Polymeric assay film for direct colorimetric detection

• US 6,001,556 A
• Filed: 01/26/1996
• Issued: 12/14/1999
• Est. Priority Date: 11/13/1992
• Status: Expired due to Fees

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Spevak, Wayne, Nagy, Jon, Charych, Deborah
• Primary Examiner(s): Caputa, Anthony C.
• Assistant Examiner(s): BAKALYAR, HEATHER
• Application Number:
  US08/592,724
• Time in Patent Office: 1,418 Days
• Field of Search: 435/518, 435/531, 435/528, 435/7.1, 435/164, 435/4, 435/5, 435/6, 422/55, 422/57, 422/58, 422/82.05, 422/82.09, 427/2, 428/441, 428/462
• US Class Current: 435/5
• CPC Class Codes: B82Y 15/00 Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: Y10T 428/31645 Next to addition polymer from unsaturated monomers
• CPC Class Codes: Y10T 428/31649 Ester, halide or nitrile of addition polymer

Abstract

A lipid bilayer with affinity to an analyte, which directly signals binding by a changes in the light absorption spectra. This novel assay means and method has special applications in the drug development and medical testing fields. Using a spectrometer, the system is easily automated, and a multiple well embodiment allows inexpensive screening and sequential testing. This invention also has applications in industry for feedstock and effluent monitoring.

Claims

1. A polymerized bilayer film comprising:

a conjugated polymer backbone, linker groups, ligands having direct affinity for an analyte, wherein said ligands are carbohydrates, and ordering head groups, wherein said linker groups are covalently conjugated to said polymer backbone, wherein said ligands are covalently conjugated to said linker groups, wherein said ordering head groups are bound to the surface of said conjugated polymer backbone in positions not occupied by said linker groups, and wherein said polymerized bilayer film undergoes a detectable color change upon binding of said analyte to said ligands.

2. The bilayer film of claim 1, wherein said analyte comprises a pathogen.

3. The bilayer film of claim 2, wherein said pathogen is selected from the group consisting of influenza virus, herpes virus, human immunodeficiency virus, coronavirus, encephalomyelitis, chlamydia, rotavirus, polyomavirus, Streptococcus, Salmonella, sendai virus, mumps virus, Newcastle Disease virus, myxovirus, E. coli, encephalomyocarditis virus, and Plasmodium.

4. The bilayer film of claim 1, wherein said conjugated polymer backbone comprises lipid monomers containing chemical moieties selected from the group consisting of diacetylenes, acetylenes, alkenes, thiophenes, pyrroles, vinylpyridinium, and combinations thereof.

5. The bilayer film of claim 1, wherein said ordering head groups are selected from the group consisting of carboxylic acid and salts thereof, esters, hydroxyl groups, amine groups, and amide groups.

6. The bilayer film of claim 1, further comprising a solid support wherein said bilayer film is attached to said solid support.

7. The bilayer film of claim 6, wherein said solid support is selected from the group consisting of plastics, metals, and ceramics.

8. The bilayer film of claim 6, wherein said solid support comprises glass.

9. The bilayer film of claim 8, wherein said glass comprises a hydrophobized glass slide.

27. A method of making the polymerized bilayer film of claim 1, comprising the steps of:

a) providing;

i) ligands having direct affinity for an analyte, wherein said ligands are carbohydrates;

ii) linker groups containing first and second terminal ends;

iii) lipid monomers;

iv) lipid monomers comprising ordering head groups; and

v) a support surface

b) attaching said ligands to said lipid monomers, wherein said ligands are attached to said first end of said linker groups and said lipid monomers are attached to said second end of said linker groups to produce monomer-linear structural unit-ligand moieties;

c) mixing said monomer-linear structural unit-ligand moieties with a plurality of said lipid monomers comprising ordering head groups to form a mixture;

d) spreading said mixture of step c) on said support surface to form a bilayer film; and

e) polymerizing said bilayer film to form said polymerized bilayer film.

28. The method of claim 27, further comprising the step of transferring said polymerized bilayer film to a solid support.

29. The method of claim 27, wherein after step d), said bilayer film is compressed.

30. The method of claim 27, wherein said support surface is an aqueous surface.

31. A method for the direct detection of an analyte, comprising:

contacting the polymerized bilayer film of claim 1 with a sample suspected of containing an analyte; and

detecting a color change in said polymerized bilayer film, wherein detection of a color change is indicative of the presence of an analyte.

10. A polymerized bilayer film comprising:

a conjugated polymer backbone, linker groups, ligands having direct affinity for an analyte, wherein said ligands are sialic acid, and ordering head groups, wherein said linker groups are covalently conjugated to said polymer backbone, wherein said ligands are covalently conjugated to said linker groups, wherein said ordering head groups are bound to the surface of said conjugated polymer backbone in positions not occupied by said linker groups, and wherein said polymerized bilayer film undergoes a detectable color change upon binding of said analyte to said ligands.

11. The bilayer film of claim 10, wherein said analyte comprises a pathogen.

12. The bilayer film of claim 11, wherein said pathogen is selected from the group consisting of influenza virus, herpes virus, human immunodeficiency virus, coronavirus, encephalomyelitis, chlamydia, rotavirus, polyomavirus, Streptococcus, Salmonella, sendai virus, mumps virus, Newcastle Disease virus, myxovirus, E. coli, encephalomyocarditis virus, and Plasmodium.

13. The bilayer film of claim 10, wherein said conjugated polymer backbone comprises lipid monomers containing chemical moieties selected from the group consisting of diacetylenes, acetylenes, alkenes, thiophenes, pyrroles, vinylpyridinium, and combinations thereof.

14. The bilayer film of claim 10, wherein said ordering head groups are selected from the group consisting of carboxylic acid and salts thereof, esters, hydroxyl groups, amine groups, and amide groups.

15. The bilayer film of claim 10, further comprising a solid support wherein said bilayer film is attached to said solid support.

16. The bilayer film of claim 15, wherein said solid support is selected from the group consisting of plastics, metals, and ceramics.

17. The bilayer film of claim 15, wherein said solid support comprises glass.

18. The bilayer film of claim 17, wherein said glass comprises a hydrophobized glass slide.

19. A polymerized bilayer film comprising:

a conjugated polymer backbone, linker groups, ligands having direct affinity for an analyte, wherein said ligands are sialic acid, and ordering head groups, wherein said conjugated polymer backbone comprises a plurality of polymerized diacetylene monomers, wherein said linker groups are covalently conjugated to said diacetylene monomers, wherein said ligands are covalently conjugated to said linker groups, wherein said ordering head groups are bound to the surface of said conjugated polymer backbone in positions not occupied by said linker groups, and wherein said film undergoes a detectable color change upon binding of said analyte to said ligands.

20. The bilayer film of claim 19, wherein said analyte comprises a pathogen.

21. The bilayer film of claim 20, wherein said pathogen is selected from the group consisting of influenza virus, herpes virus, human immunodeficiency virus, coronavirus, encephalomyelitis, chlamydia, rotavirus, polyomavirus, Streptococcus, Salmonella, sendai virus, mumps virus, Newcastle Disease virus, myxovirus, E. coli, encephalomyocarditis virus, and Plasmodium.

22. The bilayer film of claim 19, wherein said ordering head groups are selected from the group consisting of carboxylic acid and salts thereof, esters, hydroxyl groups, amine groups, and amide groups.

23. The bilayer film of claim 19, further comprising a solid support selected from the group consisting of plastics, metals, and ceramics, wherein said bilayer film is attached to said solid support.

24. The bilayer film of claim 19, wherein said solid support comprises glass.

25. The bilayer film of claim 24, wherein said glass comprises a hydrophobized glass slide.

26. A polymerized bilayer film comprising:

a conjugated polymer backbone, linker groups, sialic acid groups, ordering head groups, and a support structure, wherein said conjugated polymer backbone comprises a plurality of polymerized diacetylene monomers, wherein said linker groups are covalently conjugated to said diacetylene monomers, wherein said sialic acid groups are covalently conjugated to said linker groups, wherein said ordering head groups are bound to the surface of said conjugated polymer backbone in positions not occupied by said linker groups, and wherein said polymerized bilayer film undergoes a detectable color change upon binding of said analyte to said sialic acid groups.