JP2018500169A - Surface plasmon detection apparatus and method - Google Patents

Abstract

The present disclosure relates to methods, apparatus and systems for detecting molecules using surface plasmon resonance techniques, and more particularly to surface plasmon resonance techniques using metal nanoparticles formed on a substrate. In one aspect, a method of forming a layer of metal nanoparticles comprises providing a liquid composition comprising a binder polymer and a solvent, and at least partially immersing an article comprising a polymer surface in the liquid composition. The polymer surface includes a polymer material and does not include inorganic glass or crystalline material. The method additionally includes the step of applying a vapor phase plasma to the liquid composition to promote a chemical reaction between the binder polymer and the polymer material on the polymer surface and to form a binder layer on the polymer surface of the article. The method further includes applying metal nanoparticles to the binder layer to form the metal nanoparticle layer on the binder layer.

Description

Cross-reference of related applications Any and all applications for which foreign or national priority is identified in the application data sheet filed with this application are hereby incorporated by reference under Section 1.57 of the 37 U.S. Incorporated.

  The present disclosure relates to methods, apparatus and systems for detecting molecules using surface plasmon resonance techniques, and more particularly to surface plasmon resonance techniques using metal particles formed on a substrate.

  Surface plasmon (SP) refers to coherent electronic vibrations that propagate along an interface between a dielectric (eg, quartz glass) and a metal (eg, Ag or Au) along with electromagnetic waves, eg, light. Under certain conditions (determined by wavelength, polarization and / or angle of incidence), free electrons on the metal surface absorb incident photons and convert them to surface plasmon waves. A resonance condition called surface plasmon resonance (SPR) can be established when the photon frequency matches the natural frequency of surface electrons that oscillate against the restoring force of the positive metal nucleus. SPR conditions can be used in optical measurements such as fluorescence emission, Raman scattering, second harmonic generation, and absorption, among others.

  In one aspect, a method of forming a layer of metal nanoparticles comprises providing a liquid composition comprising a binder polymer and a solvent, and at least partially immersing an article comprising a polymer surface in the liquid composition. The polymer surface includes a polymer material and does not include inorganic glass or crystalline material. The method additionally includes the step of applying a vapor phase plasma to the liquid composition to promote a chemical reaction between the binder polymer and the polymer material on the polymer surface and to form a binder layer on the polymer surface of the article. The method further includes applying metal nanoparticles to the binder layer to form the metal nanoparticle layer on the binder layer.

  In some embodiments, the binder polymer includes a plurality of amine ends, at least a portion of the amine ends participating in a chemical reaction with the polymeric material to form an allylamine bond.

In some embodiments, the binder polymer comprises one or more molecules capable of forming an amide bond, and the binder polymer is polydiallyldimethylammonium; polydiallyldimethylammonium chloride; polyallylamine hydrochloride; poly-4 - vinylbenzyl trimethyl ammonium chloride; diethylenetriamine _{(DETA) (H 2 N-} CH 2 CH 2 -NH-CH 2 CH 2 -NH 2, diethylene glycol analogs), triethylenetetramine _{(TETA) (H 2 N-} CH 2 _{CH 2 -NH-CH 2 CH 2} -NH-CH 2 CH 2 -NH 2), tetraethylene pentamine _{(TEPA) (H 2 N-} CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH _{2 -NH-CH 2 CH 2} -NH _2), pentaethylene hexamine _{(PEHA) (H 2 N-} CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH ₂ ), polyamines derived from ethyleneamine including polyethyleneamine; hyperbranched polymers including polyamidoamine dendrimer, polypropylimine dendrimer, polyethyleneimine (PEI); or linear or multi-branched cations such as mixtures thereof Selected from the group consisting of functional polymers. In other embodiments, the binder polymer 804 includes an anionic polymer, such as polyacrylic acid, sodium poly-4-styrenesulfonate, polyvinylsulfonic acid, polysodium salt, polyamino acid, or mixtures thereof. In some embodiments, the binder polymer includes linear or hyperbranched polyethyleneimine (PEI), ethylenediamine or other crosslinkable molecules suitable for forming amide bonds on the polymer substrate surface. In some embodiments, the polymer surface does not include non-polymeric material.

In some embodiments, the polymeric material does not include SiO ₂ or Al ₂ O ₃ .

  In some embodiments, the polymer surface comprises a light transmissive polymeric material selected from the group consisting of polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), polystyrene (PS), polycarbonate (PC), and combinations thereof. Including.

  In some embodiments, providing the liquid composition includes providing the liquid composition in an electrically insulating container, the container being between two electrodes of the plasma chamber when applying a gas phase plasma. Be placed.

  In some embodiments, applying the gas phase plasma includes applying the gas phase plasma such that the liquid composition and the article immersed therein are in an electrically floating state.

  In some embodiments, the article is immersed in the liquid composition such that when a plasma is applied to the liquid composition, the entire polymer surface is submerged in the liquid composition.

  In some embodiments, applying the gas phase plasma includes applying a gas phase plasma formed from a gas mixture including a substantial amount of oxygen gas.

  In some embodiments, applying the gas phase plasma includes applying a gas phase plasma formed from a gas mixture that does not include a substantial amount of ammonia gas.

  In some embodiments, the method of forming a layer of metal nanoparticles further comprises quenching the gas phase plasma after forming the binder layer and before applying the metal nanoparticles.

  In some embodiments, the article has a shape selected from the group consisting of lamellae, strips, cavities, cylinders, cylinders, fibers, coils, U-shapes, helical windings and spirals.

  In some embodiments, forming the metal nanoparticle layer comprises forming metal nanoparticles on the polymer surface of the article facing away from the gas phase plasma.

  In some embodiments, the metal nanoparticles comprise negatively charged metal spheres, and the metal nanoparticles are bound to the free amine ends of the binder layer.

  In some embodiments, the metal nanoparticles comprise gold nanoparticles.

  In some embodiments, at least some of the metal nanoparticles comprise a metal sphere, and one or more ligands are attached to the metal sphere.

  In some embodiments, the one or more ligands include a binding moiety and a chemical moiety that has specificity for one or more target molecules.

In some embodiments, the metal nanoparticle layer has an area particle density region between about 1.0 × 10 ⁹ nanoparticles / cm ² and about 2.0 × 10 ¹¹ nanoparticles / cm ² .

  In some embodiments, the metal nanoparticles are between about 1 nm and about 10 nm, between about 5 nm and about 20 nm, between about 10 nm and about 30 nm, between about 20 nm and about 40 nm, between about 30 nm and about 50 nm. Between about 40 nm and about 60 nm, between about 50 nm and about 80 nm, between about 60 nm and about 100 nm, between about 80 nm and about 150 nm, between about 100 nm and about 200 nm, between about 150 nm and about 250 nm, It has a median diameter between about 200 nm to about 300 nm, between about 250 nm to about 400 nm, between about 300 nm to about 700 nm, between about 500 nm to about 900 nm, or between about 700 nm to about 1100 nm.

  In some embodiments, the article comprises a polymer fiber, the polymer fiber comprising at least one selected from the group consisting of a straight portion, a curved portion, and a coil portion.

  In another aspect, a test medium for detecting a target contained in a liquid sample includes a barrel including a polymer surface, the polymer surface includes a first polymer material, and an inorganic glass or crystalline material is included. Not included. The test medium additionally includes a metal nanoparticle layer formed on the polymer surface. The test medium further comprises one or more ligands attached to the metal nanoparticle layer and having specificity for one or more target molecules.

  In some embodiments, the test medium further comprises a binder polymer layer sandwiched between the polymer surface and the metal nanoparticle layer, wherein the binder layer is a second polymer material that is different from the first polymer material. including.

  In some embodiments, the barrel includes a portion having a shape selected from the group consisting of a thin plate, strip, cavity, cylinder, cylinder, fiber, coil, U-shape, helical winding, and helix.

  In some embodiments, the first polymeric material is from polycarbonate (PC), polyethylene terephthalate, polymethyl methacrylate, triacetyl cellulose (TAC), cyclic olefin, polyarylate, polybutylene terephthalate, polyimide, and combinations thereof. One or more substances selected from the group consisting of:

  In some embodiments, the barrel includes at least one surface coated with a metal nanoparticle layer.

  In some embodiments, the barrel includes at least one surface that is not covered by the metal nanoparticle layer.

  In some embodiments, the test medium includes a cuvette configured to contain a liquid sample.

  In some embodiments, the metal nanoparticle layer is formed on one or more inner surfaces of the cuvette.

  In some embodiments, the barrel takes the form of a fiber having a first end for receiving the light beam through the barrel and a second end for emitting the beam through the barrel.

  In some embodiments, the fiber includes at least one selected from the group consisting of a straight portion and a curved portion between the first end and the second end.

  In some embodiments, the barrel includes a coiled structure.

  In some embodiments, the barrel includes a U-shaped structure.

  In some embodiments, the barrel includes a cylindrical structure having a first end for receiving a light beam through the barrel and a second end for emitting the beam through the barrel.

  In some embodiments, the metal nanoparticle layer comprises negatively charged metal spheres, and the metal nanoparticles are bound to the free amine ends of the binder layer.

  In some embodiments, the metal nanoparticle layer comprises gold nanoparticles.

  In some embodiments, at least some of the metal nanoparticles comprise a metal sphere, and one or more ligands are attached to the metal sphere.

  In some embodiments, the one or more ligands comprise a binding moiety and a chemical moiety having specificity for one or more targets.

In some embodiments, the metal nanoparticle layer has an area particle density region between about 1 × 10 ⁹ nanoparticles / cm ² and about 2 × 10 ¹¹ nanoparticles / cm ² .

  In some embodiments, the metal nanoparticle layer is between about 1 nm to about 10 nm, between about 5 nm to about 20 nm, between about 10 nm to about 30 nm, between about 20 nm to about 40 nm, about 30 nm to about 50 nm. Between about 40 nm and about 60 nm, between about 50 nm and about 80 nm, between about 60 nm and about 100 nm, between about 80 nm and about 150 nm, between about 100 nm and about 200 nm, between about 150 nm and about 250 nm Having nanoparticles having a median diameter between about 200 nm and about 300 nm, between about 250 nm and about 400 nm, between about 300 nm and about 700 nm, between about 500 nm and about 900 nm, or between about 700 nm and about 1100 nm .

  In another aspect, a method of detecting a target molecule provides a test medium that includes a transparent container that contains at least one substrate that includes a polymer surface and is configured to receive a liquid solution that includes the target molecule. The process of carrying out. The polymer surface forms a binder polymer layer and a plurality of metal nanoparticles therein. The test medium further includes a capture molecule attached to at least some of the metal nanoparticles, the capture molecule being adapted to capture one or more of the target molecules. The method additionally includes the step of containing the liquid solution in a transparent container and immersing at least a portion of the polymer surface in the liquid solution, thereby capturing at least some of the target molecules with the capture molecules. The method transmits light through at least one of the first surface of the substrate and the second surface of the substrate, and from the incident light by localized surface plasmon resonance (LSPR) of the metal nanoparticles caused by the transmitted light. The method further includes detecting the changed transmitted light.

  In some embodiments, the binder polymer chain is attached to the polymer surface by a plurality of amide bonds formed between the binder polymer chain and the polymer surface.

  In some embodiments, at least one of the first surface and the second surface includes a polymer surface, and the step of transmitting includes transmitting light through the polymer surface prior to detection.

  In some embodiments, each of the first surface and the second surface includes a separate first polymer surface and a second polymer surface, and the step of penetrating the first and second surfaces before detection Transmitting light through the polymer surface.

  In some embodiments, providing the test medium includes providing a plurality of substrates each having a polymer surface, and the transmitting step transmits light through each of the polymer surfaces of the plurality of substrates. Process.

  In some embodiments, at least one of the first surface and the second surface does not include a polymer surface, and the detected light does not pass through the polymer surface before it is detected.

  In some embodiments, the substrate has a first refractive index that is higher than the second refractive index of the liquid solution, and the transmitting step receives light through the first surface, and the light is second. And transmitting under total internal reflection (TIR) or attenuated total internal reflection (ATR) conditions so that it is reflected multiple times from the polymer surface before transmitting through the surface.

  In some embodiments, providing the test medium includes providing a polymer surface having at least one of a curve, a flexure, an arc, a bend, a bow, a twist, a ring, and a bend.

  In some embodiments, the step of providing the test medium provides at least one selected from the group consisting of a straight portion and a curved portion between the first surface and the second surface of the substrate. Process.

  In some embodiments, providing the test medium includes providing a coiled structure between the first surface and the second surface.

  In some embodiments, providing the test medium includes providing a U-shaped structure between the first surface and the second surface.

  In some embodiments, providing the test medium includes providing a cylindrical structure between the first surface and the second surface.

  In another aspect, a method of forming a polymer layer on a polymer surface includes providing a liquid composition comprising a binder polymer and a solvent in a container. The binder polymer has a plurality of binder functional groups. The method additionally includes the step of at least partially sinking the article into the liquid composition, wherein the submerged portion of the article includes a polymer surface having a plurality of substrate functional groups formed thereon. The method additionally includes the step of placing a container having an article at least partially submerged in the plasma chamber. The method further includes applying energy to a quantity of gas above the liquid surface to generate a plasma from the quantity of gas, thereby causing or accelerating the formation of a binder polymer layer on the polymer surface.

  In some embodiments, a chemical reaction between some of the polymer functional groups and some of the substrate functional groups causes the formation of a binder polymer layer.

  In some embodiments, applying energy includes providing power to at least one electrode that is not in contact with the liquid composition.

  In some embodiments, the step of causing or accelerating the formation of the polymer layer comprises a polymer in which the liquid composition is sandwiched between the polymer surface and the plasma and the polymer surface is submerged so that it is not in direct contact with the plasma. Forming a polymer layer on the surface;

  In some embodiments, the polymer surface on which the polymer layer is formed faces away from the plasma.

  In some embodiments, causing or accelerating the formation of the polymer layer comprises forming a binder polymer layer on the polymer surface having at least one of a curve, a flexure, an arc, a bend, a bow, a twist, a ring, and a bend. Forming.

  In some embodiments, the step of causing or accelerating the formation of the binder polymer layer is substantially performed on a polymer surface having at least one of a curve, a flexure, an arc, a bend, a bow, a twist, a ring, and a bend. Forming a polymer layer having a uniform thickness.

  In some embodiments, causing or accelerating the formation of the binder polymer layer includes forming the binder polymer layer on a polymer surface facing away from the plasma.

In some embodiments, the polymer functional group comprises an NH ₂ group.

  In some embodiments, the substrate functional group comprises a carbonate group (—O— (C═O) —O—).

  In some embodiments, chemical reactions between some of the binder functional groups and some of the substrate functional groups cause the formation of amide bonds between them.

In some embodiments, the amount of gas does not contain nitrogen and the nitrogen atom of the NH ₂ group forms an amide bond.

  In some embodiments, the container is an electrically insulating container.

  In some embodiments, the liquid composition includes water.

  In some embodiments, the liquid further comprises NaOH dissolved therein.

In some embodiments, the binder polymer comprises PEI having a moiety of a terminal amine (—NH ₂ ) that is replaced by polyethyleneimine (PEI) or sulfur hydride (—SH).

  In some embodiments, the method of forming the polymer layer further comprises attaching the nanoparticles to the binder polymer chain after causing or accelerating a chemical reaction.

  In some embodiments, the nanoparticles are attached to some of the remaining binder functional groups of the binder polymer that are different from the binder functional groups attached to some of the substrate functional groups.

  In some embodiments, the container is an insulating substrate such that the polymer substrate and liquid composition are in an electrically floating state when energy is applied to form a plasma.

  In some embodiments, the energy is pulsed direct current energy.

  In some embodiments, the plasma is generated from a volume of gas at atmospheric pressure.

  In some embodiments, energy is supplied through the first electrode and the container is placed on another electrode that does not contact the liquid composition or polymer substrate.

It is a schematic diagram of a surface plasmon resonance (SPR) measurement system. 1 is a schematic diagram of a localized surface plasmon resonance (LSPR) measurement system according to an embodiment. FIG. 2B is a schematic diagram of the localized surface plasmon resonance (LSPR) measurement system of FIG. 2A after exposure to a target molecule, according to an embodiment. 1 is a polymer-based LSPR test medium illustrated with a case, according to one embodiment. FIG. 3B is the polymer-based LSPR test medium of FIG. 3A illustrated without the front portion of the case, according to one embodiment, showing a cross-sectional view of the container and a substrate disposed within the container. FIG. 3B is the polymer-based LSPR test medium of FIG. 3A illustrated with the case removed, showing a cross-sectional view of the container and a substrate disposed within the container, according to one embodiment. FIG. 3B is the polymer-based LSPR test medium of FIG. 3A illustrated with the case removed, illustrating a cross-sectional view of the container and a plurality of substrates disposed within the container, according to one embodiment. FIG. 3 is a schematic diagram of a polymer-based LSPR test medium having a substrate coated with nanoparticles and capture molecules, according to an embodiment. [FIG. 3F] FIG. 3F is a schematic illustration of the polymer-based LSPR test medium of FIG. 3E having target molecules supplemented by at least a subset of capture molecules, according to an embodiment. FIG. 4 shows an absorbance spectrum obtained from a polymer-based LSPR test medium with 1, 2, and 3 substrates, according to an embodiment. Figure 3 shows an absorption difference spectrum obtained from a polymer-based LSPR test medium with 1, 2, and 3 substrates, according to an embodiment. Figure 4 shows the absorption and refractive index differences obtained from polymer-based LSPR test media as a function of the number of substrates, according to an embodiment. 1 is a schematic diagram of a polymer-based attenuated total reflection (ATR) LSPR test medium under ATR mode of operation, according to an embodiment. FIG. 2 is a different configuration of a polymer-based ATR LSPR test medium, according to an embodiment. 2 is a different configuration of a polymer-based ATR LSPR test medium, according to an embodiment. 2 is a different configuration of a polymer-based ATR LSPR test medium, according to an embodiment. 2 is a flow chart illustrating a method of coating a polymer substrate with a layer of metal nanoparticles to produce a polymer-based test medium by plasma treating the polymer substrate by immersion according to an embodiment. In accordance with embodiments, various stages of coating a polymer substrate with a layer of metal nanoparticles are shown to produce a polymer-based test medium by plasma treating the polymer substrate by immersion. In accordance with embodiments, various stages of coating a polymer substrate with a layer of metal nanoparticles are shown to produce a polymer-based test medium by plasma treating the polymer substrate by immersion. In accordance with embodiments, various stages of coating a polymer substrate with a layer of metal nanoparticles are shown to produce a polymer-based test medium by plasma treating the polymer substrate by immersion. In accordance with embodiments, various stages of coating a polymer substrate with a layer of metal nanoparticles are shown to produce a polymer-based test medium by plasma treating the polymer substrate by immersion. In accordance with embodiments, various stages of coating a polymer substrate with a layer of metal nanoparticles are shown to produce a polymer-based test medium by plasma treating the polymer substrate by immersion. FIG. 9 illustrates a polymer-based test medium having a non-facing surface that is treated according to the process described in FIGS. 8A-8E, according to an embodiment. FIG. 4 illustrates an auto-calibration LSPR method that quantifies the amount of target molecule attached to a polymer-based test medium using a control medium having a known concentration of target molecule, according to an embodiment. FIG. 4 illustrates an auto-calibration LSPR method that quantifies the amount of target molecule attached to a polymer-based test medium under temperature variation conditions by using a control medium with a known concentration of target molecule, according to an embodiment.

Surface Plasmon Resonance (SPR) Measuring Device and System Surface Plasmon Resonance (SPR) conditions are determined by measuring the angle of the reflection minimum (or absorption maximum) of light, which is chemically bonded to the surface of metal (eg, gold and silver) and absorbed. Can be used to detect the presence of specific target molecules that can be or can be attached, such as polymers, DNA or proteins. For example, the presence of a target molecule can be detected by utilizing a specific capture molecule that is configured to capture or interact with, bind to or bind to the target molecule. When a capture molecule that can be immobilized on the metal surface captures the target molecule, a perturbation of the metal surface is caused, which can subsequently induce a change in SPR conditions. Such changes can be measured as changes in the reflectance of the test medium and form the basis for some SPR-based measurement techniques that are adapted to measure the presence of a wide variety of target molecules.

Planar SPR Measurement System FIG. 1 illustrates a planar metal thin film based SPR measurement system 100 for detecting the presence of a target molecule using a test medium 120, according to one embodiment. The planar SPR measurement system 100 includes a light source 110 for illuminating incident light 114 (eg, polarized incident light) onto a test medium 120, and a detector 130 for receiving reflected light 118 at a specific range of wavelengths. For example, a photodetector. The test medium 120 includes a substrate 124, a thin metal film 128 formed on one side of the substrate 124, and a glass block 136 or suitable monochromator disposed on the other side of the substrate 124. Test medium 120 additionally includes a channel 132 for supplying solution 142 to the surface of metal film 128. The solution 142 includes an analyte that may contain the target molecule 140 that is detected by the measurement system 100. On the surface of the metal film 128, a capture molecule 144 (referred to as a ligand) configured to capture (interact with, bind to, or bind to) the target molecule 140 that may be present in the solution. Is also attached. Thus, by contacting the surface of the metal film 128 with the solution 142, at least some capture molecules 144 can capture and bind to the target molecule 140, thereby resonating the surface plasmon of the metal film 128. Conditions can be changed.

  In operation, the light source 110 illuminates one side of the glass block 136 with incident light 114. In some configurations, the metal thin film 128 (eg, gold) may allow the evanescent wave of the incident light 114 to interact with the plasma wave on the surface of the metal film 128, thereby exciting the plasmons of the metal film 128. For example, the glass block 136 is disposed in close proximity to and in contact with the glass block 136. In the planar SPR measurement system 100 of FIG. 1, perturbations at the metal surface 128 that induce a change in the plasmon resonance conditions of the metal film 128 cause the target molecule 140 to bind to or be captured by at least some of the capture molecules 144. Can occur when Subsequently, the perturbation can induce a change in reflectance that can be measured by the detector 130, and the signal of the detector 130 can then be analyzed by the analyzer 148. In the illustrated system 100, some target molecules 140 chemically bind to the capture molecules when the solution 142 is injected through the channel 132, thereby increasing the refractive index proportional to the target molecule 140 binding concentration. Can cause. In this way, the illustrated SPR measurement system 100 allows measurement of the interaction between the capture molecule 144 and the target molecule 140.

  Metal thin film based SPR measurement techniques remain difficult and / or expensive to implement for several reasons. One reason is related to the fact that many existing techniques for forming metal films may limit the shape and surface of the substrate on which the metal film is formed. For example, thermal chemical vapor deposition often requires temperatures that are not suitable for substrates such as polymer substrates. In other vapor deposition methods such as physical vapor deposition or plasma enhanced chemical vapor deposition, the shadowing effect may result in a non-uniform thickness. Some techniques that may be conformal, such as plating, may require a special seed layer. Still other conformal techniques such as atomic layer deposition may be slow in deposition. Furthermore, producing test media having complex shapes to increase sensitivity and / or versatility can be relatively expensive. Another reason that many metal film based SPR techniques remain difficult to implement is related to obtaining an accurate and reliable quantitative signal in the face of changes in environmental factors such as temperature. Accordingly, in the following, the various disclosed embodiments relate to test media, systems, and methods that improve the sensitivity, versatility, and reliability of SPR-based measurement techniques using nanoparticle-based localized SPR (LSPR).

Localized Surface Plasmon Resonance (LSPR) Measurement System FIGS. 2A and 2B show a localized surface plasmon resonance (LSPR) measurement system 200a / b for detecting target molecules that may adhere to the surface of a test medium, according to an embodiment. Show. FIG. 2A shows the LSPR measurement system 200a before introducing the target molecule to be detected, and FIG. 2B shows the LSPR measurement system 200b after introducing the target molecule to be detected. The LSPR measurement system 200a / b was configured to detect the transmitted light 238a / 238b at a specific range of wavelengths, and a light source 210 configured to illuminate the test media 220a / 220b and transmit the incident light 214 A photodetector 230 is included. Unlike the SPR measurement system described with respect to FIG. 1, where the detected light is reflected light, the LSPR measurement system of FIGS. 2A and 2B extracts target molecules from light 238a, 238b that is transmitted through the test media 220a / 220b. Configured to detect.

  Referring to FIGS. 2A and 2B, the test medium 220a / 220b includes a container 250 configured to hold a solution 232 and a substrate 224. Container 250 and substrate 224 have inner surfaces 250S and 224S configured to contact solution 232 when solution 232 is present. At least a portion of the inner surface 250S and / or a portion of the inner surface 224S has a layer 8 of metal nanoparticles 22 formed on the inner surface. In addition, at least a portion of the inner surface 250S and / or a portion of the inner surface 224S has a capture molecule (sometimes referred to as a ligand) 244 formed on the inner surface. In some embodiments, capture molecule 244 is immobilized to inner surface 250S and / or inner surface 224S either directly (eg, chemically coupled) or indirectly (eg, via a capture antibody). Capture molecule 244 is adapted to capture a specific target molecule 240 by chemically binding to it. Referring to FIG. 2B, when the target molecule 240 is introduced into the solution 232, at least some of the target molecule 240 is chemically attached to the capture molecule 244.

  In operation, after exposing the capture molecule 244 and the polarized incident light 214 from the light source 210 passing through the test medium 220a prior to exposing the capture molecule 244 to the target molecule 240 to detect the presence of the target molecule 240. The polarized incident light 214 from the light source 210 that is transmitted through the test medium 220b is compared. Without being bound by any theory, when target molecule (or analyte) 240 is introduced into test medium 220a, some of target molecule 240 binds to capture molecule (or ligand) 244, thereby , Causing a perturbation on the surface of the metal nanoparticle 228 that induces a change in resonance conditions. This change may be measured based on the difference between the transmitted light 238a transmitted through the test medium 220a before being exposed to the target molecule 240 and the transmitted light 238b transmitted through the test medium 220b after being exposed to the target molecule 240. It causes a change in absorbance. The bound target molecule 240 causes an increase in refractive index on a scale proportional to the concentration of bound target molecule 240. Therefore, the bound target molecule 240 induces a change in absorbance that is detected via the photodetector 230 and analyzed by the analysis unit 248. Thus, the disclosed LSPR measurement system allows a quantitative measurement of the concentration of the target molecule 240.

  Without being bound by any theory, it is understood that the layer of metal nanoparticles 228 of FIG. 2B has distinctly different optical response characteristics compared to the metal thin film of FIG. 2A. Under certain circumstances, metal nanoparticles exhibit a stronger optical response compared to metal thin films. As a result, plane waves that impinge on a single metal particle (eg, tens of nanometers) are strongly “focused” on that particle, resulting in a high electric field density in the region of a few nanometers to tens of nanometers around the particle The size of the region may generally increase as the size of the particle increases. For example, in the case of 40 nm gold nanoparticles, the region can extend from the surface of the nanoparticles to 60 nm. In addition, without being bound by any theory, densely packed and / or regularly spaced nanoparticles are further reduced as a result of plasmon coupling between adjacent particles. May show enhanced field strength. Furthermore, by changing the nanoparticle shape or morphology, the SPR frequency can be tuned over a wide spectral range. Thus, according to the various embodiments disclosed herein, the ability to achieve extreme local field strengths enhances the versatility of the LSPR technique compared to the SPR technique.

  In the following, examples of target molecules 240 to be detected without loss of generality include amino acids, peptides, polypeptides, proteins, glycoproteins, lipoproteins, nucleosides, nucleotides, oligonucleotides, nucleic acids, sugars, Carbohydrates, oligosaccharides, polysaccharides, fatty acids, lipids (eg lipid vesicles), hormones, metabolites, cytokines, neurotransmitters, antigens, allergens, antibodies, inhibitors, drug molecules, toxins, toxins, insecticides, bacteria, Mention molecules such as viruses, radioisotopes, vitamins, amphetamines, deoxyribonucleic acid (DNA), ribonucleic acid (RNA), metal ions, chemical residues in foods such as antibiotics in meat, and contaminants in water it can.

  In the following, without loss of generality, examples of capture molecules 244 include antigens, antibodies, proteins, peptides, deoxyribonucleic acid (DNA), ribonucleic acid (RNA), peptide nucleic acid (PNA), enzymes, hormones. Or suitable molecules suitable for capturing the target substance to be detected, including hormone receptors.

In the following, without losing generality, examples of the metal nanoparticles 228 include aluminum (Al), bismuth (Bi), cobalt (CO), copper (Cu), gold (Au), iron (Fe). ), Indium (In), molybdenum (Mo), nickel (Ni), chromium (Cr), silver (Ag), palladium (Pd), platinum (Pt), ruthenium (Ru), rhodium (Rh), tin (β Examples thereof include metal elements such as -Sn), tantalum (Ta), titanium (Ti), tungsten (W), and zinc (Zn). Examples of the metal nanoparticles may include oxides or nitrides of metallic elements such as TiN, TaN, TaCN, and RuO ₂ . Metal nanoparticles may additionally include semiconductor materials having a sufficiently high doping concentration so that the properties are metallic. For example, silicon (Si), germanium (Ge), tin (α-Sn), gallium arsenide (GaAs), indium arsenide (InAs), cadmium selenide (CdSe) can be cited as examples of highly doped semiconductor nanoparticles. ), Cadmium sulfide (CdS), cadmium telluride (CdTe), zinc sulfide (ZnS), zinc selenide (ZnSe), and lead sulfide (PbS). Metal oxides, metal nitrides, and semiconductor nanoparticles having metallic properties can have a carrier density (eg, electron density) greater than, for example, 1 × 10 ¹⁸ / cm ³ .

  In FIG. 2B, metal nanoparticles 228 are illustrated as having a spherical shape, for purposes of illustration only. However, the metal nanoparticles 228 can have various other shapes including, for example, spheroidal, ellipsoidal, pyramidal, rod, linear, polygonal, and multi-podded. . In addition, the metal nanoparticles 228 can have voids enclosed therein or have a core-shell structure, and at least the shell of the core-shell structure includes the metal or metal oxide or semiconductor described above.

Polymer-Based LSPR Test Medium Outer Case and Sample Container Having Substrate Coated with Nanoparticles Referring to FIGS. 3A-3D, an LSPR test medium 300 according to one embodiment is illustrated. With reference to FIGS. 3A and 3B, the test medium 300 includes an outer case 304 that includes a front portion 310 and a rear portion 320 configured to enclose the container 350. The test medium 300 is configured to receive incident light 314 traveling in the x direction on the front side and to transmit transmitted light 338 from the opposite side.

  In various embodiments, at least a portion of the outer case 304 and the container 350 are made of a light transmissive material, such as a polymer material that transmits light in the visible wavelength range. As described herein, visible light refers to photons of light having a wavelength between about 380 and about 1100 nm. In some embodiments, at least a portion of the outer case 304 is made using a material that is optically opaque to visible light. In some embodiments, some portions of the container 350 are made using an opaque material. In some embodiments, at least a portion of the sides of the outer case 304 and the container 350 through which incident light 314 passes as it enters and exits the test medium is light transmissive so that light can pass therethrough. It can be manufactured using a material. In some embodiments, at least a portion of the sides of the outer case 304 and the container 350 through which incident light 314 passes as it enters and exits the test medium is light transmissive so that light passes through the test medium. It can be manufactured using a material. The remaining portion can also be made using a transparent material, an opaque material, or both. In some embodiments, the side of the container 350 through which the incident light 314 passes as it enters and exits the test medium, whereas the outer case 304 is substantially entirely made of an opaque material. At least a portion of can be made using a transparent material. The remaining portion of the container 350 can also be made using a transparent material, an opaque material, or both.

  In some embodiments, the front portion 310 of the outer case 304 has a light entrance window 340 adapted to allow incident light to pass unimpeded. In the illustrated embodiment, the light entrance window 340 is a slot, dimple, or notch from which the material of the outer case 304 has been removed or cut to maximize the transmission of light passing therethrough. . In other embodiments, the light entrance window 340 may be in the form of a transparent or partially transparent window. In still other embodiments, the light entrance window 340 may be in the form of an optical filter configured to selectively pass a larger fraction of a particular wavelength. Although not shown, the rear portion 320 may have a light exit portion that is similar to the light entrance window 340 and may be at least partially aligned in the x direction. Good.

  Referring to FIGS. 3B to 3D, the container 350 has a light incident window 354 for receiving incident light 314 without being disturbed. In the illustrated embodiment, the light entrance window 354 is a light transmissive window that is optimized in material and thickness to maximize the transmittance of light passing therethrough. For example, in embodiments where substantially the entire container 350 is made using a light transmissive material, the light entrance window 354 may have a thickness that is less than the remainder of the container 350. In yet other embodiments, the light entrance window 354 may be in the form of an optical filter configured to selectively pass a larger fraction of a particular wavelength. In yet other embodiments, the light entrance window 354 may be in the form of an optical lens configured to focus light passing therethrough. Although not shown, the container 350 may have a light exit window 358, which may be similar to the light entrance window 354 and is at least partially in the x direction. It may be aligned.

  In the illustrated embodiment, the light entrance window 354 and the light exit window 358 are such that the distance between them is short compared to the distance between the remaining front and back surfaces of the container 350. They are in a deep position relative to each other. Having such a configuration may be advantageous in optimizing the light path and / or optimizing, eg reducing, the total volume of the solution held by the container 350.

  In some embodiments, the container 350 is configured as a cuvette that includes a tubular internal cavity and is configured to hold a liquid sample. In the illustrated embodiment, the container 350 and the outer case 304 have a square cross section. However, it is understood that the container 350 and / or the outer case 304 may have other suitable cross-sectional shapes, such as a circular cross-section. The container 350 may have a lateral dimension that has dimensions that allow easy calculation of various parameters, such as 10 mm over the length of the x direction traversed by the light beam.

Polymer-Based Substrate for Target Molecule Detection With continued reference to FIGS. 3B-3D, the container 350 is configured to hold one sample substrate 360, or multiple sample substrates 360a, 360b, and 360c. Has been. In the embodiment shown in FIG. 3D, the container 350 holds three substrates 360a, 360b and 360c. However, in other embodiments, the container 350 may hold fewer or more substrates. Each substrate 360, 360a-360c has a handle region 362 and an analysis region 368. Each substrate has a front side facing incident light 314 and a back side facing transmitted light 338. At least a part of the sample substrates 360, 360a to 360c is formed using a transparent material, for example, a transparent polymer material.

  Referring back to FIGS. 3A and 3B, the outer case front portion 310 has a front opening 330 and the rear portion 320 has a rear opening 334. In addition, referring to FIGS. 3B-3D, the substrates 360, 360a-360c also have an opening 364 in the handle region 362. We have such an opening formed in the gap between the substrate and the adjacent inner surface of the container 350, or in the gap between adjacent substrates when multiple substrates are present. It has been found that it can be beneficial to reduce the number of bubbles obtained. Under some circumstances, bubbles formed in such gaps can rise to the opening where they are broken, thereby reducing or removing the bubbles. In some embodiments, the front and rear openings 330 and 334 and the opening 364 are substantially aligned in both the horizontal and vertical directions so that we minimize bubbles. I found out to maximize the effect.

  As described herein, a light transmissive material in the visible wavelength range refers to a material that transmits at least about 80% of incident light at visible wavelengths. Transparent polymer materials that can be used in any or all of cases, containers, and substrates without losing generality include polycarbonate (PC), polyethylene terephthalate, polymethyl methacrylate, triacetyl cellulose (TAC), cyclic olefin, poly Examples include arylate, polybutylene terephthalate, polyimide, or a combination thereof. Transparent non-polymeric or inorganic materials that can be used include silicon oxide based materials (eg, amorphous silica or quartz) or aluminum oxide based materials (eg, sapphire).

Target Molecule Detection via Nanoparticles and Capture Molecules Referring to FIGS. 3E and 3F, container 350 is configured to receive and hold solution 332 therein. At least a portion of the front surface 368F and the back surface 368R of the analysis region 368 of the substrate 360 has a layer of metal nanoparticles 328 formed thereon. In addition, at least a portion of the front and back surfaces of the analysis region 368 has a layer of capture molecules 344 formed thereon. The capture molecule 344 may be immobilized. In some embodiments, the substrate 360 has an uncoated portion 362 that does not have one or both of the metal nanoparticles 328 and the layer of capture molecules 344 formed thereon. Capture molecule 344 is adapted to capture a specific target molecule by chemically binding to it. Referring to FIG. 3E, when target molecule 340 is introduced into solution 332, at least some of target molecule 340 chemically attaches to capture molecule 344. Although the illustrated embodiment shows only nanoparticles 328 and capture molecules 344 formed on the surface of the analysis region 368 of the substrate 360, in other embodiments, at least a portion of the inner surface of the container 350, eg, The portion of the inner surface of the light entrance window 354 and the light exit window 358 can also have nanoparticles 328 and capture molecules 344 formed therein.

  Referring to FIGS. 3E and 3F, in operation, incident light 314 passes through the test medium 300a (FIG. 3E) before the substrate 360 having capture molecules 344 is exposed to the target molecule 340, and targets the substrate 360. After exposure to the molecule 340, it penetrates the test medium 300b (FIG. 3F). Next, to detect the presence of the target molecule 340, the absorbance of light transmitted through the test media 300a and 300b is compared. In the illustrated embodiment, incident light 314 is directed toward analysis region 368 of surface 360 such that substantially all of incident light 314 exits analysis region 368 as transmitted light 338a / b with substantially no reflection. It is done. In some embodiments, the incident light 314 is directed at an angle that is substantially perpendicular to the surface of the analysis region that receives the incident light 314. Without being bound by any theory, some of the target molecules 340 bind to the capture molecules 344, thereby causing perturbations on the surface of the metal nanoparticles 328 that induce a change in resonance conditions, and then Of absorbance that can be measured based on the difference between transmitted light 338a transmitted through test medium 300a before being exposed to target molecule 340 and transmitted light 338b transmitted through test medium 300b after being exposed to target molecule 340. Bring change. The bound target molecule 340 causes an increase in refractive index on a scale proportional to the concentration of bound target molecule 340. Thus, the bound target molecule 340 induces a change in absorbance that can be detected in the quantitative analysis described below.

Absorbance and Absorption Difference Spectrum FIG. 4A uses an LSPR measurement system similar to that described above with respect to FIGS. 2A and 2B, and using an LSPR test medium similar to that described above with respect to FIGS. 3 is a graph 400 showing a set of acquired absorbance spectra. The y-axis of the graph 400 represents the measured absorbance, and the x-axis represents the wavelength (λ) when the absorbance is measured. The illustrated absorbance spectra 410a, 420a, and 430a represent, for example, measured absorbance values for test media having 1, 2, and 3 substrates, respectively, each substrate corresponding to the test media described above with respect to FIG. 3E. Each of the substrates that are similarly arranged and have nanoparticles formed thereon are immersed in a solution to form capture molecules therein prior to introduction of the target molecule. The illustrated absorbance spectra 410b, 420b, and 430b represent, for example, measured absorbance values for 1, 2, and 3 substrates, respectively, with each substrate positioned similarly to the test medium described above with respect to FIG. 3F. Each of the substrates having nanoparticles formed therein is immersed in a solution after the introduction and attachment of the target molecule to the capture molecule to form the capture molecule therein. In the illustrated absorbance spectra 410a, 420a, and 430a, each of the substrates has approximately the same coverage for nanoparticles and capture molecules within the analysis region (368, 368a, 368b, and 368c in FIGS. 3B-3D). have. In addition, in the illustrated absorbance spectra 410b, 420b and 430b, each of the substrates is subjected to approximately the same concentration of target molecules in the solution in which the analysis region is immersed.

  FIG. 4B is a graph 450 illustrating a set of absorption difference spectra that can be obtained based on a set of absorbance data sets similar to that of FIG. 4A, according to an embodiment. The y-axis of the graph 450 represents the difference in absorbance value between the absorbance measured before and after the substrate is subjected to a specific concentration of the target molecule in the solution in which the analysis region of the substrate is immersed. The x-axis represents the wavelength (λ) when the absorbance is measured. The illustrated absorption difference spectra 460, 470, and 480 were measured before and after subjecting the substrate to target molecules at various wavelengths, for example, for test media having 1, 2, and 3 substrates, respectively. It represents the difference in absorbance values. That is, the absorbance spectra 460, 470, and 480 may include, for example, the intensity of light (eg, similar to 410a, 420a, and 430a) before the substrate is subjected to the target molecule, and the substrate is subjected to the target molecule. The difference from the intensity of the light after (eg, similar to 410b, 420b and 430b) can be expressed. As shown, the difference values of the absorption difference spectra 460, 470 and 480 increase in proportion to the number of substrates.

  In some practical implementations, instead of obtaining a spectrum of absorbance or absorbance change as shown in FIGS. 4A and 4B, the absorbance or absorbance change (Δ absorbance, especially at a selected wavelength (eg, peak wavelength)). ) May be more practical. FIG. 5 shows exposure of a substrate to a target molecule measured at a single wavelength (eg, peak wavelength) in the y-axis as a function of the number of substrates (x-axis) placed in the test medium. 5 is a graph 500 showing a change in absorbance resulting from the operation. Similar to the description above for FIG. 4B, the magnitude of the change in absorbance increases in proportion to the number of substrates through which light is transmitted. In addition, since the change in absorbance is directly proportional to the corresponding change in refractive index, the y-axis can be interchangeably expressed as a change in Δ refractive index.

  Thus, as shown, in FIGS. 4A and 4B, the absorbance spectra 410b, 420b and 430b of the LSPR test medium show the capture molecules formed on the nanoparticles relative to the nanoparticles and target molecules. And shows an increase in the absorbance value after exposing the substrate to the target molecule compared to the respective absorbance spectra 410a, 420a and 430a before exposing the substrate to the target molecule. The magnitude of the increase in absorbance value is proportional to the surface concentration of the attached target molecule, among others. Thus, based on known values of the nanoparticle surface density and capture molecule surface density, and based on the measured difference in absorbance before and after exposing the substrate to the target molecule, the concentration of the target molecule in solution Can be determined. In addition, since the absorbance increases in proportion to the number of substrates, the test medium is configured to hold multiple substrates as shown in FIG. 3D as described in FIGS. 4B and 5 Can be used to increase the detection sensitivity (eg, signal to noise ratio) of the target molecule.

Test medium based on attenuated total internal reflection A light beam passes through a first medium (eg, glass) having a _first refractive index n ₁ and has a second refractive index n ₂ that is less than n ₁ . When directed to a medium (eg, liquid or air), the light beam bends at the interface between the first medium and the second medium away from the normal having a direction perpendicular to the interface. It has been known. The magnitude of the bending of the light beam is determined by Snell's law indicating n ₁ × sin θ ₁ = n ₂ × sin θ ₂ , where n ₁ and n ₂ are the refractive indices of the first and second media, respectively. , Θ ₁ represents the first angle of the incident beam in the first medium relative to the normal, and θ ₂ represents the _second angle of the refracted beam in the second medium relative to the normal.

When the light beam traveling through the first medium hits the interface between the two media at a sufficiently large angle known as the critical angle θ _c , its refraction direction is parallel to the interface (90 degrees with respect to the normal). At a larger angle, the luminous flux is totally reflected and returns to the first medium. This condition is known as total internal reflection. A structure in which a parallel light beam propagating through a light-guiding structure (for example, a fiber) including a first medium having a first refractive index has an angle larger than θ _c reaching the interface. When reaching the interface between the body and an external medium, for example a substance comprising a second medium formed on the surface of the structure and having a second refractive index smaller than the first refractive index, By undergoing a series of total internal reflections at the interface between the media, it can be induced to pass through the structure under total internal reflection mode. The number of reflections can be changed by changing the incident angle. The technique is sometimes referred to as attenuated total internal reflection, or ATR. In some embodiments of the LSPR test medium, LSPR measurement method and LSPR measurement system disclosed herein, an ATR can be advantageously used as described below.

Polymer-Based Optical Structure for LSPR Measurement Based on ATR FIG. 6A shows a polymer-based LSPR test medium 600 configured to utilize attenuated total reflection (ATR). The test medium 600 includes a light structure 660 having a length L extending in the z direction and having a first refractive index n ₁ . The illustrated portion of the structure 660 represents an analysis region similar to the analysis region 368 of FIGS. 3A-3F, for example, and may include a light transmissive material, such as a light transmissive polymer material.

  In some embodiments, the light structure 660 has one or more surfaces. The structure 660 is illustrated as having a first surface 660S1 and a second surface 660S2 configured to face each other and reflect the light beam in the ATR mode. In some embodiments, the first surface 660S1 and the second surface 660S2 represent different portions of a single surface, described below with respect to FIGS. 6B-6C.

  At least a portion of the first surface 660S1 and the second surface 660S2 has a layer of metal nanoparticles 628 formed thereon. In addition, at least a portion of the first surface 660S1 and the second surface 660S2 have capture molecules 644 formed thereon. Capture molecule 644 is adapted to capture a specific target molecule by chemically binding to it.

Similar to the test medium 300 described above with respect to FIGS. 3A-3F, the surfaces 660S1 and 660S2 may be at least partially immersed in the structure 660 in a similar manner as described with respect to the substrate 360 of FIGS. 3A-3F. It is configured to come into contact with a solution 632 that can be held in a container. The solution 632 has a second refractive index n ₂ that is less than the first refractive index n ₁ of the structure 660, whether or not it contains dissolved target molecules. Similar to the capture molecules described above with respect to FIGS. 3E-3F, at least some of the target molecules 640 chemically attach to the capture molecules 644 when the target molecules 640 are introduced into the solution.

Continuing to refer to FIG. 6A, unlike the test medium described above with respect to FIGS. 3E and 3F, where the light beam passes through the substrate in a single pass, the incident light beam 614, eg, the light beam, passes through the end surface 660S3 and is guided through The structure 660 is entered and proceeds along the z direction as a whole corresponding to the longitudinal direction of the structure 660. The light beam 614 is directed to either the surface 660S1 or 660S2 of the structure 660 at an angle θ relative to the normal (x-axis) greater than the critical angle θ _c due to the attenuated total internal reflection (ATR) condition described above. The light beam 614 does not exit through either the surface 660S1 or 660S2, but instead is reflected internally and returns to the other side of the surface 660S1 or 660S2 back into the structure 660. The internal reflection event is such that by independently selecting the values of n ₁ , n ₂ , L and θ, a suitable number of internal reflection events can occur that may be suitable for a given system. Can be repeated multiple times.

  In some implementations, it is understood that the light beam 614, which may be collimated, is directed to either the surface 660S1 or 660S2 of the structure 660 such that the ATR condition is met as described above. In some other implementations, the light beam 614 may be uncollimated or partially collimated when entering the first end of the light structure 660. Upon entry, some photons satisfy the ATR condition and are totally internally reflected, thereby reaching the second end of the light structure 660, while other photons that do not meet the ATR condition are either on the surface 660S1 or 660S2. It is transmitted or diffusely scattered. In this way, in some implementations, the polymer-based LSPR test medium 600 “self-selects” the light beam that satisfies the ATR condition, eliminating the need for a high degree of pre-collimation of the light beam 614.

Although not bound by any theory, even if the light beam 614 is totally internally reflected, the reflected light generates a limited electromagnetic field adjacent to the surfaces 660S1 and 660S2, and an evanescent field is generated. The evanescent field decays exponentially as it moves away from the surfaces 660S1 and 660S2 in the x direction. The characteristic total reflection evanescent attenuation length, which can be expressed as d ₀ , for example, 1 / e attenuation length (not shown) of the amplitude of the field wave, is about 1/3 of the wavelength of the incident light, for example 100 It can be extended to ~ several nanometers, in one example about 200 nm.

In the LSPR test medium 600 of FIG. 6A under ATR conditions according to embodiments, free electrons on the surface of the metal nanoparticles 628 can be excited by an evanescent field, thereby inducing localized surface plasmon resonance (LSPR). Without being bound by any theory, due to the exponential decrease in evanescent field strength within d ₀ , surface free electron excitation is typically less than 100 nanometers thick (eg, about 20 nm). ), Which is also limited to a region also known as LSPR field decay length (λ ₀ ). Compared to many other SPR techniques, the excitation of the metal nanoparticles 628 can be relatively localized in a relatively thin region, and the light flux undergoes multiple internal reflection events, so A much higher signal-to-noise ratio can be achieved when using.

  Thus, in accordance with the embodiments disclosed herein, capture molecules 644 are based on perturbations caused by target molecules 640 bound to capture molecules 644 on the surface of metal nanoparticles 628 that induce a change in resonance conditions. By comparing the light flux transmitted through the light structure 660 under ATR conditions before and after exposure to the target molecule 648, the presence of the target molecule in the solution 632 can be detected. The change in resonance conditions is then in a similar manner as described above with respect to FIGS. 3A-3F, but before and after exposing structure 660 to target molecule 640 with higher sensitivity and higher signal-to-noise ratio. This results in a change in absorbance that can be measured based on the difference in transmitted light 638 passing through the test medium 600.

Configuration of Polymer-Based Optical Structure for LSPR Measurement Based on ATR FIGS. 6B, 6C, and 6D were each configured to utilize attenuated total reflection (ATR) in a similar manner as described above with respect to FIG. 6A. Polymer based LSPR test media 670, 680 and 690 are shown. In particular, FIGS. 6B-6D show a test medium having an analysis region that is uniformly and continuously coated with metal nanoparticles and capture molecules. The analysis regions of the test media shown in FIGS. 6B-6D are understood to represent embodiments that can be used as independent analysis regions for a given test medium. In addition, the analysis region and various regions or portions thereof can be used in combination or repeatedly.

Each of the test media 670, 680, and 690 includes optical structures 660a, 660b, and 660c, respectively, each of which includes a respective analysis region 668a-668c having a _first refractive index n1. Each of the light structures 660a-660c has a first end and a second end and a respective analysis region 668a-668c between the first end and the second end. Each of these analysis regions includes a light transmissive material, such as a light transmissive polymer material. Each of the analysis regions 668a-668c has a respective analysis region surface 668S1, 668S2, and 668S3 that includes a transparent polymeric material. In the illustrated embodiment, the transparent polymeric material has a first refractive index n ₁ and each of the analysis region surfaces 668S1-668S3 is configured to reflect the light beam in the ATR mode, as described above with respect to FIG. 6A. Has been.

  Similar to the previous description for surfaces 660S1 and 660S2 of light structure 660 described above with respect to FIG. 6A, in the illustrated embodiment of FIGS. 6B-6D, at least a portion of analysis region surfaces 668S1-668S3 are formed therein. A layer of metal nanoparticles 628 (not shown for clarity). In addition, at least a portion of the analysis region surfaces 668S1-668S3 has capture molecules 644 (not shown for clarity) formed thereon. Capture molecule 644 is adapted to capture a specific target molecule by chemically binding to it.

  With continued reference to FIGS. 6B-6D, any one or more of the light structures 660a-660c comprises the same transparent polymeric material as the analysis regions 668a-668c, and the metal nanoparticles 628 and the capture molecules formed thereon There may be uncovered portions 664a, 664b, and 664c having respective surfaces 664S1, 664S2, and 664S3 that do not have one or both of the 644 layers. In addition, any one or more of the light structures 660a-660c includes the same transparent polymeric material as the analysis regions 668a-668c and is disposed near at least one of the first and second ends of the light structures. Each optical structure portion 662a, 662b, and 662c may be included.

  With continued reference to FIGS. 6B-6D, each of the test media 670, 680, and 690 has a respective container 628a, 628b, and 628c configured to hold a liquid solution 632 therein. The liquid is adapted to dissolve the target molecule 648 (not shown) as described above with respect to FIG. 6A. When the liquid 632 is present, each of the light structures 660 a-660 c is configured such that at least a portion of the analysis regions 668 a-668 c can be immersed in the liquid 632.

As described above with respect to FIG. 6A, each of the analysis region surfaces 668S1-668S3 is configured to contact the solution 632 in the containers 628a-628c. The solution 632 has a second refractive index n ₂ that is smaller than the first refractive index n ₁ of the analysis regions 668 a-668 c, whether or not it contains dissolved target molecules. Similar to the capture molecules described above with respect to FIGS. 3E-3F, when target molecules 640 (not shown) are introduced into solution 632, at least some of those target molecules are transferred to capture molecules 644 (not shown). It adheres chemically and induces a change in absorbance.

  With continued reference to FIGS. 6B-6D, in operation, each of the test media 670, 680, and 690 causes the incident polarized light beam 614 emitted from the light source 610 at the first end of the light structures 660a, 660b, and 660c. It is configured to receive light, and is further configured to at least partially transmit light through the respective light guide structures 660a, 660b, and 660c. Each of the test media 670, 680, and 690 is further configured such that transmitted light is emitted at the second ends of the light structures 660a, 660b, and 660c and detected using the detector 630. In some embodiments, one or more lenses 672 and / or one or more optical filters 674 are used to focus and / or selectively pass the traveling light through the second of the light structure. It may be arranged between the end of the detector and the detector 630.

Each of the test media 670, 680, and 690 allows light transmitted through the light structures 660a, 660b, and 660c to be second from the first end of the light structure under the attenuated total internal reflection (ATR) condition described above. It is configured to be guided to the end. That is, since the second refractive index n ₂ of the solution 632 is smaller than the first refractive index n ₁ of the analysis regions 668 a, 668 b, and 668 c, the second refractive index n ₂ travels through the optical structures 660 a to 660 c and passes through the interface with the solution 632. The light beams reaching the respective surfaces 668S1, 668S2 and 668S3 to be formed do not exit the surface 668S1 at a specific angle, but instead are internally reflected and return to the inside of the light structures 660a to 660c toward the opposite surface. . In particular, the light flux that reaches the interface and forms an angle θ greater than the critical angle θ _{c with} respect to the normals of the surfaces 668S1, 668S2, and 668S3 at the point of light impingement is the attenuation described above. Satisfies total internal reflection (ATR) conditions. The internal reflection events are selected by independently selecting the values of n ₁ , n ₂ , θ, and the light path length so that a suitable number of internal reflection events that may be suitable for a given system occurs. It can be repeated as many times as desired.

  Referring to FIG. 6B, according to one embodiment, a test medium 670 including a light structure 660a configured to receive incident light 614 from one side and collect transmitted light 638 from the opposite side of the test medium 670 in the vertical direction. Is shown. Test medium 670 includes a light portion 662a, an uncoated portion 664a, and an analysis portion 668a arranged in a substantially co-linear column configuration. At least the analysis portion 668a is formed using a polymeric material so that the analysis region surface 668S1 forms a layer of metal nanoparticles (not shown for clarity) and is captured in the layer of metal nanoparticles It has a polymer surface that is adapted to attach molecules (not shown for clarity), and the capture molecules are adapted to capture specific target molecules. At least a portion of the analysis portion 668 a is configured to be immersed in the solution 632. The solution is adapted to dissolve the target molecule to be detected therein. Advantageously, the substantially collinear configuration of the light structure 660a is suitable for an LSPR measurement system in which the light source 610 and the photodetector 630 are arranged on both sides of the light structure 660a in the light traveling direction.

It is understood that the various dimensions of the various portions of the light guide 660a can be adjusted to the desired detection characteristics. For example, the z-direction length of the analysis region 688, the y- or x-direction diameter of the analysis region 688, and the refractive index n1 of the polymer material of the light structure 660a may be other physical properties such as, for example, the size of the container 628a. The size of the container 628a can be selected based on the total number of internal reflections desired based on constraints, and the availability of the amount of target molecules that can be dissolved in the container 628a, to name a few, of the solution 632 It can be selected based on the refractive index n ₂ and the physical constraints of the nanoparticle coating apparatus and process described below.

  In the embodiment illustrated in FIG. 6B, the various portions of the light structure 660a, including at least the analysis portion 668a, have a substantially circular cross-sectional shape (in the xy plane). However, possible embodiments are not so limited. For example, polygonal (triangular, square, rectangular, pentagonal, hexagonal, octagonal, etc.), elliptical or other suitable cross-sectional shapes are possible.

  In the embodiment illustrated in FIG. 6B, the various portions of the light structure 660a including at least the analysis portion 668a of the light structure 660a do not substantially deviate from being linear in the light travel direction (z direction). That is, at least the analysis portion 668a of the light structure 660a does not have a substantial curvature, deflection, arc, bend, bow, twist, ring or bend that deviates from the z direction. In the following, embodiments having such deviations from linearity will be described with respect to FIGS. 6C and 6D.

  Referring to FIG. 6C, the test medium 680 includes a light structure 660 b having at least one curved portion 682. Unlike the light structure 660a of FIG. 6B, the light structure 660b of the test medium 680 receives incident light 614 from one side (eg, the top) of the test medium 680 and is the same as the test medium 680, according to one embodiment. The transmitted light 638 is emitted to the vertical side.

  Similar to the test medium 670 of FIG. 6B, at least the analysis portion 668b of the test medium 680 is formed using a polymer material so that the analysis region surface 668S2 is a layer of metal nanoparticles (for clarity). Having a polymer surface adapted to form a capture molecule (not shown for clarity) attached to the metal nanoparticle layer, wherein the capture molecule is a specific target molecule. It is suitable to capture. Further, similar to FIG. 6B, at least a portion of the analysis portion 668b is configured to be immersed in a solution 632 adapted to dissolve the target molecule.

  Unlike FIG. 6B, the test medium 680 has light portions 662 b formed at the light incident end and the light exit end, respectively, and both the light incident end and the light exit end are disposed outside the solution 632. It is configured as follows. Further, unlike FIG. 6B, at least the analysis portion 668b of the light structure 660b deviates substantially from being linear and includes at least a curved portion 682. In the illustrated embodiment, the curved portion 682 includes a U-shaped bend region so that the light propagation direction is reversed from down to up. As a result, the total internal reflection light 672 emitted from the light detection end of the light structure 660b is on the same vertical side as the light incident end of the light structure 660b, unlike the light structure 660a of FIG. 6B. Advantageously, the non-linear configuration of the optical structure 660b may be suitable for an LSPR measurement system in which the light source 610 and the photodetector 630 are disposed on both sides of the optical structure 660a in the light traveling direction.

  For illustrative purposes only, an analysis region 668b is shown that includes a single curved portion 682 that includes a U-shaped bend region. However, it is understood that the embodiments described herein are not so limited and that a plurality of curved portions 682 may be included as part of the analysis region 668b. For example, the analysis region 668b may include a plurality of curved portions 682 having U-shaped bend regions with alternating irregularities connected in series to increase the overall effective length of the analysis region 668b.

  Further, each curved portion 682 may include other shapes of curvature. For example, the curved portion 682 includes one or more of flexures, arcs, bends, and bows, among other curved shapes, so that light is directed in different directions away from the original travel direction (downward). You may go out. In addition, such curvature may be optimized to have a curvature radius selected to achieve the desired ATR absorption signal or signal to noise ratio.

  In addition, in the same manner as described above with reference to FIG. 6B, the cross-sectional shape of the various portions of the optical structure 660a may be a polygon (triangle, square, rectangle, pentagon, hexagon, octagon, etc.) in addition to the illustrated circle. ), Other shapes such as oval or other suitable shapes.

  Referring to FIG. 6D, according to one embodiment, test medium 690 includes a plurality of convoluted portions 692 configured to receive incident light 614 from one side and collect transmitted light 638 from the same vertical side of test medium 680. The optical structure 660c which has is included.

  Similar to the test medium 680 of FIG. 6B, at least the analysis portion 668c of the test medium 690 is formed using a polymer material so that the analysis region surface 668S3 has a layer of metal nanoparticles (for clarity) Having a polymer surface adapted to form a capture molecule (not shown for clarity) attached to the metal nanoparticle layer, wherein the capture molecule is a specific target molecule. It is suitable to capture. Further, similar to FIG. 6B, at least a portion of the analysis portion 668c is configured to be immersed in a solution 632 adapted to dissolve the target molecule. In addition, the test medium 690 has light portions 662 c formed at the light incident end and the light exit end, respectively, and both the light incident end and the light exit end are arranged outside the solution 632. Has been. Further, similar to FIG. 6C, at least the analysis portion 668c of the light structure 660c deviates substantially from being linear and includes a plurality of convoluted portions 692. In the illustrated embodiment, the analysis region 668c includes a plurality of convoluted portions 692 whose light propagation direction changes continuously in a spiral, horizontal direction, and downward, vertically. The analysis region 668c has an optical return path portion 694 connected to the end of the lower maximum winding portion 692 so that the light beam changes its path upward and is transmitted at the exit end. In addition. As a result, the total internal reflection light 672 emitted from the light detection end of the optical structure 660c is on the same vertical side as the light incident end of the optical structure 660c. Advantageously, the rotational configuration of the light structure 660c may be suitable for an LSPR measurement system in which the light source 610 and the photodetector 630 are disposed on both sides of the light structure 660c in the light traveling direction.

  For illustrative purposes only, an analysis region 668c that includes six convoluted portions 692 is shown. However, it is understood that the embodiments described herein are not so limited and that any desired number of convoluted portions 692 may be included as part of the analysis region 668c. Further, although the illustrated optical return path portion 694 is relatively straight, embodiments are not so limited. For example, the return path may include a plurality of convoluted portions having a radius of curvature that is less than or greater than the convoluted portion 692, such that the light travel path is further extended.

  Further, each curved portion 682 may include other shapes of curvature. For example, the curved portion 682 includes one or more of flexures, arcs, bends, and bows, among other curved shapes, so that light is directed in different directions away from the original travel direction (downward). You may go out. In addition, such curvature may be optimized to have a curvature radius selected to achieve the desired ATR absorption signal or signal to noise ratio.

  In addition, in the same manner as described above with reference to FIG. 6B, the cross-sectional shape of the various portions of the optical structure 660a may be a polygon (triangle, square, rectangle, pentagon, hexagon, octagon, etc.) in addition to the illustrated circle. ), Other shapes such as oval or other suitable shapes.

Plasma-assisted coating of polymer substrates with nanoparticles The plasma process finds many applications in the manufacturing industry, including electronics, aerospace, automotive, steel, biomedical and toxic waste management, to name a few. . A gas phase plasma is an electrically neutral mixture containing neutral molecules, electrons, ions and radicals. In a gas phase plasma, energy (eg, RF or DC) is applied to a certain amount of gas in the chamber through one or more electrodes, so that electrons acquire sufficient kinetic energy to generate a certain amount of gas. It can be generated by colliding with atoms or molecules, leading to the formation of a gas phase plasma containing electrons, ions and radicals. As an example, when a certain amount of oxygen gas (O ₂ plasma) is subjected to sufficient energy to initiate plasma generation, the generated plasma may be electrons, oxygen radicals, O ₂ , O ₃ , O ^−. , O ²⁻ , O ⁺ , O ²⁺ and O ^{+2 and} other species. Thus, the generated reactive radical species can be utilized to perform various chemical operations, and the ionized atoms and molecular species can interact with the target surface of the article, eg, the substrate, through various interactions. It can be used to perform chemical and / or physical work. In most gas phase plasma processes, reactive radical species and / or ionized species come into contact with the target surface of the article to be modified.

  Some plasma processes, referred to herein as liquid-based plasma processes, generate a discharge in the liquid or use the liquid as an electrode. In some liquid-based plasma processes, a discharge is directly generated in the liquid between two electrodes placed inside the liquid. For example, in a process called Solution Plasma Processing (SPP), two electrodes are placed directly in a solution that may contain a chemical, eg, a precursor, and a high voltage is applied between the electrodes. Is applied to cause dielectric breakdown (eg, arc) of the liquid. Some other liquid-based plasma processes use a liquid that serves as one of the electrodes and the other electrode placed outside (eg, above) the liquid to generate a discharge on the liquid. . The liquid serves as an electrode by having conductive ions dissolved therein, and when connected to the immersed electrode, a discharge current flows through the liquid due to the ions in the liquid. In yet other liquid-based plasma processes, the discharge is generated in bubbles and cavities in the conductive liquid and is thus completely surrounded by the liquid in which the two electrodes are arranged. Accordingly, in a liquid-based plasma process according to the prior art, a discharge is generated by applying energy to a certain amount of liquid itself using one or more conductive electrodes submerged in the liquid.

  In the following, according to an embodiment, a gas phase plasma treatment is disclosed in which the article to be treated is at least partially submerged. Unlike other gas phase plasma processes, the article to be modified is not in direct contact with the gas phase plasma. In addition, unlike other liquid-based plasma processes, the liquid is not in direct contact with the electrode. Instead, the vapor phase plasma generated on the surface-immersed liquid composition causes or accelerates the deposition of material (eg, binder polymer) on the surface of the article.

In an embodiment, the plasma treatment method relates to the coating of an article with a material layer, for example a polymer layer and / or a nanoparticle layer. The method includes providing a liquid composition comprising a binder polymer chain and a solvent in a container, eg, an electrically insulating container. The binder polymer may have multiple functional groups, such as amine (NH ₂ ) groups. An article to be treated that includes a polymer surface is at least partially submerged in the liquid composition. The container with the at least partially submerged article is then placed in the plasma reactor chamber. Next, sufficient energy to generate a gas phase plasma is applied to a volume of gas over the surface of the liquid composition. A gas phase plasma generated from a quantity of gas and persisting over the surface of the liquid composition causes or accelerates the formation of a binder polymer layer on the polymer surface of the article. For example, chemical reactions between polymer functional groups (eg, NH ₂ groups) and polymer surface atoms (eg, oxygen atoms) can cause or accelerate deposition and can be accelerated. In some embodiments, the nanoparticles may then be attached to the binder polymer layer. In various embodiments, the gas phase plasma is not in direct contact with the polymer surface of the article on which the binder polymer layer is formed. Furthermore, the liquid composition, article and container are not electrically connected, resulting in an electrically floating state while deposition is triggered or accelerated.

Plasma Assisted Nanoparticle Coating Method Referring to FIG. 7, a method 700 for coating a polymer substrate with a layer of metal nanoparticles according to various embodiments is described. The method 700 includes, in process 710, providing a liquid composition that includes a binder polymer and a solvent. The method 700 additionally includes, in process 720, at least partially immersing the article comprising the polymer surface in the liquid composition. The polymer surface includes a polymeric material and does not include inorganic glass or crystalline material. The method 700 additionally includes, in process 730, applying a vapor phase plasma to the liquid composition to promote a chemical reaction between the binder polymer and the polymer surface to form a binder layer on the polymer surface of the article. . The method 700 further includes, after forming the binder layer, applying metal nanoparticles to the binder layer in process 740 to form the metal nanoparticle layer on the binder layer. In the following, with reference to FIGS. 8A-8E, the various stages of the process of coating a polymer substrate with a layer of metal nanoparticles will be described in more detail.

Liquid Composition FIG. 8A illustrates a process for preparing a liquid composition 800A comprising a binder polymer and a solvent, according to an embodiment, wherein the binder polymer has a plurality of functional groups. Liquid composition 800A includes a mixture or solution that includes solvent 808 and binder polymer 804. Binder polymer 804 may be dissolved in solvent 808 to form a solution, partially dissolved in solvent 808 to form a partial solution / partial mixture, or dissolved in solvent 808 Without forming, a mixture may be formed. In the illustrated embodiment, the liquid composition 800A is prepared in a container 850 that is electrically insulating. For example, the container 850 may be a dielectric container, for example, a light transmissive dielectric container such as a Petri dish. Binder polymer 804 is a polymer-based material that functions to immobilize nanoparticles on a substrate, and more specifically, to immobilize metal nanoparticles on a polymer substrate at a later stage in the process. A functional polymer-based material. The binder polymer 804 is a nanoparticle, for example, by binding to surface atoms of the polymer substrate at several positions in the chain of the binder polymer 804 and simultaneously binding to metal particles at several other positions in the chain. Can be fixed (details will be described later). In some embodiments, the binder polymer 804 includes a plurality of binder functional groups 816 that, in the illustrated embodiment, are amine terminated (NH ₂ ). There may also be other binder functional groups 816 such as thiol (SH), phosphonic acid (—PO (OH) ₂ or —PO (OR) ₂ ).

In some embodiments, the binder polymer 804 includes polydiallyldimethylammonium; polydiallyldimethylammonium chloride; polyallylamine hydrochloride; poly-4-vinylbenzyltrimethylammonium chloride; diethylenetriamine (DETA) (H ₂ N—CH ₂ CH _2- NH—CH ₂ CH ₂ —NH ₂ , an analog of diethylene glycol), triethylenetetramine (TETA) (H ₂ N—CH ₂ CH ₂ —NH—CH ₂ CH ₂ —NH—CH ₂ CH ₂ —NH ₂ ), tetraethylenepentamine _{(TEPA) (H 2 N-} CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH 2), pentaethylenehexamine (PEHA) _{(H 2} N-CH 2 _{CH 2 -NH-CH 2 CH 2} -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH 2), polyamines derived from ethylene amines including polyethylene amine; polyamidoamines And cationic polymers such as hyperbranched polymers including dendrimers, polypropylimine dendrimers and polyethyleneimine (PEI); or mixtures thereof. In other embodiments, the binder polymer 804 includes an anionic polymer, such as polyacrylic acid, sodium poly-4-styrenesulfonate, polyvinylsulfonic acid, polysodium salt, polyamino acid, or mixtures thereof. In some embodiments, the binder polymer includes linear or hyperbranched polyethyleneimine (PEI), ethylenediamine or other crosslinkable molecules suitable for forming amide bonds on the polymer substrate surface. In the embodiment illustrated in FIG. 8A, for purposes of illustration, the binder polymer 804 is an amine functionalized organic molecule comprising a polymer.

  The solvent can include any suitable solvent that can dissolve the binder molecules to form a solution. Suitable solvents include, for example, water, sodium hydroxide, ammonium hydroxide, or mixtures thereof, among other suitable solvents that can dissolve or retain the binder polymer 804 in the liquid composition 800A as a mixture. Is mentioned.

  Advantageously, in some embodiments, a desired optimization that is optimized in a subsequent process for the binding reaction between the binder polymer 804 and the surface atoms of the substrate and between the binder polymer 804 and the nanoparticles. The pH level of the liquid composition 800A can be controlled within the range, and a specific amount of the binder polymer 804 and the solvent 808 can be mixed at an appropriate volume ratio. In some embodiments, prior to mixing, solvent 808 may be prepared to have a pH level between about 5-9, between about 6-8, such as 7. After mixing, the binder polymer and solvent combination may be controlled to have a pH between about 8-12, between about 9-11, for example about 10. For example, if the binder polymer comprises TETA, the pH can be optimized using a volume ratio of 0.01 to 10% between water and TETA.

We also note that a binder polymer 804 having a specific molecular weight can be advantageous in optimizing the density of bonding sites between the binder polymer chain and the surface atoms and nanoparticles of the substrate. I found it. The molecular weight of the binder polymer 804 ranges from between about 100 daltons to about 1 × 10 ⁷ daltons, between about 1000 daltons to about 1 × 10 ⁶ daltons, or between about 1000 daltons to about 1 × 10 ⁵ daltons, for example, about 10,000 daltons. You can choose to have

Insulating Container Referring still to FIG. 8A, a liquid composition 800A is provided in a dielectric container 850 that is electrically insulating. Dielectric container 850 can be any suitable material that does not conduct electricity, such as polymer materials including polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), polystyrene (PS), polycarbonate (PC), to name a few. It can be formed from any insulating material. In some embodiments, the dielectric container 850 includes an inorganic glass or crystalline material, such as, for example, SiO ₂ or Al ₂ O ₃ .

  Liquid composition 800A may be prepared in a non-dielectric container, such as a conductive container, but prior to subjecting liquid composition 800A to plasma, as described in detail below with respect to FIG. It is transferred to the dielectric container 850. Providing a dielectric container 850 that includes an insulating material that does not conduct electricity may provide several advantages. For example, the dielectric container 850 includes a liquid composition 800A against arcing and / or dielectric breakdown when the dielectric container 850 containing the liquid composition 800A is subjected to plasma conditions (described in detail with respect to FIG. 8C). And the protection of the dielectric container 850 can be improved. That is, when the dielectric container 850 is installed on a substrate holder that can function as an electrode of the plasma processing chamber, the dielectric container 850 electrically stores the liquid composition and the article that are later put in the liquid composition 800A. Can be suspended.

Immersion of Polymer Surface into Liquid Composition FIG. 8B illustrates the step of at least partially immersing or submerging an article, eg, a polymer substrate 860, in the liquid composition 800B, according to an embodiment. According to an embodiment, the polymer substrate 860 has a polymer surface 860s. The portion of the polymer surface 860S that is exposed to the liquid solution 800B (eg, immersed in the liquid composition 800B) is in direct contact with the liquid composition 800B. According to embodiments, the polymer surface 860S can be, for example, polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), polystyrene (PS), polycarbonate (PC), and cyclic olefin polymer (COC), to name a few. , Cyclic olefin copolymers), etc., which have exposed carbon-based chains. In some embodiments, the polymer surface 860S does not include inorganic glass or crystalline material, such as, for example, SiO ₂ or Al ₂ O ₃ .

  In the illustrated embodiment, the substrate 860 is immersed in a liquid composition 800B that includes a mixture or solution comprising a solvent 808 and a binder polymer 804, as described above with respect to FIG. 8A. A polymer surface 860S having a plurality of substrate functional groups 832 contacts a binder polymer 804 having a binder functional group 816. The polymer surface 860S has a plurality of substrate functional groups 832 that are adapted to chemically react with the binder functional groups 816. In the illustrated embodiment, the substrate functional group 832 is a carbonate group (—O— (C═O) —O—). There may be other functional groups 832 on the polymer surface 860S, such as isothiocyanate ends, isocyanate ends and amine ends.

  In FIG. 8B, for purposes of illustration, the substrate 860 is illustrated as fully immersed in the liquid composition 800B such that the entire surface of the polymer surface 860S is in contact with the liquid solution 808. However, it is understood that in other embodiments, some portions of the substrate 860 may be immersed and other portions may remain outside the liquid composition 800B. As will be described later, the portion that contacts the liquid composition 808 reacts with the binder polymer 804, and then the other portion that is covered by the nanoparticles but does not contact the liquid composition 808 reacts with the binder polymer 804. It is understood that there is no coating with nanoparticles.

  It is further understood that in other embodiments, the substrate 860 may have a non-active surface 860NS. In these embodiments, the non-active surface 860NS is a substrate in which metal nanoparticles may not be desired so that the metal nanoparticles can be selectively formed only on the polymer surface 860S, as described below with respect to FIG. 8E. There may be 860 surfaces. For purposes of illustration only, the non-active surface 860NS is formed on the side surface of the substrate 860. However, it is understood that the non-active surface 860NS can be formed anywhere on the surface of the substrate 860. For example, both the non-active surface 860NS and the polymer surface 860 can be formed on the top surface of the substrate 860. The non-active surface 860NS may be formed from a different material compared to the polymer surface 860S, may be formed from the same material as the polymer surface 860S, but may be functionalized with a different functional group than the substrate functional group 832 Alternatively, it may be unfunctionalized or deactivated such that reaction with the functional group 816 of the binder polymer 804 is prevented in a later process (FIG. 8C).

Plasma Reactor for Vapor Phase Plasma Treatment Under Solution Immersion FIG. 8C illustrates applying a vapor phase plasma to a liquid composition after submerging at least a portion of a polymer substrate 860 into the liquid composition, according to an embodiment. And a step of causing a chemical reaction between the binder functional group of the binder polymer and the substrate functional group of the polymer surface, thereby forming a binder layer on the polymer surface 860S. The container 850 is installed inside the plasma reactor 880.

  The plasma reactor 880 is configured for gas phase plasma treatment of articles under solution immersion according to an embodiment. The plasma reactor 880 includes at least one electrode that can be energized to add energy to gas phase atoms or molecules above the solvent surface and initiate plasma generation. In the illustrated embodiment, the reactor 880 includes an upper electrode 862 and a lower electrode 864 and is configured to accommodate a vessel 850. Reactor 880 is configured to receive at least one gas species for generating gas phase plasma through one or more of gas inlets 868 connected to reactor 880. The reactor may be connected to a vacuum pump (not shown) to control the pressure inside the reactor. In particular, the reactor controls the pressure inside the reactor so that the components do not completely evaporate during the gas phase plasma treatment, and all of the components of the liquid composition 800C, including solvent 808 and binder polymer 836, are controlled. It is configured to maintain a suitable partial pressure. For example, the reactor may be controlled at and maintained in subatmospheric and atmospheric conditions by, for example, controlling the pressure between about 100 mtorr and about 780 torr, between about 1 torr and about 760 torr, or between about 100 torr and about 760 torr. It is configured to generate and maintain a plasma below.

  After receiving at least one gas species through at least one valve 868, a quantity of gas between the surface of the liquid composition 800C and the upper electrode 862 through at least one of the upper electrode 862 and the lower electrode 864, Energy 876 is applied, thereby generating a plasma 872 between the surface of the liquid composition 800C and the upper electrode 862. Although the upper electrode 862 and the lower electrode 864 are illustrated as being disposed within the reactor 880, one or both of the upper electrode 862 and / or the lower electrode 864 are disposed outside the reactor 880. It is understood that

  As defined in this specification, an electrode of a plasma reactor is an element that can add energy to a certain amount of gas, and examples thereof include a capacitor plate and an inductor coil. Without loss of generality, in one embodiment, the plasma reactor may be configured such that the DC power or the first electrode 862 or the second electrode 864 may form a capacitively coupled plasma discharge. A DC or AC plasma reactor to which AC power is applied. In some embodiments, where the plasma reactor is a DC plasma reactor, the plasma reactor is configured as a pulsed DC plasma reactor where DC power can be applied in a pulsed form. The pulsed DC voltage may be bipolar or unipolar. If it is bipolar, the DC voltage may be symmetric or asymmetric in amplitude of opposite polarity. DC or AC power can be applied through one or both of the first electrode 862 and the second electrode 864 and can be driven by a power source 866. In the illustrated embodiment, both the first electrode 862 and the second electrode 864 are connected to a power source 866, but in other embodiments, only one of the two electrodes is “hot” and the other Is understood to be electrically grounded or floating. In addition, if one of the two electrodes is “hot” and receives pulsed DC or AC power, the other electrode is biased, eg, DC bias, so that the charged species can accelerate toward the liquid composition 800C. Can be placed below.

  Other types of plasma generation may be used. For example, the plasma reactor 880 can be energized by a current generated by a time-varying magnetic field that can increase the density of the plasma under some circumstances, an inductively coupled plasma (ICP) reactor or electron cyclotron resonance (ECR). A plasma reactor may also be used.

Gas Phase Plasma Treatment to Form a Polymer Layer Under Solution Immersion Referring still to FIG. 8C, energy 876 is transmitted through at least one of the upper electrode 862 and the lower electrode 864 to generate a liquid phase plasma 872. A certain amount of gas is applied between the surface of the object 800 </ b> C and the upper electrode 862.

  In the illustrated embodiment, the substrate 860 is fully submerged in the liquid composition 800C such that the liquid composition 800C separates the polymer surface 860S and the plasma 872. That is, the polymer surface 860S to which the binder polymer 836 is attached is not directly exposed to the plasma 872.

  In the illustrated embodiment, the entire substrate 860 is submerged so that the entire polymer surface 860S is submerged in the liquid composition 800C. However, in other embodiments, the substrate 860 is partially submerged so that only a portion of the substrate 860 is submerged in the liquid composition 800C.

In some embodiments, the composition of the gas in the plasma reactor 880 is introduced by introducing one or more gases into the plasma reactor 880 through the one or more gas inlets 868 prior to generating the plasma 872. Is adjusted. In some embodiments, the amount of gas may include an inert gas such as He, Ar, Ne and Xe, or mixtures thereof. In some embodiments, the amount of gas is selected from among other gases: O ₂ , O ₃ , N ₂ , H ₂ , NH ₃ , N ₂ O, and NO, and mixtures thereof. A seed or a plurality of gases may be included. However, since the nitrogen atom is provided by the binder functional group (for example, NH ₂ ) itself, in order to cause a chemical reaction between the base functional group and the binder functional group containing nitrogen, N ₂ or NH ₃ or the like It is understood that no separate nitrogen containing any gas is required.

  With continued reference to FIG. 8C, a plasma 872 is generated above the surface of the liquid composition 800C, for example, just above the liquid composition 800C. In the illustrated embodiment, using one of the power supply methods described above, by applying energy to a certain amount of gas disposed between the first electrode 862 and the liquid composition 800C surface, Plasma 872 is generated between the two. For example, in some embodiments, one or both of electrode 860 and / or electrode 862 may have a regulated frequency of between about 10 KHz and about 1 MHz, such as 100 KHz, under alternating or pulsed DC conditions. Can be driven. In other embodiments, the frequency may be higher, for example, a regulated RF frequency of 13.56 MHz, or a regulated microwave frequency of 2.45 GHz. The maximum and minimum amplitude of the applied bias may be, for example, between about 100 V and about 100 kV, or between about 1 kV and about 10 kV.

Various optical and chemical processes of the plasma 872 glow discharge can contribute to the reaction between the binder functionality and the substrate functionality. For example, without being bound by any theory, the reaction can be at least partially facilitated by the diffusion of various species, such as radicals, from the plasma into the liquid composition 800C. In addition, without being bound by any theory, the reaction may occur in various atomic transitions between species states (eg, excited and ground states) of the plasma 872 and / or in visible and ultraviolet light. It can also be facilitated by dissociation and recombination reactions between various neutral and charged species in plasma 872, which can cause photon dissipation. Thus, without being bound by such theory, the plasma species and photons generated from the sustained plasma 872 can cause binder functional groups 816 of the binder polymer 836, eg, NH ₂ and at least a portion of the polymer surface 860S. A chemical reaction is caused between oxygen atoms of surface functional groups, for example carbonate groups (—O— (C═O) —O—), and, as in the embodiment illustrated in FIG. 8C, a binder polymer. An 836 layer is formed on the polymer surface 860S.

  Other embodiments are possible where the plasma does not sustain a glow discharge. Still other embodiments are possible where the plasma is generated outside the chamber space above the surface of the liquid composition 800C and then transferred to the space above the surface of the liquid composition 800C.

  We have plasma 872 for between about 1 millisecond and about 1 hour, between about 1 second and about 1 hour, between about 1 second and about 10 minutes, or between about 1 second and about 5 minutes. It has been found that a chemical reaction is triggered when sustained over the liquid composition 800C for the duration of the plasma treatment in between. The pressure in the reactor 880 during the duration may be applied under an average pressure between about 1 and about 760 torr and maintained in a steady state by a constant flow of gas or by a pressure servomechanism.

  As described above with respect to FIG. 8A, the container 850 is an electrically insulating container, such as a dielectric container. Thus, in the illustrated embodiment, the substrate 860 and the liquid composition 800C are not in direct contact with the lower electrode 864. In addition, no other direct external electrical connection is made to the substrate 860 or the liquid composition 800C, so that in the illustrated embodiment, the substrate 860 is electrically floating. doing. However, in other embodiments, the substrate 860 may be grounded or placed under an independent bias, such as a DC bias. However, even when the substrate 860 is electrically floating, the lower electrode 864 is used to alter (eg, accelerate) the charged species in the plasma 872 toward the liquid composition 800C. Is independently biased, for example, may be DC biased.

The combination of the various plasma conditions described above results in an enhanced chemical reaction between the binder polymer 804 (FIG. 8B) and the polymer surface 860S to form the binder layer 836 on the polymer surface 860S. The binder layer 836 is chemically bonded to the polymer surface 860S through a reactive functional group 840 resulting from a chemical reaction between the binder functional group 816 and the substrate functional group 832. In the illustrated embodiment, the NH ₂ groups (FIG. 8B) of the binder polymer 804 react with the oxygen atoms of the carbonate groups (—O— (C═O) —O—) of the polymer surface 860S, causing the binder layer 836 to Form.

Preparation of Nanoparticle Solution Referring to FIG. 8D, after forming a binder layer 836 that causes a chemical reaction between the binder functional group and the substrate functional group to chemically adhere to the polymer surface 860S, the unreacted binder polymer. The liquid solution 800C (FIG. 8C) containing the liquid may be discarded from the container and the substrate 860 may be washed. Thereafter, the substrate 860 may be submerged in a nanoparticle solution 800D comprising a nanoparticle solvent 818 and a plurality of metal nanoparticles 828.

  The nanoparticle solution 800D may be a suitable solvent, such as an aqueous solution, such as distilled water. The metal nanoparticles 828 may include one of the suitable metal materials having the form described above with respect to FIGS. 2A and 2B and stabilized by a surfactant 832 that can prevent the nanoparticles 828 from solidifying. . Surfactant 832 is exemplified by sodium citrate, ascorbic acid, 4-mercaptobenzoic acid, meso-2,3-dimercaptosuccinic acid, mercaptosuccinic acid, succinic acid, sodium dodecyl sulfate, and sodium octyl sulfate. , Sodium decanesulfonate, lysine, glucose, cetyltrimethylammonium bromide (CTAB), hexadecyltrimethylammonium bromide, tetradecylammonium bromide, tetraoctylammonium bromide, tetrahexylammonium bromide, dodecyltrimethylammonium bromide, and cetylpyridinium chloride Suitable compounds may be used.

  In nanoparticle solution 800D, nanoparticles 828 are between about 1 nm and about 10 nm, such as about 5 nm; between about 5 nm and about 20 nm, such as about 10 nm; between about 10 nm and about 30 nm, such as about 20 nm; Between about 40 nm, such as about 30 nm; between about 30 nm and about 50 nm, such as about 40 nm; between about 40 nm and about 60 nm, such as about 50 nm; between about 50 nm and about 80 nm, such as about 60 nm; Between 100 nm, for example about 80 nm; between about 80 nm and about 150 nm, for example about 100 nm; between about 100 nm and about 200 nm, for example about 150 nm; between about 150 nm and about 250 nm, for example about 200 nm; about 200 nm to about 300 nm Between, for example, about 250 nm; between about 250 nm and about 400 nm, such as about 300 nm; Having a median diameter selected to be between 300 nm and about 700 nm, such as about 500 nm; between about 500 nm and about 900 nm, such as about 700 nm; or between about 700 nm and about 1100 nm, such as about 900 nm. Good. The median diameter may be, for example, the minimum lateral dimension of nanoparticles 828 measured along 10204 in the direction of symmetry of nanoparticles 828.

  In some embodiments, the median size of the nanoparticles 828 can be, for example, about 0.1% to about 2%, about 2% to about 4%, about 4% to about 6%, about 6% to about 8%. A relatively narrow standard deviation of about 8% to about 10%.

  It is understood that the specific median diameter of the nanoparticles 828 can be selected such that the resulting LSPR peak wavelength is in the specific wavelength band desired. In various embodiments, the median diameter is such that the LSPR peak is between about 515 nm and about 525 nm, such as 520 nm; between about 525 nm and about 535 nm, such as 530 nm; between about 535 nm and about 545 nm, such as 540 nm; May be selected to be between about 555 nm, such as 550 nm; between about 555 nm and about 565 nm, such as 560 nm; between about 565 nm and about 575 nm, such as 570 nm; or between about 575 nm and about 585 nm, such as 580 nm. .

  The particle size can be selected to have a specific LSPR peak absorption wavelength, and the standard deviation of the particle size is to have a LSPR spectrum of a specific width, eg, less than 100 nm or less than 80 nm, as described above with respect to FIGS. 4A and 4B. It is understood that it can be adjusted to.

Step of Attaching Metal Nanoparticles to Binder Polymer Referring to FIG. 8E, after submerging the substrate 860 in a nanoparticle solution 800E that includes a nanoparticle solvent 818 and metal nanoparticles 828, the nanoparticles 828 are bound to a binder functional group 816. Of which remains unreacted or remains. According to an embodiment, the chemical reaction may be unreacted or remaining binder functional groups 816 present after attachment of the binder polymer 836 to the polymer surface 860S, such as NH, as described above with respect to FIG. 8D. ₂ , thereby forming a layer of metal nanoparticles on the polymer surface 860S. In the illustrated embodiment, the binder functional group 816 attached to the substrate functional group 832 (FIG. 8B) is the same as the binder functional group 816 attached to the nanoparticle 828, but the binder functional group is different. Other embodiments are also possible that selectively attach only to one of the substrate functional groups 832 or nanoparticles 828.

When formed, the nanoparticle layer may have any of the particle size ranges, standard deviations, and peak wavelengths described above with respect to FIG. 8D. In addition, the layer of metal nanoparticles has a surface density that can be optimized for LSPR based at least in part on the surface density of the nanoparticles and the distance between the nanoparticles. As an example, for nanoparticles having a particle size range between about 1 nm and about 50 nm, the surface density is between about 1 × 10 ⁸ / cm ² and about 1 × 10 ¹³ / cm ² , between about 5 × 10 ⁸ / cm ² and It can be between about 5 × 10 ¹² / cm ² or between about 1 × 10 ⁹ / cm ² and about 2 × 10 ¹² / cm ² .

  In addition, the nanoparticles are median nanoparticles between 1 nm and about 10 nm, between 10 nm and about 100 nm, between 100 nm and about 1000 nm, between 1000 nm and about 10,000 nm, or between 10,000 nm and about 100,000 nm. It may have a median inter-nanoparticle distance. In addition, the nanoparticles may be between about 0.1% and about 2%, between about 2% and about 4%, between about 4% and about 6%, or between about 6% and about 8%. It may have a median nanoparticle distance standard deviation.

Substrate Surface Orientation In FIG. 8C, for purposes of illustration only, plasma 872 is generated above the surface of liquid composition 800C, thereby forming a layer of binder polymer 836 on polymer surface 860S facing plasma 872. It is causing or accelerating. However, other embodiments are possible where the polymer surface on which the polymer layer is formed does not face the plasma 872, or at least partially faces away from the plasma, and the substrate 860 is curved Can include surfaces having deflections, arcs, bends, bows, twists, rings, bends, and the like. As an illustrative example, in FIG. 8F, the polymer transparent substrate 861 has cavities having internal (top, bottom, and side) surfaces. The substrate 861 may have an opening so that the cavity is filled with the solvent 818 / liquid composition 800F. Similar to the substrate 860 of FIG. 8C, the inner surface of the cavity of the substrate 861 contacts the plasma (similar to FIG. 8C, not shown) when submerged in the solvent 818 / liquid composition 800F. Without being separated from the plasma by solvent 818 / liquid composition 800F. In addition, in addition to being separated by the solvent 818 / liquid composition 800F, the inner surface is further separated from the plasma by the upper portion of the substrate 861 itself. The lower surface inside the cavity faces the plasma, the upper surface inside the cavity faces away from the plasma, and the side surface inside the cavity is at a certain angle, eg 90 °, from the horizontal surface of the solvent 818 / liquid composition 800F. It is oriented in the direction forming When processed under process conditions similar to those described above with respect to FIGS. 8A-8E, the top, bottom, and side surfaces advantageously have a binder polymer 836 layer having a substantially uniform thickness. On average, the layer thicknesses of the binder polymers 836 on the different surfaces are within about 10% of each other, or within about 5% of each other. In addition, the difference in surface density of the nanoparticles on the top, bottom, and side surfaces can be less than about 10%, or less than about 5%.

Autocalibration LSPR Measurement System and Method When performing LSPR measurements on the samples described above with respect to FIGS. 2A and 2B, quantitative information is often required regarding the amount of target molecules attached to the polymer substrate. However, since the signal from the LSPR measurement may fluctuate greatly due to fluctuations in external conditions such as temperature, for example, quantification of the amount of the target molecule is often hindered. In the following, an automatic calibration LSPR measurement system and related measurement methods are described, where a standard that varies by the same amount as the sample as the environment changes is advantageously used, and each time an LSPR measurement is performed, the LSPR signal Increase and decrease are automatically calibrated.

Self-calibrating LSPR Measurement System FIG. 9 shows an automatic calibration localized surface plasmon resonance (LSPR) measurement system 900 for detecting target molecules that may adhere to the surface of a test medium, according to an embodiment. The automatic calibration LSPR system 900 includes a first reference test medium 902, a sample measurement test medium 904, and a second reference test medium 906. Similar to the LSPR measurement system and method described above with respect to FIGS. 3E and 3F and FIGS. 6B-6D, the auto-calibration LSPR system 900 was configured to illuminate each of the media 902, 904, and 906 with incident light 914, respectively. A plurality of light sources 910 and a plurality of photodetectors 930 configured to detect light 938 transmitted through each of media 902, 904, and 906 are included.

  Each of media 902, 904, and 906 has a respective substrate 940, 960, and 980 that has a first reference coverage analysis region 948, a sample coverage analysis region 968, and a second reference coverage analysis region 988. Each of media 902, 904, and 906 may have uncoated portions 942, 962, and 982. Each of the analysis regions 948, 968 and 988 has a respective analysis region surface comprising the transparent polymer material described above. At least a portion of the surface of the coating analysis regions 948, 968, and 988 has a layer of metal nanoparticles (not shown for clarity) formed thereon using the method described above with respect to FIGS. 8A-8F. . In addition, each of the media 902, 904, and 906 holds and sinks at least the coating analysis regions 948, 968, and 988 of the respective substrates 940, 960, and 980 in the liquid solution 936 that contains the target molecule to be detected. It is comprised so that it may be installed in the container 932 comprised.

  The sample coverage analysis region 968 of the sample measurement test medium 904 is similar to that described above with respect to FIGS. 3E and 3F and FIGS. 6B-6D so that the target molecules in the liquid solution 936 can be captured by the capture molecules. It has capture molecules (not shown for clarity) formed on at least some of the nanoparticles on the coating analysis region 968. Unlike the coating analysis region 968, the first and second coating reference analysis regions 948 and 988 are not configured to capture target molecules. Instead, the first and second coating reference analysis regions 948 and 988 have first and second concentrations of reference molecules attached to the nanoparticles.

  In some embodiments, all coating analysis regions 948, 968 and 988 have the same or similar standard type and concentration of nanoparticles, which can be of the material, shape and concentration described above with respect to FIG. 8E. In other embodiments, each of the coating analysis regions 948, 968, and 988 has a different standard type and / or concentration of nanoparticles.

  The first reference coating analysis region 948 has a first predetermined concentration of reference molecules attached to the nanoparticles in the region, and the first reference refractive index measured from the region is from the sample coating analysis region 968. It is different from, for example, smaller than the measured refractive index. The second reference coating analysis region 988 has a second predetermined concentration of reference molecules attached to the nanoparticles in the region, and the second reference refractive index measured from the region is from the sample coating analysis region 968. It differs from the measured refractive index, for example, becomes larger. In some embodiments, the reference molecules attached to the first and second reference coating analysis regions 948 and 988 include the same type of reference molecule. However, in other embodiments, the reference molecules attached to the first and second reference coverage analysis regions 948 and 988 may be heterogeneous reference molecules. In addition, the reference molecule attached to one or both of the first analysis region 948 and the second analysis region 988 may include the same or different types of molecules compared to the target molecule. However, regardless of the type of molecules attached to the coating analysis regions 948, 968 and 988, all three coating analysis regions are configured to contact the solution 936 that may contain the target molecule, while the sample Only the coating analysis region 968 is configured to capture target molecules, and the first and second reference coating analysis regions 948 and 988 have a predetermined concentration of reference molecules already formed therein, and target molecules Is not configured to capture further.

Test media 902, 904, and 906 are configured as a permeable LSPR measurement system similar to that described with respect to FIGS. 3E and 3F, but each of test media 902, 904, and 906 is associated with FIGS. 6A-6D. It will be understood that it may be configured as any one of the aforementioned attenuated total internal reflection (ATR) LSPR measurement test media. In these embodiments, each of the test media 902, 904, and 906 allows the light transmitted through the light structure (similar to the light structures 660a-660c in FIGS. 6B-6D) to be attenuated total internal reflection as described above. It is configured to be induced between a first end and a second end under (ATR) conditions. That is, since the second refractive index n ₂ of the solution 936 is smaller than the first refractive index n ₁ of the coating analysis regions 948, 968 and 988, the light beam traveling in the optical structure is totally internally reflected, and the optical Return to the inside of the structure.

LSPR Self-calibration Measurement Method Referring to FIG. 10, the concentration of target molecules attached to the sample coating analysis region 968 based on the effective refractive index and absorbance or peak wavelength values measured from the coating analysis regions 948, 968 and 988, respectively. Can be determined. FIG. 10 shows a graph 1000 illustrating a method for quantifying the amount of target molecules attached to a polymer-based test medium under temperature variation conditions by using a control medium having a known concentration of target molecules, according to an embodiment. . The x-axis represents the effective refractive index (n _eff ), and the y-axis represents the absorbance or peak absorption wavelength (λ _max ). Curves 1010a and 1010b represent absorbance or λ _max versus effective refractive index curves measured at two different temperatures, a first temperature T ₁ and a second temperature T ₂ > T ₁ , respectively.

The first curve 1010a measured at T ₁ has a first reference coverage analysis region 948, a sample coverage analysis region 968, and a second reference coverage analysis region 988, respectively, described above with respect to FIG. , 904 and 906, corresponding to the measured refractive index, a first reference cold absorbance 1012a, a sample cold absorbance 1014a, and a second reference cold absorbance 1016a. As described above with respect to FIG. 9, the first reference coverage analysis region 948 and the second reference coverage analysis region 988 each have a predetermined concentration of reference molecules that is lower or higher than the concentration of target molecules in the sample coverage analysis region 968. Have.

  The inventors have found that the concentration of the target molecule is linearly dependent on the refractive index, regardless of whether the reference molecule is the same or different from the target molecule. Accordingly, based on the measured refractive index value of the sample coating analysis region 968, the quantification of target molecules attached to the sample coating analysis region 968 can be determined.

The inventors have also found that the measured absorbance and / or λ _max is higher at a given refractive index value when measured at elevated temperatures. Furthermore, the inventors have found that the amount of absorbance and / or λ _max upward shift is independent of the concentration of the target or reference molecule. That is, the slopes 1018a and 1018b are substantially the same between the first curve 1010a and the second curve 1010b. Accordingly, the second curve 1010b measured at _{T 2} are, first reference hot absorbance 1012b, a sample hot absorbance 1014b and the second reference hot absorbance 1016b, a first reference low absorbance 1012a, samples low absorbance 1014a Compared with the second reference low-temperature absorbance 1016a, the absorbance and / or λ _max are shifted by the same value, respectively. Thus, a quantitative measurement of the concentration of the target molecule is accurately determined from the sample coating analysis region 968 at different temperatures without independently determining the reference concentration of the reference coating analysis regions 948 and 988 at different temperatures. obtain.

  While the invention has been described in terms of particular embodiments, other embodiments will be apparent to those skilled in the art, including embodiments that do not realize all of the features and advantages described herein. Embodiments are also within the scope of the present invention. Furthermore, the various embodiments described above can be combined to provide further embodiments. In addition, certain features shown in the context of one embodiment can be incorporated into other embodiments. Accordingly, the scope of the invention is defined only by reference to the appended claims.

DESCRIPTION OF SYMBOLS 100 Planar metal thin film base SPR measurement system 110 Light source 114 Incident light 118 Reflected light 120 Test medium 124 Base material 128 Metal thin film 130 Detector 132 Channel 136 Glass block 140 Target molecule 142 Solution 144 Capture molecule 148 Analytical part 200 Localized surface plasmon Resonance (LSPR) measurement system 210 Light source 214 Incident light 220 Test medium 250 Container 250S inner surface 224 Substrate 224S inner surface 228 Metal nanoparticle layer 230 Photodetector 232 Solution 238 Transmitted light 240 Target molecule 244 Capture molecule 250S Inner surface 300 LSPR test medium 300a Test medium 300b Test medium 304 Outer case 310 Front part 314 Incident light 320 Rear part 328 Metal nanoparticles 330 Front opening 332 Solution 334 Rear opening 33 Transmitted light 338a transmitted light 338b transmitted light 340 light incident window (Figure 3A)
340 Target molecule (Figure 3E / D)
344 Capture molecule 350 Container 354 Light entrance window 358 Light exit window 360 Sample substrate 360a Sample substrate 360b Sample substrate 360c Sample substrate 362 Handle region 364 Opening 368 Analysis region 368F Front surface 368R Rear surface 400 Graph 410a Absorbance spectrum 420a Absorbance spectrum 430a Absorbance spectrum 450 Graph 460 Absorption difference spectrum 470 Absorption difference spectrum 480 Absorption difference spectrum 600 LSPR test medium 610 Light source 614 Incident light 628 Metal nanoparticle layer 628a Container 628b Container 628c Container 630 Light detector 632 Solution 638 Target molecule 644 Capture molecule 660 Light structure 660a Light structure 660b Light structure 660c Light structure 660S End surface 660S1 First Surface 660S2 Second surface 662a Optical portion 662b Optical portion 662c Optical portion 664a Uncoated portion 664b Uncoated portion 664c Uncoated portion 664S1 Surface 664S2 Surface 664S3 Surface 668a Analysis region 668b Analysis region 668c Analysis region 668c Analysis region 668c Analysis region 668c 668S3 Analysis region surface 670 Test medium 672 Total internal reflection light 680 Test medium 682 Curved part 690 Test medium 692 Rotating part 694 Optical return part 800A Liquid composition 800B Liquid composition 800C Liquid composition 800D Nanoparticle solution 800E Nanoparticle solution 800F Liquid Composition 804 Binder polymer 808 Solvent 816 Binder functional group 818 Nanoparticle solvent 828 Metal nanoparticle 832 Substrate functional group (FIG. 8B)
832 Surfactant (Figure 8E)
836 Binder polymer 840 Reactive functional group 850 Dielectric container 860 Polymer substrate 860NS Inactive surface 860S Polymer surface 861 Substrate 862 Upper electrode 862 First electrode 864 Lower electrode 864 Second electrode 866 Power supply 868 Gas inlet 868 Valve 872 Plasma 876 Energy 900 Automatic calibration LSPR system 902 First reference test medium 904 Sample measurement test medium 906 Second reference test medium 910 Light source 914 Incident light 930 Photodetector 932 Container 936 Liquid solution 938 Light 940 Substrate 942 Uncoated portion 948 First reference coverage analysis region 948 Reference coverage analysis region 960 Base material 962 Uncovered portion 968 Sample coverage analysis region 968 Sample coverage analysis region 980 Base material 982 Uncovered portion 988 Second reference coverage analysis region 1 10a first curve 1010b second curve 1012a first reference low temperature absorbance 1012a first reference low temperature absorbance 1012b first reference high temperature absorbance 1014a sample low temperature absorbance 1014a sample low temperature absorbance 1014b sample high temperature absorbance 1016a second reference low temperature absorbance 1016a Second reference low temperature absorbance 1016b Second reference high temperature absorbance 1018a slope 1018b slope

Claims (35)

Providing a liquid composition comprising a binder polymer and a solvent, wherein the binder polymer comprises amine ends;
Providing an article comprising a polymer surface having one or more functional groups that react with amines, wherein the polymer surface does not contain inorganic glass or crystalline material;
Contacting the liquid composition with the polymer surface;
Applying a gas phase plasma to the liquid composition in contact with the polymer surface for a period of 1 millisecond to 1 minute so that the liquid composition is sandwiched between the gas phase plasma and the polymer surface; Forming a polymer layer comprising a cross-linked binder polymer bonded to the polymer surface by a chemical reaction between an amine end of the binder polymer and one or more functional groups on the polymer surface, wherein the gas phase plasma comprises A method of forming a polymer layer on a polymer substrate, comprising the steps of free of ammonia. Binder polymer
Polydiallyldimethylammonium; polydiallyldimethylammonium chloride; polyallylamine hydrochloride; poly-4-vinylbenzyltrimethylammonium chloride; diethylenetriamine (DETA), an analog of diethylene glycol, H ₂ N—CH ₂ CH ₂ —NH—CH ₂ CH ₂ -NH _2, triethylenetetramine _{(TETA), H 2 N-} CH 2 CH 2 -NH-CH 2 CH 2 -NH-CH 2 CH 2 -NH 2, tetraethylenepentamine _{(TEPA), H} 2 N —CH ₂ CH ₂ —NH—CH ₂ CH ₂ —NH—CH ₂ CH ₂ —NH—CH ₂ CH ₂ —NH ₂ , pentaethylenehexamine (PEHA), H ₂ N—CH ₂ CH ₂ —NH—CH _{2 CH 2 -NH-CH 2 CH 2} -NH- _{H 2 CH 2 -NH-CH 2} CH 2 -NH 2, polyamines derived from ethylene amines including polyethylene amine; polyamidoamine dendrimer, poly propyl dendrimers, hyperbranched polymers including polyethylene imine (PEI); or The method of claim 1, comprising one or more selected from the group consisting of these mixtures.   The polymer surface comprises at least one light transmissive polymeric material selected from the group consisting of polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), polystyrene (PS), and polycarbonate (PC). The method described.   The method according to claim 1, wherein the liquid composition is contained in an electrically insulating container while the gas phase plasma is applied, and the container is disposed between two electrodes of the plasma chamber.   The method of claim 1, wherein the liquid composition further comprises water and NaOH.   The step of applying the gas phase plasma includes the step of generating the gas phase plasma using a plasma generator having at least one electrode, and at least one of the plasma generators while applying the gas phase plasma. The method of claim 1, wherein none of the electrodes is in electrical contact with the liquid composition.   Contacting the liquid composition with the polymer surface comprises immersing the article in the liquid composition such that the polymer surface is submerged in the liquid composition when the gas phase plasma is applied; The method of claim 1.   The method of claim 1, wherein the article is in a shape selected from the group consisting of laminae, strips, cavities, cylinders, cylinders, fibers, coils, U-shapes, vine windings and spirals.   The method of claim 1, wherein the article comprises another surface facing away from the polymer surface, wherein the liquid composition is not in contact with another surface and no polymer layer is formed on the other surface.   The polymer surface is referred to as the first polymer surface, the polymer layer is referred to as the first polymer layer, the article includes a second polymer surface facing away from the first polymer surface, and the liquid composition is A second polymer layer is formed on the second polymer surface when also in contact with the second polymer surface and the gas phase plasma is also applied to the liquid composition in contact with the second polymer surface. Item 2. The method according to Item 1.   The polymer surface is referred to as the first surface, and the article further includes a second surface, the first surface is in contact with the liquid composition, but the second surface is liquid while applying the gas phase plasma. The method of claim 1, wherein the method is not in contact with the composition.   The method of claim 11, wherein the article further comprises a thin film on the second surface such that the liquid composition does not contact the second surface during application of the gas phase plasma.   The article includes a cavity, the first surface is the inner surface of the cavity, the second surface faces away from the first surface and is located outside the cavity, and the liquid composition is the second surface Contacting the liquid composition with the polymer surface such that the liquid composition contacts the first surface while preventing contact with the liquid composition comprises filling at least a portion of the cavity with the liquid composition. Item 12. The method according to Item 11. A method of forming a nanoparticle layer comprising:
The method of claim 1 for forming a polymer layer on a polymer surface of an article;
Applying the metal nanoparticles to the polymer layer after forming the polymer layer and forming a single layer of metal nanoparticles on the polymer layer.   15. The method of claim 14, wherein the step of applying metal nanoparticles is performed after completion of the step of applying gas phase plasma.   15. The method of claim 14, wherein the article is in a shape selected from the group consisting of a sheet, strip, cavity, cylinder, cylinder, fiber, coil, U-shape, vine winding and helix.   15. The method of claim 14, wherein the article comprises another surface facing away from the polymer surface, the liquid composition is not in contact with the other surface, and no polymer layer is formed on the other surface.   The polymer surface is referred to as the first polymer surface, the polymer layer is referred to as the first polymer layer, the article includes a second polymer surface facing away from the first polymer surface, and the liquid composition is A second polymer layer is formed on the second polymer surface when the second polymer surface is also contacted and a gas phase plasma is also applied to the liquid composition contacting the second polymer surface. 15. The method of claim 14, wherein the metal nanoparticles are also applied to the second polymer layer to form a single layer of metal nanoparticles on the layer.   The polymer surface is referred to as the first surface, and the article further includes a second surface, the first surface is in contact with the liquid composition, but the second surface is liquid while applying the gas phase plasma. Without contact with the composition, a single layer of metal nanoparticles is formed on the polymer layer on the first surface, but no layer of such metal nanoparticles is formed on the second surface; The method according to claim 14.   The method of claim 19, wherein the article further comprises a thin film on the second surface such that the liquid composition does not contact the second surface during application of the gas phase plasma.   The article includes a cavity, the first surface is the inner surface of the cavity, the second surface faces away from the first surface and is located outside the cavity, and the liquid composition is the second surface Contacting the liquid composition with the polymer surface such that the liquid composition contacts the first surface while preventing contact with the liquid composition comprises filling at least a portion of the cavity with the liquid composition. Item 20. The method according to Item 19.   15. The method of claim 14, wherein the metal nanoparticles comprise negatively charged metal spheres and the metal nanoparticles are bound to the free amine ends of the polymer layer.   The method of claim 14, wherein at least a portion of the metal nanoparticles comprises a metal sphere having one or more ligands attached thereto.   15. The method of claim 14, further comprising one or more ligands attached to the metal nanoparticle comprising a chemical moiety having specificity for one or more target molecules.   The metal nanoparticles are between about 1 nm to about 10 nm, between about 5 nm to about 20 nm, between about 10 nm to about 30 nm, between about 20 nm to about 40 nm, between about 30 nm to about 50 nm, about 40 nm to about 60 nm. Between about 50 nm and about 80 nm, between about 60 nm and about 100 nm, between about 80 nm and about 150 nm, between about 100 nm and about 200 nm, between about 150 nm and about 250 nm, between about 200 nm and about 300 nm 15. The method of claim 14, having a median diameter between about 250 nm to about 400 nm, between about 300 nm to about 700 nm, between about 500 nm to about 900 nm, or between about 700 nm to about 1100 nm. A barrel including at least one sidewall defining a cavity configured to hold liquid therein;
Sidewalls made including the inner surface of a cavity made of plastic material;
Bonding layer of a cross-linked polymer material other than the inner surface plastic material, wherein the cross-linked polymer material of the bonding layer forms a covalent bond with the inner base plastic material, and the cross-linked polymer material of the bonding layer has a free amine end With layers;
A single layer of metal nanoparticles formed on a tie layer, wherein the single layer of metal nanoparticles is bonded to at least a portion of the free amine ends of the cross-linked polymeric material of the tie layer. A test device for use in detecting at least one target molecule.   27. The apparatus of claim 26, wherein the cavity does not include an inner surface made from inorganic glass or crystalline material.   The inner surface is referred to as the first inner surface, the side wall further includes a second inner surface facing the first surface, the second inner surface is also made of a plastic material, and the bonding layer extends over the second inner surface. 27. The device of claim 26, wherein a single layer of metal nanoparticles extends over the second inner surface.   30. The apparatus of claim 28, wherein the first inner surface and the second inner surface opposite the first inner surface are curved to form a single curved surface. A method for detecting a target molecule comprising:
Providing an apparatus according to claim 26;
Placing the test sample in the cavity of the device;
Directing the light beam to the device so that the light beam passes through the body including the inner surface;
Analyzing the light flux passing through the sidewall. A body including a first optical terminal, a second optical terminal, and an optical path between the first optical terminal and the second optical terminal, and receives an incident light flux through the first optical terminal. A body portion configured to transmit the light beam through the optical path portion by total internal reflection and to emit the light beam through the second optical terminal;
An optical passage section made of a plastic material and including at least one curved outer surface of the plastic material;
A tie layer of a cross-linked polymeric material other than at least one curved outer surface plastic material, wherein the cross-linked polymer material of the tie layer forms a covalent bond with the base plastic material of at least one curved outer surface, Having a free amine end, a tie layer;
A single layer of metal nanoparticles formed on a tie layer, wherein the single layer of metal nanoparticles is bonded to at least a portion of the free amine ends of the cross-linked polymeric material of the tie layer. A test device for use in detecting at least one target molecule.   32. The apparatus of claim 31, wherein the optical passage portion does not include an outer surface made from inorganic glass or crystalline material.   32. The apparatus of claim 31, wherein the optical path portion is in the form of a fiber.   32. The apparatus of claim 31, wherein the optical path portion is a vine winding, spiral or coil configuration. A method for detecting a target molecule comprising:
Providing an apparatus according to claim 31;
Inserting at least a portion of the optical path portion of the device into the test sample;
Directing the light beam to the first optical terminal and allowing the light beam to travel to the second optical terminal via the optical path;
Analyzing the light beam emitted from the second optical terminal.

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