Method for enhanced bio-conjugation events

• US 6,602,400 B1
• Filed: 06/15/2000
• Issued: 08/05/2003
• Est. Priority Date: 06/15/2000
• Status: Expired due to Term

• Current Assignee: Osmetech Technology Inc. (Roche Holding AG)
• Original Assignee: Motorola, Inc. (Motorola Solutions, Inc.)
• Inventors: Li, Changming, Shi, Song, Choong, Vi-En, Maracas, George
• Primary Examiner(s): Nguyen, Nam
• Assistant Examiner(s): Olsen, Kaj K.
• Application Number:
  US09/595,381
• Time in Patent Office: 1,146 Days
• Field of Search: 205/775, 205/777.5, 205/792, 204/403.01, 204/412, 324/600, 324/658, 324/663, 324/691, 324/692, 435/6, 435/7.1, 435/7.2, 422/82.01, 422/82.02
• US Class Current: 506/9
• CPC Class Codes: B01J 2219/00578 electrophoric
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00653 the compounds being bound to electrodes embedded in or on the solid supports
• CPC Class Codes: B01J 2219/00704 integrated with the reactor apparatus
• CPC Class Codes: G01N 27/3277 being a redox reaction, e.g. detection by cyclic voltammetry voltammetry per se G01N27/42, G01N27/48
• CPC Class Codes: G01N 33/5438 Electrodes

Abstract

The present invention provides an apparatus and methods for efficient, high-throughput electrical or electrochemical detection of biomolecules. More specifically, the invention provides an apparatus in which an independent set of electrodes is used to increase the occurrence of a desired bio-conjugation event at a test site.

Claims

1. A method for the electrical or electrochemical detection of molecular interactions between an immobilized probe molecule and a target molecule, comprising:

(a) providing a substrate comprising at least one test site electrode comprising immobilized probe molecules;

(b) contacting said test site electrode with a sample mixture containing a particular target molecule;

(c) concentrating said target molecules by applying an electrical potential between a pair of manipulation electrodes such that bioconjugation between said immobilized probe molecules and said target molecule is enhanced, and wherein said bioconjugation forms a conjugated complex, wherein said conjugated complex does not comprise a reporter group;

(d) detecting an electrical signal at said test site electrode; and

(e) determining whether said target molecule is present.

2. The method of claim 1 wherein said detection is an electrical detection method selected from the group consisting of impedance spectroscopy, AC impedance, cyclic voltammetry, AC voltammetry, pulse voltammetry, square wave voltammetry, hydrodynamic modulation voltammetry, conductance, potential step method, potentiometric measurements, amperometric measurements, and current step method.

3. A method according to claim 2 wherein said method is AC impedance.

4. A method according to claim 3, wherein said AC impedance is measured over a range of frequencies.

5. A method according to claim 3, wherein said AC impedance is measured by transient methods with AC signal perturbation superimposed upon a DC potential applied to an electrochemical cell.

6. A method according to claim 3, wherein said AC impedance is measured by impedance analyzer, lock-in amplifier, AC bridge, AC voltammetry, or combinations thereof.

7. A method according to claim 1 wherein the polarity of said manipulation electrodes is periodically alternated.

8. A method according to claim 1 wherein said immobilized probe molecules are immobilized with linker moieties.

9. A method according to claim 8 wherein said linker moieties comprise a conjugated polymer.

10. A method according to claim 9 wherein said conjugated polymer is chosen from the group consisting of polypyrrole, polythiphene, polyaniline, polyfuran, polypyridine, polycarbazole, polyphenylene, poly(phenylenvinylene), polyfluorene, and polyindole.

11. A method according to claim 9 wherein said conjugated polymer is polypyrrole.

12. A method according to claim 8 wherein said linker moieties comprise a neutral pyrrole matrix.

13. A method according to claim 1 wherein said substrate comprises an array of test site electrodes.

14. A method according to claim 1 or 13 wherein said immobilized probe molecules are oligonucleotides.

15. A method according to claim 1 or 13 wherein said target molecule is an oligonucleotide.

16. A method according to claim 1 wherein said immobilized probe molecules are antigens.

17. A method according to claim 1 wherein said immobilized probe molecules are peptides.

18. A method according to claim 1 wherein said immobilized probe molecules are proteins.

19. A method according to claim 1 wherein said target molecule is a protein.

20. A method according to claim 1 wherein said target molecule is an antibody.