody to labeled binding protein is from about 500:1 to about 1:1.

Details for Patent 4,469,787

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Nps Pharmaceuticals, Inc.	NATPARA	parathyroid hormone	For Injection	125511	01/23/2015		2039-03-29