SUBSTRATES FOR MULTIPLEXED ASSAYS AND USES THEREOF

• US 20090253586A1
• Filed: 02/20/2009
• Published: 10/08/2009
• Est. Priority Date: 02/21/2008
• Status: Abandoned Application

• Current Assignee: Intuitive Biosciences, Inc.
• Original Assignee: Gentel Biosciences Inc. (Intuitive Biosciences, Inc.)
• Inventors: Bart, John C., Garcia, Bradley H., Nelson, Bryce P.
• Application Number:
  US12/390,074
• Publication Number:
  US 20090253586A1
• Time in Patent Office: Days
• US Class Current: 506/9
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 21/6452 Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/54306 Solid-phase reaction mechanisms
• CPC Class Codes: G01N 33/54393 Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding

Abstract

The present invention relates to novel methodologies for performing multiplexed assays for biological molecules such as proteins and nucleic acids. In particular, the present invention provides multiplexed assays using precipitating reagents and optically clear nitrocellulose-coated solid supports.

Claims

1. A method for performing a multiplexed assay, comprising:

a) contacting a substrate with a sample comprising a target molecule under conditions such that said target molecule binds to a capture molecule, wherein said substrate comprises an array of said capture molecules affixed to an optically clear coating of nitrocellulose on said substrate;

b) contacting said arrays with reagents under conditions such that a precipitate is formed where said target molecule is bound to said capture molecule.

2. The method of claim 1, further comprising the step of c) determining the presence of said precipitate in discrete regions on said array, wherein the presence of said precipitate is indicative of the presence of said target molecule in said sample.

3. The method of claim 2, further comprising the step of quantifying the level of said target molecule in said sample.

4. The method of claim 1, wherein said substrate is plastic.

5. The method of claim 1, wherein said substrate is glass.

6. The method of claim 1, wherein said precipitate is formed from the precipitate of a metallic compound upon the complex of said target molecule and said capture molecule.

7. The method of claim 6, wherein said metallic compound is a magnetic metallic compound.

8. The method of claim 6, wherein said precipitate is formed via a chemical reduction of silver in the presence of colloidal gold particles coupled to the bound target compound.

9. The method of claim 2, wherein said determining the presence of said precipitate comprises the use of a colorimetric reader.

10. The method of claim 9, wherein said reader is CCD or CMOS based.

11. The method of claim 1, wherein said array is selected from the group consisting of a 3″

×

1″

slide, a 96-well array plate, and a 384-well plate.

12. The method of claim 2, wherein said determining the presence of said precipitate comprises a detection assay selected from the group consisting of gold-catalyzed silver deposition, horseradish peroxidase, AP, and tyramide signal amplification.

13. The method of claim 1, wherein said capture molecule is selected from the group consisting of a nucleic acid, a protein, and a small molecule.

18. The substrate of claim 1, wherein said capture molecule is selected from the group consisting of a nucleic acid, a protein, and a small molecule.

14. The method of claim 14, wherein said protein is an antibody.

15. A substrate comprising an array of capture molecules affixed to an optically clear coating of nitrocellulose on said substrate.

16. The substrate of claim 15, wherein said substrate is plastic.

17. The substrate of claim 15, wherein said substrate is glass.

19. A system, comprising:

a) a substrate comprising an array of capture molecules affixed to an optically clear coating of nitrocellulose on said substrate; and

b) a device for detection of target molecules bound to said capture molecules.

20. The system of claim 19, further comprising reagents that form a precipitate where said target molecule is bound to said capture molecule.

21. The system of claim 19, wherein said device detects the presence of a precipitate of said capture molecule and said target molecule on said array.

22. The system of claim 19, wherein said device quantifies the level of said target molecule.

23. The system of claim 19, wherein said substrate is plastic.

24. The system of claim 19, wherein said substrate is glass.

25. The system of claim 19, wherein said device is a calorimetric reader.

26. The system of claim 25, wherein said reader is CCD or CMOS based.

27. The system of claim 19, wherein said array is selected from the group consisting of a 3″

×

1″

slide, a 96-well array plate, and a 384-well plate.

28. The system of claim 19, wherein said capture molecule is selected from the group consisting of a nucleic acid, a protein, and a small molecule.

29. The system of claim 28, wherein said protein is an antibody.

30. A kit, comprising:

a) a substrate comprising an array of capture molecules affixed to an optically clear coating of nitrocellulose on said substrate; and

b) a device for detection of target molecules bound to said capture molecules, and

c) reagents that form a precipitate where said target molecule is bound to said capture molecule.