Sensing systems

Patent metadata

Field	Value
Publication number	US-12050194-B2
Application number	US-202017257464-A
Country	US
Kind code	B2
Filing date	Jan 24, 2020
Priority date	Feb 15, 2019
Publication date	Jul 30, 2024
Grant date	Jul 30, 2024

Abstract

In an example, a sensing system includes a pH sensor. The pH sensor includes two electrodes and a conductive channel operatively connected to the two electrodes. A complex is attached to the conductive channel of the pH sensor. The complex includes a polymerase linked to at least one pH altering moiety that is to participate in generating a pH change within proximity of the conductive channel from consumption of a secondary substrate in a fluid that is exposed to the pH sensor. The at least one pH altering moiety is selected from the group consisting of an enzyme, a metal coordination complex, a co-factor, and an activator.

First claim

What is claimed is:

1. A sensing system, comprising:
a pH sensor, including:
two electrodes; and
a conductive channel operatively connected to the two electrodes; and

a complex attached to the conductive channel of the pH sensor, the complex including a polymerase linked to at least one pH altering moiety that is to participate in generating a pH change within proximity of the conductive channel from consumption of a secondary substrate in a fluid that is exposed to the pH sensor, the at least one pH altering moiety being selected from the group consisting of an enzyme, a metal coordination complex, a co-factor, and an activator.

2. The sensing system as defined in claim 1 , wherein the at least one pH altering moiety is the enzyme, and wherein the enzyme generates an acid or a base in a reaction with the secondary substrate.

3. The sensing system as defined in claim 2 , wherein the enzyme is selected from the group consisting of hydrolases and oxidases.

4. The sensing system as defined in claim 1 , wherein kinetics of the at least one pH altering moiety are at least 10 fold faster than kinetics of the polymerase.

5. The sensing system as defined in claim 1 , wherein the at least one pH altering moiety is the enzyme, and wherein the complex further comprises a nucleic acid hairpin-enzyme inhibitor conjugate attached to the enzyme.

6. The sensing system as defined in claim 1 , wherein:
the at least one pH altering moiety is the enzyme; and
the complex further includes a second enzyme attached to the polymerase.

7. The sensing system as defined in claim 1 , wherein the complex is a fusion protein or a protein chimera.

8. The sensing system as defined in claim 1 , wherein the conductive channel of the pH sensor is selected from the group consisting of a semi-conducting nanostructure, a graphene nanostructure, a metallic nanostructure, and a conducting polymer nanostructure.

9. The sensing system as defined in claim 1 , further comprising:
a support including a plurality of depressions separated by interstitial regions, wherein at least the conductive channel of the pH sensor is at a bottom of one of the plurality of depressions; and
a plurality of additional pH sensors, wherein at least a conductive channel of each of the plurality of additional pH sensors is at a bottom of a respective one of the plurality of depressions.

10. The sensing system as defined in claim 9 , wherein each of the plurality of depressions includes sidewalls, and wherein the sidewalls include a pH buffer material.

11. A kit, comprising:
a pH sensor, including:
two electrodes; and
a conductive channel operatively connected to the two electrodes; and

a fluid, including:
a liquid carrier; and
a complex in the liquid carrier, the complex including a polymerase linked to at least one enzyme that is to create a pH change within proximity of the conductive channel from consumption of a secondary substrate in a second fluid that is exposed to the pH sensor.

12. The kit as defined in claim 11 , further comprising the second fluid, including:
a second liquid carrier; and
a labeled nucleotide, including:
a nucleotide;
a linking molecule attached to a terminal phosphate group of the nucleotide; and
a label attached to the linking molecule, the label being selected from the group consisting of a first group that enhances kinetics of the enzyme and a second group that slows kinetics of the enzyme.

13. The kit as defined in claim 12 , wherein:
the secondary substrate is in the second fluid and is a separate molecule from the labeled nucleotide; and
the first group or the second group is to alter kinetics of an acid or base generating reaction involving the enzyme and the secondary substrate.

14. The kit as defined in claim 12 , wherein:
the secondary substrate is in the second fluid and is a separate molecule from the labeled nucleotide;
the label is the second group that slows kinetics of the enzyme; and
the second group is selected from the group consisting of an allosteric inhibitor, a steric exclusion group, and a buffering group.

15. The kit as defined in claim 12 , wherein:
the secondary substrate is in the second fluid and is a separate molecule from the labeled nucleotide;
the label is the first group that that enhances kinetics of the enzyme; and
the first group is a co-factor of the enzyme.

16. The kit as defined in claim 11 , further comprising the second fluid, including:
a second liquid carrier; and
a labeled nucleotide, including:
a nucleotide; and
the secondary substrate attached to a base or a sugar of the nucleotide, wherein kinetics of the secondary substrate are at least 10 fold faster than kinetics of the polymerase.

17. A method, comprising:
introducing a fluid to a sensor array including a plurality of individually addressable conductive channels, thereby attaching a complex to at least some of the plurality of individually addressable conductive channels, the complex including:
a polymerase; and
a pH altering moiety linked to the polymerase, the pH altering moiety being selected from the group consisting of an enzyme that is to catalyze consumption of a secondary substrate in a solution that is to be exposed to the sensor array, a metal coordination complex that is to catalyze consumption of the secondary substrate in the solution that is to be exposed to the sensor array, and a co-factor or an activator of a catalyst label attached to a labeled nucleotide that is to be introduced to the sensor array.

18. A method, comprising:
selecting a pH altering moiety from the group consisting of an enzyme that is to catalyze consumption of a secondary substrate in a solution, a metal coordination complex that is to catalyze consumption of the secondary substrate in the solution, and a co-factor or an activator of a catalyst label attached to a labeled nucleotide;
conjugating a polymerase to the pH altering moiety to generate a complex; and
attaching the complex to a conductive channel operatively connected to two electrodes.

Assignees

• Illumina Inc

Inventors

• Fisher Jeffrey S
• Mather Brian D
• Pugliese Kaitlin M
• Mandell Jeffrey G
• Rogert Bacigalupo Maria Candelaria
• Boyanov Boyan

Classifications

• C12Q1/6825

  Nucleic acid detection involving sensors · CPC title

• C12Q1/48

  involving transferase · CPC title

• C12Q1/001

  Enzyme electrodes · CPC title

• C12Q2521/101

  DNA polymerase · CPC title

• G01N27/4167

  pH (electrodes therefor G01N27/302, G01N27/36) · CPC title