Biomolecular sensors and methods

• US 10,036,064 B2
• Filed: 10/27/2016
• Issued: 07/31/2018
• Est. Priority Date: 06/25/2015
• Status: Active Grant

• Current Assignee: Roswell Me Incorporated
• Original Assignee: Roswell Biotechnologies Inc.
• Inventors: Merriman, Barry L., Mola, Paul W.
• Primary Examiner(s): Kessel, Maris R
• Application Number:
  US15/336,557
• Publication Number:
  US 20170044605A1
• Time in Patent Office: 642 Days
• CPC Class Codes: B82Y 15/00 Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
• CPC Class Codes: B82Y 40/00 Manufacture or treatment of nanostructures
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2521/101 DNA polymerase
• CPC Class Codes: C12Q 2537/157 A reaction step characterised by the number of molecules incorporated or released
• CPC Class Codes: C12Q 2563/116 electrical properties of nucleic acids, e.g. impedance, conductivity or resistance
• CPC Class Codes: C12Q 2565/133 conformational analysis
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: G01N 27/26 by investigating electrochemical variables; by using electrolysis or electrophoresis
• CPC Class Codes: G01N 27/327 Biochemical electrodes , e.g. electrical or mechanical details for in vitro measurements
• CPC Class Codes: G01N 27/3272 Test elements therefor, i.e. disposable laminated substrates with electrodes, reagent and channels optical biosensors G01N33/52
• CPC Class Codes: G01N 27/3276 being a hybridisation with immobilised receptors using a FET type sensor G01N27/4145; concerning the hybridisation C12Q1/68
• CPC Class Codes: G01N 27/3278 involving nanosized elements, e.g. nanogaps or nanoparticles nanopores G01N33/48721; magnetic beads G01N27/745
• CPC Class Codes: G01N 27/4145 specially adapted for biomolecules, e.g. gate electrode with immobilised receptors

Abstract

Electronic sensors configured to detect single molecule targets and methods of using and manufacturing such electronic sensors are disclosed. A sensor may include a first electrode and a second electrode separated by a sensor gap. The first and second electrodes can be coupled by a sensor complex that can include a biopolymer bridge molecule and a probe. The probe can interact with a target molecule, and interaction of the probe and target molecule can produce a signal suitable to provide detection of the target molecule.

Claims

1. A sensor device comprising:

a first contact coupled to a first electrode;

a second contact coupled to a second electrode;

a sensor gap defined between one of the first contact and the first electrode and one of the second contact and the second electrode;

a bridge molecule comprising a first end and a second end;

wherein the bridge molecule comprises a biopolymer, and wherein the bridge molecule is coupled to the first contact at the first end and coupled to the second contact at the second end; and

a probe molecule coupled to the biopolymer to form a sensor complex,

wherein the probe molecule is coupled to the biopolymer by a precisely positioned linker comprising a streptavidin-biotin complex,

wherein the sensor complex is configured to interact with at least one target molecule.

2. The sensor device of claim 1, wherein the sensor gap has a sensor gap dimension of between about 5 nm and about 30 nm.

3. The sensor device of claim 1, wherein the first end comprises a first self-assembling anchor and the second end comprises a second self-assembling anchor.

4. The sensor device of claim 1, wherein the biopolymer comprises a linear biopolymer.

5. The sensor device of claim 4, wherein the linear biopolymer comprises an end-to-end length that is less than a persistence length of the linear biopolymer.

6. The sensor device of claim 4, wherein the linear biopolymer comprises an end-to-end length configured to approximate a sensor gap dimension of between about 5 nm and about 30 nm.

7. The sensor device of claim 1, wherein the probe molecule comprises an enzyme configured to engage the target molecule.

8. The sensor device of claim 7, wherein the enzyme is a polymerase or a reverse transcriptase.

9. A sensor device comprising:

a first electrode overlying a substrate surface;

a second electrode overlying the substrate surface;

a sensor gap defined between the first electrode and the second electrode;

a bridge molecule comprising a first end and a second end;

wherein the sensor gap comprises a sensor gap dimension of between about 5 nm and about 30 nm;

wherein the bridge molecule comprises a biopolymer bridge molecule; and

wherein the bridge molecule is coupled to the first contact at the first end and coupled to the second contact at the second end; and

a probe molecule attached to the biopolymer bridge molecule,

wherein the probe molecule is attached to the biopolymer bridge molecule by a precisely positioned linker comprising a streptavidin-biotin complex,

wherein the probe molecule is configured to engage at least one target molecule.

10. The sensor device of claim 9, wherein the first end comprises a first self-assembling anchor and the second end comprises a second self-assembling anchor.

11. The sensor device of claim 10, wherein one of the first self-assembling anchor and the second self-assembling anchor comprises a 5′

-thiol modification.

12. The sensor device of claim 10, wherein the biopolymer bridge comprises a peptide sequence, and wherein one of the first self-assembling anchor and the second self-assembling anchor comprises a cysteine residue or a peptide configured to selectively bind gold, aluminum, silicon dioxide, palladium, platinum, or other metals or material contacts capable of thiol-metal binding.

13. The sensor device of claim 9, wherein the biopolymer bridge molecule comprises a linear biopolymer.

14. The sensor device of claim 13, wherein the linear biopolymer comprises an end-to-end length that is less than a persistence length of the linear biopolymer.

15. The sensor device of claim 14, wherein the linear biopolymer comprises an end-to-end length configured to approximate the sensor gap.

16. The sensor device of claim 9, wherein the probe molecule comprises an enzyme configured to engage the target molecule.

17. The sensor device of claim 16, wherein the enzyme is a polymerase or a reverse transcriptase.

18. The sensor device of claim 16, wherein the enzyme is configured to engage the target molecule during a reaction in a solution comprising a plurality of different target molecules, wherein the reaction comprises a time period t, and wherein contacting the target molecule produces a plurality of conformational changes in the enzyme in response to the plurality of target molecule features, wherein each of the plurality of conformational changes modulates an electrical measurement in the sensor to produce a signal feature.

19. The sensor device of claim 18, further comprising a signal processing system configured to detect the signal feature.

20. The sensor device of claim 18, wherein the sensor device is configured to produce a signal trace comprising a plurality of signal features detected over time period t.

21. The sensor device of claim 9, wherein the biopolymer bridge molecule comprises a nucleic acid duplex.

22. The sensor device of claim 21, wherein the nucleic acid duplex comprises a DNA duplex, a DNA-RNA hybrid duplex, a DNA-PNA hybrid duplex, a PNA-PNA duplex, or a DNA-LNA hybrid duplex.

23. The sensor device of claim 21, wherein the nucleic acid duplex comprises a thiol-modified oligonucleotide.

24. The sensor device of claim 21, wherein the nucleic acid duplex further comprises an internal modification.

25. The sensor device of claim 9, further comprising a signal interpretation device, wherein the signal interpretation device comprises a signal interpretation map, and wherein the signal interpretation map is calibrated against a signal trace from a known target sequence.