US4548908A
Expired / Lapsed
US
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Competitive immunofluorescence assay and test kit
Assignee: PARAGON DIAGNOSTICS
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Filing: 1983-05-27
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Granted: 1985-10-22
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Expires: 2003-05-27
Abstract
Abstract Competitive immunofluorescence assays for antigens in which immune complexes are precipitated with a nonfluorescent, nonlight-scattering precipitant such as polyethylene glycol and the resulting immunoprecipitate is dissolved with a nonfluorescent solvent of low ionic strength that maintains the pH of the solution substantially constant for immunofluorescence intensity reading. The assays are carried out by incubating the sample with fluorescent-labeled antigen, anti-antigen antibody, and a secondary antibody to the anti-antigen antibody followed by addition of the precipitant to form an immunoprecipitate. The precipitate is separated by centrifuging, dissolved in the solvent, and the immunofluorescence intensity of the solution is read with a fluorometer and compared to a standard curve.
Inventors
- 👤 Naomi Kameda
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-03 21:58
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/58; G01N33/582; G01N33/53; G01N33/536; G01N33/537; G01N33/539; G01N33/94; Y; Y10; Y10S; Y10S436/00; Y10S436/80; Y10S436/807; Y10S436/808; Y10S436/815; Y10S436/817; Y10S436/826
Claims Analysis
Total Claims: 16
Claim 1
Independent Claim
104 words
A competitive immunofluorescence assay for determining the amount of an antigen in a sample suspected of containing the antigen comprising: (a) incubating the sample with a solution of a fluorescent-labeled antigen, anti-antigen antibody, and an antibody against the anti-antigen antibody; (b) adding polyethylene glycol to the incubation mixture in amount effective to form an immunoprecipitate; (c) separating the immunoprecipitate and dissolving the immunoprecipitate in a nonfluorescent solvent that has a low ionic strength and maintains the pH of the resulting solution substantially constant; and (d) measuring the fluorescence intensity of the solution of step (c) and comparing said fluorescence intensity to a standard curve.
Claim 2
Dependent Claim
104 words
A competitive immunofluorescence assay for determining the amount of an antigen in a sample suspected of containing the antigen comprising: (a) incubating the sample with a solution of a fluorescent-labeled antigen, anti-antigen antibody, and an antibody against the anti-antigen antibody; (b) adding polyethylene glycol to the incubation mixture in amount effective to form an immunoprecipitate; (c) separating the immunoprecipitate and dissolving the immunoprecipitate in a nonfluorescent solvent that has a low ionic strength and maintains the pH of the resulting solution substantially constant; and (d) measuring the fluorescence intensity of the solution of step (c) and comparing said fluorescence intensity to a standard curve.
Claim 3
Dependent Claim
25 words
The competitive immunofluorescence assay of claim 1 wherein the antigen is T 3 , T 4 , digoxin, gentamicin, amikacin, tobramycin, kanamycin, netilmicin, or theophylline.
Claim 4
Dependent Claim
14 words
The immunofluorescence assay of claim 1 wherein the fluorescent-labeled antigen is a fluorescein-labeled antigen.
Claim 5
Dependent Claim
19 words
The competitive immunofluorescence assay of claim 1 wherein the solvent maintains the pH of the solution above about 8.
Claim 6
Dependent Claim
13 words
The competitive immunofluorescence assay of claim 1 wherein the solvent is a buffer.
Claim 7
Dependent Claim
24 words
The competitive immunofluorescence assay of claim 1 wherein the solvent is an aqueous solution of an alkali metal hydroxide or an alkali metal borate.
Claim 8
Dependent Claim
22 words
The competitive immunofluorescence assay of claim 1 wherein the solvent is aqueous sodium hydroxide having an ionic strength less than about 0.04.
Claim 9
Dependent Claim
14 words
The competitive immunofluorescence assay of claim 7 wherein the ionic strength is approximately 0.03.
Claim 10
Dependent Claim
25 words
The competitive immunofluorescence assay of claim 1 wherein the solvent is an aqueous solution of sodium borate having an ionic strength less than about 0.2.
Claim 11
Dependent Claim
20 words
The competitive immunofluorescence assay of claim 1 wherein a minimum volume of the solvent is used to dissolve the immunoprecipitate.
Claim 12
Dependent Claim
47 words
The competitive immunofluorescence assay of claim 2 wherein the fluorescent labeled antigen is fluorescein isothiocyanate labeled antigen, the solvent is aqueous sodium hydroxide having an ionic strength less than about 0.04 or sodium borate having an ionic strength less than about 0.2, said pH is above 8.
Claim 13
Dependent Claim
77 words
A test kit for carrying out the assay of claim 1, comprising: (a) a container containing a fluorescent-labeled antigen reagent; (b) a container containing anti-antigen antibody reagent; (c) a container containing an antibody against the anti-antigen antibody; (d) a container containing a polyethylene glycol; and (e) a container containing a nonfluorescent solvent for dissolving immunoprecipitates of the immunoprecipitant that has a low ionic strength and the ability to maintain the pH of the immunoprecipate solution substantially constant.
Claim 14
Dependent Claim
19 words
The test kit of claim 12 wherein the solvent has the ability to maintain said pH above about 8.
Claim 15
Dependent Claim
36 words
The test kit of claim 13 wherein the solvent is aqueous sodium hydroxide having an ionic strength less than about 0.04 or an aqueous solution of sodium borate having an ionic strength less than about 0.2.
Claim 16
Dependent Claim
14 words
The test kit of claim 14 wherein the fluorescent-labeled antigen is a fluorescein-labeled antigen.
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) | 1988-10-21 | 1989-04-22 | 1989-10-22 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1992-10-21 | 1993-04-22 | 1993-10-22 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1996-10-21 | 1997-04-22 | 1997-10-22 | $7700.00 | Window Closed / Lapsed |