DEVICES AND METHODS FOR SAMPLE ANALYSIS

• US 20180188230A1
• Filed: 10/03/2017
• Published: 07/05/2018
• Est. Priority Date: 04/03/2015
• Status: Active Grant

• Current Assignee: Abbott Laboratories Incorporated
• Original Assignee: Abbott Laboratories Incorporated
• Granted Patent: US 11,022,598 B2
• Time in Patent Office: Days
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2400/0406 capillary forces
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• 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: G01N 2458/10 Oligonucleotides as tagging agents for labelling antibodies
• CPC Class Codes: G01N 33/48721 Investigating individual macromolecules, e.g. by translocation through nanopores Coulter counters in general G01N15/12; fabrication methods for nanoscale apertures B81B1/00; sequencing of nucleic acids C12Q1/68
• CPC Class Codes: G01N 33/5308 for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites
• CPC Class Codes: G01N 33/54306 Solid-phase reaction mechanisms

Abstract

Methods, devices, and systems for analyte analysis using a nanopore are disclosed. The methods, devices, and systems utilize a first and a second binding member that each specifically bind to an analyte in a biological sample. The method further includes detecting and/or counting a cleavable tag attached to the second binding member and correlating the presence and/or the number of tags to presence and/or concentration of the analyte. Certain aspects of the methods do not involve a tag, rather the second binding member may be directly detected/quantitated. The detecting and/or counting may be performed by translocating the tag/second binding member through a nanopore. Devices and systems that are programmed to carry out the disclosed methods are also provided.

Claims

1. (canceled)

2. (canceled)

3. (canceled)

4-6. -6. (canceled)

7-11. -11. (canceled)

12. (canceled)

13-14. -14. (canceled)

15-17. -17. (canceled)

18. (canceled)

19. (canceled)

20. (canceled)

21. (canceled)

22-23. -23. (canceled)

24. (canceled)

25. (canceled)

26. (canceled)

27-28. -28. (canceled)

29. (canceled)

30-139. -139. (canceled)

140-145. -145. (canceled)

146-216. -216. (canceled)

217. An integrated digital microfluidics nanopore device comprising:

a microfluidics module; and

a nanopore module in fluid communication with the microfluidics module and comprising at least one nanopore,

wherein the microfluidics module is configured to introduce a liquid droplet to the at least one nanopore.

218. The device of claim 217, wherein the at least one nanopore is a solid state nanopore or a biological nanopore.

219. The device of claim 217, wherein the microfluidics module is configured to move the liquid droplet via a fluid manipulation force selected from an electrical force pressure, electrowetting, dielectrophoresis, electrode-mediated force, opto-electrowetting, electric-field mediated force, electrostatic actuation, and surface acoustic waves.

220. The device of claim 217, wherein the microfluidics module comprises:

a first substrate comprising an array of electrodes to generate electrical actuation forces on a liquid droplet disposed thereon; and

a second substrate spaced apart from the first substrate,

wherein a first electrode in the array of electrodes forms an anode and a second electrode in the array of electrodes forms a cathode, to drive current through the nanopore.

221. The device of claim 220, wherein the at least one nanopore is disposed in a nanopore layer attached to at least one of the first substrate and the second substrate, and wherein the first electrode and the second electrode flank the nanopore layer.

222. The device of claim 220, wherein at least one of the first substrate and the second substrate is substantially transparent.

223. The device of claim 220, wherein the array of electrodes in the array are disposed in a grid pattern.

224. The device of claim 220, wherein the electrodes in the array of electrodes are interdigitated.

225. The device of claim 217, wherein the microfluidics module comprises a layer of surface acoustic wave (SAW) generation material.

226. The device of claim 217, further comprising at least one reagent disposed on the microfluidics module and configured to be carried by the liquid droplet.

227. The device of claim 226, wherein the reagent is selected from the group consisting of a binding member, a cleaveable tag, and a combination thereof.

228. The device of claim 226, wherein the cleavable tag comprises an anionic polymer, a cationic polymer, a dendrimer, or a nanoparticle and the binding member comprises a binding member or a receptor.

229. A method for detecting an analyte present in a biological sample, the method comprising:

contacting the sample with a first binding member, wherein the first binding member is immobilized on a solid support and wherein the first binding member specifically binds to the analyte;

contacting the analyte with a second binding member, wherein the second binding member specifically binds to the analyte and wherein the second binding member comprises a cleavable tag attached thereto;

removing second binding member not bound to the analyte bound to the first binding member;

cleaving the cleavable tag attached to the second binding member bound to the analyte bound to the first binding member;

translocating the cleavable tag through a nanopore; and

detecting the cleavable tag translocating through the nanopore.

230. The method of claim 229, wherein the nanopore is disposed in a device comprising:

a microfluidics module; and

a nanopore module in fluid communication with the microfluidics module and comprising the nanopore,

wherein the microfluidics module is configured to introduce a liquid droplet to the nanopore.

231. The method of claim 230, wherein the nanopore is formed in situ within the device.

232. The method of claim 229, wherein the cleavable tag is selected from the group consisting of an anionic polymer, a cationic polymer, a dendrimer, and a nanoparticle.

233. The method of claim 229, wherein the cleavable tag is negatively charged and the translocating comprises applying a positive potential across the nanopore thereby translocating the cleavable tag through the nanopore.

234. The method of claim 229, wherein the cleavable tag is positively charged and the translocating comprises applying a negative potential across the nanopore thereby translocating the cleavable tag through the nanopore.

235. The method of claim 229, wherein each cleavable tag translocating through the nanopore is a translocation event and the method further comprises measuring the translocation events to measure an amount of analyte present in the sample, wherein the amount of analyte present in the sample is determined by:

counting a number of translocation events during a set period of time and correlating the number of translocation events to a control;

measuring an amount of time for a set number of translocation events to occur and correlating to a control;

or

determining an average time between translocation events and correlating to a control,

wherein the control is a reference standard comprising a calibration curve, standard addition, or digital polymerase chain reaction.

236. The method of claim 235, wherein measuring the translocation events comprises observing a change in current induced by an interaction of the cleavable tag with the nanopore.