US4287300A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Charge effects in enzyme immunoassays
Assignee: Siemens Healthcare Diagnostics GmbH Germany
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Filing: 1979-07-26
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Granted: 1981-09-01
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Expires: 1999-07-26
Abstract
Abstract A method for determining a member of a specific binding pair-ligand and receptor (antiligand). Reagents employed include a first modified member which provides an electrical field due to the presence of a plurality of ionic charges and a second modified member labeled with a component of a signal producing system, which system may have one or more components. The average proximity in the assay medium of the first and second modified members is related to the amount of analyte, where the observed signal from the signal producing system is related to the effect of the electrical field on the signal producing system. Also, compositions are provided, as well as reagents, in predetermined ratios for optimizing the signal response to variations in analyte concentration.
Inventors
- 👤 Ian Gibbons
- 👤 Gerald L. Rowley
- 👤 Edwin F. Ullman
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-24 22:38
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/53; G01N33/536; G01N33/542; Y; Y10; Y10S; Y10S435/00; Y10S435/81; Y10S436/00; Y10S436/80; Y10S436/805; Y10S436/806; Y10S436/811; Y10S436/815; Y10S436/82
Claims Analysis
Total Claims: 18
Claim 1
Independent Claim
220 words
A method for determining the presence of an analyte in a sample suspected of containing said analyte, said analyte being a member of a specific binding pair consisting of ligand and antiligand, wherein a detectible signal is obtained related to the amount of analyte by employing a signal producing system comprising an enzyme labeled specific binding member and a polyionic charged specific binding member having sufficient charge to attract an oppositely charged macromolecular substrate or cofactor, where the detectible signal level is related to the average proximity of said charged member to said enzyme labeled member, said method comprising: combining in an aqueous assay medium; (1) said sample; (2) said charged specific binding member; (3) said enzyme labeled specific binding member; (4) substrates and cofactors for said enzyme wherein at least one substrate or cofactor for said enzyme is a charged macromolecular of opposite charge from said polyionic charged member, wherein at least one of (1), (2) and (3) is ligand and at least one of the remaining of (1), (2) and (3) is antiliand, so that a complex may be formed between said charged specific binding member and said enzyme labeled specific binding member; and determining the detectible signal produced by said enzyme as compared to the signal produced in an assay medium having a known amount of analyte.
Claim 2
Independent Claim
220 words
A method for determining the presence of an analyte in a sample suspected of containing said analyte, said analyte being a member of a specific binding pair consisting of ligand and antiligand, wherein a detectible signal is obtained related to the amount of analyte by employing a signal producing system comprising an enzyme labeled specific binding member and a polyionic charged specific binding member having sufficient charge to attract an oppositely charged macromolecular substrate or cofactor, where the detectible signal level is related to the average proximity of said charged member to said enzyme labeled member, said method comprising: combining in an aqueous assay medium; (1) said sample; (2) said charged specific binding member; (3) said enzyme labeled specific binding member; (4) substrates and cofactors for said enzyme wherein at least one substrate or cofactor for said enzyme is a charged macromolecular of opposite charge from said polyionic charged member, wherein at least one of (1), (2) and (3) is ligand and at least one of the remaining of (1), (2) and (3) is antiliand, so that a complex may be formed between said charged specific binding member and said enzyme labeled specific binding member; and determining the detectible signal produced by said enzyme as compared to the signal produced in an assay medium having a known amount of analyte.
Claim 3
Dependent Claim
17 words
A method according to claim 1, wherein said substrate is said charged macromolecule having enzymatically hydrolyzable bonds.
Claim 4
Dependent Claim
15 words
A method according to claim 2, wherein said charged macromolecular substrate has a positive charge.
Claim 5
Independent Claim
27 words
A method according to any of claims 1 to 3, wherein said aqueous assay medium is at a pH in the range of about 5 to 10.
Claim 6
Independent Claim
20 words
A method according to any of claims 1 to 3, wherein said charged member has a plurality of carboxy groups.
Claim 7
Independent Claim
20 words
A method according to any of claims 1 to 3, wherein said charged member has a plurality of phenolic groups.
Claim 8
Dependent Claim
99 words
A composition useful for the immunoassay of claim 1 comprising, a macromolecular charged substrate or coenzyme and modified members of a specific binding pair consisting of ligand and antiligand, wherein the modified members are an enzyme labeled member and a charged member substituted with a plurality of functionalities which under the conditions of the assay are sufficiently charged opposite to said charged macromolecular substrate or cofactor to attract said charged macromolecular substrate or cofactor, the relative amounts of said charged member and said enzyme-labeled member substantially optimize the response of the detectible signal level to variations in analyte concentration.
Claim 9
Dependent Claim
24 words
An assay composition according to claim 7, wherein said charged member is polycarboxyl substituted antiligand and said signal labeled member is β-galactosidase substituted antiligand.
Claim 10
Dependent Claim
26 words
An assay composition according to claim 7, wherein said macromolecular charged substrate is a positively charged substrate for said β-galactosidase having a plurality of o-nitrophenylgalactoside groups.
Claim 11
Dependent Claim
25 words
An assay composition according to claim 7, wherein said charged member is a polyphenolic substituted antiligand and said signal labeled member is β-galactosidase labeled antiligand.
Claim 12
Dependent Claim
26 words
An assay composition according to claim 10, wherein said charged macromolecular substrate is a positively charged substrate for said β-galactosidase having a plurality of o-nitrophenylgalactoside groups.
Claim 13
Independent Claim
167 words
A method for determining the presence of an antigenic protein in a sample suspected of containing said antigenic protein, said antigenic protein being a member of a specific binding pair consisting of said antigenic protein and antibody to said antigenic protein, wherein a detectible signal is obtained related to the amount of antigenic protein by employing a signal producing system including a labeled member, said labeled member being enzyme conjugated to said antibody, and a polyionic charged member, wherein said charged member is said antibody substituted with a plurality of carboxyl groups, where the detectible signal level is related to the average proximity of said charged member to said labeled member, said method comprising: combining in an aqueous assay medium: (1) said sample; (2) said charged member; (3) said signal labeled member; (4) said enzyme substrates and co-factors; and determining the detectible signal produced by said signal producing system as compared to the signal produced in an assay medium having a known amount of said protein antigen.
Claim 14
Independent Claim
167 words
A method for determining the presence of an antigenic protein in a sample suspected of containing said antigenic protein, said antigenic protein being a member of a specific binding pair consisting of said antigenic protein and antibody to said antigenic protein, wherein a detectible signal is obtained related to the amount of antigenic protein by employing a signal producing system including a labeled member, said labeled member being enzyme conjugated to said antibody, and a polyionic charged member, wherein said charged member is said antibody substituted with a plurality of carboxyl groups, where the detectible signal level is related to the average proximity of said charged member to said labeled member, said method comprising: combining in an aqueous assay medium: (1) said sample; (2) said charged member; (3) said signal labeled member; (4) said enzyme substrates and co-factors; and determining the detectible signal produced by said signal producing system as compared to the signal produced in an assay medium having a known amount of said protein antigen.
Claim 15
Dependent Claim
26 words
A method according to claim 12, wherein said enzyme is β-galactosidase and said substrate is a positively charged macromolecular substrate having a plurality of o-nitrophenylgalactoside groups.
Claim 16
Independent Claim
167 words
A method for determining the presence of an antigenic protein in a sample suspected of containing said antigenic protein, said antigenic protein being a member of a specific binding pair consisting of said antigenic protein and antibody to said antigenic protein, wherein a detectible signal is obtained related to the amount of antigenic protein by employing a signal producing system including a labeled member, said labeled member being enzyme conjugated to said antibody, and a polyionic charged member, wherein said charged member is said antibody substituted with a plurality of phenolic groups, where the detectible signal level is related to the average proximity of said charged member to said labeled member, said method comprising: combining in an aqueous assay medium: (1) said sample; (2) said charged member; (3) said signal labeled member; (4) said enzyme substrates and co-factors; and determining the detectible signal produced by said signal producing system as compared to the signal produced in an assay medium having a known amount of said protein antigen.
Claim 17
Independent Claim
167 words
A method for determining the presence of an antigenic protein in a sample suspected of containing said antigenic protein, said antigenic protein being a member of a specific binding pair consisting of said antigenic protein and antibody to said antigenic protein, wherein a detectible signal is obtained related to the amount of antigenic protein by employing a signal producing system including a labeled member, said labeled member being enzyme conjugated to said antibody, and a polyionic charged member, wherein said charged member is said antibody substituted with a plurality of phenolic groups, where the detectible signal level is related to the average proximity of said charged member to said labeled member, said method comprising: combining in an aqueous assay medium: (1) said sample; (2) said charged member; (3) said signal labeled member; (4) said enzyme substrates and co-factors; and determining the detectible signal produced by said signal producing system as compared to the signal produced in an assay medium having a known amount of said protein antigen.
Claim 18
Dependent Claim
26 words
A method according to claim 14, wherein said enzyme is β-galactosidase and said substrate is a positively charged macromolecular substrate having a plurality of o-nitrophenylgalactoside groups.
References Cited (Backward Citations)
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1984-08-31 | 1985-03-02 | 1985-09-01 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1988-08-31 | 1989-03-02 | 1989-09-01 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1992-08-31 | 1993-03-02 | 1993-09-01 | $7700.00 | Window Closed / Lapsed |