Methods and arrays for detecting biological molecules

• US 7,429,466 B2
• Filed: 06/18/2002
• Issued: 09/30/2008
• Est. Priority Date: 01/24/2000
• Status: Expired due to Term

• Current Assignee: Hypromatrix Incorporated
• Original Assignee: Hypromatrix Incorporated
• Inventors: Wang, Yingjian
• Primary Examiner(s): SHIBUYA, MARK LANCE
• Application Number:
  US10/173,991
• Publication Number:
  US 20020155493A1
• Time in Patent Office: 2,296 Days
• Field of Search: 435/6, 435/7.9, 435/40.51, 435/7.1, 435/7.2, 435/7.92, 435/40.5, 435/40.52
• US Class Current: 435/40.51
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2565/515 characterised by the interaction between or sequential use of two or more arrays
• CPC Class Codes: C40B 30/04 by measuring the ability to specifically bind a target molecule, e.g. antibody-antigen binding, receptor-ligand binding
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/6842 Proteomic analysis of subsets of protein mixtures with reduced complexity, e.g. membrane proteins, phosphoproteins, organelle proteins
• CPC Class Codes: G01N 33/6845 Methods of identifying protein-protein interactions in protein mixtures

Abstract

Arrays and methods for detecting one or more biological molecules, where the methods generally comprise the steps of: providing a first support immobilized with one or more reagents; providing a second support immobilized with one or more of ligands; contacting the reagents immobilized to the first support with the ligands immobilized on the second support whereby one or more of the reagents bind to one or more of the ligands; and separating the first support from the second support so that one or more of the bound reagents remain bound to one or more ligands on the second support after separation. In one preferred method, proteins are immobilized on a support with adequate strength so that the proteins can be dissociated from the support under certain conditions, such as after binding with other proteins immobilized on another support. For example, antibody arrays produced according to the present invention may be used to detect protein expressions in a protein lysate and may be used in immunostaining to reveal the presence and location of proteins in cells.

Claims

1. A method for staining cells to detect one or more biological molecules in said cells, comprising the steps of:

immobilizing a plurality of antibodies on a first support, wherein each of said antibodies is provided at predetermined positions on said first support such that each of said antibodies can be identified by the position where it is immobilized;

(b) placing said cells on a second support;

(c) fixing said cells with a fixation solution;

(d) contacting said antibodies on said first support with said cells on said second support to allow said antibodies to bind to any corresponding interacting biological molecules in said cells;

(e) separating said first support from said second support whereby one or more of said bound antibodies remain bound to the corresponding biological molecules after separation;

(f) detecting said biological molecules by detecting said antibodies bound to said biological molecules in said cells.

2. The method of claim 1, wherein said fixation solution comprises chemicals selected from the group consisting of formaldehyde, glutaldehyde, acetone and methanol.

3. The method of claim 1, wherein said cells are one or more tissue sections.

4. The method of claim 1, wherein one or more of said antibodies are specific for one or more posttranslationally modified proteins.

5. The method of claim 1, wherein one or more of said antibodies are specific for one or more phosphorylated proteins.

6. The method of claim 1, wherein 5 to 100,000 different kinds of antibodies are each immobilized at one or more predetermined positions on said first support.

7. The method of claim 1, wherein 200 to 10,000 different antibodies are each immobilized at one or more predetermined positions on said first support.

8. The method of claim 1, wherein 50 to 1,000 different antibodies are each immobilized at one or more predetermined positions on said first support.

9. The method of claim 1, wherein 5 to 500 different antibodies are each immobilized at one or more predetermined positions on said first support.

10. The method of claim 1, wherein 5 to 100 different antibodies are each immobilized at one or more predetermined positions on said first support.

11. The method of claim 1, wherein 5 to 50 different antibodies are each immobilized at one or more predetermined positions on said first support.

12. The method of claim 1, wherein said first support comprises materials selected from the group consisting of nitrocellulose, nylon, polyvinylidene difluordie, glass, plastic, and their derivatives.

13. The method of claim 1, wherein said first support comprises materials selected from a- the group consisting of nylon and its derivatives.

14. The method of claim 1, wherein said reagents are immobilized on said first support in one or more shapes selected from the group consisting of circular, elongated, and polygonal.

15. The method of claim 1, wherein said first support comprises an array.

16. The method of claim 15, wherein said array comprises a plurality of whole or partial capillary tubes.

17. The method of claim 1, wherein said step of immobilizing said antibodies comprises the steps of, applying one or more intermediates to said first support;

and applying one or more of said antibodies to said first support so that one or more of said antibodies interacts with one or more of said intermediates.

18. The method of claim 1, wherein said step of detecting comprises the one or more steps of detecting selecting from the group consisting of, determining whether one or more of said antibodies is present on one or both supports, identifying one or more locations of said antibodies after the separating step, determining one or more quantities of said antibodies, and identifying one or more types of antibodies.

19. The method of claim 1, wherein said antibodies are immobilized on said first support in circular shape.

20. The method of claim 1, wherein said step of immobilizing said antibodies comprises the step of applying one or more of said antibodies to said first support.

21. The method of claim 1, wherein said step of detecting comprises the step of determining the quantities of one or more of said antibodies.