# US 10,768,177 B2 · Bacteriophage-based electrochemical biosensor > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,768,177 B2 - Document ID: US10768177B2 - Status: Granted - Kind: B2 - Application number: 16/115,957 - Filed: 2018-08-29 - Published: 2020-03-05 - Granted: 2020-09-08 - Earliest claimed: 2018-08-29 - Assignees: The United States of America as represented by the Secretary of Agriculture - Applicants: The United States of America, as represented by the Secretary of Agriculture - Inventors: Vivian C. Wu; Irwin Quintela - Web page: https://patentgrep.com/patent/US10768177B2 ## CPC - G01N 33/56911: {Bacteria} - G01N 27/3275: {Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction} - G01N 33/542: with steric inhibition or signal modification, e.g. fluorescent quenching - G01N 33/585: {with a particulate label, e.g. coloured latex} - G01N 27/327: Biochemical electrodes {, e.g. electrical or mechanical details for in vitro measurements} - 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) ## Abstract A biosensor for detecting bacteria may use bacteriophages in a sandwich-assay system. The biosensor may include a capture element and a detection element. The capture element may include a substrate and a bacteriophage. The detection element may include a bacteriophage and a signal amplification element. The biosensor may be utilized such that the target bacterium is sandwiched between the capture element and the detection element, and a quantifiable signal may be generated to measure the amount of bacteria in a sample. The biosensor of the present invention utilizes direct sensing to detect the bacteria in the sample as opposed to indirect sensing methods. ## Claims ### Claim 1 A biosensor, comprising: a capture element comprising a substrate, a surface of the substrate being functionalized with at least one capturing bacteriophage; and a detection element comprising at least one detection bacteriophage conjugated to a signal amplification element, wherein the capturing bacteriophage and the detection bacteriophage are configured to bind to the same strain or species of a target bacterium, and wherein the substrate is one of a printed electrode, a printed circuit board, a silicon-based board, a carbon-based board, or a glass substrate with metal interdigitated electrodes. ### Claim 2 (depends on 1) The biosensor of claim 1, wherein the capturing bacteriophage is functionalized with a first conjugating element, and wherein the detection bacteriophage is functionalized with a second conjugating element. ### Claim 3 (depends on 2) The biosensor of claim 2, wherein the first conjugating element is one of biotin, histadine-tagged nitriloacetic acid, or dithiobis succinimidyl propionate, and wherein the second conjugating element is one of biotin, histadine-tagged nitriloacetic acid, or dithiobis succinimidyl propionate. ### Claim 4 (depends on 1) The biosensor of claim 1, wherein the capture element further comprises a linking molecule disposed to affix the capturing bacteriophage to the substrate. ### Claim 5 (depends on 4) The biosensor of claim 4, wherein the linking molecule is one of streptavidin or ethanolamine. ### Claim 6 (depends on 1) The biosensor of claim 1, wherein the signal amplification element comprises a particle, the particle being one of a metal particle, a metal oxide particle, or a semiconductor particle. ### Claim 7 (depends on 1) The biosensor of claim 1, wherein the signal amplification element comprises a catalyst. ### Claim 8 (depends on 7) The biosensor of claim 7, wherein the catalyst is one of horseradish peroxidase, soybean peroxidase, glucose oxidase, acetyl cholinesterase, or galactose oxidase. ### Claim 9 (depends on 1) The biosensor of claim 1, further comprising a signal detection device communicatively connected to the capture element. ### Claim 10 (depends on 1) The biosensor of claim 1, wherein the target bacterium is one of E. coli, Listeria monocytogenes, Campylobacter spp., and Staphylococcus spp. ### Claim 11 (depends on 1) The biosensor of claim 1, wherein the biosensor is configured to be operable using direct sensing of the target bacterium. ### Claim 12 A kit for the detection of a target bacterium, the kit comprising: a capture element comprising a substrate, a surface of the substrate being functionalized with at least one capturing bacteriophage; a detection element comprising at least one detection bacteriophage conjugated to a signal amplification element; and a signal detection device communicatively connected to the capture element, wherein the capturing bacteriophage and the detection bacteriophage are configured to bind to the target bacterium. ### Claim 13 (depends on 12) The kit of claim 12, wherein the signal amplification element comprises a particle and a catalyst, the particle being one of a metal particle, a metal oxide particle, or a semiconductor particle. ### Claim 14 (depends on 12) The kit of claim 12, wherein the target bacterium is one of E. coli, Listeria monocytogenes, Campylobacter spp., and Staphylococcus spp. ### Claim 15 (depends on 12) The kit of claim 12, wherein the kit is configured to be operable using direct sensing of the target bacterium. ### Claim 16 A method of detecting a target bacterium in a sample, the method comprising: performing a capture incubation step, the step comprising introducing a capture element to the sample and incubating for a pre-determined capture time; following the capture incubation step, performing a sandwich incubation step, the step comprising introducing a detection element to the capture element and incubating for a pre-determined sandwich time; and following the sandwich incubation step, generating a signal to detect the target bacterium, wherein the capture element comprises a substrate and a capturing bacteriophage functionalized thereon, and wherein the detection element comprises a detection bacteriophage conjugated to a signal amplification element. ### Claim 17 (depends on 16) The method of claim 16, wherein the pre-determined capture time is between 8-16 minutes. ### Claim 18 (depends on 16) The method of claim 16, wherein the pre-determined sandwich time is between 6-14 minutes. ### Claim 19 (depends on 16) The method of claim 16, further comprising introducing hydrogen peroxide. ### Claim 20 (depends on 16) The method of claim 16, wherein the sandwich incubation step generates a sandwich complex, the sandwich complex comprising the target bacterium disposed between the capture element and the detection element. ### Claim 21 (depends on 16) The method of claim 16, further comprising introducing a mediator.