# US 2020/0399693 A1 · Nanopore assemblies and uses thereof > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2020/0399693 A1 - Document ID: US20200399693A1 - Status: Published application - Kind: A1 - Application number: 16/969,518 - Filed: 2019-02-11 - Published: 2020-12-24 - Earliest claimed: 2018-02-12 - Assignees: Oxford Nanopore Technologies PLC - Applicants: Oxford Nanopore Technologies, Ltd. - Inventors: Farzin Haque; Shaoying Wang - Web page: https://patentgrep.com/patent/US20200399693A1 ## CPC - B01J 20/2808: {being less than 2 nm, i.e. micropores or nanopores} - G01N 33/6818: {Sequencing of polypeptides} - B01J 20/3204: {Inorganic carriers, supports or substrates} - B01J 20/3206: {Organic carriers, supports or substrates} - B01J 20/3274: {Proteins, nucleic acids, polysaccharides, antibodies or antigens} - C12Q 1/6825: Nucleic acid detection involving sensors - G01N 33/54373: {involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings} - G01N 33/573: for enzymes or isoenzymes - B82Y 15/00: Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors - C12Q 1/6869: Methods for sequencing - G01N 2333/96455: Kallikrein (3.4.21.34; 3.4.21.35) - B01J: CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) - C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES - B82Y: SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES ## Abstract The disclosure provides a nanopore system assembled with non-membrane proteins for detecting analytes. Also disclosed are the methods, kits, and detection devices employing the disclosed nanopore system. The nanopore system has a wide variety of applications, including single molecule detection, DNA/RNA/peptide sequencing, sensing of chemicals, biological reagents, and polymers, and disease diagnosis. ## Claims ### Claim 1 A nanopore assembly for detecting an analyte, the nanopore assembly comprising a channel formed of a plurality of subunits, wherein each of the subunits comprises a non-membrane protein capable of forming a protein channel. ### Claim 2 (depends on 1) The nanopore assembly of claim 1, wherein each of the subunits comprises a polypeptide having a polypeptide sequence at least 75% identical to a polypeptide sequence selected from the group consisting of SEQ ID NOs: 1-35, preferably SEQ ID NOs: 4-12. ### Claim 3 (depends on 2) The nanopore assembly of claim 2, wherein the polypeptide comprises at least one residue substituted with cysteine. ### Claim 4 (canceled) ### Claim 5 (depends on 1) The nanopore assembly of claim 1, further comprising a probe for detecting an analyte, the probe being operably linked to at least one of the subunits. ### Claim 6 (depends on 5) The nanopore assembly of claim 5, wherein the probe is selected from the group consisting of chemicals, carbohydrates, aptamers, nucleic acids, peptide, protein, antibodies, and receptors. ### Claim 7 (depends on 5) The nanopore assembly of claim 5, wherein the probe comprises a sequence at least 75% identical to a sequence selected from the group consisting of SEQ ID NOs: 36-79. ### Claim 8 (depends on 5) The nanopore assembly of claim 5, wherein the probe is an anti-PSA antibody. ### Claim 9 (depends on 5) The nanopore assembly of claim 5, wherein the analyte is selected from the group consisting of nucleic acids, amino acids, peptides, proteins, polymers, and chemical molecules. ### Claim 10 (depends on 5) The nanopore assembly of claim 5, wherein the analyte is selected from the group consisting of PSA, CEA, AFP, VCAM, MiR-155, MiR-22, MiR-7, MiR-92a, MiR-122, MiR-192, MiR-223, MiR-26a, MiR-27a, MiR-802, and PSA or a fragment thereof. ### Claim 11 (canceled) ### Claim 12 (depends on 5) The nanopore assembly of claim 5, wherein the probe is operably linked via covalent bonding to at least one of the subunits, and wherein the covalent bonding optionally comprises a disulfide linkage, an ester linkage, or a sulfhydryl linkage. ### Claim 13 (canceled) ### Claim 14 (depends on 5) The nanopore assembly of claim 5, wherein the probe is operably linked to a location in proximity to an entrance of the channel or a location at an interior side of the channel. ### Claim 15 (canceled) ### Claim 16 (depends on 1) The nanopore assembly of claim 1, wherein the channel is embedded in a polymersome. ### Claim 17 (depends on 1) The nanopore assembly of claim 1, further comprising a membrane, wherein the channel is inserted in the membrane, and optionally at least one of cholesterol and porphyrin. ### Claim 18 (depends on 17) The nanopore assembly of claim 17, wherein the membrane comprises a polymer membrane or a lipid membrane. ### Claim 19 (depends on 18) The nanopore assembly of claim 18, wherein the polymer membrane comprises an alternating copolymer, a periodic copolymer, a block copolymer, a di-block copolymer, a tri-block copolymer, a terpolymer, or a combination thereof. ### Claim 20 (depends on 19) The nanopore assembly of claim 19, wherein the polymer membrane comprises PMOXA-PDMS-PMOXA. ### Claim 21 (canceled) ### Claim 22 (depends on 1) An apparatus for detecting an analyte, comprising the nanopore assembly of claim 1 and optionally a support for the nanopore assembly. ### Claim 23 (depends on 1) A kit comprising the nanopore assembly of claim 1 and optionally instructions for using the nanopore assembly. ### Claim 24 (depends on 1) A method of detecting an analyte, comprising: (a) contacting a sample containing an analyte with the nanopore assembly of claim 1 optionally placed on a support; (b) applying an electrical current across the channel of the nanopore assembly; (c) determining the electrical current passing through the channel at one or more time intervals; and (d) comparing the electrical current measured at one or more time intervals with a reference electrical current, wherein a change in electrical current relative to the reference electrical current indicates a presence of the analyte in the sample. ### Claim 25 (depends on 24) The method of claim 24, wherein the reference electrical current is measured with a sample that does not contain the analyte. ### Claim 26 (canceled)