Immobilization strategies for enhancing sensitivity of electrochemical aptamer-based sensors

Patent metadata

Field	Value
Publication number	US-11796499-B2
Application number	US-202218065118-A
Country	US
Kind code	B2
Filing date	Dec 13, 2022
Priority date	Dec 13, 2021
Publication date	Oct 24, 2023
Grant date	Oct 24, 2023

Abstract

The subject invention provides methods for fabricating electrochemical aptamer-based (E-AB) sensors with enhanced sensitivity, signal-to-noise ratios, LOD, and improved stability and reproducibility. The subject invention also provides methods for aptamer immobilization on the surface of the electrode, which favors sufficient spacing between aptamers at the microscale to achieve optimal target recognition, folding, and signal transduction. The E-AB sensors of the subject invention provide superior sensing regardless of the sequence or structure of the bound aptamers or the physiochemical properties of the target.

First claim

We claim:

1. A method for fabricating an electrochemical aptamer-based (E-AB) sensor, the method comprising:
providing an electrode;
incubating the electrode with an immobilizing solution comprising aptamer-target complexes each comprising an aptamer bound to an analyte, the immobilizing solution having a low ionic strength;
contacting the electrode with a solution comprising a backfiller; and
removing the target from each of the aptamer-target complexes.

2. The method according to claim 1 , the electrode being a solid electrode or paper-based electrode, the solid electrode being made of gold, silver, copper, platinum, palladium, an alloy, or a mixture thereof.

3. The method according to claim 1 , the electrode being a gold electrode.

4. The method according to claim 1 , the aptamer having structure-switching functionality.

5. The method according to claim 1 , the ionic strength being from about 1 mM to about 50 mM.

6. The method according to claim 1 , the aptamer being modified with a redox tag at one end and a functional group at the other end.

7. The method according to claim 6 , the redox tag being methylene blue.

8. The method according to claim 6 , the functional group being a thiol, sulfide, disulfide, amide, ester, alkenyl, alkynyl, carbonyl, aldehyde, carboxylate, carboxyl, or carbonate ester group.

9. The method according to claim 6 , the functional group being linked to the aptamer via a linker.

10. The method according to claim 9 , the linker comprising 1-10 carbon atoms.

11. The method according to claim 1 , the aptamer being labeled with a thiol group at the 5′ end, and methylene blue redox tag at the 3′ end.

12. The method according to claim 1 , the backfiller being selected from 6-mercapto-1-hexanol (MCH), dithiothreitol (DTT), 9-mercapto-1-nonanol, 2-{2-[2-(2-mercaptoethoxy)ethoxy]ethoxy}lethanol, and combination thereof.

13. A method for spatial distribution of aptamers on an electrode, the method comprising:
incubating the electrode with an immobilizing solution comprising aptamer-target complexes each comprising an aptamer bound to an analyte;
contacting the electrode with a solution comprising a backfiller; and
removing the target from each of the aptamer-target complexes.

14. The method according to claim 13 , the electrode being a solid electrode or paper-based electrode, the solid electrode being made of gold, silver, copper, platinum, palladium, an alloy, or a mixture thereof.

15. The method according to claim 13 , the electrode being a gold electrode.

Assignees

• Xiao Yi
• Liu Yingzhu
• Canoura Juan
• The Florida International Univ Board Of Trustees

Inventors

• Xiao Yi
• Liu Yingzhu
• Canoura Juan

Classifications

• G01N27/3276 Primary

  being a hybridisation with immobilised receptors (using a FET type sensor G01N27/4145; concerning the hybridisation C12Q1/68) · CPC title

• G01N27/3277 Primary

  being a redox reaction, e.g. detection by cyclic voltammetry (voltammetry per se G01N27/42, G01N27/48) · CPC title

• G01N33/5308

  for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites · CPC title

• G01N33/5438

  Electrodes · CPC title