Apparatus including analyzer unit

There is provided an apparatus including a chip containing metal bodies capable of exciting localized surface plasmon resonance at a first surface, and an analyzer unit that performs a scan of the first surface of the chip, in a state where the first surface is in contact with a sample, with a laser in at least a one-dimensional direction and records scattered light, which has been enhanced at the first surface, in association with the scan. The chip includes a substrate, a first layer where concave and convex structures are repeatedly provided on the first surface of the substrate; and a second layer that contains the metal bodies and is provided via the first layer.

TECHNICAL FIELD

The present invention relates to an apparatus that includes an analyzer unit.

BACKGROUND ART

Japanese Laid-open Patent Publication No. 2015-152492 describes the provision of an analyzer apparatus capable of a high level of intensity enhancement in an enhancement spectrum, which enables target substances to be detected and analyzed with high sensitivity. This analyzer apparatus is equipped with: an electric field enhancing element including a metal layer, a translucent layer that is provided on the metal layer and transmits excitation light, a plurality of metal particles that are provided on the translucent layer and are arranged in a first direction and a second direction that intersects the first direction; a light source that irradiates the electric field enhancing element with at least one of linearly polarized light that is polarized in the first direction, linearly polarized light that is polarized in the second direction, and circularly polarized light as the excitation light; and a detector that detects the light emitted from the electric field enhancing element. In this analyzer apparatus, there is electromagnetic interaction between localized surface plasmons that are excited at the metal particles and propagating surface plasmons excited at the interface between the metal layer and the translucent layer.

Japanese Laid-open Patent Publication No. 2017-181308 describes the provision of a metal nanostructure array and an electric field enhancing device that achieves a high level of intensity enhancement for Raman scattering. In this device, a plurality of convex nanostructures are formed on a substrate at predetermined intervals, and for a metal nanostructure array used in an electric field enhancing device such as a substrate for surface-enhanced Raman spectroscopy, the base parts of the convex nanostructures are formed of metal and a polycrystalline metal film composed of crystal grains with shape anisotropy is formed from the same type of metal as the base parts or a different type of metal so as to cover the base parts.

SUMMARY OF INVENTION

One aspect of the present invention is an apparatus including: a chip containing metal bodies capable of exciting localized surface plasmon resonance at a first surface; and an analyzer unit that performs a scan of the first surface of the chip, in a state where the first surface is in contact with a sample including an analysis target substance, with a laser in at least a one-dimensional direction and records scattered light, which has been enhanced at the first surface, in association with the scan. The analyzer unit may record a spectrum of the scattered light in association with the scan or may record a fingerprint of the scattered light in association with the scan.

By scanning the first surface of the chip with a laser in at least a one-dimensional direction including at least one of a thickness direction and a planar direction, it becomes highly probable that scattered light, typically SERS (Surface Enhanced Raman Scattering) light in Raman spectroscopy that has been enhanced due to hot spots that have a high probability of being present on the first surface of the chip and are suited to detection of an analysis target substance, the presence of the analysis target substance in the sample, and a laser spot that excites the scattered light that are matched, can be acquired during a scan. By associating the SERS light with the scan, such as by synchronously recording the SERS light, it is possible to identify points on the chip that are suited to detecting the analysis target substance and to increase the detection sensitivity for the analysis target substance. Also, by statistically processing the SERS light recorded in association with the scan, it is possible to improve the measurement accuracy of the concentration of an analysis target substance contained in the sample, which improves reproducibility. The scan may move the spot, and it is also possible to perform measurement by switching between a large number of laser spots like a multifocal-type device.

The chip may include: a substrate; a first layer where concave and convex structures are repeatedly provided on the first surface of the substrate; and a second layer that contains metal bodies capable of exciting localized surface plasmon resonance and is provided via the first layer. By disposing the metal bodies, which are capable of exciting localized surface plasmon resonance, as the second layer via the first layer in which nano-level concave and convex structures are repeatedly provided, it is possible to control the intervals between the metal bodies at the nano level. This means that there is a high probability that hot spots (hot sites), which are suited to local surface plasmon excitation that forms an enhanced electric field which contributes to the measurement of the analysis target substance in the sample, will be present on the first surface of the chip.

The first layer and/or the second layer may include a structure that captures microorganisms and/or proteins. When analyzing or detecting microorganisms or proteins, by capturing the detection target, it becomes easier to acquire SERS light caused by the detection target. The first layer and the second layer may independently or cooperatively (jointly) have a micron-level structure, a submicron structure, a nanostructure, or a sub-nanostructure. This means that a structure suited to capturing microorganisms and proteins may be introduced. As examples, the first layer and the second layer may independently or cooperatively have a structure suited to capturing viruses of several tens to several hundreds of nm, a structure suited to capturing bacteria of around several μm, or a structure suited to capturing cells of several μm to several hundreds of μm. The first layer and the second layer may independently or cooperatively have a structure suited to capturing proteins that are several nm to several tens of nm and contain antibodies or the like.

The chip may include a plurality of sectors in which at least one of a configuration of the first layer and a configuration of the second layer differs. The chip may include a third layer which contains one or more affinity ligands and adheres to at least part of a surface of the metal bodies of the second layer, and the plurality of sectors may include sectors where at least one of the configuration of the first layer, the configuration of the second layer, and the affinity ligand differs. The chip may include a fourth layer that covers the second layer or covers the third layer and contains a structure that captures microorganisms and/or proteins. The fourth layer may be nanostructures using a nanomaterial such as carbon nanotubes. The fourth layer may be microstructures or nanostructures formed by a method, such as nanoimprinting or etching, on a different substrate. The fourth layer may include different structures that capture different microorganisms and proteins, either on a sector-by-sector basis or by dividing sectors.

The analyzer unit may include a unit that scans the plurality of sectors in order. The second layer may contain metal bodies respectively provided at the front ends or tips of convex portions of the first layer, and may include a region where the intervals or distances between the metal bodies are narrower than the intervals or distances between the convex portions of the first layer. The first layer may include one or more regions where the intervals between the convex portions differ. It is also possible to design a second layer that prioritizes formation of hot spots, to design a first layer that prioritizes reflection or transmission of the SERS light, and to design a first layer for the purpose of dispersion, concentration, or the like of the analysis target substances in the sample.

The analyzer unit may include a unit that irradiates (emits) the laser onto the first surface via the sample. The analyzer unit may include a unit that focuses (collimates) at least two laser beams onto a same spot (common spot) on the first surface. This facilitates control of the position of the laser spot that excites scattered light at the first surface of the chip, and makes it possible to improve the scanning accuracy. CARS (Coherent Anti-Stokes Raman Scattering, Coherent Anti-Stokes Raman Spectroscopy) analysis, SRS

(Stimulated Raman Scattering) analysis, and a time-division-based CARS can be given as examples of measurement methods that use at least two laser beams (laser lights).

The sample may be a solid, may be a gas, or may be a liquid, and may have a property or shape that enables contact with the first surface of the chip. The apparatus may be a device or a monitor that detects the presence/absence and/or concentration of components in the sample being measured. The apparatus may further include a translucent (light transmitting) holder in which a fixed amount of sample is internally held or flows and that has a translucent chip mounted on or embedded in a wall surface thereof. The apparatus may further include a route through which a sample including a liquid waste (excreted liquid, effluent, drainage, waste water) from a living body flows and has the chip mounted onto and embedded in a wall surface thereof.

Another aspect of the present invention is an electric field enhancing chip including: a substrate; and metal bodies that are capable of exciting localized surface plasmon resonance and are provided on a first surface of the substrate. The chip may include a first layer and a second layer that are laminated or stacked on the substrate. Additionally, the chip may include a third layer and may include a plurality of sectors where a configuration of at least one of the layers differs.

Yet another aspect of the present invention is a method including detecting presence/absence and/or concentration of a target in a sample using the chip described above. The method includes, during detecting, recording scattered light generated by performing a scan of the first surface in a state where the first surface is in contact with a sample, with a laser in at least a one-dimensional direction, and enhanced at the first surface, in association with the scan. When the chip includes a plurality of sectors, the recording may include scanning the plurality of sectors in order.

Yet another aspect of the present invention is a program of an apparatus that uses the chip described above to detect the presence/absence and/or concentration of a target in a sample. The program (or program product) includes instructions that cause an apparatus to perform a scan of a first surface in a state where the first surface is in contact with a sample, with a laser in at least a one-dimensional direction and to record scattered light, which has been enhanced at the first surface, in association with the scan. This program (or program product) may be provided having been recorded on a computer readable recording medium.

DESCRIPTION OF EMBODIMENTS

FIG.1depicts one of embodiments of an apparatus that measures or detects an analysis target in a sample, such as in a liquid. This measurement apparatus (detection apparatus, detector)1includes a translucent holder (light transmitting holder)10where a sample3, which contains an analysis target (analysis target substance, or measurement target), is internally held in a predetermined amount (quantity) or flows at a predetermined flow rate, and a circulating system13that circulates or supplies the sample3between a source12of the sample3and the holder10. The measurement apparatus1further includes an electric field enhancing chip (chip for detection chip, chip for measurement, measurement chip)20, which is mounted on or embedded in one wall surface (wall)15of the holder10, and an analyzer unit (analyzer, spectroscopic analyzer unit)30, which is disposed or arranged so as to irradiate or emit lasers31and32onto a surface (first surface)21of the chip20that contacts the sample3. One example of a spectroscopic analyzer module30is a Raman analyzer apparatus, and as specific examples, it is possible to use a CARS (Coherent Anti-Stokes Raman Scattering, Coherent Anti-Stokes Raman Spectroscopy) analyzer apparatus, an SRS (Stimulated Raman Scattering) analyzer apparatus, a time-division-based CARS analyzer, or the like suited to microanalysis.

The CARS analyzer unit30according to the present embodiment includes a laser light source35, a head module36that irradiates (focuses, collimates) pump light (pump beam)31and Stokes light (Stokes beam)32obtained from the laser light source35via the sample3onto the same position (region, laser spot, or simply, spot)34on the surface21of the enhancing chip20, a scanning unit37that moves the laser spot34formed by the pump light31and the Stokes light32across the surface21of the chip20, a detection module (spectrometer)38that detects scattered light (surface-enhanced Raman scattering (SERS))33, which has been enhanced at the surface21of the enhancing chip20, through the rear surface22of the enhancing chip20, and an analysis control unit39that records the scattered light33detected by the detection module38in association with the scan and analyzes.

The analysis control unit (or simply control unit)39may be implemented as software (that is, a program or program product)41on a PC40equipped with computer resources including a CPU and a memory. The program41may be provided having been recorded on an appropriate type of memory (or recording medium) that is computer readable. The analysis control unit39may record the SERS light detected (measured) by the detection module38as a spectrum including the SERS light, may record feature components (called a “fingerprint”) extracted from the spectrum, or may record in the form of information that has been compressed using an appropriate method, such as differences from a base waveform (spectrum).

The sample3, which is in the form of a fluid and contains a liquid (which includes an aqueous solution or other solution) or a gas to be measured by the measurement apparatus1, may be any fluid, such as a liquid, containing a substance (or analyte) to be measured, examples of which include a fluid used during a manufacturing process, a waste fluid discharged during a manufacturing process, atmospheric air, river water, liquid waste (drainage, excreted liquid, waste water), blood, serum, a bodily fluid, a culture, or an ampule liquid. The sample3may be matter that has solidified with the chip20inside or may be solid matter where the chip20adheres to part of the surface. The sample3may be a substance or form capable of contacting or tightly adhering to the surface (first surface)21of the chip20.

The sample3may be excreted from a living body. One example of the sample3is a urine sample. The sample3may be dialysis effluent. The sample3may be exhaled breath (that is, exhaled gas). The measurement apparatus1is capable of detecting trace components contained in these fluid samples3using surface-enhanced Raman spectroscopy. As the laser light source35, it is possible to use a pulse laser. As one example, a pulse laser of several kHz, such as around 5 kHz, can be used. Examples of the spectra that are enhanced and detected by this method include emission spectroscopy, fluorescence, Raman scattered light, and nonlinear Raman.

FIG.2depicts the structures of the electric field enhancing chip20by way of enlarged cross sections. The chip20includes a substrate25and a first layer26which is provided on the first surface (or simply “surface”)21of the substrate25on the side of the substrate25in contact with the sample3that contains the analysis target. The first layer26includes concave and convex (concavo-convex, undulating, uneven) structures26athat are repeatedly provided. The chip20further includes a second layer27that is stacked (layered, laminated) on the first layer26and contains metal bodies27athat can excite localized surface plasmon resonance. One example of the substrate25is a glass plate, but it is also possible to use another material, such as a silicon substrate.

With this chip20, the surface21of the substrate (base or board)25is irradiated with the pump light31and the Stokes light32through the sample3, and the SERS light33that is emitted in the forward direction due to the incident light may be detected from or through the rear surface22. Here, it is desirable for the substrate25to have sufficient transparency for the wavelength range of the SERS light33, as one example, electromagnetic waves (light) with a wavelength of 600 to 800 nm. The substrate25does not have to be transparent for the wavelength range of the pump light31and the Stokes light32, as one example, electromagnetic waves (light) with a wavelength of 1000 to 1300 nm. With this chip20, it is also possible to irradiate the surface21of the substrate25with the pump light31and the Stokes light32through the sample3and to detect the SERS light33emitted in a rearward (epi-) direction due to the incident light. Also, the chip20may be irradiated with the pump light31and the Stokes light32through the rear surface22of the substrate25. With this configuration, it is desirable for the substrate25to have sufficient transparency for electromagnetic waves (light) of the wavelength range of the pump light31and the Stokes light32.

As the fine concave and convex structures26aof the first layer26, it is possible to use a resist or other highly heat-resistant resin, such as polyether sulfone, and as one example, the concave and convex structures26amay be formed by a method suited to manufacturing nanostructures, such as nanoimprinting or etching. The concave and convex structures26amay be micron structures, submicron structures, nanostructures, or sub-nano structures. As one example of when nanostructures are formed, the thickness of the first layer26, that is, the height26hof the concave and convex structures26amay be 10 to 1000 nm or 100 to 300 nm. Also, the distance, pitch or intervals26pof the concave and convex structures26amay be 1 to 1000 nm, or may be 50 to 300 nm.

The metal bodies27aof the second layer27are supported by the concave and convex structures26aof the first layer26, and are provided so as to be distanced from or floating above the substrate25due to the concave and convex structures26aby a distance26hof several tens to several hundreds of nm. The metal bodies27a, which are electric field enhancing elements capable of exciting localized surface plasmon resonance, may be a metal such as gold, silver, copper, aluminum, platinum, chromium, nickel, palladium, tungsten, rhodium, or tellurium, or may include an alloy containing any of these metals. The metal bodies27amay be a metalloid, such as silicon, polysilicon, gallium, arsenide, or the like, or may be a mixture containing any of these substances or an alloy of any metalloid and any of the metals mentioned above.

The individual metal bodies27a, which are supported by the concave and convex structures26a, may be shapes produced by extending the concave and convex structures26a, as one example, rod shapes, or may be shapes with a predetermined volume in a direction that is perpendicular to the concave and convex structures26a, such as spherical shapes or ellipsoidal shapes. The diameter of apex portions when the metal bodies27ahave a certain volume and are spherical or ellipsoidal may be 50 nm or less. Gaps27pmay be provided between adjacent metal bodies27a. Gaps27pthat are suited to hot spots may be several nm in size, as one example, 10 nm or less.

As depicted inFIG.2(a), the metal bodies27amay be configured so that expanded apex portions26bof the concave and convex structures26aare covered with a thin metal film27bwith a thickness of around several nm. As depicted inFIG.2(b), it is also possible to use a configuration where metal nanostructures27care supported by convex (protruding) portions of the concave and convex structures26a. The structure of the metal bodies27athat have a predetermined volume and are capable of controlling the gaps27pbetween adjacent metal bodies27ais not limited to a shape composed of curved surfaces, such as a sphere or ellipsoid, and may be a polyhedron or any shape with acute or obtuse corners or steps.

The first layer26and the second layer27of the chip20may be integrally formed of the same metal material. The first layer26and the second layer27of the chip20may be formed of different materials. By using a configuration where these layers26and27are separate, it is possible for the first layer26to have a structure that can be stably and accurately manufactured as nanostructures, and by using these nanostructures of the first layer26which has been stably produced, for the second layer27to have a metal27aselected based on suitability as hot spots rather than stability as a structure. This means that it is possible to provide a chip20which as a whole is equipped with the metal27athat is suited to hot spots and that has metal nanostructures with a stable shape. Accordingly, it is possible to provide a chip20capable of forming hot sites for electric field enhancement more efficiently and uniformly using metal nanostructures.

The chip20may include a region where the intervals27pbetween the metal bodies27ain the second layer27is narrower than the distance26pbetween the convex portions in the first layer26. In the second layer27, the metal bodies27athat are suited to enhancing an electric field can be arranged at narrow intervals (gaps, distances)27pof nm level that are suited to forming hot spots. On the other hand, in the first layer26, structures26amade of a highly transparent material can be disposed at a relatively wide intervals (gaps, distances) so as to guide the scattered light33that has been enhanced by the second layer27to the rear surface side22of the substrate25. Accordingly, it is possible to provide a structure that is suited to a chip20where the front surface21of the substrate25is irradiated with the pump light31and the Stokes light32via the sample3and the SERS light33emitted in the forward direction due to this light is detected through the rear surface22of the substrate25. This configuration is also expected to suppress the transmission of the pump light31and the Stokes light32, which are incident on the front surface21, to the rear surface22.

As depicted inFIG.2(c), the chip20may further include a third layer28containing one or more affinity ligands28athat are attached to at least part of the surfaces of the metal bodies27aof the second layer27. The affinity ligands28aare ligands containing capture molecules including arbitrary molecules capable of binding to a plurality of target substances to be analyzed. Although antibodies, antibody fragments, recombinant antibodies, single-stranded antibodies, receptor proteins, binding proteins, enzymes, inhibitor proteins, lectins, cell adhesion proteins, oligonucleotides, polynucleotides, nucleic acids, and aptamers can be given as examples of capture molecules, the capture molecules in the present embodiment are not limited to these. The affinity ligands28acan improve the concentration of the analysis target substances in the vicinity of the metal bodies27athat have a nanostructure suited to enhancing an electric field, and can thereby further improve detection sensitivity for the analysis target substances.

The first layer26and the second layer27may independently or jointly (cooperatively) construct micron-level structures, submicron structures, nanostructures, or sub-nanostructures. Accordingly, by introducing a structure that physically captures the detection target using these layers26and27, the detection efficiency when using the chip20can be further improved. As examples, the first layer26and the second layer27may independently or cooperatively have one or more structures suited to capturing a virus of several tens to several hundreds of nm, one or more structures suited to capturing bacteria of around several μm, or one or more structures suited to capturing cells of several μm to several hundreds of μm. The first layer26and the second layer27may independently or cooperatively have one or more structures suited to capturing proteins such as antibodies or the like that are several nm to several tens of nm. The first layer26and the second layer27may independently or cooperatively have one or more structures suited to capturing a particular molecule, for example, genes (DNA).

FIG.3schematically depicts the configuration of the surface21of the electric field enhancing chip20. As depicted inFIG.3(a), the chip20may include a plurality of sectors (areas or regions)29in which at least one of the configuration of the first layer26and the configuration of the second layer27differs. When the chip20further includes a third layer28containing an affinity ligand28a, it is possible to include a plurality of sectors29where at least one of the configuration of the first layer26, the configuration of the second layer27, and the affinity ligand28adiffers.

As depicted inFIG.3(b), the chip20may also include regions or sectors29where the intervals (gaps or pitch)26pbetween the convex (protruding) portions in the concave and convex structures26aof the first layer26differs, resulting in the intervals (gaps or pitch)27pbetween the metal bodies27aalso differing. The intervals27pbetween the metal bodies27aand the intervals26pbetween the concave and convex structures26amay differ between a direction where the formation of hot spots is prioritized and a direction where the effect on the transmission of the scattered light33is prioritized. The intervals26pand27pmay differ with respect to the direction in which the sample3flows, may differ with consideration to the adhesion of the analysis target substance to be measured in each sector29, and may differ with consideration to the concentration and detection sensitivity of the analysis target substance. The intervals26pand27pmay differ with consideration to manufacturing tolerances of the chip20, may differ with consideration to various other factors, may differ in sector29units, or may differ in regions included in the sectors29.

Enhancement of an electric field by surface plasmons is typically used in surface-enhanced Raman scattering. As causes of surface-enhanced Raman scattering, aside from the enhancement effect (electromagnetic effect) on an electric field caused by surface plasmons, a chemical effect involving chemical adsorption of molecules is also conceivable. In this case, it is believed that enhancement is due to a resonance Raman effect achieved through the charge transfer state produced between the chemically adsorbed molecules and the metal. Although it is believed that the most important factor determining the enhancement is the surface structure of the metal that produces the surface plasmons, the electric field at the surface will change or vary in various ways depending on the structure of the metal and, in the case of a fine-grained metal, will depend on the size and shape. Accordingly, the enhancement of Raman scattering can change depending on a variety of factors, such as parameters like the morphology and size of metal nanostructures, localization of the electromagnetic field, adsorption of the measurement target substances, and focusing intensity of one or more laser beams. The enhancement is also related to manufacturing tolerances of the nanostructures formed on the surface21of the chip20.

For this reason, in this measurement apparatus1, a plurality of sectors29with different conditions for enhancing an electric field using surface plasmons are provided on the surface21of the chip20, and the analyzer unit30scans these sectors29with a laser spot34formed by the pump light31and the Stokes light32. By providing a plurality of sectors29with different conditions for enhancing an electric field using surface plasmons in units of sectors29, surface-enhanced Raman scattered light (SERS)33, which is related to the measurement target substance or is unique to each measurement target substance, can be obtained with a high probability in sector29units (sector-by-sector basis). The analyzer unit30(that is, the control unit39) records the SERS light33in association with the scan of the surface21of the chip20, for example, in synchronization with the scanning position or timing, and by doing so, it is possible to identify specify timing and/or positions indicating hot spots, which are suited to detecting the measurement target substances contained in the sample3, in units of the sectors29and also in units of the positions of spots34formed in the sectors29.

This means the measurement apparatus1is capable of detecting the measurement target substances contained in the sample3with high accuracy and also with high reproducibility (repeatability) using the chip20that is provided with a plurality of sectors29. Also, the analyzer unit30(the control unit39) may have a function of statistically processing the SERS light33that has been recorded in association with a scan. This makes it possible to improve the measurement accuracy of the concentration of the analysis target substances contained in the sample3and to also improve the reproducibility. The measurement apparatus1may include a cell10capable of controlling the volume or flow rate of the sample3during measurement, which makes it possible to improve the accuracy of quantitative measurements.

The scanning direction may be controlled by the analysis control unit39of the analyzer unit30via the scanning unit37. The scanning direction may be one-dimensional or two-dimensional along the arrangement of sectors29on the chip20. In addition, the state of the intervals between the metal bodies27aof the second layer27, the form or shape of the metal bodies27a, adhesion of the measurement target substances, and the like may change or vary according to the thickness direction (depth direction, longitudinal direction) of the chip20. Accordingly, a scanning direction may also be set in the thickness direction of the chip20, and the analyzer unit30may have a function of scanning in three-dimensional directions.

As the method of moving (scanning) the laser spot34with the scanning unit37, it is possible to physically move the head36, or the optical path may be controlled using a polygon mirror or a micromirror device. The analyzer unit30may also use a multifocal-type head36and scan by switching between a large number of laser spots.

The SERS light33obtained at the surface21of the chip20may be detected as reflected light (backscattered light, Epi-light) from the surface21of the chip20. In the apparatus1according to the present embodiment, by making the substrate25of the chip20transparent for the SERS light33, it becomes possible to detect the SERS light33transmitted through the chip20, that is, forward scattered light, using the spectrometer (light detector)38. By detecting the SERS light33that has been transmitted, it is possible to suppress a decrease in intensity due to the SERS light33propagating in the sample3and the generation of noise due to interaction with the sample3, which makes it possible to measure the measurement target substances even more accurately.

As described earlier, the present apparatus1enables a highly sensitive and quantitative method of measurement by increasing signal intensity with surface plasmon polaritons that are directional and are produced using laser beams. A typical method that uses surface plasmon polaritons detects only reflected light produced by omnidirectional scattering. The present method utilizes nanostructures which are formed on a transparent substrate (or base)25as an electric field enhancing device (chip)20and are uniform as a whole or are uniform in sector29units (sector basis). In addition, by adjusting the focal sizes of exiting lights with high directivity to appropriate sizes, it is possible to achieve a high utilization efficiency of the excited light to efficiently detect an enhanced signal as reflected light and also as transmitted light. In addition, by using the chip20provided with the translucent (light transmissive) substrate25, the present method is capable of detecting the SERS light33generated at the surface21using the detector38through the rear surface22where the distance is constant and without the SERS light33passing through the solvent (sample)3. This means that the sensitivity and reproducibility (repeatability) of the SERS light33obtained during measurement are improved. Accordingly, it is possible to perform reproducible (repeatable) and quantitative spectroscopic measurements.

In particular, when CARS is used as the excited light, an increase in intensity of 106to 107times is expected, and a further enhancement of the signal by surface plasmon polaritons of 102to 1012times is expected. The light to be measured is not limited to CARS, and may be induced stimulated Raman scattering (SRS), coherent Stokes Raman scattering (CSRS), or the like.

Note that when irradiation with a laser over an extended period is required, a resin with high heat resistance (polyether sulfone or the like) may be used for the resist that forms the first layer26. When durability is required, the metal surfaces may be coated (with silica glass or the like). The appropriate wavelength range may be adjusted by changing the nanostructures and/or changing the thickness of the silica glass layer on the metal surfaces. Since the chip20including the substrate25also serves as an output source of CARS light or Raman light, it is desirable to use materials whose outputted spectra do not overlap the spectra (fingerprint regions) of the measurement targets. In particular, since the substrate25makes up the majority of the volume of the chip20, it is desirable for the substrate25to be a substance that does not overlap the fingerprint region, and effective for the substrate25to be a glass substrate as opposed to a silicon substrate.

FIG.4depicts, by way of a flowchart, an example measurement method that uses the chip20in the measurement apparatus1. This measurement method may be provided by a program41as a control method of the measurement apparatus1. When the measurement of the sample3has started in step81, in step82, the analysis control unit39scans the plurality of sectors29provided on the first surface21of the chip20in a predetermined pattern with laser light (that is, the pump light31and the Stokes light32) using the scanning unit37. Together with this, in step83, the analysis control unit39measures, using the detection module38, the scattered light (SERS light)33that has been enhanced in each sector29of the first surface21and records the detected scattered light33in association with the scan.

In step82, if the chip20is provided with a plurality of sectors29in one-dimension, as one example, in a row, the scanning unit37may continuously move the pump light31and the Stokes light32in the one-dimensional direction at a constant speed. The scanning unit37may intermittently move the pump light31and the Stokes light32so that the spot34is continuously formed for a predetermined time at the center of each sector29or at a predetermined position. If the chip20is provided with a plurality of sectors29in two dimensions, the scanning unit37may move the pump light31and the Stokes light32in two dimensions continuously or intermittently at a constant speed, or may move the light continuously or intermittently in three dimensions.

In step83, the analysis control unit39may record the spectrum of the SERS light33obtained by the detection module38, or the intensity of a predetermined wavelength, in association with the scan, as examples in association with the positions or identification information of the sectors29where the SERS light33is obtained. Detection results for the SERS light33may be recorded in association with more detailed positions of the spot34that moves inside the sectors29.

When a scan of the sectors29provided on the surface21of the chip20has been completed in step84, in step85the analysis control unit39analyzes the detection result of the SERS light33obtained by the scan. All of the sectors29provided on the surface21of the chip20may be scanned, or a limited number of sectors29that are decided in advance by pre-measurement, a test measurement, or the like may be scanned before proceeding to the analysis process. Out of the scanning results, the analysis control unit39may select and analyze the SERS light33that has an intensity or spectrum suited to quantitative or qualitative measurement of a measurement target substance. The control unit39may statistically process the scanning results to obtain the presence/absence and/or concentrations of measurement target substances.

In step86, the analysis control unit39outputs the analysis result to an application on a higher level. As one example, if the measurement apparatus1is a urine monitoring apparatus or part of one, by using the chip20, the measurement apparatus1can simultaneously detect multiple components including uric acid, urea, and creatinine, which are trace components in urine, with high sensitivity. This makes it possible for example for an application running on the monitoring apparatus to give health management advice to the monitored user based on the urine analysis result of the measurement apparatus1.

FIG.5depicts different embodiment of apparatus that uses the chip20to measure analysis targets present in a sample. This apparatus60includes a route (or tube or pipe)11through which a sample3including excreted liquid (liquid waste)66from a living body65such as a human body flows, with the chip20being attached to a part of a wall surface15of the route11so that the first surface21contacts the sample3. The apparatus60includes a monitoring apparatus50that irradiates the chip20with one or more laser lights (laser beams) to measure the presence/absence and/or concentration of an analysis target substance (measurement target substance) contained in the sample3. The monitoring device50includes the analyzer unit30that irradiates (focuses, collimates) the pump light31and the Stokes light32onto the chip20and acquires the scattered light (surface-enhanced Raman scattering or SERS)33that has been enhanced at the surface21of the chip20. The basic configuration of this analyzer unit (spectrum analyzer unit)30is the same as the analyzer unit30included in the measurement apparatus1described earlier.

The analyzer unit30in this embodiment is a type that detects SERS light (which is backward CARS, Epi-CARS)33that is outputted to the opposite (counter) direction of the pump light31and the Stokes light32incident on the chip20. The head module36also functions as a module that collects the SERS light33generated at the surface21of the chip20.

One example of the excreted liquid (waste liquid)66from the living body65is urine mentioned earlier. Another example of waste liquid66is effluent (dialysis effluent) discharged from the human body via the blood purifier (dialyzer) of a dialysis apparatus. The effluent monitor50can detect the concentration of a predetermined substance in the effluent66(for example, the concentration of a substance such as urea or uric acid), and can accurately identify the end of a dialysis treatment.

These apparatuses and methods are suited to applications that measure a liquid sample3, and instead of having the fluid flow by, or held in, the cell10, it is also possible to perform measurement with the chip20inserted into a flow (line)11. These apparatuses and methods enable highly sensitive simultaneous detection of multiple components including uric acid, urea, and creatinine in aqueous solutions (such as bodily fluids, dialysate, effluent, food components, and other aqueous solution components). During measurement, calibration may be performed for measurement target substances in anticipation of errors.

FIG.6depicts an example of a virus testing apparatus that uses the chip20. This apparatus70is an apparatus (platform) that places a liquid (test liquid or sample)3containing an analyte collected from a human body or the like into a translucent vessel16that has the chip20mounted on a part of a wall surface15and identifies viruses contained in the sample3. The sample3may be a liquid obtained by culturing the analyte. On top of the substrate25, the chip20includes, in addition to the first layer26, second layer27and the third layer28described earlier, a fourth layer24that contains nanostructures for capturing viruses. The chip20according to this embodiment includes a second substrate23, which has sufficient transparency (light transmissive) for the pump light31and the Stokes light32, and a fourth layer24, which is formed on the second substrate23and contains structures, such as undulations or concave and convex of several nm to several hundred nm or several tens of nm, that are suited to capturing viruses. The first substrate25and the second substrate23are laminated or stacked so as to sandwich these layers. The fourth layer24may contain nanostructures using a nanomaterial such as carbon nanotubes and may be configured so as to be laminated or layered on the third layer28or the second layer27without using the second substrate23.

In the virus detection apparatus (virus detection platform)70, the virus contained in the sample3is captured by the fourth layer24of the chip20, and by irradiating (focusing, collimating) the pump light31and the Stokes light32onto the chip20, CARS light (scattered light) relating to the structure or components constructing the virus can be detected as scattered light (surface-enhanced Raman light, SERS)33that has been enhanced at the surface (first surface)21sandwiched between the two substrates25and23of the chip20. The virus detection apparatus70can optically detect the presence/absence and/or concentration of a virus. This means that the presence/absence of a virus and/or the concentration of the virus can be detected (measured) with high accuracy without destroying the virus during detection or mixing the components constituting the virus.

The virus detection apparatus70includes an analyzer unit (spectrum analyzer unit)30, which has the same basic configuration as the analyzer unit30included in the measurement apparatus1described earlier. The virus detection apparatus70is capable of acquiring SERS light33, which indicates one or a plurality of viruses that have already been identified, or an unknown virus, and by comparing with the fingerprints of known viruses, the presence and concentration of these viruses can be identified in real time. Also, for an unknown virus, by comparing an obtained fingerprint with the fingerprints of known viruses, it is possible to determine the type, danger, and the like of the virus more quickly. The virus detection apparatus70has a simple configuration and can be made compact, and can be provided as an easily portable apparatus.

The chip20may include structures where SERS light33including fingerprints for identifying viruses respectively can be easily obtained in units of sectors (or regions or segments)29, and the fourth layer24may include structures that facilitate the capturing of viruses to be detected. Structures that capture viruses may also be provided in the first layer26and/or the second layer27. By forming the first layer26and the second layer27, whose main purpose is to provide a configuration suited to hotspots, and the fourth layer24whose main purpose is to capture viruses, using different layers, it is possible to provide a chip20with one or more structures that are more suited to the respective purposes. In addition, by providing structures that physically capture the target viruses together with or separately to the affinity ligands28a, it is possible to detect the targets more quickly and more accurately.

It is also possible to provide a cell analysis platform with a configuration that captures predetermined cells in place of the above configurations that capture viruses, and to provide a bacteria analysis platform with a configuration that captures predetermined bacteria. It is also possible to provide an antibody detection platform with a configuration for capturing a predetermined protein including an antibody, and a DNA analysis platform for capturing and analyzing genes.

Although specific embodiments of the present invention have been described above, various other embodiments and modifications will be conceivable to those of skill in the art without departing from the scope and spirit of the invention. Such other embodiments and modifications are addressed by the scope of the patent claims given below, and the present invention is defined by the scope of these patent claims.