System and method for electrochemical detection of biological compounds

• US 7,820,030 B2
• Filed: 04/16/2004
• Issued: 10/26/2010
• Est. Priority Date: 04/16/2003
• Status: Active Grant

• Current Assignee: HandyLab Incorporated (Becton, Dickinson & Co)
• Original Assignee: HandyLab Incorporated (Becton, Dickinson & Co)
• Inventors: Handique, Kalyan, Brahmasandra, Sundaresh N., Namasivayam, Vijay, Kyonghoon, Lee, Althaus, John S.
• Primary Examiner(s): Michener; Jennifer K
• Assistant Examiner(s): DIETERLE, JENNIFER M
• Application Number:
  US10/553,584
• Publication Number:
  US 20060207891A1
• Time in Patent Office: 2,384 Days
• Field of Search: 205/787, 205/792, 435/6, 4352831-2887, 204/403.01
• US Class Current: 205/792
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 2563/173 staining/intercalating agent, e.g. ethidium bromide
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

The present invention relates to an electrochemical method for detecting a target polynucleotide. An electrode comprising an electrode surface is provided. The electrode surface includes at least one probe molecule reversibly immobilized with respect to the electrode surface. A first electrochemical signal indicative of an amount of probe molecule immobilized with respect to the electrode surface is obtained. The electrode surface is contacted with a liquid comprising the target polynucleotide. Upon the contacting step, at least some of the probe molecule immobilized with respect to the electrode surface dissociates therefrom. A second electrochemical signal indicative of an amount of probe molecule immobilized with respect to the electrode surface is obtained. The presence of the target polynucleotide is determined at least partially on the basis of the first and second electrochemical signals.

Claims

1. An electrochemical method for detecting a polynucleotide, comprising:

obtaining a first electrochemical signal from a first amount of probe molecule in the presence of a first polynucleotide and a second polynucleotide, the first and second polynucleotides being sufficiently complementary to form a duplex;

subjecting the first and second polynucleotides to at least one of an annealing step or a melting step in the presence of the first amount of probe molecule;

and then, obtaining a second electrochemical signal from the probe molecule.

2. The electrochemical method of claim 1, wherein the first electrochemical signal is obtained at a temperature below the melting point of the duplex region and the second electrochemical signal is obtained at a temperature at least as great as the melting point of the duplex.

3. The electrochemical method of claim 1, wherein subjecting the first and second polynucleotides to at least one of an annealing step or a melting step further comprises subjecting the first and second polynucleotides to at least one amplification step intermediate obtaining the first and second electrochemical signals and in the presence of the probe molecule.

4. The electrochemical method of claim 1, wherein the first electrochemical signal is obtained using an electrode and the method comprises contacting the electrode with a liquid prior to obtaining the first electrochemical signal, the electrode being dry prior to being contacted with the liquid.

5. The electrochemical method of claim 1, wherein the first electrochemical signal is obtained using an electrode and the method comprises contacting the electrode with a liquid prior to obtaining the first electrochemical signal, prior to the contacting step, the electrode comprising at least a portion of the first amount of probe molecule reversibly immobilized with respect thereto.

6. The electrochemical method of claim 1, wherein the probe molecule is substantially free of polynucleotides having a length of at least 8 bases.

7. The electrochemical method of claim 6, wherein the first and second electrochemical signals arise from an electrochemically active-moiety of the probe molecule that is free of purines.

8. The electrochemical method of claim 6, wherein the first and second electrochemical signals are substantially free of a contribution arising from an oxidation or a reduction of guanine.

9. The electrochemical method of claim 1, wherein detection of the polynucleotide is determined based upon a difference between the first and second electrochemical signals.