Granted Patent B2

US 12,704,503 · App. 18/406,279 · Granted Aug 11, 2026

Inventors: Holger Schmidt (Capitola, CA); Aaron Roe Hawkins (Provo, UT); David W. Deamer (Santa Cruz, CA)

Assignee: The Regents of the University of California

G01N33/5302B01L3/50273C12Q1/6825C12Q1/70G01N33/48721G01N33/54333G01N35/00B01L2200/0668G01N2035/00247

Abstract

Claims (41)

1 . A method of detecting a plurality of target molecules, comprising:

applying a plurality of microbeads to a microfluidic chip, wherein the plurality of target molecules are specifically bound to one or more of the plurality of microbeads, wherein:

the microfluidic chip comprises a membrane,

the membrane comprises a first side, a second side, and a pore, and

the pore comprises a first opening on the first side of the membrane and a second opening on the second side of the membrane,

such that an ionic current flows from the first side of the pore to the second side of the pore;

trapping the plurality of microbeads within a capture volume of the pore;

while the plurality of microbeads are trapped within the capture volume of the pore, releasing the plurality of target molecules from the plurality of microbeads, such that a first target molecule and a second target molecule of the plurality of target molecules each pass through the same pore; and

measuring the ionic current as the first and second target molecules each pass through the pore, thereby detecting the first and second target molecules.

2 . The method of claim 1 , wherein the plurality of target molecules comprises a polypeptide or nucleic acid.

3 . The method of claim 1 , wherein each microbead of the plurality of microbeads comprises a plurality of capture molecules bound to a surface of the microbead, and wherein the target molecules are specifically bound to the microbeads via the capture molecules.

4 . The method of claim 3 , wherein the plurality of capture molecules comprises an antibody or an antigen binding fragment thereof.

5 . The method of claim 3 , wherein the plurality of target molecules comprises a nucleic acid and wherein the plurality of capture molecules comprises a complementary nucleic acid.

6 . The method of claim 1 , wherein the sample is selected from blood or any fraction thereof, urine, sweat, sputum, saliva, feces, or semen.

7 . The method of claim 1 , further comprising:

contacting a sample suspected of containing the plurality of target molecules with the plurality of microbeads under conditions that allow binding of the plurality of target molecules to one or more of the plurality of microbeads.

8 . The method of claim 1 , wherein the capture volume defines a volume around the pore in which the ionic current creates an electric field that is strong enough to pull the released target molecule through the pore.

9 . The method of claim 1 , wherein the plurality of microbeads are trapped on the first side of the membrane and where the first opening of the pore on the first side of the membrane is greater than 1 μm in diameter and where the second opening of the pore on the second side of the membrane is less than 1 μm in diameter.

10 . The method of claim 1 , wherein the plurality of microbeads are trapped on the first side of the membrane and where the first opening of the pore on the first side of the membrane is less than 1 μm in diameter and where the second opening of the pore on the second side of the membrane is less than 1 μm in diameter.

11 . The method of claim 1 , wherein the second opening of the pore on the second side of the membrane is less than 100 nm.

12 . The method of claim 1 , wherein the second opening of the pore on the second side of the membrane is less than 50 nm.

13 . The method of claim 1 , wherein releasing the plurality of target molecules comprises applying heat to the plurality of microbeads.

14 . The method of claim 1 , wherein releasing the plurality of target molecules comprises changing a pH in a solution near the microbeads or changing a salt concentration in the solution near the microbeads.

15 . The method of claim 1 , wherein releasing the plurality of target molecules comprises changing a salt concentration in a solution near the microbeads.

16 . The method of claim 1 , wherein releasing the plurality of target molecules comprises photocleaving the target molecules from the plurality of microbeads.

17 . A system, comprising:

a plurality of microbeads, wherein each microbead of the plurality of microbeads is configured to specifically bind to one or more of a plurality of target molecules; and

a microfluidic chip comprising a membrane, the membrane comprising a first side, a second side, and a pore, the pore comprises a first opening on the first side of the membrane and a second opening on the second side of the membrane, wherein an ionic current flows from the first side of the pore to the second side of the pore;

wherein the microfluidic chip is configured to:

trap the plurality of microbeads within the capture volume of the pore;

while the plurality of microbeads are trapped within the capture volume of the pore, release the plurality of target molecules from the plurality of microbeads, such that a first target molecule and a second target molecule of the plurality of target molecules each pass through the same pore; and

measure the ionic current as the first and second target molecules each pass through the pore, thereby detecting the first and second target molecules.

18 . The system of claim 17 , wherein each microbead of the plurality of microbeads comprises a plurality of capture molecules bound to a surface of the microbead, and wherein the plurality of microbeads specifically bind to the plurality of target molecules via the capture molecules.

19 . The system of claim 17 , wherein the first opening of the pore on the first side of the membrane is greater than 1 μm in diameter and where the second opening of the pore on the second side of the membrane is less than 1 μm in diameter.

20 . The system of claim 17 , wherein the plurality of microbeads are trapped on the first side of the membrane and where the first opening of the pore on the first side of the membrane is less than 1 μm in diameter and where the second opening of the pore on the second side of the membrane is less than 1 μm in diameter.

21 . The system of claim 17 , wherein the second opening of the pore on the second side of the membrane is less than 100 nm.

22 . The system of claim 17 , wherein the second opening of the pore on the second side of the membrane is less than 50 nm.

23 . The system of claim 17 , wherein the microfluidic chip further comprises a heating device configured to apply heat to cause the release of the plurality of target molecules from the plurality of microbeads.

24 . The system of claim 17 , wherein the microfluidic chip is configured to change a salt concentration in a solution near the microbeads to cause the release of the plurality of target molecules from the plurality of microbeads.

25 . The system of claim 17 , wherein the microfluidic chip is configured to change a pH in a solution near the microbeads to cause the release of the plurality of target molecules from the plurality of microbeads.

26 . The system of claim 17 , wherein the microfluidic chip further comprises a light source configured to apply light to cause the release of the plurality of target molecules from the plurality of microbeads by photocleaving.

Assignments (2)

Continuity (3)

References Cited (213)