Methods for creating bilayers for use with nanopore sensors

Patent metadata

Field	Value
Publication number	US-11299781-B2
Application number	US-201916396284-A
Country	US
Kind code	B2
Filing date	Apr 26, 2019
Priority date	Feb 16, 2012
Publication date	Apr 12, 2022
Grant date	Apr 12, 2022

Abstract

The present disclosure provides biochips and methods for making biochips. A biochip can comprise a nanopore in a membrane (e.g., lipid bilayer) adjacent or in proximity to an electrode. Methods are described for forming the membrane and insert-ing the nanopore into the membrane. The biochips and methods can be used for nucleic acid (e.g., DNA) sequencing. The present disclosure also describes methods for detecting, sorting, and binning molecules (e.g., proteins) using biochips.

First claim

The invention claimed is:

1. A method for forming a membrane for use in a nanopore sensor, comprising:
(a) introducing a buffer solution over a plurality of wells, each well comprising an opening and an electrode and a material layer disposed over the opening of the well, wherein the buffer solution is electrically conductive, wherein the material layer comprises one or more polymers, and wherein the material layer does not comprise a lipid component;
(b) bringing the buffer solution in contact with the material layer;
(c) measuring an electrical property of the material layer with the electrode to characterize the material layer, wherein the electrical property is selected from the group consisting of current, capacitance and resistance; and
(d) based on the measured electrical property of step (c), applying a stimulus to the material layer to induce at least a portion of the material layer to form a membrane over the opening of the well, wherein the membrane has a thickness that is less than a length of a nanopore such that the nanopore when inserted into the membrane extends through the membrane and wherein the material layer has a thickness that is greater than the length of the membrane such that the nanopore cannot extend through the material layer if it is inserted into the material layer.

2. The method of claim 1 , wherein, in (c), one or more voltages are applied to the electrode.

3. The method of claim 2 , wherein the voltage is selected to break or disrupt the membrane over the electrode.

4. The method of claim 1 , wherein the stimulus comprises at least one of a liquid flow over the surface of the electrode, a sequential flow of one or more different liquids over the surface of the electrode, the flow of one or more bubbles over the surface of the electrode, an electrical pulse, sonication pulse, pressure pulse, or sound pulse.

5. The method of claim 1 , wherein the material layer comprises at least two types of polymers.

6. The method of claim 1 , further comprising, after (d), applying an electrical stimulus through the electrode to facilitate an insertion of a pore protein in the membrane.

7. The method of claim 6 , wherein the membrane and the pore protein together exhibit a resistance of about 1 GΩ or less.

8. The method of claim 1 , wherein the membrane without a pore protein exhibits a resistance greater than about 1 GΩQ.

9. The method of claim 1 , further comprising, prior to (a), directing a material solution comprising a material for forming the membrane over the plurality of wells; and depositing the material layer over the opening of each well.

10. The method of claim 9 , wherein the material solution comprises one or more polymers and an organic solvent.

11. The method of claim 10 , wherein the organic solvent comprises decane.

12. The method of claim 1 , wherein the buffer solution comprises an ionic solution.

13. The method of claim 12 , wherein the ionic solution comprises a chloride anion.

14. The method of claim 12 , wherein the ionic solution comprises sodium acetate.

15. The method of claim 1 , wherein a surface of the electrode is hydrophilic and wherein the hydrophilic surface is exposed to a flow channel associated with the well.

16. A method for forming a membrane for use in a nanopore sensor, comprising:
(a) introducing a buffer solution over a plurality of wells, each well comprising an opening and an electrode and a material layer disposed over the opening of the well, wherein the buffer solution is electrically conductive, wherein a hydrophilic surface of the electrode is exposed to a flow channel associated with the well, wherein the material layer comprises one or more polymers, and wherein the material layer does not comprise a lipid component;
(b) bringing the buffer solution in contact with the material layer;
(c) measuring an electrical property of the material layer with the electrode to characterize the material layer, wherein the electrical property is selected from the group consisting of current, capacitance and resistance; and
(d) based on the measured electrical property of step (c), applying a stimulus to the material layer to induce at least a portion of the material layer to form a membrane over the opening of the well.

17. The method of claim 16 , wherein the stimulus comprises at least one of a liquid flow over the surface of the electrode, a sequential flow of one or more different liquids over the surface of the electrode, the flow of one or more bubbles over the surface of the electrode, an electrical pulse, sonication pulse, pressure pulse, or sound pulse.

18. The method of claim 16 , further comprising, after (d), applying an electrical stimulus through the electrode to facilitate an insertion of a pore protein in the membrane.

19. The method of claim 16 , further comprising, prior to (a), directing a material solution comprising a material for forming the membrane over the plurality of wells; and depositing the material layer over the opening of each well.

20. The method of claim 16 , wherein the buffer solution comprises an ionic solution.

Assignees

• Genia Tech Inc
• Roche Sequencing Solutions Inc

Inventors

• Davis Randall
• Chen Roger

Classifications

• B01J2219/00734

  Lipids · CPC title

• B01J2219/00313

  the reactor vessels being formed by arrays of wells in blocks · CPC title

• B01J2219/00653

  the compounds being bound to electrodes embedded in or on the solid supports · CPC title

• G01N33/48721

  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 title

• G01N33/5432

  Liposomes or microcapsules · CPC title