IP Library › Granted Patent US 12,735,712
Granted Patent B2
US 12,735,712 · App. 18/158,466 · Granted Sep 15, 2026

Electrochemical detection nanostructure, systems and uses thereof

Inventors: Christopher J. Easley (Auburn, AL); Subramaniam Somasundaram (Auburn, AL)
Assignee: Auburn University
C12N15/115C12Q1/6816C12N2310/122C12Q2565/518
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Quick Facts
Patent No.
US 12,735,712
App. No.
18/158,466
Granted
Sep 15, 2026
Kind
B2
Abstract

Described herein are DNA-nanostructures that can be used in an assay to detect and/or quantify an analyte of interest. Aspects of the DNA-nanostructure can include a single DNA molecule composed of hairpin structural motifs, an anchor recognition moiety, and a signal moiety, where the anchor recognition moiety and the signal moiety are in effective proximity to each other such that the tethered diffusion of the signal molecule can be altered based upon binding status of the anchor recognition moiety. Also described herein are methods of making and using the DNA-nanostructures.

Claims (33)

1 . A method for detecting an analyte, the method comprising:

contacting a sample containing the analyte to an anchor recognition moiety coupled to a DNA nanostructure, wherein the DNA nanostructure comprises:

a single continuous DNA molecule comprising: a first hairpin structural motif, a second hairpin structural motif, a first segment of single stranded DNA, and a second segment of single stranded DNA, wherein the first hairpin structural motif and the second hairpin structural motif are attached to each other via the first segment of single stranded DNA, wherein the second segment of single stranded DNA is attached to the second hairpin structural motif such that the second segment of single stranded DNA forms a single stranded tether region at one end of the single continuous DNA molecule;

the anchor recognition moiety, wherein the anchor recognition moiety is covalently coupled to a region of the single continuous DNA molecule between the first hairpin structural motif and the second hairpin structural motif; and

a signal moiety coupled to an end of the single continuous DNA molecule opposite from the tether region, wherein the signal moiety and the anchor recognition moiety on the single continuous DNA molecule are in effective proximity to each other such that binding of the analyte to the anchor recognition moiety changes a tethered diffusion rate of the signal moiety,

wherein a terminal base of the second segment of single stranded DNA of the DNA nanostructure is coupled to a surface of an electrode; and

detecting a change in the tethered diffusion rate of the signal moiety relative to the surface of the electrode when the analyte binds to the anchor recognition moiety wherein said detecting the change in the tethered diffusion rate of the signal moiety comprises detecting a change in an electrochemical current at the surface of the electrode.

2 . The method of claim 1 , further comprising binding the analyte to the anchor recognition moiety.

3 . The method of claim 1 , further comprising assembling the DNA nanostructure on the electrode.

4 . The method of claim 3 , wherein said assembling the DNA nanostructure on the electrode comprises immobilizing a thio-DNA comprising the second hairpin structural motif and the second segment of single stranded DNA to the surface of the electrode by the terminal base of the second segment of single stranded DNA, ligating an anchor recognition unit comprising the anchor recognition moiety, the first segment of single stranded DNA and the first hairpin structural motif to the second hairpin structural motif such that the second segment of single stranded DNA is attached to one end of the second hairpin structural motif, and the first segment of single stranded DNA is attached to other end of the second hairpin structural motif and one end of the first hairpin structural motif, and ligating a third segment of single stranded DNA coupled to the signal moiety to other end of the first hairpin structural motif of the anchor recognition unit.

5 . The method of claim 1 , wherein the DNA nanostructure further comprises a linker having a reactive group capable of attaching to the surface of the electrode, wherein the linker is attached to the terminal base of the second segment of single stranded DNA.

6 . The method of claim 5 , wherein the reactive group is selected from the group consisting of: a carboxyl group, amino group, aromatic amine group, a chloromethyl group, an amide group, a hydrazide group, a hydroxyl group, a thiol group, an epoxy group, and combinations thereof.

7 . The method of claim 1 , wherein the single continuous DNA molecule has a sequence that is 1-100% identical to one of SEQ ID NOs: 7-8.

8 . The method of claim 1 , wherein the signal moiety is a redox molecule.

9 . The method of claim 1 , wherein the signal moiety is methylene blue.

10 . A method for detecting an analyte, the method comprising:

contacting a sample containing the analyte to an anchor recognition moiety coupled to a DNA nanostructure, wherein the DNA nanostructure comprises:

a single continuous DNA molecule comprising: a first hairpin structural motif, a second hairpin structural motif, a first segment of single stranded DNA, and a second segment of single stranded DNA, wherein the first hairpin structural motif and the second hairpin structural motif are attached to each other via the first segment of single stranded DNA, wherein the second segment of single stranded DNA is attached to the second hairpin structural motif such that the second segment of single stranded DNA forms a single-stranded tether region at one end of the single continuous DNA molecule;

the anchor recognition moiety, wherein the anchor recognition moiety is covalently coupled to the single continuous DNA molecule and extends from the single continuous DNA molecule; and

a signal moiety coupled to an end of the single continuous DNA molecule opposite from the tether region, wherein the signal moiety and the anchor recognition moiety on the single continuous DNA molecule are in effective proximity to each other such that binding of the analyte to the anchor recognition moiety changes a tethered diffusion rate of the signal moiety,

wherein a terminal base of the second segment of single stranded DNA of the DNA nanostructure is coupled to the surface of the electrode; and

detecting a change in the tethered diffusion rate of the signal moiety relative to the surface of the electrode when the analyte binds to the anchor recognition moiety wherein said detecting the change in the tethered diffusion rate of the signal moiety comprises detecting the change in an electrochemical current at the surface of the electrode.

11 . The method of claim 10 , further comprising binding the analyte to the anchor recognition moiety.

12 . The method of claim 10 , further comprising assembling the DNA nanostructure on the electrode.

13 . The method of claim 10 , wherein the DNA nanostructure further comprises a linker having a reactive group capable of attaching to the surface of the electrode, wherein the linker is attached to the terminal base of the second segment of single stranded DNA.

14 . The method of claim 13 , wherein the reactive group is selected from the group consisting of: a carboxyl group, amino group, aromatic amine group, a chloromethyl group, an amide group, a hydrazide group, a hydroxyl group, a thiol group, an epoxy group, and combinations thereof.

15 . The method of claim 10 , wherein the signal moiety is a redox molecule.

16 . A method for detecting an analyte, the method comprising:

contacting a sample containing the analyte to an anchor recognition moiety coupled to a DNA nanostructure, wherein the DNA nanostructure comprises:

a single continuous DNA molecule comprising: a first hairpin structural motif, a second hairpin structural motif, a first segment of single stranded DNA, and a second segment of single stranded DNA, wherein the first hairpin structural motif and the second hairpin structural motif are attached to each other via the first segment of single stranded DNA, wherein the second segment of single stranded DNA is attached to the second hairpin structural motif such that the second segment of single stranded DNA forms a single-stranded tether region at one end of the single continuous DNA molecule;

the anchor recognition moiety, wherein the anchor recognition moiety is covalently coupled to the single continuous DNA molecule and extends from the single continuous DNA molecule; and

a signal moiety, wherein the signal moiety is coupled to an end of the single continuous DNA molecule opposite from the tether region, wherein the signal moiety and the anchor recognition moiety on the single continuous DNA molecule is in effective proximity to each other, wherein the DNA nanostructure is coupled to the surface of the electrode by a terminal base of the second segment of single stranded DNA of the DNA nanostructure such that binding of the analyte to the anchor recognition moiety changes a tethered diffusion rate of the signal moiety and a change in the tethered diffusion rate of the signal moiety changes an electrochemical current at the surface of the electrode; and

detecting a change in the tethered diffusion rate of the signal moiety relative to the surface of the electrode when the analyte binds to the anchor recognition moiety wherein said detecting the change in the tethered diffusion rate of the signal moiety comprises detecting the change in an electrochemical current at the surface of the electrode.

Assignments (2)
CONFIRMATORY LICENSE Recorded Feb 10, 2025
From: AUBURN UNIVERSITY
To: NATIONAL SCIENCE FOUNDATION
Reel/Frame 070160/0431 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded May 8, 2023
From: EASLEY, CHRISTOPHER J.; SOMASUNDARAM, SUBRAMANIAM
To: AUBURN UNIVERSITY
Reel/Frame 063571/0102 →
Continuity (3)
Continuation 16440113 · Jun 13, 2019
Provisional Application 62684227 · Jun 13, 2018
Related Publication 20230257755A1 · Aug 17, 2023
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