Counteracting osmotic imbalance in a sequencing cell

Patent metadata

Field	Value
Publication number	US-2017369944-A1
Application number	US-201715630342-A
Country	US
Kind code	A1
Filing date	Jun 22, 2017
Priority date	Jun 27, 2016
Publication date	Dec 28, 2017
Grant date	—

Abstract

A method of analyzing a molecule is disclosed. A lipid bilayer is formed such that it divides a first reservoir characterized by a first reservoir osmolarity from a second reservoir characterized by a second reservoir osmolarity. An electrolyte solution is flowed to the first reservoir that tends to make a first change to a ratio of the first reservoir osmolarity to the second reservoir osmolarity. A voltage is applied across the lipid bilayer, wherein the lipid bilayer is inserted with a nanopore, and wherein a net transfer of ions between the first reservoir and the second reservoir tends to make a second change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity, and wherein the first change to the ratio and the second change to the ratio tends to counter-balance each other.

First claim

What is claimed is:

1 . A method of analyzing a molecule, the method comprising:
forming a lipid bilayer that divides a first reservoir from a second reservoir, wherein the first reservoir has a first reservoir osmolarity, and wherein the second reservoir has a second reservoir osmolarity; flowing an electrolyte solution to the first reservoir, wherein the electrolyte solution has an electrolyte solution osmolarity that differs from the first reservoir osmolarity, thereby making a first change to a ratio of the first reservoir osmolarity to the second reservoir osmolarity; and applying a voltage across the lipid bilayer, wherein the lipid bilayer includes a nanopore, and wherein the voltage causes a net transfer of ions between the first reservoir and the second reservoir, thereby making a second change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity, wherein the first change to the ratio and the second change to the ratio substantially counterbalance each other.

2 . The method of claim 1 , wherein a volume of the first reservoir has an initial value before flowing the electrolyte solution, wherein the first change the ratio of the first reservoir osmolarity to the second reservoir osmolarity causes water to flow across the lipid bilayer, thereby causing a first change to the volume of the first reservoir, wherein the second change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity causes water to flow across the lipid bilayer, thereby causing a second change to the volume of the first reservoir, and wherein the first change to the volume of the first reservoir substantially counterbalances the second change to the volume of the first reservoir.

3 . The method of claim 1 , wherein the net transfer of ions between the first reservoir and the second reservoir comprises a net efflux of ions from the second reservoir to the first reservoir.

4 . The method of claim 3 , wherein the net efflux of ions from the second reservoir to the first reservoir increases the ratio of the first reservoir osmolarity to the second reservoir osmolarity, and wherein flowing the electrolyte solution to the first reservoir decreases the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

5 . The method of claim 4 , wherein the electrolyte solution osmolarity is lower than the second reservoir osmolarity before the electrolyte solution is flowed to the first reservoir.

6 . The method of claim 5 , further comprising:
progressively reducing the electrolyte solution osmolarity from an initial electrolyte solution osmolarity to a final electrolyte solution osmolarity to make the first change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

7 . The method of claim 1 , wherein the net transfer of ions between the first reservoir and the second reservoir comprises a net influx of ions into the second reservoir from the first reservoir.

8 . The method of claim 7 , wherein the net influx of ions into the second reservoir from the first reservoir decreases the ratio of the first reservoir osmolarity to the second reservoir osmolarity, and wherein flowing the electrolyte solution to the first reservoir increases the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

9 . The method of claim 8 , wherein the electrolyte solution osmolarity is higher than the second reservoir osmolarity before the electrolyte solution is flowed to the first reservoir.

10 . The method of claim 9 , further comprising:
progressively increasing the electrolyte solution from an initial electrolyte solution osmolarity to a final electrolyte solution osmolarity to make the first change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

11 . The method of claim 1 , further comprising:
inserting the nanopore into the lipid bilayer before the electrolyte solution is flowed to the first reservoir.

12 . The method of claim 1 , further comprising:
inserting the nanopore into the lipid bilayer after the electrolyte solution is flowed to the first reservoir.

13 . The method of claim 1 , wherein the lipid bilayer spans across the second reservoir, wherein the first reservoir is external to the second reservoir, wherein the first reservoir has a first reservoir volume, wherein the second reservoir has a second reservoir volume, and wherein the first reservoir volume is larger than the second reservoir volume.

14 . The method of claim 1 , wherein the voltage applied across the lipid bilayer is an alternating current voltage.

15 . A system for analyzing molecules in a sequencing chip, the system comprising:
a sequencing chip comprising an array of cells, wherein each of the cells comprises a well; a reservoir coupled to the sequencing chip; and a processor or a circuitry configured to:
form a lipid bilayer that divides the reservoir from the well of one of the array of cells, wherein the reservoir has a first reservoir osmolarity, and wherein the well has a second reservoir osmolarity;
flow an electrolyte solution to the reservoir, wherein the electrolyte solution has an electrolyte solution osmolaritythat differs from the first reservoir osmolarity, thereby making a first change to a ratio of the first reservoir osmolarity to the second reservoir osmolarity; and
apply a voltage across the lipid bilayer, wherein the lipid bilayer includes a nanopore, and wherein the voltage causes a net transfer of ions between the reservoir and the well, thereby making a second change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity, wherein the first change to the ratio and the second change to the ratio substantially counterbalance each other.

16 . The system of claim 15 , wherein the net transfer of ions between the reservoir and the well comprises a net efflux of ions from the well to the reservoir.

17 . The system of claim 16 , wherein the net efflux of ions from the well to the reservoir increases the ratio of the first reservoir osmolarity to the second reservoir osmolarity, and wherein flowing the electrolyte solution to the reservoir decreases the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

18 . The system of claim 17 , wherein the electrolyte solution osmolarity is lower than the second reservoir osmolarity before the electrolyte solution is flowed to the reservoir.

19 . The system of claim 18 , further comprising:
progressively reducing the electrolyte solution osmolarity from an initial electrolyte solution osmolarity to a final electrolyte solution osmolarity to make the first change to the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

20 . The system of claim 15 , wherein the net transfer of ions between the reservoir and the well comprises a net influx of ions into the well from the reservoir.

21 . The system of claim 20 , wherein the net influx of ions into the well from the reservoir decreases the ratio of the first reservoir osmolarity to the second reservoir osmolarity, and wherein flowing the electrolyte solution to the reservoir increases the ratio of the first reservoir osmolarity to the second reservoir osmolarity.

22 . The system of claim 21 , wherein the electrolyte solution osmolarity is higher than the second reservoir osmolarity before the electrolyte solution is flowed to the reservoir.

23 . The s

Assignees

• Roche Sequencing Solutions Inc

Inventors

• Barrall Geoffrey
• Komadina Jason
• ROJEK Marcin

Classifications

• C12Q1/6869 Primary

  Methods for sequencing · CPC title

• G01N27/44782

  of a plurality of samples · 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

• C12Q1/6874 Primary

  involving nucleic acid arrays, e.g. sequencing by hybridisation · CPC title

• G01N27/44791

  Microapparatus (sample containers with integrated microfluidic structures B01L3/5027) · CPC title