Hybridoma cell line and its monoclonal antibody to erythropoletin

Claim 2

1 . As a new composition, a hybridoma cell line A.T.C.C. HB8209.

Claim 3

As a new composition, the monoclonal antibody produced from hybridoma cell line A.T.C.C. H B8209 and specifically immunoreactive with erythropoietin.

Claim 4

As a new composition, the monoclonal antibody produced from hybridoma cell line A.T.C.C. H B8209 and specifically immunoreactive with erythropoietin.

Claim 5

The process of using the monoclonal antibody of claim 2 in an immunological procedure for isolation of biologically active erythropoietin from a biological fluid.

Claim 6

4 . The process of claim 3'wherein said fluid is a human fluid.

Claim 7

5 . The process of claim 3 wherein said fluid is a member of the group comprising whole blood, serum, urine, cystic fluid and cerebrospinal fluid.

Claim 8

The process of using the monoclonal antibody of claim 2 in an immunological procedure for the quantitative detection of erythropoietin in a biological fluid.

Claim 9

The process of claim 6 wherein said fluid is a human fluid.

Claim 10

The process of claim 6.wherein said fluid is a member of the group-comprising whole blood, serum, urine, cystic fluid and cerebrospinal fluid.

Claim 11

The process of claim 6 wherein said immunological procedure is an immunobinding assay.

Claim 12

As a new composition, a monoclonal antibody specifically immunoreactive with erythropoietin and with a polypeptide substantially duplicative of the amino acid sequence extant in erythropoietin.

Claim 13

As a new composition, a monoclonal antibody specifically immunoreactive with erythropoietin and with a polypeptide substantially duplicative of the amino acid sequence extant in erythropoietin.

Claim 14

The product of claim 10 wherein the amino acid sequence of the polypeptide to which said antibody is immunoreactive comprises: NH 2 -Ala-Pro-Pro-Arg-Leu-Ile-Cys-Asp-Ser-Arg-Val-Leu-Glu-Arg-Tyr-Leu-Leu-Glu-Aia-Lys-COOH.

Claim 15

An immunological assay for quantitative detection of erythropoietin in a biological fluid sample comprising the steps of: (1) contacting said fluid with a first, immobilized antibody which reacts with a first antigenic determinant of erythropoietin in said fluid to form an immunological complex of erythropoietin and said first antibody; (2) contacting the complex formed in step (1) with a second antibody which reacts with an antigenic determinant of erythropoietin other than said first antigenic determinant to form an immunological complex of erythropoietin and said second antibody; and (3) quantifying the amount of said second antibody bound to said immunological complex formed in step (2), either said first or said second antibody comprising a monoclonal antibody of claim 10.

Claim 16

The assay of claim 12 wherein said first antibody or said second antibody is a polyvalent, serum-derived antibody.

Claim 17

In an immunoassay for quantitative detection of a selected proteinaceous material in a fluid sample wherein said sample is contacted with an antibody capable of an immunobinding reaction with said proteinaceous material and wherein the amount of said proteinaceous material in said sample is determined by comparison of (a) the extent of immunobinding reaction of said antibody with sample components, to (b) the extent of immunobinding reaction of said antibody with a fixed quantity of a standard substance, the improvement comprising: employing as said antibody a monoclonal antibody immunoreactive with both the selected proteinaceous material and with a polypeptide substantially duplicative of an amino acid sequence extant in said proteinaceous material; and employing said polypeptide-as said standard substance.

Claim 18

In an immunoassay for quantitative detection of a selected proteinaceous material in a fluid sample wherein said sample is contacted with an antibody capable of an immunobinding reaction with said proteinaceous material and wherein the amount of said proteinaceous material in said sample is determined by comparison of (a) the extent of immunobinding reaction of said antibody with sample components, to (b) the extent of immunobinding reaction of said antibody with a fixed quantity of a standard substance, the improvement comprising: employing as said antibody a monoclonal antibody immunoreactive with both the selected proteinaceous material and with a polypeptide substantially duplicative of an amino acid sequence extant in said proteinaceous material; and employing said polypeptide-as said standard substance.

Claim 19

An improved process according to claim 14 for quantitative detection of erythropoietin.

Claim 20

An improved process according to claim 14 wherein said amino acid sequence of said polypeptide comprises: wherein R and R' are the same or different and are selected from the group consisting of hydrogen or one or more amino acid residues.

Claim 21

An improved process according to claim 14 wherein said amino acid sequence of said polypeptide comprises: RNH-Ala-Pro-Pro-Arg-Leu-Ile-Cys-Asp-Ser-Arg-Val-Leu-Glu-Arg-Tyr-Leu-Leu-Glu-Ala-Lys-COOR', wherein R and R' are the same or different and are selected from the group consisting of hydrogen or one or more amino acid residues.

Claim 22

A process for isolation of a selected proteinaceous material from a fluid, said process comprising: (1) immobilizing on a solid substrate a monoclonal antibody immunoreactive with said selected proteinaceous material and with a polypeptide which is substantially duplicative of a sequence of amino acids extant in said proteinaceous material, said antibody characterized as having a higher immunological affinity for said polypeptide than for said proteinaceous material; (2) contacting said fluid with said immobilized antibody, whereby the selected proteinaceous material is immobilized through immunobinding to said antibody; and (3) eluting said proteinaceous material from immunobinding association with said immobilized antibody by means of contact with a solution of said polypeptide for which said antibody has a higher immunological affinity.

Claim 23

A process for isolation of a selected proteinaceous material from a fluid, said process comprising: (1) immobilizing on a solid substrate a monoclonal antibody immunoreactive with said selected proteinaceous material and with a polypeptide which is substantially duplicative of a sequence of amino acids extant in said proteinaceous material, said antibody characterized as having a higher immunological affinity for said polypeptide than for said proteinaceous material; (2) contacting said fluid with said immobilized antibody, whereby the selected proteinaceous material is immobilized through immunobinding to said antibody; and (3) eluting said proteinaceous material from immunobinding association with said immobilized antibody by means of contact with a solution of said polypeptide for which said antibody has a higher immunological affinity.

Claim 24

A process according to claim-18 wherein said proteinaceous material is erythropoietin.

Claim 25

A process according to claim 18 wherein the amino acid sequence of said polypeptide comprises: RNH-Ser-Arg-Val-Leu-Glu-Arg-COOR', wherein R and R ' are the same or different and are selected from the group consisting of hydrogen or one or more amino acid residues.

Claim 26

A process according to claim 18 wherein the amino acid sequence of said polypeptide comprises: RN H-Ala-Pro-Pro-Arg-Leu-Ile-Cys-Asp-Ser-Arg-Val-Leu-Glu-Arg-Tyr-Leu-Leu-Glu-Ala-Lys-COOR', wherein R and R ' are the same or different and are selected from the group consisting of hydrogen or one or more amino acid residues.