# US 8,785,352 B2 · Metal nanoparticle structures for enhancing fluorescence-based assays > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 8,785,352 B2 - Document ID: US8785352B2 - Status: Granted - Kind: B2 - Application number: 13/643,875 - Filed: 2011-04-29 - Published: 2013-05-16 - Granted: 2014-07-22 - Earliest claimed: 2010-04-30 - Assignees: Pharmaseq, Inc. - Applicants: Wlodek Mandecki; Ji Li; Zhuying Wang - Inventors: Wlodek Mandecki; Ji Li; Zhuying Wang - Web page: https://patentgrep.com/patent/US8785352B2 ## CPC - G01N 21/6486: {Measuring fluorescence of biological material, e.g. DNA, RNA, cells (G01N21/6428 takes precedence)} - G01N 33/587: {Nanoparticles} - Y10T 428/265: 1 mil or less - 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) - Y10T: TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION ## Abstract Provided, among other things, is a multiplex assay comprising: conducting a fluorescence-developing assay on microtabs having at least one surface that shows plasmonic enhancement, wherein a plurality of the microtabs have unique probes affixed to their plasmonically enhanced surfaces; and measuring the fluorescence associated with the substrates and identifying the correlated probe by for the microtab. The microtabs can be, for example, MTPs that send a unique identifier, and the correlated probe can be identified by querying the MTPs for their identifier. ## Claims ### Claim 1 A fluorescence enhancing substrate configured to covalently attach assay reagent comprising: a substrate; a first coating comprising AS and ES (AS/ES) on the substrate, wherein the coating is thicker than monolayer; and metal particles deposited or formed on the AS/ES coating, said particles effective to enhance a surface-mediated fluorescence assay by 5-fold or more, wherein AS is a reactive amine that is not a tertiary amine, and wherein the substrate with its first coating and any additional AS/ES coating(s) has amine and hydroxy functionality from AS and ES available for attaching an assay useful amount of the assay reagent. ### Claim 2 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein the metal particles are stabilized or protected with a coating or further coating of polymer. ### Claim 3 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein the substrate is a microtab that is a light-triggered transponder. ### Claim 4 (depends on 1) An assay comprising: conducting a fluorescence-developing assay in a plurality of wells of a microtiter plate or with a nucleic acid or protein array, the microtiter wells or different regions of the array comprising substrates according to claim 1, wherein a plurality of the substrates have unique probes affixed thereto; and measuring the fluorescence associated with the substrates and identifying the correlated probe by location. ### Claim 5 (depends on 1) A multiplex assay comprising: conducting a fluorescence-developing assay on microtabs having substrates according to claim 1, wherein a plurality of the microtabs have unique probes affixed to their substrates; and measuring the fluorescence associated with the substrates and identifying the correlated probe. ### Claim 6 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein AS is according to formula I: wherein R is a moiety of carbon, hydrogen and optionally oxygen such that the linkage to the nitrogen is C—N, and the linkage to the silicon is C—Si; and wherein R1, R2 and R3 are independently moieties of carbon, hydrogen and optionally oxygen such that the linkage to the illustrated oxygen is C—O and the so-defined silicon ethers are reactive with silicate glass; and ES is according to formula II: wherein R4 is a moiety of carbon, hydrogen and optionally oxygen such that the linkage to the silicon is C—Si; wherein R5, R6 and R7 are independently moieties of carbon, hydrogen and optionally oxygen such that the linkage to the illustrated oxygen is C—O and the so-defined silicon ethers are reactive with silicate glass; and wherein R8 and R9 are independently H or C1 to C4 alkyl. ### Claim 7 (depends on 6) An assay comprising: conducting a fluorescence-developing assay in a plurality of wells of a microtiter plate or with a nucleic acid or protein array, the microtiter wells or different regions of the array comprising substrates according to claim 6, wherein a plurality of the substrates have unique probes affixed thereto; and measuring the fluorescence associated with the substrates and identifying the correlated probe by location. ### Claim 8 (depends on 6) A multiplex assay comprising: conducting a fluorescence-developing assay on microtabs having substrates according to claim 6, wherein a plurality of the microtabs have unique probes affixed to their substrates; and measuring the fluorescence associated with the substrates and identifying the correlated probe. ### Claim 9 (depends on 8) The multiplex assay of claim 8, wherein the microtabs are MTPs that send a unique identifier, and the correlated probe is identified by querying the MTPs for their identifiers. ### Claim 10 (depends on 1) A method of forming a fluorescence enhancing substrate of claim 1, comprising: applying a first coating of AS/ES on the substrate, wherein the coating is thicker than monolayer; and depositing metal particles on the AS/ES coating by reducing metal salts, wherein said reduction-formed particles are effective to enhance a surface-mediated fluorescence assay by 5-fold or more. ### Claim 11 (depends on 10) A method of forming a fluorescence enhancing substrate of claim 10, wherein applying a first coating of AS/ES on the substrate comprises: reacting AS and ES with the substrate, wherein AS is according to formula I: wherein R is a moiety of carbon, hydrogen and optionally oxygen such that the linkage to the nitrogen is C—N, and the linkage to the silicon is C—Si; and wherein R1, R2 and R3 are independently moieties of carbon, hydrogen and optionally oxygen such that the linkage to the illustrated oxygen is C—O and the so-defined silicon ethers are reactive with silicate glass; and ES is according to formula II: wherein R4 is a moiety of carbon, hydrogen and optionally oxygen such that the linkage to the silicon is C—Si; wherein R5, R6 and R7 are independently moieties of carbon, hydrogen and optionally oxygen such that the linkage to the illustrated oxygen is C—O and the so-defined silicon ethers are reactive with silicate glass; and wherein R8 and R9 are independently H or C1 to C4 alkyl. ### Claim 12 (depends on 11) The method of claim 11, wherein the reacting is maintained or repeated until a thickness of 0.5 micron or more is achieved. ### Claim 13 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein the first coating is formed with AS and ES used in an AS:ES ratio of 2:1 molar or higher. ### Claim 14 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein the first coating has a thickness of 0.1 micron or more. ### Claim 15 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein the first coating has a thickness of 0.1 micron or more, and wherein the first coating has a porous structure. ### Claim 16 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein the first coating has a thickness of 0.2 micron or more. ### Claim 17 (depends on 1) The fluorescence enhancing substrate of claim 1, wherein said particles are effective effective to enhance a surface-mediated fluorescence assay by 10-fold or more.