US4254095A
Expired / Lapsed
US
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Radioimmunoassay for erythropoietin
Assignee: Research Corp
•
Filing: 1978-04-27
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Granted: 1981-03-03
•
Expires: 1998-04-27
Abstract
Abstract A kit and method for the radioimmunoassay (RIA) for erythropoietin. Also, a method for purifying erythropoietin and purified labelled erythropoietin-conjugate for use as a tracer in the RIA.
Inventors
- 👤 James W. Fisher
- 👤 Arvind B. Rege
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-26 11:39
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/74; G01N33/746; C; C07; C07K; C07K14/00; C07K14/435; C07K14/475; C07K14/505; A; A61; A61K; A61K38/00; Y; Y10; Y10S; Y10S436/00; Y10S436/804; Y10S436/807; Y10S436/808; Y10S436/815; Y10S436/827
Claims Analysis
Total Claims: 59
Claim 1
Independent Claim
79 words
In a radioimmunoassay of erythropoietin comprising (a) reacting binder containing a fixed number of antibody binding sites with a mixture of labelled and unlabelled erythropoietin; (b) separating bound erythropoietin from free erythropoietin; and (c) determining the percent labelled erythropoietin bound; the improvement comprising the use of labelled erythropoietin prepared by the conjugation labelling technique wherein the labelled erythropoietin has been purified by immunosorption with anti-erythropoietin antiserum that has been preadsorbed with human proteins that are low in erythropoietin content.
Claim 2
Dependent Claim
79 words
In a radioimmunoassay of erythropoietin comprising (a) reacting binder containing a fixed number of antibody binding sites with a mixture of labelled and unlabelled erythropoietin; (b) separating bound erythropoietin from free erythropoietin; and (c) determining the percent labelled erythropoietin bound; the improvement comprising the use of labelled erythropoietin prepared by the conjugation labelling technique wherein the labelled erythropoietin has been purified by immunosorption with anti-erythropoietin antiserum that has been preadsorbed with human proteins that are low in erythropoietin content.
Claim 3
Dependent Claim
25 words
A radioimmunoassay in accordance with claim 1 wherein the human proteins are selected from the group consisting of human serum proteins and human urinary proteins.
Claim 4
Dependent Claim
14 words
A radioimmunoassay according to claim 2 wherein the human proteins are human serum proteins.
Claim 5
Dependent Claim
16 words
A radioimmunoassay according to claim 3 wherein the human serum proteins are substantially free of erythropoietin.
Claim 6
Dependent Claim
16 words
A radioimmunoassay according to claim 4 wherein the human serum protein is polycythemia vera serum protein.
Claim 7
Dependent Claim
21 words
A radioimmunoassay in accordance with claim 1 wherein the erythropoietin antiserum used is selected from whole erythropoietin antiserum and antierythropoietin immunoglobulins.
Claim 8
Dependent Claim
14 words
A radioimmunoassay according to claim 6 wherein the human protein is human urinary protein.
Claim 9
Dependent Claim
14 words
A radioimmunoassay in accordance with claim 7 wherein the erythropoietin antiserum is antierythropoietin immunoglobulin.
Claim 10
Dependent Claim
19 words
A radioimmunoassay in accordance with claim 8 wherein the erythropoietin antiserum is the immunoglobulin fraction prepared by Rivanol precipitation.
Claim 11
Dependent Claim
27 words
A radioimmunoassay in accordance with claim 8 wherein the erythropoietin antiserum is the IgA fraction prepared by fractionation on dextran 2-(diethylamino)ethyl 2[[-(diethylamino)-ethyl]-diethylammonio]ethyl ether chloride hydrochloride epichlorohydrin cross-linked.
Claim 12
Dependent Claim
21 words
A radioimmunoassay in accordance with claim 5 wherein the erythropoietin antiserum used is selected from whole erythropoietin antiserum and antierythropoietin immunoglobulins.
Claim 13
Dependent Claim
14 words
A radioimmunoassay in accordance with claim 11 wherein the erythropoietin antiserum is antierythropoietin immunoglobulins.
Claim 14
Dependent Claim
19 words
A radioimmunoassay in accordance with claim 12 wherein the erythropoietin antiserum is the immunoglobulin fraction prepared by Rivanol precipitation.
Claim 15
Dependent Claim
21 words
A radioimmunoassay in accordance with claim 12 wherein the erythropoietin antiserum is the IgA fraction prepared by fractionation on DEAE sephadex.
Claim 16
Dependent Claim
13 words
A radioimmunoassay in accordance with claim 1 wherein the erythropoietin conjugate is lyophillized.
Claim 17
Dependent Claim
13 words
A radioimmunoassay in accordance with claim 5 wherein the erythropoietin conjugate is lyophillized.
Claim 18
Dependent Claim
13 words
A radioimmunoassay in accordance with claim 12 wherein the erythropoietin conjugate is lyophillized.
Claim 19
Dependent Claim
43 words
An erythropoietin-conjugate having the structure. ##STR1## wherein ESF is erythropoietin, lys is lysine and 125 I is iodine-125, being substantially free of urinary proteins, having a specific activity of between 40 and 55 microcuri/microgram and an immunoreactivity of between 75 and 90 percent.
Claim 20
Dependent Claim
43 words
An erythropoietin-conjugate having the structure. ##STR1## wherein ESF is erythropoietin, lys is lysine and 125 I is iodine-125, being substantially free of urinary proteins, having a specific activity of between 40 and 55 microcuri/microgram and an immunoreactivity of between 75 and 90 percent.
Claim 21
Dependent Claim
36 words
An antigen-antibody complex comprising (1) purified erythropoietin that has been conjugated with 125 Iodinated p-hydroxyphenylpropionic acid N-hydroxysuccinimidyl ester and (2) antiserum to erythropoietin that has been adsorbed with human proteins that are low in erythropoietin content.
Claim 22
Dependent Claim
36 words
An antigen-antibody complex comprising (1) purified erythropoietin that has been conjugated with 125 Iodinated p-hydroxyphenylpropionic acid N-hydroxysuccinimidyl ester and (2) antiserum to erythropoietin that has been adsorbed with human proteins that are low in erythropoietin content.
Claim 23
Dependent Claim
36 words
An antigen-antibody complex according to claim 19 wherein the human proteins are selected from the group consisting of human serum proteins that are low in erythropoietin content and human urinary proteins are low in erythropoietin content.
Claim 24
Dependent Claim
15 words
An antigen-antibody complex according to claim 20 wherein the human proteins are human serum protein.
Claim 25
Dependent Claim
17 words
An antigen-antibody complex according to claim 20 wherein the human serum proteins are substantially free of erythropoietin.
Claim 26
Dependent Claim
17 words
An antigen-antibody complex according to claim 22 wherein the human serum protein is polycythemia vera serum protein.
Claim 27
Dependent Claim
16 words
An antigen-antibody complex according to claim 23 wherein the polycythemia vera serum proteins are erythropoietin free.
Claim 28
Dependent Claim
26 words
An antigen-antibody complex according to claim 19 wherein the antiserum is selected from the group consisting of whole erythropoietin antiserum and the immunoglobulin fraction of antiserum.
Claim 29
Dependent Claim
16 words
An antigen-antibody complex according to claim 25 wherein the erythopoietin antiserum is the antierythropoietin immunoglobulin fraction.
Claim 30
Dependent Claim
19 words
An antigen-antibody complex according to claim 26 wherein the erythropoietin antiserum is the immunoglobulin fraction prepared by Rivano precipitation.
Claim 31
Dependent Claim
27 words
An antigen-antibody complex according to claim 26 wherein the erythropoietin antiserum is the IgA fraction prepared by fractionation on dextran 2-(diethylamino)ethyl 2-[[-(diethylamino)-ethyl]-diethylam(monio]ethyl ether chloride hydrochloride epichlorohydrin cross-linked.
Claim 32
Dependent Claim
26 words
An antigen-antibody complex according to claim 23 wherein an antiserum is slected from the group consisting of whole erythropoietin antiserum and the immunoglobulin fraction of antiserum.
Claim 33
Dependent Claim
16 words
An antigen-antibody complex according to claim 29 wherein the erythropoietin antiserum is the anti-erythropoietin immunoglobulin fraction.
Claim 34
Dependent Claim
19 words
An antigen-antibody complex according to claim 30 wherein the erythropoietin antiserum is the immunoglobulin fraction prepared by Rivanol precipitation.
Claim 35
Dependent Claim
27 words
An antigen-antibody complex according to claim 30 wherein the erythropoietin antiserum is the IgA fraction prepared by fractionation on dextran 2-(diethylamino)ethyl 2-[[(diethylamino)-ethyl]-diethylamino]ethyl ether chloride hydrochloride epichlorohydrin cross-linked.
Claim 36
Dependent Claim
64 words
A process for purifying a erythropoietin-conjugate having the formula ##STR2## wherein ESF is erythropoietin, lys is lysine and 125 I is iodine -125, which comprises adsorption and dissorption of said conjugate with erythropoietin antiserum selected from the group consisting of whole erythropoietin anti-serum and the immunoglobulin fraction of erythropoietin antiserum, said antiserum having been absorbed with human proteins that are low in erythropoietin content.
Claim 37
Dependent Claim
64 words
A process for purifying a erythropoietin-conjugate having the formula ##STR2## wherein ESF is erythropoietin, lys is lysine and 125 I is iodine -125, which comprises adsorption and dissorption of said conjugate with erythropoietin antiserum selected from the group consisting of whole erythropoietin anti-serum and the immunoglobulin fraction of erythropoietin antiserum, said antiserum having been absorbed with human proteins that are low in erythropoietin content.
Claim 38
Dependent Claim
25 words
A process according to claim 33 wherein the human serum proteins are selected from the group consisting of human serum proteins and human urinary proteins.
Claim 39
Dependent Claim
14 words
A process according to claim 34 wherein the human proteins are human serum proteins.
Claim 40
Dependent Claim
17 words
A process according to claim 35 wherein the human serum proteins are human polycythemic vera serum proteins.
Claim 41
Dependent Claim
15 words
A process according to claim 36 wherein the polycythemic vera serum protein is erythropoietin free.
Claim 42
Dependent Claim
24 words
A process according to claim 37 wherein the erythopoietin antiserum used is selected from the group consisting of whole erythropoietin antiserum and antierythropoietin immunoglobulins.
Claim 43
Dependent Claim
13 words
A process according to claim 38 wherein the erythropoietin antiserum is antierythropoietin immunoglobulins.
Claim 44
Dependent Claim
18 words
A process according to claim 39 wherein the erythropoietin antiserum is the immunoglobulin fraction prepared by Rivanol precipitation.
Claim 45
Dependent Claim
26 words
A process according to claim 39 wherein the erythropoietin antiserum is the IgA fraction prepared by fractionation on dextran 2-(diethylamino)ethyl 2-[[-(diethylamino)ethyl]-diethylammonio]ethyl ether chloride hydrochloride epichlorohydrin cross-linked.
Claim 46
Dependent Claim
23 words
A process according to claim 34 wherein the erythropoietin antiserum has been adsorbed with human urinary proteins that are low in erythropoietin content.
Claim 47
Dependent Claim
35 words
An antigen-binder complex comprising (a) binder-labelled erythropoietin and (b) binder-unlabelled erythropoietin. wherein the binder-labelled erythropoietin has been purified by immunosorption with antiserum that has been preadsorbed with human proteins that are low in erythropoietin content.
Claim 48
Dependent Claim
35 words
An antigen-binder complex comprising (a) binder-labelled erythropoietin and (b) binder-unlabelled erythropoietin. wherein the binder-labelled erythropoietin has been purified by immunosorption with antiserum that has been preadsorbed with human proteins that are low in erythropoietin content.
Claim 49
Dependent Claim
28 words
An antigen-binder complex according to claim 43 wherein the labelled erythropoietin has the structure ##STR3## wherein ESF is erythropoietin, lys is lysine and 125 I is iodine -125.
Claim 50
Dependent Claim
14 words
An antigen binder complex according to claim 44 wherein the binder is anti-erythopoietin antiserum.
Claim 51
Dependent Claim
18 words
An antigen-binder complex according to claim 45 wherein the binder consists of the immunoglobulin fraction of erythropoietin antiserum.
Claim 52
Dependent Claim
133 words
A kit for the radioimmunoassay of erythropoietin comprising a container having within the container a multiplicity of smaller containers and contents said contents comprising (a) lyophillized assay buffer concentrate; (b) lyophillized rabbit anti-erythropoietin antiserum; (c) goat anti-rabbit gamma-globulin; (d) erythropoietin standards; and (e) erythropoietin that has been conjugated with a moniodinated succimidyl ester of 3-(4-hydroxyphenyl)-propionic acid wherein the erythropoietin conjugate has been purified by immunosorption with antiserum that has been preadsorbed with human proteins that are low in erythropoietin content and with the proviso that each item is of such quantity that when all the items are brought together at a proper pH, dilution and temperature in the presence of an erythropoietin sample of human blood serum or plasma, antibodies in the antiserum bind to both labelled erythropoietin and erythropoietin in the sample.
Claim 53
Dependent Claim
133 words
A kit for the radioimmunoassay of erythropoietin comprising a container having within the container a multiplicity of smaller containers and contents said contents comprising (a) lyophillized assay buffer concentrate; (b) lyophillized rabbit anti-erythropoietin antiserum; (c) goat anti-rabbit gamma-globulin; (d) erythropoietin standards; and (e) erythropoietin that has been conjugated with a moniodinated succimidyl ester of 3-(4-hydroxyphenyl)-propionic acid wherein the erythropoietin conjugate has been purified by immunosorption with antiserum that has been preadsorbed with human proteins that are low in erythropoietin content and with the proviso that each item is of such quantity that when all the items are brought together at a proper pH, dilution and temperature in the presence of an erythropoietin sample of human blood serum or plasma, antibodies in the antiserum bind to both labelled erythropoietin and erythropoietin in the sample.
Claim 54
Dependent Claim
24 words
A kit according to claim 47 wherein the human proteins are selected from the group consisting of human serum proteins and human urinary proteins.
Claim 55
Dependent Claim
14 words
A kit according to claim 48 wherein the human proteins are human serum proteins.
Claim 56
Dependent Claim
16 words
A kit according to claim 49 wherein the human serum protein is polycythemic vera sera proteins.
Claim 57
Dependent Claim
15 words
A kit according to claim 50 wherein the polycythemic vera serum proteins are erythropoietin free.
Claim 58
Dependent Claim
40 words
A kit according to claim 47 wherein a means for holding instructions for use of the smaller container contents in a radioimmunoassay for determining the level of erythropoietin in an unknown sample is within or affixed to the said container.
Claim 59
Dependent Claim
15 words
A kit according to claim 47 wherein a top wholly or partially covers the container.
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1984-03-02 | 1984-09-01 | 1985-03-03 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1988-03-02 | 1988-09-01 | 1989-03-03 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1992-03-02 | 1992-09-01 | 1993-03-03 | $7700.00 | Window Closed / Lapsed |