Dipstick immunoassay device

• US 5,656,448 A
• Filed: 01/14/1994
• Issued: 08/12/1997
• Est. Priority Date: 01/10/1990
• Status: Expired due to Fees

• Current Assignee: Princeton Biomeditech Corporation
• Original Assignee: Princeton Biomeditech Corporation
• Inventors: Lee, Dong Joon, Colanduoni, John A., Ok, Chiyoung, Kang, Walter J., Kang, Jemo, Youn, Byungwoo
• Primary Examiner(s): Housel, James C.
• Assistant Examiner(s): Wolski, Susan C.
• Application Number:
  US08/182,512
• Time in Patent Office: 1,306 Days
• Field of Search: 422/56, 422/57, 435/7.9, 435/7.92-7.95, 435/18, 435/21, 435/25-26, 435/28, 435/962, 435/963, 435/970, 435/975, 435/287.1, 435/287.2, 435/287.7, 435/805, 435/810, 436/518, 436/527, 436/530, 436/531, 436/808, 436/810, 436/169
• US Class Current: 435/7.94
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/54386 Analytical elements
• CPC Class Codes: G01N 33/76 Human chorionic gonadotropin including luteinising hormone, follicle stimulating hormone, thyroid stimulating hormone or their receptors
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 435/81 Packaged device or kit
• CPC Class Codes: Y10S 435/962 Prevention or removal of interfering materials or reactants or other treatment to enhance results, e.g. determining or preventing nonspecific binding
• CPC Class Codes: Y10S 435/963 Methods of stopping an enzyme reaction or stabilizing the test materials
• CPC Class Codes: Y10S 435/97 Test strip or test slide
• CPC Class Codes: Y10S 435/975 Kit
• CPC Class Codes: Y10S 436/808 Automated or kit
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick

Abstract

The invention pertains to dipstick immunoassay devices. The device comprises a base member and a single, combined sample contact zone and test zone, wherein the test zone incorporates the use of symbols to detect analytes in a sample of biological fluid. A first immunological component, an anti-immunoglobulin capable of binding to an enzyme-labeled antibody, is immobilized in a control indicia portion. A second immunological component, capable of specifically binding to a target analyte which is bound to the enzyme-labeled antibody to form a sandwich complex, is immobilized in a test indicia portion. The enzyme-labeled antibody produces a visual color differential between a control indicia portion and a non-indicia portion in the test zone upon contact with a substrate. The device additionally includes a first polyol and a color differential enhancing component selected from the group consisting of an inhibitor to the enzyme and a competitive secondary substrate for the enzyme distributed throughout the non-indicia portion of the test zone.

Claims

1. A dipstick immunoassay device comprising:

a base member;

a single, combined sample contact zone and test zone, defined on said base member, said test zone including a control indicia portion, a test indicia portion and a non-indicia portion;

a first immobilized immunological component being disposed solely in said control indicia portion, said first immunological component being an anti-immunoglobulin which is capable of binding to an enzyme-labeled antibody, said enzyme-labeled antibody producing a visual color differential between said control indicia portion and said non-indicia portion upon contact with a substrate;

a second immobilized immunological component being disposed solely in said test indicia portion, said second immunological component being capable of specifically binding to a target analyte which is bound to said enzyme-labeled antibody to form a sandwich complex, said enzyme-labeled antibody producing a visually discernable difference between said indicia portions and said non-indicia portion, upon contact with a substrate;

a first polyol distributed throughout said non-indicia portion of said test zone; and

a color differential enhancing component selected from the group consisting of an inhibitor to said enzyme and a competitive secondary substrate for said enzyme, said component distributed throughout said non-indicia portion of said test zone.

2. The assay device according to claim 1 wherein said second immobilized component is an antibody, antigen or hapten.

3. The assay device according to claim 2 wherein said second immobilized component is an antibody.

4. The assay device according to claim 1 wherein said enzyme is alkaline phosphatase, horse radish peroxidase, lysozyme, glucose-6-phosphate dehydrogenase, lactate dehydrogenase, urease, glucose oxidase, or cholesterol oxidase.

5. The assay device according to claim 4 wherein said enzyme is alkaline phosphatase.

6. The assay device according to claim 1 wherein said enzyme inhibitor is an amino acid or a metal chelator.

7. The assay device according to claim 1 wherein said inhibitor is selected from the group consisting of cysteine, histidine, L-phenylalanine, β

-phenyl-β

-alanine, p-fluorophenylalanine, tyrosine, iodobenzoate, iodoacetamide, citrate, ethylene diamine tetraacetic acid, borate, arsenate, phosphate, orthophosphate, carbonate, polyestradiol phosphate, pyrophosphate, and polyphloretin phosphate.

8. The assay device according to claim 1 wherein said competitive secondary substrate is p-nitrophenyl phosphate.

9. The assay device according to claim 1 wherein the target analyte is an antigen, an antibody, or a hapten.

10. The assay device according to claim 1, wherein said test indicia portion and said control indicia portion are each in the form of a bar or a dot.

11. The assay device according to claim 1 wherein said base member is comprised of a member selected from the group consisting of plastic and glass.

12. The assay device according to claim 1 wherein said test zone is comprised of a member selected from the group consisting of nylon, nitrocellulose, cellulose, fiberglass, polysulfone, polyvinylidene difluoride, and polyester.

13. The assay device according to claim 1 wherein said second immunological component is an antibody specific for said analyte and being conjugated to alkaline phosphatase and said inhibitor is a member selected from the group consisting of cysteine, histidine, L-phenylalanine, β

-phenyl-β

-alanine, p-fluorophenylalanine and tyrosine.

14. A dipstick immunoassay device comprising:

a base member;

a single, combined sample contact zone and test zone, defined on said base member, said test zone including a control indicia portion, a test indicia portion and a non-indicia portion;

a first immobilized antibody being disposed solely in said control indicia portion, said first immobilized antibody being an anti-immunoglobulin antibody which is capable of binding to an alkaline phosphatase-labeled antibody, said alkaline phosphatase-labeled antibody producing a visual color differential between said control indicia portion and said non-indicia portion upon contact with a substrate;

a second immobilized antibody being disposed solely in said test indicia portion, said second antibody being capable of specifically binding to a target analyte which is bound to said alkaline phosphatase-labeled antibody to form a sandwich complex, said alkaline phosphatase-labeled antibody producing a visually discernable difference between said indicia portions and said non-indicia portion, upon contact with a substrate;

a first polyol distributed throughout said non-indicia portion of said test zone; and

a color differential enhancing component selected from the group consisting of an inhibitor to said alkaline phosphatase and a competitive secondary substrate for said alkaline phosphatase, said component distributed throughout said non-indicia portion of said test zone.

15. The assay device according to claim 14 wherein said inhibitor is an amino acid or a chelating agent.

16. The assay device according to claim 14 wherein said inhibitor is cysteine, histidine, L-phenylalanine, β

-phenyl-β

-alanine, p-fluorophenylalanine, tyrosine, iodobenzoate, iodoacetamide, ethylenediamine tetraacetic acid, citrate, orthophosphate, arsenate pyrophosphate, borate, carbonate, polyestradiol phosphate, polyphloretin phosphate or phosphate.

17. The assay device according to claim 14 wherein said competitive secondary substrate is p-nitrophenyl phosphate.

18. The assay device according to claim 14 wherein the target analyte is an antigen, an antibody, or a hapten.

19. The assay device according to claim 14 wherein said base member is comprised of a member selected from the group consisting of plastic and glass.

20. The assay device according to claim 14 wherein said test zone is comprised of a member selected from the group consisting of nylon, nitrocellulose, cellulose, fiberglass, polysulfone, polyvinylidene difluoride, and polyester.

21. The assay device according to claim 14 wherein said first polyol is polyvinylalcohol, carbohydrate, polyethylene glycol, sorbitol, polypropylene glycol, dextran, methyl cellulose, milk, or corn starch.

22. The assay device according to claim 14 wherein said first polyol is polyvinylalcohol.

23. The assay device according to claim 14, wherein said test indicia portion and said control indicia portion are each in the form of a bar or a dot.

24. A kit comprising

an assay device according to claim 14;

a first solution comprising alkaline phosphatase labeled antibody; and

a second solution comprising a substrate operable to form an insoluble colored product and phosphate ions upon contact with said alkaline phosphatase and a second polyol operable to bind said phosphate ions.

25. The kit according to claim 14 wherein said second polyol is propylene glycol, ethane glycol, 2-amino-2-methyl-1-propanol, or butane diol.

26. The kit according to claim 25 wherein said second polyol is 2-amino-2-methyl-1-propanol.