WO2022236905A1 - 实现fp-wa耦合模式的生物传感器及其制备方法和应用 - Google Patents

实现fp-wa耦合模式的生物传感器及其制备方法和应用 Download PDF

Info

Publication number
WO2022236905A1
WO2022236905A1 PCT/CN2021/099061 CN2021099061W WO2022236905A1 WO 2022236905 A1 WO2022236905 A1 WO 2022236905A1 CN 2021099061 W CN2021099061 W CN 2021099061W WO 2022236905 A1 WO2022236905 A1 WO 2022236905A1
Authority
WO
WIPO (PCT)
Prior art keywords
groove
metal
metal layer
dielectric
biosensor
Prior art date
Application number
PCT/CN2021/099061
Other languages
English (en)
French (fr)
Inventor
金崇君
沈杨
何凯
邢珊
Original Assignee
中山大学
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 中山大学 filed Critical 中山大学
Publication of WO2022236905A1 publication Critical patent/WO2022236905A1/zh

Links

Images

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/55—Specular reflectivity
    • G01N21/552—Attenuated total reflection
    • G01N21/553—Attenuated total reflection and using surface plasmons
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/55—Specular reflectivity
    • G01N21/552—Attenuated total reflection
    • G01N21/553—Attenuated total reflection and using surface plasmons
    • G01N21/554—Attenuated total reflection and using surface plasmons detecting the surface plasmon resonance of nanostructured metals, e.g. localised surface plasmon resonance
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G01N2021/0106—General arrangement of respective parts
    • G01N2021/0112—Apparatus in one mechanical, optical or electronic block

Definitions

  • the invention relates to the technical field of sensors, in particular to a biosensor for realizing FP-WA coupling mode and its preparation method and application.
  • SPR surface plasmon resonance
  • PSPR propagating surface plasmon resonance
  • LSPR localized surface plasmon resonance
  • PSPR has good sensitivity, it needs to rely on complex equipment and high cost, making it difficult to transform from "centralization” to "decentralization”.
  • the structure of LSPR is relatively simple, but the line width of its resonance peak is relatively large and the quality factor is low.
  • the technical solution of the present invention is:
  • a biosensor for realizing the FP-WA coupling mode comprising a dielectric layer and a metal layer;
  • the dielectric layer has a plurality of dielectric grooves, and a plurality of the dielectric grooves are periodically distributed at equal intervals, and the dielectric grooves are automatically The opening width from the notch to the groove bottom gradually decreases;
  • the metal layer is arranged on the dielectric layer and forms a metal groove corresponding to each of the dielectric grooves;
  • a represents the period of the metal groove
  • h represents the depth of the metal groove
  • w 0 represents the notch width of the metal groove
  • w 1 represents the groove bottom width of the metal groove
  • i represents the WA mode
  • ⁇ WA represents the resonant wavelength of the WA mode
  • m represents the order of the FP mode
  • ⁇ WG represents the resonant wavelength of the FP mode
  • k represents the resonant wavenumber of the FP mode, represents the sum of reflection phases of the FP mode at the notch of the metal groove and the groove bottom of the metal groove
  • ⁇ d represents the dielectric constant of the environment where the biosensor is located
  • ⁇ m represents the dielectric constant of the metal layer.
  • the period of the metal groove is 600nm-1500nm
  • the depth of the metal groove is 300nm-800nm
  • the slot width of the metal groove is 400nm-600nm
  • the metal groove The groove bottom width is 200nm-400nm.
  • the resonance linewidth of the biosensor is 3nm-9nm.
  • the metal layer has a thickness of 200nm-500nm.
  • the root mean square of the surface roughness of the metal layer is 0.2 nm ⁇ 1.9 nm.
  • the plurality of dielectric grooves are distributed in multiple rows and columns, each row is parallel to each other, each column is parallel to each other, and each row and each column are perpendicular to each other.
  • the dielectric grooves of each row are connected to form a communication groove of uniform width; or, the dielectric grooves of each column are connected to the dielectric grooves of each row to form a communication groove of uniform width.
  • the biosensor further includes a substrate, and the substrate is provided on the surface of the dielectric layer away from the metal layer.
  • the metal layer is at least one of a gold layer, a platinum layer and a silver layer.
  • the medium layer is a heat-cured epoxy resin medium layer or a light-cured epoxy resin medium layer.
  • Auxiliary bumps distributed periodically and equally spaced are formed on the template
  • the template is a silicon wafer.
  • the application includes the following steps:
  • the incident light is incident on the surface of the metal layer along the direction perpendicular to the groove bottom of the metal groove, and the initial optical parameters of the reflected light are measured;
  • the plurality of dielectric grooves are distributed in multiple rows and columns, each row is parallel to each other, each column is parallel to each other, and each row and each column are perpendicular to each other; the polarization direction of the incident light is perpendicular to the row direction, or the polarization direction of the incident light is perpendicular to the row direction, or the The polarization direction of the incident light is perpendicular to the column direction.
  • the dielectric grooves in each row are connected to form a connected groove with a uniform width; or, the dielectric grooves in each column are connected to form a connected groove with a uniform width; the polarization direction of the incident light and The extending direction of the communication groove is vertical.
  • the target is salt, organic solvent, antigen, antibody, DNA or RNA.
  • a plurality of metal grooves distributed at regular intervals are formed on the dielectric layer.
  • Each metal groove is equivalent to a resonator, and a metal groove with a relatively wide line width is supported in the metal groove.
  • Wide Fabry-Perot cavity modes (FP modes) meanwhile, the periodicity of metal grooves induces Wood'anomaly modes (WA modes).
  • the period of the metal groove, the depth of the metal groove, the width of the notch of the metal groove and the width of the bottom of the metal groove are set.
  • biosensors At the same time, the above-mentioned biosensor has a simple structure, and the FP-WA coupling mode can be excited under normal incidence, thus avoiding the high-precision angle adjustment components required by biosensors based on oblique incidence excitation, which is more conducive to device integration, easy to transport and use, and can Effectively promote the transformation of biological detection from "centralization” to "decentralization".
  • the preparation method of the above-mentioned biosensor by first forming periodic auxiliary protrusions on the template, then forming a metal layer on the surface of the auxiliary protrusions to form metal grooves, and then forming a dielectric layer on the surface of the metal layer, and then Separate the metal layer and template.
  • Adopting the above preparation method can make the metal layer have the same smooth surface as the template, and can effectively improve the smoothness of the metal layer, so that the resonance peak line width can be further reduced and the quality factor can be improved.
  • adopting the above preparation method can effectively avoid problems such as changes in the thickness of the metal layer and asymmetry of the metal layer affecting the optical response of the biosensor.
  • biosensor in the detection of the parameters of interest of the target object for the purpose of non-disease diagnosis is convenient and easy to operate, does not need to rely on detectors with high wavelength resolution, and the detection cost is low.
  • Fig. 1 is a schematic structural diagram of a biosensor in an embodiment of the present invention
  • Fig. 2 is the size mark of the metal groove in the biosensor corresponding to Fig. 1;
  • FIG. 3 is a schematic structural diagram of a mask template in the preparation process of the biosensor corresponding to FIG. 1;
  • Fig. 4 is a schematic structural diagram of auxiliary protrusions in the preparation process of the biosensor corresponding to Fig. 1;
  • Fig. 5 is a schematic structural diagram of forming a metal layer during the preparation process of the biosensor corresponding to Fig. 1;
  • Fig. 6 is a structural schematic diagram of a dielectric layer and a substrate formed during the preparation of the biosensor corresponding to Fig. 1;
  • FIG. 7 is a physical diagram of a biosensor in an embodiment of the present invention.
  • Fig. 8 is a top view electron micrograph of periodic metal grooves in the biosensor corresponding to Fig. 1;
  • Fig. 9 is a side-view electron micrograph of periodic metal grooves in the biosensor corresponding to Fig. 1;
  • Fig. 10 is an atomic force microscope image of the metal layer in the biosensor corresponding to Fig. 1;
  • Figure 12 is the reflection spectrum of the FP-WA coupling mode with a resonance linewidth of 2nm;
  • Figure 13 is the reflectance spectra of 6 biosensors replicated on the same silicon template
  • Fig. 14 (a) is the reflection spectrum of the gold nano groove array of different groove depths, (b) the spectrum under the optimal groove depth parameter;
  • Fig. 15 is a schematic diagram of avoiding the asymmetric effect and thickness error of the gold layer during the gold plating process by using the preparation method in an embodiment of the present invention
  • Figure 16 is the reflectance spectrum when the AFP antigen solution of different concentrations is dripped when using the biosensor corresponding to Figure 1;
  • Figure 19 is the reflectance spectrum of metal grooves with different working wavelengths
  • FIG. 20 (a) is a schematic diagram of a one-dimensional array of metal grooves, and (b) is a schematic diagram of a two-dimensional array of metal grooves.
  • biosensor 101. medium layer; 102. metal layer; 1021. metal groove; 103. substrate; 200. template; 201. auxiliary protrusion; 300. mask material.
  • first and second are used for descriptive purposes only, and cannot be interpreted as indicating or implying relative importance or implicitly specifying the quantity of indicated technical features.
  • the features defined as “first” and “second” may explicitly or implicitly include at least one of these features.
  • “plurality” means at least two, such as two, three, etc., unless otherwise specifically defined.
  • an embodiment of the present invention provides a biosensor 100 implementing FP-WA coupling mode.
  • the biosensor 100 includes a dielectric layer 101 and a metal layer 102; the dielectric layer 101 has a plurality of dielectric grooves, the plurality of dielectric grooves are periodically distributed at equal intervals, and the opening of the dielectric groove in the direction from its notch to the bottom of the groove The width gradually decreases; the metal layer 102 is disposed on the dielectric layer 101 and forms a metal groove 1021 corresponding to each dielectric groove;
  • a represents the period of the metal groove
  • h represents the depth of the metal groove
  • w 0 represents the notch width of the metal groove
  • w 1 represents the groove bottom width of the metal groove
  • i represents the order of the WA mode
  • ⁇ WA represents the WA
  • m represents the order of the FP mode
  • ⁇ WG represents the resonant wavelength of the FP mode
  • k represents the resonant wave number of the FP mode, represents the sum of reflection phases of the FP mode at the notch of the metal groove and the bottom of the metal groove
  • ⁇ d represents the dielectric constant of the environment where the biosensor is located
  • ⁇ m represents the dielectric constant of the metal layer.
  • the period of the metal groove is 600nm-1500nm, the depth of the metal groove is 300nm-800nm, the width of the notch of the metal groove is 400nm-600nm, and the width of the bottom of the metal groove is 200nm-400nm .
  • the period of the metal groove can be but not limited to 600nm, 650nm, 700nm, 750nm, 800nm, 850nm, 900nm, 950nm, 1000nm, 1050nm, 1100nm, 1150nm, 1200nm, 1250nm, 1300nm, 1350nm, 1400nm, 1450nm or 1500nm.
  • the depth of the metal groove can be but not limited to 300nm, 330nm, 350nm, 375nm, 380nm, 400nm, 425nm, 450nm, 475nm, 500nm, 505nm, 530nm, 555nm, 580nm, 600nm, 610nm, 640nm, 665nm, 695nm, 710nm , 725nm, 755nm, 785nm or 800nm.
  • the slot width of the metal groove can be but not limited to 400nm, 420nm, 440nm, 450nm, 460nm, 470nm, 480nm, 490nm, 500nm, 510nm, 520nm, 530nm, 540nm, 550nm, 560nm, 570nm, 580nm, 590nm or 600nm .
  • the groove bottom width of the metal groove can be but not limited to 200nm, 210nm, 220nm, 230nm, 240nm, 250nm, 260nm, 270nm, 280nm, 290nm, 300nm, 3100nm, 320nm, 330nm, 340nm, 350nm, 360nm, 370nm, 380nm , 390nm or 400nm.
  • the resonance linewidth of the biosensor is 3nm-9nm.
  • the resonance linewidth of the biosensor can be 3.1nm, 3.2nm, 3.3nm, 3.4nm, 3.5nm, 3.6nm, 3.7nm, 3.8nm, 3.9nm, 4.0nm, 4.1nm, 4.2nm, 4.3nm, 4.4nm nm, 4.5nm, 4.6nm, 4.7nm, 4.8nm, 4.9nm, 5.0nm, 5.1nm, 5.2nm, 5.3nm, 5.4nm, 5.5nm, 5.6nm, 5.7nm, 5.8nm, 5.9nm, 6.0nm, 6.1nm, 6.2nm, 6.3nm, 6.4nm, 6.5nm, 6.6nm, 6.7nm, 6.8nm, 6.9nm, 7.0nm, 7.1nm, 7.2nm, 7.3nm,
  • each metal groove is equivalent to a resonator, and a metal groove is supported in the metal groove.
  • a wide linewidth Fabry-Perot cavity mode (FP mode) meanwhile, the periodicity of metal grooves induces Wood'anomaly (Wood'anomaly) mode (WA mode).
  • FP mode Fabry-Perot cavity mode
  • WA mode Wood'anomaly
  • the period of the metal groove, the depth of the metal groove, the width of the notch of the metal groove and the width of the bottom of the metal groove are set.
  • biosensors At the same time, the structure of the biosensor in this embodiment is simple, and the FP-WA coupling mode can be excited under vertical incidence, thus avoiding the high-precision angle adjustment components required by the biosensor based on oblique incidence excitation, which is more conducive to device integration, and is convenient for transportation and Using it can effectively promote the transformation of biological detection from "centralization" to "decentralization".
  • a metal groove 1021 may be formed in a partial area of the metal layer 102 . Further, the area of the metal layer 102 can be made larger than the area of the surface of the dielectric layer with grooves.
  • the metal layer covers the surface of the dielectric layer to form metal grooves on the surface of each dielectric groove, which means that the metal layer covers the entire dielectric groove so that the metal groove covers the surface of the dielectric groove. At this time, the groove bottom and side surfaces of the dielectric groove and the connection surfaces between two adjacent dielectric grooves are all covered by the metal layer.
  • each metal groove can see a resonator, and a kind of FP mode with a wider line width is supported in the metal groove (as shown in Fig. 11 (a)), the FP mode is a localized mode, which has a strong localized electric field and a large radiation loss.
  • the width of the metal groove and the Adjusting the bottom width of the metal groove can adjust the resonant wavelength of the FP mode.
  • the periodicity of the metal groove induces the WA mode.
  • WA is a propagating mode with extremely low radiation loss and extremely narrow line width (as shown in (b) in Fig. 11).
  • the period can tune the resonant wavelength of the WA mode. Based on this, the period of the metal groove, the depth of the metal groove, the notch width of the metal groove and the groove bottom of the metal groove are determined according to the relationship between the structural parameters and the resonance wavelength in formula (1) and formula (2). The width is set so that the wavelengths of the FP mode and the WA mode are close to each other, and a coupling mode with an extremely narrow linewidth and an extremely deep reflection valley is obtained.
  • the radiative decay rate represents the rate at which the surface plasmons of the system are converted into free space photons
  • the internal decay rate represents the rate at which the surface plasmons of the system are converted into heat energy inside the system.
  • the thickness of the metal layer is 200nm ⁇ 500nm. If the thickness of the metal layer is too small, there may be a risk of light passing through the metal layer and entering the dielectric layer, resulting in a decrease in detection accuracy. If the thickness of the metal layer is too large, the manufacturing cost of the sensor will be increased.
  • the thickness of the metal layer may be, but not limited to, 200nm, 250nm, 300nm, 350nm, 400nm, 450nm or 500nm.
  • the thickness of the metal layer is preferably 300 nm.
  • the metal layer is a stable metal layer.
  • the metal layer is at least one of a gold layer, a platinum layer and a silver layer.
  • the metal layer may be a single-layer structure of a stable metal layer, or may be a stacked structure composed of different stable metal layers.
  • the metal layer may be a single layer structure of gold layer, platinum layer or silver layer, or may be a stacked structure composed of at least two of gold layer, platinum layer and silver layer.
  • the root mean square of the surface roughness of the metal layer is 0.2 nm to 1.9 nm.
  • the metal layer has a very smooth surface, which can effectively reduce the scattering loss caused by the particle effect on the surface of the metal layer, and further reduce the resonant peak linewidth of the FP-WA coupling mode.
  • the root mean square of the surface roughness of the metal layer does not exceed 0.5 nm, further preferably, the root mean square of the surface roughness of the metal layer does not exceed 0.36 nm.
  • the root mean square of the surface roughness of the metal layer is 0.2nm-1.9nm, which means that the root mean square roughness of the surface of the gold layer away from the dielectric layer is 0.2nm-1.9nm.
  • the quality factor of the biosensor can be further improved.
  • the quality factor of the biosensor can be up to 285. More specifically, when the metal layer is a gold layer, the root mean square of the surface roughness of the metal layer is preferably 0.2 nm to 0.8 nm. When the metal layer is a silver layer, the root mean square of the surface roughness of the metal layer is preferably 0.4 nm to 1.5 nm.
  • the medium layer is a heat-cured epoxy resin medium layer or a light-cured epoxy resin medium layer.
  • the light-cured epoxy resin medium layer is an ultraviolet light-cured epoxy medium layer.
  • the plurality of dielectric grooves are distributed in multiple rows and columns, each row is parallel to each other, each column is parallel to each other, and each row and each column are perpendicular to each other.
  • the notch of the medium groove is a square, and the width of the notch is the side length of the square.
  • the notch of the medium groove is a circle, and the width of the notch is the diameter of the circle.
  • the dielectric grooves in each row are connected to form a communication groove with a uniform width; or, the dielectric grooves in each column are connected to form a communication groove with a uniform width.
  • the structure of the metal groove corresponds to that shown in FIG. 1 , FIG. 2 , FIG. 8 and FIG. 9 .
  • each row is parallel to each other
  • each column is parallel to each other
  • a two-dimensional array distribution of metal grooves can be formed (in Figure 20 (b) shown).
  • a one-dimensional array distribution of metal grooves can be formed (as shown in (a) in FIG. 20 ).
  • the biosensor 100 further includes a substrate 103 disposed on the surface of the dielectric layer 101 away from the metal layer 102 .
  • the substrate 103 is a glass substrate.
  • FIG. 3 to FIG. 6 another embodiment of the present invention provides a preparation method of the above-mentioned biosensor.
  • the preparation method includes the following steps: forming auxiliary protrusions 201 distributed periodically and at equal intervals on the template 200; forming a metal layer 102 on the surface of the template, and the metal layer 102 forms metal grooves 1021 corresponding to each auxiliary protrusion 201; Form a dielectric layer 101 on the 102; separate the metal layer 102 from the template.
  • the metal layer and the template can have the same smooth surface, and the smoothness of the metal layer can be effectively improved, so that the resonance peak line width can be further reduced and the quality factor can be improved.
  • the template is a silicon wafer.
  • the silicon wafer has better lubricity, which can further improve the lubricity of the metal layer.
  • the optical response of the metal groove is mainly determined by the interface between the silicon wafer and the metal layer, when the thickness of the metal layer is as shown in (a) in Figure 15
  • the interface between the silicon wafer and the metal layer is consistent and thus does not affect the biosensor optical response.
  • the metal layer with a root mean square roughness of 0.2 nm to 1.9 nm can be obtained by using the preparation method in this embodiment, which can greatly improve the smoothness of the metal layer. Further reduce the resonant peak linewidth of the FP-WA coupling mode.
  • the root-mean-square roughness of the surface of the traditional direct evaporation or sputtering metal film is greater than 2nm.
  • the root mean square roughness of the surface of the traditional directly evaporated or sputtered gold film is greater than 2 nm
  • the root mean square roughness of the surface of the traditional directly evaporated or sputtered silver film is greater than 4 nm. That is, this embodiment provides a method for preparing a metal layer that can effectively reduce the root-mean-square surface roughness of the metal layer, which can be applied to the preparation of a metal layer with low surface roughness.
  • forming the periodic auxiliary protrusions 201 on the template includes the following steps: covering the surface of the template with a mask material; exposing and developing the mask material, and forming the auxiliary protrusions 201 on the surface of the template.
  • the matching mask material 300 is obtained to obtain a mask template (the mask template is shown in FIG. 3 ); the mask template is etched to form auxiliary protrusions 201 ; the mask material on the mask template is removed.
  • the mask material may be, but not limited to, polymethyl methacrylate.
  • a layer of polymethyl methacrylate film is spin-coated on the surface of the silicon template.
  • the thickness of the polymethyl methacrylate film is controlled to be 850nm-950nm, more preferably 900nm.
  • the mask material is processed by exposure and development to obtain the mask template shown in FIG. 3 .
  • the mask template is etched, and then the remaining mask material on the mask template is removed to obtain a template with periodic auxiliary protrusions 201 as shown in FIG. 4 .
  • a metal layer is formed on the template to obtain a template with a metal layer as shown in Figure 5, and then a dielectric layer is formed on the template with a metal layer.
  • the surface of the layer forms the substrate, and then the structure shown in Figure 6 is obtained. It can be understood that standard electron beam lithography processes can be used for exposure, development, and etching.
  • the metal layer can be formed on the template by means of metal deposition, preferably, the thickness of the metal layer is controlled to be 200nm-500nm. After obtaining a template with a metal layer as shown in FIG. 5 , the metal layer and the template are separated to obtain a metal layer with a smooth surface and periodic grooves.
  • forming the dielectric layer on the surface of the metal layer includes the following steps: transferring heat-cured epoxy resin or heat-cured epoxy resin to the surface of the metal layer, and then curing and molding. Furthermore, the substrate is covered on the surface of the dielectric layer before curing and molding to facilitate the molding of the dielectric layer.
  • the substrate is glass.
  • cleaning the template includes the following steps: sequentially cleaning the template with acetone, iodine/potassium iodide mixed solution and ethanol. Further, the mass ratio of iodine, potassium iodide and water in the iodine/potassium iodide mixed solution is 3:(8-12):(135-145). Furthermore, the mass ratio of iodine, potassium iodide and water in the iodine/potassium iodide mixed solution is 3:10:140.
  • Still another embodiment of the present invention provides an application of the above-mentioned biosensor in detecting parameters of interest of a target. Specifically, the application includes the following steps:
  • the incident light is incident on the surface of the metal layer along the direction perpendicular to the groove bottom of the metal groove, and the initial optical parameters of the reflected light are measured;
  • another embodiment of the present invention provides an application of the above-mentioned biosensor in the detection of parameters of interest of a target object for non-disease diagnosis purposes. Specifically, the application includes the following steps:
  • the incident light is incident on the surface of the metal layer along the direction perpendicular to the groove bottom of the metal groove, and the initial optical parameters of the reflected light are measured;
  • the target substance is a salt, an organic solvent, an antigen, an antibody, DNA or RNA.
  • the parameter of interest may be a parameter that can be reflected by optical parameters such as concentration or refractive index.
  • the above-mentioned biosensor is used in the detection of the concentration of biological targets for non-disease diagnosis purposes.
  • the application includes the following steps:
  • the incident light is incident on the surface of the metal layer along the direction perpendicular to the groove bottom of the metal groove, and the initial optical parameters of the reflected light are measured;
  • the biological target is an antigen, antibody, DNA or RNA.
  • the optical parameter is at least one of resonance wavelength, reflectivity and reflected light intensity.
  • the resonant wavelength is the wavelength at which the lowest reflection valley point of the biosensor is located.
  • Reflectance is the ratio of the light intensity reflected by the biosensor to the light intensity reflected by the silver mirror (100% reflectance calibration). It can also be understood that when calculating the difference between optical parameters, the difference between the same optical parameters is calculated.
  • the application is the application of the biosensor in the detection of antigen concentration. Specifically, the application includes the following steps:
  • the incident light is incident on the surface of the metal layer along the direction perpendicular to the groove bottom of the metal groove, and the initial optical parameters of the reflected light are measured;
  • the antibody immobilization includes the following steps: after adding the antibody solution dropwise, incubating at 37° C., then washing with phosphate buffer saline and deionized water for 3 times, and drying with nitrogen.
  • the specific binding of antigen and antibody includes the following steps: after adding the antigen solution dropwise, incubate at 37°C, then wash with phosphate buffer saline and deionized water for 3 times, and blow dry with nitrogen.
  • the following steps are further included: performing carboxylation treatment on the surface of the metal layer, and then performing carboxyl activation treatment.
  • the carboxylation treatment included the following steps: contacting the metal layer with a phosphate buffer solution of mercaptopropionic acid, followed by washing with phosphate buffer and deionized water for 3 times, and drying with nitrogen.
  • the carboxyl group activation treatment includes the following steps to mix the metal layer with a mixed solution of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) contact, and then washed three times with phosphate buffered saline and deionized water, and blown dry with nitrogen.
  • EDC 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride
  • NHS N-hydroxysuccinimide
  • the following step is also included: performing hydrophilic treatment on the surface of the metal layer.
  • the hydrophilization treatment includes the step of performing plasma etching on the metal layer.
  • the following step is also included: performing free carboxyl group blocking treatment on the surface of the metal layer after antibody immobilization treatment.
  • the free carboxyl group blocking treatment includes the following steps: contact the metal layer with bovine serum albumin (BSA) solution, then wash with phosphate buffer saline and deionized water for 3 times, and blow dry with nitrogen.
  • BSA bovine serum albumin
  • Still another embodiment of the present invention provides an application of the above-mentioned biosensor in measuring the refractive index of a solution. Specifically, the application includes the following steps:
  • the incident light is incident on the surface of the metal layer along the direction perpendicular to the groove bottom of the metal groove, and the initial optical parameters of the reflected light are measured;
  • the plurality of dielectric grooves are distributed in multiple rows and columns, each row is parallel to each other, each column is parallel to each other, and each row and each column are perpendicular to each other; the polarization direction of the incident light is perpendicular to the row direction, or the polarization direction of the incident light is perpendicular to the column Direction is vertical.
  • the metal grooves are distributed in a two-dimensional array as shown in (b) in FIG. 20 , the polarization direction of the incident light is perpendicular to the row direction, or the polarization direction of the incident light is perpendicular to the column direction.
  • the dielectric grooves of each row are connected to form a connected groove with a uniform width; or, the dielectric grooves of each column are connected to form a connected groove with a uniform width; the polarization direction of the incident light and the extending direction of the connected groove vertical.
  • the metal grooves are distributed in a one-dimensional array as shown in (a) of FIG. 20 , the polarization direction of the incident light is perpendicular to the extending direction of the metal grooves.
  • the metal grooves of the biosensor in this embodiment are distributed in a one-dimensional array as shown in (a) of FIG. 20 .
  • S101 spin coating a layer of polymethyl methacrylate film on the silicon wafer. Control the thickness of the polymethyl methacrylate film to 900nm.
  • S103 Using the polymethyl methacrylate grating structure as a mask, perform reactive ion beam etching on the masked silicon wafer to form auxiliary protrusions on the silicon wafer, and the gap between two adjacent auxiliary protrusions is a groove structure , wherein, the slot width and bottom width of the groove are 450nm and 260nm respectively, the groove depth is 350nm, and the array period is 700nm.
  • S104 Etching the silicon wafer obtained in S103 with oxygen plasma to remove residual polymethyl methacrylate to form a silicon template with auxiliary bumps.
  • S105 Deposit gold with a vertical thickness of 200 nm on the silicon template obtained in S104 by using a magnetron sputtering apparatus to obtain a gold layer.
  • the sputtering current is 35mA
  • the time is 500s.
  • S107 Cut the interface between the gold layer and the silicon template with a clean knife to separate the two, forming a biosensor with a smooth surface of the gold layer, periodic gold grooves, and a one-dimensional array of gold grooves.
  • the structural parameters of the periodic gold groove are: the period is 700nm, the groove depth is 350nm, the groove width is 440nm, the groove bottom width is 250nm, and the vertical thickness of the gold layer is 200nm.
  • the atomic force microscope image of the gold layer is shown in FIG. 10 . It can be seen from FIG. 10 that the root mean square of the surface roughness of the gold layer is 0.36 nm, indicating that this method can obtain ultra-smooth gold grooves.
  • the biosensor in this embodiment has a very narrow resonance linewidth, which is only 4.7nm, and has excellent sensing performance.
  • the extremely narrow resonance linewidth and the extremely large resonance intensity are achieved at the same time, so it is very Ideal structural parameters.
  • S201 Hydrophilicizing the surface of the gold layer: etching the biosensor obtained in S107 with oxygen plasma for 30 seconds with a power of 200W.
  • S202 Carboxylation treatment on the surface of the gold layer: at room temperature, soak the biosensor after S201 treatment in 10mM mercaptopropionic acid phosphate buffer for 12 hours, and then sequentially wash with phosphate buffer and deionized water Wash 3 times and blow dry with nitrogen.
  • S203 Carboxyl activation treatment is carried out on the surface of the gold layer: at room temperature, the biosensor after S202 treatment is dissolved in 400 mM 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride ( EDC) and 100 mM N-hydroxysuccinimide (NHS) mixed aqueous solution for 2 hours, then washed with phosphate buffer saline and deionized water for 3 times, and dried with nitrogen.
  • EDC 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride
  • NHS N-hydroxysuccinimide
  • S204 Add antibody solution dropwise on the surface of the metal layer of the biosensing and carry out antibody immobilization: drop 100 ⁇ g/ml alpha-fetoprotein (AFP) antibody solution on the surface of the gold layer of the biosensor treated in S203, in a 37°C incubator Incubate for 1 hour, then wash with phosphate buffered saline and deionized water three times sequentially, and blow dry with nitrogen gas.
  • AFP alpha-fetoprotein
  • S205 Soak the biosensor treated in S204 in bovine serum albumin (BSA) solution for 40 minutes, then wash with phosphate buffer saline and deionized water for 3 times, and blow dry with nitrogen.
  • BSA bovine serum albumin
  • AFP alpha-fetoprotein
  • S208 Use the micro-area optical measurement system to vertically irradiate the incident light onto the gold layer of the biosensor processed in S207, the incident direction of the incident light is perpendicular to the groove bottom of the gold layer, the polarization direction of the incident light is perpendicular to the incident direction, and record this
  • the reflectance spectrum of the biosensor at time is shown by the dotted lines in (a)-(e) in FIG. 16 .
  • the reusability of the silicon template during the preparation method in Example 1 is verified.
  • the reflectance spectra of the six biosensors were measured with a microspectrometer, and the results are shown in Figure 13.
  • the 6 sensors have almost the same spectrum, the standard error of the resonance wavelength is only 0.06nm, and the standard error of the reflected light intensity at the wavelength of 702.5nm is only 0.54%, which illustrates the preparation method in Example 1
  • the obtained biosensor has a good optical response consistency.
  • the metal grooves of the biosensor in this embodiment are distributed in a one-dimensional array as shown in (a) of FIG. 20 .
  • the structural parameters of the periodic gold grooves in this embodiment are: the period is 700nm, the groove depth is 350nm, the groove width is 450nm, the groove bottom width is 260nm, and the vertical thickness of the gold layer is 300nm.
  • the full width at half maximum of the biosensor in the coupling mode in this embodiment is only 2nm, where the full width at half maximum is defined as the spectral line width corresponding to half of the sum of the light intensity at the highest point and the lowest point of the reflection valley.
  • Incident light is incident on the surface of the metal layer along a direction perpendicular to the groove bottom of the gold groove, and the polarization direction of the incident light is perpendicular to the extension direction of the gold groove, and the initial resonance wavelength of the reflected light is measured.
  • S303 Dropping glycerol aqueous solutions with different concentrations in S301 respectively on the surface of the metal layer, and measuring the corresponding resonant wavelength of reflected light. It is understood that after each measurement, the sensor was rinsed with deionized water.
  • S305 Drop the solution to be tested on the surface of the metal layer, test the resonant wavelength of the reflected light corresponding to the solution to be tested, and calculate the difference between the resonant wavelength and the initial resonant wavelength, and then calculate the difference between the resonant wavelength and the initial resonant wavelength according to the difference between the difference and that shown in (b) in Figure 18 Obtain the refractive index of the solution to be tested using the change curve shown.
  • the relationship between the reflectance of reflected light and the resonance wavelength can also be measured under the condition of different concentrations of glycerol aqueous solution, as shown in (a) in FIG. 18 .
  • the biosensor has a sensitivity (S) of 681 nm/RIU and a wavelength figure of merit (FOM ⁇ ) as high as 285.
  • S sensitivity
  • FOM ⁇ wavelength figure of merit
  • the optimal size of the gold groove at different working wavelengths is designed.
  • the metal grooves of the biosensor in this embodiment are distributed in a one-dimensional array as shown in (a) of FIG. 20 .
  • the period of the fixed gold groove is 600nm
  • the groove width and groove bottom width are 420nm and 220nm respectively
  • the resonance wavelength of the first-order WA is determined to be 600nm according to formula (1).
  • the resonant wavelength of the second-order FP mode is in the range of 200-400 nm in the groove depth near 600 nm.
  • the reflection spectra of gold grooves with different groove depths (200-400nm) were simulated by the finite difference time domain method, and the FP-WA coupling mode with the narrowest line width and deepest reflection valley was searched, as shown in #1 in Figure 19.
  • S402 Repeat S401, respectively fix the period of the gold groove (from 600nm to 1500nm), and then use the finite difference time domain method to scan the reflection spectra in the groove depth corresponding to different periods, and search for the FP- WA coupling mode. Finally, the optimal structural parameters of several gold nanogroove arrays with different working wavelengths are obtained, as shown in the table below. The reflectance spectra of the gold grooves at different working wavelengths are shown in Fig. 19 .

Landscapes

  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Nanotechnology (AREA)
  • Engineering & Computer Science (AREA)
  • Investigating Or Analysing Materials By Optical Means (AREA)

Abstract

一种实现FP-WA耦合模式的生物传感器（100）及其制备方法和应用。生物传感器（100）包括介质层（101）和金属层（102）；介质层（101）具有多个介质凹槽，多个介质凹槽周期性等间距分布，介质凹槽在自槽口到槽底的方向上的开口宽度逐渐减小；金属层（102）设在介质层（101）之上并对应各介质凹槽形成金属凹槽（1021）；金属凹槽（1021）的周期满足式(1)；金属凹槽（1021）的深度、金属凹槽（1021）的槽口宽度以及金属凹槽（1021）的槽底宽度满足式(2)；其中，（III），a：金属凹槽的周期，h：金属凹槽的深度，w0：金属凹槽的槽口宽度，w1：金属凹槽的槽底宽度，i：WA模式的阶数，λWA：WA模式的谐振波长，m：FP模式的阶数，λWG：FP模式的谐振波长，k：FP模式的谐振波数，φr：FP模式在金属凹槽的槽口和金属凹槽的槽底的反射相位之和，εd：生物传感器所处环境的介电常数，εm：金属层的介电常数。通过对金属凹槽的周期、金属凹槽的深度、金属凹槽的槽口宽度以及金属凹槽的槽底宽度进行设定，得到了一种极窄线宽和极深反射谷的耦合模式，即FP-WA耦合模式，进而得到了一种谐振峰线宽小、品质因子高的生物传感器。

Description

实现FP-WA耦合模式的生物传感器及其制备方法和应用 技术领域
本发明涉及传感器技术领域，尤其是涉及一种实现FP-WA耦合模式的生物传感器及其制备方法和应用。
背景技术
随着全球医疗模式开始由“集中化”向“分散化”转变以及数字化远程医疗的普及，生物医学检测的应用场景也逐渐从大型医院扩大到了社区、交通枢纽和家庭。传统的生物医学检测主要通过将收集的样本(血液、尿液或遗传样本等)送到现场以外的分析实验室进行检验。而如今，利用基于生物传感器的即时检测(point-of-care testing,POCT)设备，在采样现场即刻进行分析，可以省去标本在实验室检验时的复杂处理程序，快速得到检验结果。
在这种趋势下，高灵敏、低成本、快速和便携性成为新一代生物传感器的追求目标。其中，表面等离激元(Surface plasmon resonance，SPR)传感作为上世纪90年代初新兴的一种生物分子传感技术，可以提供一种灵敏、简捷、快速、集成化的生物医学检测解决方案。在SPR传感技术中，主要可以分为两种：传统的基于传播表面等离激元(Propagating surface plasmon resonance，PSPR)，以及基于局域表面等离激元(Localized surface plasmon resonance，LSPR)。然而，尽管PSPR具有良好的灵敏性，但是其需要依赖复杂的设备和高昂的成本，难以由“集中化”向“分散化”转变。LSPR结构较为简单，但是其谐振峰的线宽比较大、品质因子较低。
发明内容
基于此，有必要提供一种结构简单、谐振峰线宽小以及品质因子高的实现FP-WA耦合模式的生物传感器及其制备方法和应用。
为了解决以上技术问题，本发明的技术方案为：
一种实现FP-WA耦合模式的生物传感器，包括介质层和金属层；所述介质层具有多个介质凹槽，多个所述介质凹槽周期性等间距分布，所述介质凹槽在自其槽口到槽底的方向上的开口宽度逐渐减小；所述金属层设在所述介质层之上并对应各所述介质凹槽形成金属凹槽；
所述金属凹槽的周期满足式(1)：
Figure PCTCN2021099061-appb-000001
所述金属凹槽的深度、所述金属凹槽的槽口宽度以及所述金属凹槽的槽底宽度满足式(2)：
Figure PCTCN2021099061-appb-000002
其中，
Figure PCTCN2021099061-appb-000003
a代表所述金属凹槽的周期，h代表所述金属凹槽的深度，w 0代表所述金属凹槽的槽口宽度，w 1代表所述金属凹槽的槽底宽度，i代表WA模式的阶数，λ WA代表WA模式的谐振波长，m代表FP模式的阶数，λ WG代表FP模式的谐振波长，k代表FP模式的谐振波数，
Figure PCTCN2021099061-appb-000004
代表FP模式在金属凹槽的槽口和金属凹槽的槽底的反射相位之和，ε d代表所述生物传感器所处环境的介电常数，ε m代表所述金属层的介电常数。
在其中一个实施例中，所述金属凹槽的周期为600nm～1500nm，所述金属凹槽的深度为300nm～800nm，所述金属凹槽的槽口宽度为400nm～600nm，所述金属凹槽的槽底宽度为200nm～400nm。
在其中一个实施例中，所述生物传感器的谐振线宽为3nm～9nm。
在其中一个实施例中，所述金属层的厚度为200nm～500nm。
在其中一个实施例中，所述金属层的表面粗糙度的均方根为0.2nm～1.9nm。
在其中一个实施例中，多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直。
在其中一个实施例中，各行的介质凹槽相连通以形成宽度均匀的连通凹槽；或者，各列的介质凹槽相连通各行的介质凹槽相连通以形成宽度均匀的连通凹槽。
在其中一个实施例中，所述生物传感器还包括衬底，所述衬底设于所述介质层的远离所述金属层的表面。
在其中一个实施例中，所述金属层为金层、铂层以及银层中至少一种。
在其中一个实施例中，所述介质层为热固化环氧树脂介质层或光固化环氧树脂介质层。
上述任一实施例中所述的生物传感器的制备方法，包括如下步骤：
在模板上形成周期性等间距分布的辅助凸起；
在所述模板的表面形成金属层，所述金属层对应各所述辅助凸起形成所述金属凹槽；
在所述金属层之上形成所述介质层；
分离所述金属层与所述模板。
在其中一个实施例中，所述模板为硅片。
上述任一实施例中所述的生物传感器以非疾病诊断目的在目标物感兴趣参数检测中的应用。
在其中一个实施例中，所述应用包括如下步骤：
将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
在金属层表面分别滴加不同已知感兴趣参数的目标物溶液，并测量该溶液对应的反射光的标准光学参数；
分别计算各不同已知感兴趣参数的溶液对应的标准光学参数与所述初始光学参数的差值，并基于该差值与对应的已知感兴趣参数获取光学参数随溶液感兴趣参数的变化关系；
在金属层表面滴加待测溶液，测试所述待测溶液对应的反射光的样品光学参数，并计算该样品光学参数与所述初始光学参数的差值，进而根据该差值与所述变化关系获取所述待测溶液的感兴趣参数。
在其中一个实施例中，多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直；所述入射光的偏振方向与行方向垂直，或者所述入射光的偏振方向与列方向垂直。
在其中一个实施例中，各行的介质凹槽相连通以形成宽度均匀的连通凹槽；或者，各列的介质凹槽相连通以形成宽度均匀的连通凹槽；所述入射光的偏振方向与所述连通凹槽的延伸方向垂直。
在其中一个实施例中，所述目标物为盐、有机溶剂、抗原、抗体、DNA或RNA。
上述生物传感器中，通过对结构进行设置，在介质层上形成多个周期性等间距分布的金属凹槽，每个金属凹槽相当于一个谐振器，在金属凹槽里面支持一种线宽较宽的法布里-珀罗腔模式(FP模式)，同时，金属凹槽的周期性诱导了伍德异常(Wood’anomaly)模式(WA模式)。根据式(1)和式(2)中的结构参数和谐振波长的关系来对金属凹槽的周期、金属凹槽的深度、金属凹槽的槽口宽度以及金属凹槽的槽底宽度进行设定，使得FP模式和WA模式的波长互相靠近，得到一种极窄线宽和极深反射谷的耦合模式，即FP-WA耦合模式，进而得到了一种谐振峰线宽小、品质因子高的生物传感器。同时，上述生物传感器结构简单，FP-WA耦合模式可以在垂直入射下激发，因此避免了基于斜入射激发的生物传感器所需的高精度角度调节组件，更加利于器件集成，便于转运和使用，能够有效促进生物检测由“集中化”向“分散化”转变。
上述生物传感器的制备方法中，通过先在模板上形成周期性的辅助凸起，然后在辅助凸起的表面形成金属层，以形成金属凹槽，再在金属层的表面形成介质层，接着再分离金属层和模板。采用上述制备方法能够使金属层与模板具有同样的光滑表面，能够有效提高金属层的光滑性，这样就可以进一步降低谐振峰线宽、提高品质因子。同时采用上述制备方法能够有效避免金属层厚度变化和金属层不对称而影响生物传感器的光学响应等问题。
上述生物传感器以非疾病诊断目的在目标物感兴趣参数检测中的应用操作方便易行，不需要依赖高波长分辨率的探测器，检测成本低。
附图说明
图1为本发明一实施例中生物传感器的结构示意图；
图2为图1对应的生物传感器中金属凹槽的尺寸标记；
图3为图1对应的生物传感器的制备过程中掩膜模板的结构示意图；
图4为图1对应的生物传感器的制备过程中辅助凸起的结构示意图；
图5为图1对应的生物传感器的制备过程中形成金属层的结构示意图；
图6为图1对应的生物传感器的制备过程中形成介质层和衬底的结构示意图；
图7为本发明一实施例中生物传感器的实物图；
图8为图1对应的生物传感器中周期性金属凹槽的俯视电镜图；
图9为图1对应的生物传感器中周期性金属凹槽的侧视电镜图；
图10为图1对应的生物传感器中金属层的原子力显微镜图；
图11中(a)为FP模式的反射谱、(b)为WA模式的反射谱、(c)为FP-WA耦合模式的反射谱；
图12为谐振线宽为2nm的FP-WA耦合模式的反射谱；
图13为6个由同一硅模板上复制的生物传感器的反射谱；
图14中(a)为不同凹槽深度的金纳米凹槽阵列的反射谱、(b)最佳槽深参数下的光谱；
图15为采用本发明一实施例中制备方法对镀金过程中出现金层非对称效应和厚度误差的规避示意图；
图16为采用图1对应的生物传感器时滴加不同浓度的AFP抗原溶液时的反射谱；
图17中(a)为图1对应的生物传感器波长移动和AFP浓度之间的关系曲线、(b)为图1对应的生物传感器相对反射率改变和AFP浓度之间的关系曲线；
图18中(a)为图1对应的生物传感器在不同浓度丙三醇水溶液中的反射谱、(b)为图1对应的生物传感器的波长移动和不同浓度丙三醇水溶液的折射率的关系曲线；
图19为不同工作波长的金属凹槽的反射谱；
图20中(a)为金属凹槽一维阵列示意图、(b)为金属凹槽二维阵列示意图。
100、生物传感器；101、介质层；102、金属层；1021、金属凹槽；103、衬底；200、模板；201、辅助凸起；300、掩膜材料。
具体实施方式
为使本发明的上述目的、特征和优点能够更加明显易懂，下面对本发明的具体实施方式做详细的说明。在下面的描述中阐述了很多具体细节以便于充分理解本发明。但是本发明能够以很多不同于在此描述的其它方式来实施，本领域技术人员可以在不违背本发明内涵的情况下做类似改进，因此本发明不受下面公开的具体实施例的限制。
此外，术语“第一”、“第二”仅用于描述目的，而不能理解为指示或暗示相对重要性或者隐含指明所指示的技术特征的数量。由此，限定有“第一”、“第二”的特征可以明示或者隐含地包括至少一个该特征。在本发明的描述中，“多个”的含义是至少两个，例如两个，三个等，除非另有明确具体的限定。
在本发明中，除非另有明确的规定和限定，术语“安装”、“相连”、“连接”、“固定”等术语应做广义理解，例如，可以是固定连接，也可以是可拆卸连接，或成一体；可以是机械连接，也可以是电连接；可以是直接相连，也可以通过中间媒介间接相连，可以是两个元件内部的连通或两个元件的相互作用关系，除非另有明确的限定。对于本领域的普通技术人员而言，可以根据具体情况理解上述术语在本发明中的具体含义。
除非另有定义，本文所使用的所有的技术和科学术语与属于本发明的技术领域的技术人员通常理解的含义相同。本文中在本发明的说明书中所使用的术语只是为了描述具体的实施例的目的，不是旨在于限制本发明。本文所使用的术语“和/或”包括一个或多个相关的所列项目的任意的和所有的组合。
请参阅图1、图2、图8和图9，本发明一实施例提供了一种实现FP-WA耦合模式的生物传感器100。该生物传感器100包括介质层101和金属层102；介质层101具有多个介质凹槽，多个介质凹槽周期性等间距分布，介质凹槽在自其槽口到槽底的方向上的开口宽度逐渐减小；金属层102设在介质层101之上并对应各介质凹槽形成金属凹槽1021；
金属凹槽1021的周期满足式(1)：
Figure PCTCN2021099061-appb-000005
金属凹槽1021的深度、金属凹槽1021的槽口宽度以及金属凹槽1021的槽底宽度满足式(2)：
Figure PCTCN2021099061-appb-000006
其中，
Figure PCTCN2021099061-appb-000007
a代表金属凹槽的周期，h代表金属凹槽的深度，w 0代表金属凹槽的槽口宽度，w 1代表金属凹槽的槽底宽度，i代表WA模式的阶数，λ WA代表WA模式的谐振波长，m代表FP模式的阶数，λ WG代表FP模式的谐振波长，k代表FP模式的谐振波数，
Figure PCTCN2021099061-appb-000008
代表FP模式在金属凹槽的槽口和金属凹槽的槽底的反射相位之和，ε d代表所述生物传感器所处环境的介电常数，ε m代表所述金属层的介电常数。
在其中一个实施例中，金属凹槽的周期为600nm～1500nm，金属凹槽的深度为300nm～800nm，金属凹槽的槽口宽度为400nm～600nm，金属凹槽的槽底宽度为200nm～400nm。可以理解的是，金属凹槽的周期可以是但不限定为600nm、650nm、700nm、750nm、800nm、850nm、900nm、950nm、1000nm、1050nm、1100nm、1150nm、1200nm、1250nm、1300nm、1350nm、1400nm、1450nm或1500nm。金属凹槽的深度可以是但不限定为300nm、330nm、350nm、375nm、380nm、400nm、425nm、450nm、475nm、500nm、505nm、530nm、555nm、580nm、600nm、610nm、640nm、665nm、695nm、710nm、725nm、755nm、785nm或800nm。金属凹槽的槽口宽度可以是但不限定为400nm、420nm、440nm、450nm、460nm、470nm、480nm、490nm、500nm、510nm、520nm、530nm、540nm、550nm、560nm、570nm、580nm、590nm或600nm。金属凹槽的槽底宽度可以是但不限定为200nm、210nm、220nm、230nm、240nm、250nm、260nm、270nm、280nm、290nm、300nm、3100nm、320nm、330nm、340nm、350nm、360nm、370nm、380nm、390nm或400nm。
在一个具体的示例中，生物传感器的谐振线宽为3nm～9nm。比如，生物传感器的谐振线宽可以为3.1nm、3.2nm、3.3nm、3.4nm、3.5nm、3.6nm、3.7nm、3.8nm、3.9nm、4.0nm、4.1nm、4.2nm、4.3nm、4.4nm、4.5nm、4.6nm、4.7nm、4.8nm、4.9nm、5.0nm、5.1nm、5.2nm、5.3nm、5.4nm、5.5nm、5.6nm、5.7nm、5.8nm、5.9nm、6.0nm、6.1nm、6.2nm、6.3nm、6.4nm、6.5nm、6.6nm、6.7nm、6.8nm、6.9nm、7.0nm、7.1nm、7.2nm、7.3nm、7.4nm、7.5nm、7.6nm、7.7nm、7.8nm、7.9nm、8.0nm、8.1nm、8.2nm、8.3nm、8.4nm、8.5nm、8.6nm、8.7nm、8.8nm、8.9nm或9.0nm。
在本实施例的生物传感器中，通过对结构进行设置，在介质层上形成多个周期性等间距分布的金属凹槽，每个金属凹槽相当于一个谐振器，在金属凹槽里面支持一种线宽较宽的法布里-泊罗腔模式(FP模式)，同时，金属凹槽的周期性诱导了伍德异常(Wood’anomaly)模式(WA模式)。根据式(1)和式(2)中的结构参数和谐振波长的关系来对金属凹槽的周期、金属凹槽的深度、金属凹槽的槽口宽度以及金属凹槽的槽底宽度进行设定，使得FP模式和WA模式的波长互相靠近，得到一种极窄线宽和极深反射谷的耦合模式，即FP-WA耦合模式，进而得到了一种谐振峰线宽小、品质因子高的生物传感器。同时，本实施例中生物传感器结构简单，FP-WA耦合模式可以在垂直入射下激发，因此避免了基于斜入射激发的生物传感器所需的高精度角度调节组件，更加利于器件集成，便于转运和使用，能够有效促进生物检测由“集中化”向“分散化”转变。
可以理解的是，如图7所示，在实际设计中，可以在金属层102的部分区域形成金属凹槽1021。进一步地，可以使金属层102的面积大于介质层的具有凹槽的表面的面积。
还可以理解的是，金属层覆盖于介质层的表面以在各介质凹槽的表面对应形成金属凹槽，表示金属层将介质凹槽整体覆盖，使得金属凹槽覆盖在介质凹槽的表面。此时，介质凹槽的槽底、侧面以及相邻两个介质凹槽之间的连接面都被金属层覆盖。
在对生物传感器进行设计时，发明人发现，在本实施例的生物传感器中，每个金属凹槽可以看一个谐振器，在金属凹槽中支持一种线宽较宽的FP模式(如图11中(a)所示)，FP模式是一种局域模，具有较强的局域电场，也具有较大的辐射损耗，通过对金属凹槽的深度、金属凹槽的槽口宽度和金属凹槽的槽底宽度进行调节可以调整FP模式的谐振波长。同时，金属凹槽的周期性诱导了WA模式，WA是一种传播模，具有极低的辐射损 耗和极窄的线宽(如图11中(b)所示)，通过调整金属凹槽的周期可以调节WA模式的谐振波长。基于此，根据式(1)和式(2)中的结构参数和谐振波长的关系来对金属凹槽的周期、金属凹槽的深度、金属凹槽的槽口宽度以及金属凹槽的槽底宽度进行设定，使得FP模式和WA模式的波长互相靠近，得到一种极窄线宽和极深反射谷的耦合模式，此时可以达到一种临界耦合条件，使FP模式和WA模式的谐振波长靠近甚至相等，辐射衰减率等于内部衰减率，产生一种谐振峰线宽小、高品质因子、高光学反差以及强局域场的FP-WA耦合模式(如图11中(c)所示)。此处辐射衰减率表示的是体系的表面等离激元转化为自由空间光子的速率，而内部衰减率表示的是体系的表面等离激元在体系内部转化成热能的速率。
在一个具体的示例中，金属层的厚度为200nm～500nm。金属层厚度太小可能会存在光透过金属层进入介质层的风险，导致检测准确性降低，金属层厚度太大则会增加传感器的制造成本。可选地，金属层的厚度可以是但不限定为200nm、250nm、300nm、350nm、400nm、450nm或500nm。金属层的厚度优选为300nm。
进一步地，金属层为稳定金属层。更进一步地，金属层为金层、铂层以及银层中至少一种。可以理解的是，金属层可以是稳定金属层的单层结构，也可以是不同的稳定金属层构成的叠层结构。比如，金属层可以是金层、铂层或银层的单层结构，也可以是金层、铂层以及银层中至少两种组成的叠层结构。
作为金属层的表面粗糙度的一个优选范围，金属层的表面粗糙度的均方根为0.2nm～1.9nm。此时金属层具有十分光滑的表面，可以有效减少金属层表面颗粒效应造成的散射损耗，进一步减小FP-WA耦合模式的谐振峰线宽。优选地，金属层的表面粗糙度的均方根不超过0.5nm，进一步优选地，金属层的表面粗糙度的均方根不超过0.36nm。可以理解的是，金属层的表面粗糙度的均方根为0.2nm～1.9nm，表示金层远离介质层的表面的粗糙度的均方根为0.2nm～1.9nm。通过将金属层的表面粗糙度的均方根设置为0.2nm～1.9nm，可以进一步提高生物传感器的品质因子。在一个具体的示例中，生物传感器的品质因子可达285。更具体地，金属层为金层时，金属层的表面粗糙度的均方根优选为0.2nm～0.8nm。金属层为银层时，金属层的表面粗糙度的均方根优选为0.4nm～1.5nm。
可以理解的是，介质层为热固化环氧树脂介质层或光固化环氧树脂介质层。进一步地，光固化环氧树脂介质层为紫外光固化环氧介质层。
在一个具体的示例中，多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直。具体地，介质凹槽的槽口为正方形，槽口宽度为正方形的边长。在另一个具体的示例中，介质凹槽的槽口为圆形，槽口宽度为圆形的直径。
在另一个具体的示例中，各行的介质凹槽相连通以形成宽度均匀的连通凹槽；或者，各列的介质凹槽相连通以形成宽度均匀的连通凹槽。此时金属凹槽的结构对应图1、图2、图8和图9所示。
请参阅图20，当多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直时，能够形成金属凹槽的二维阵列分布(图20中(b)所示)。当各行的介质凹槽相连通；或者，各列的介质凹槽相连通时，能够形成金属凹槽的一维阵列分布(图20中(a)所示)。
在一个具体的示例中，生物传感器100还包括衬底103，衬底103设于介质层101的远离金属层102的表面。可选地，衬底103为玻璃衬底。
请参阅图3～图6，本发明还有一实施例提供了一种上述生物传感器的制备方法。该制备方法包括如下步骤：在模板200上形成周期性等间距分布的辅助凸起201；在模板的表面形成金属层102，金属层102对应各辅助凸起201形成金属凹槽1021；在金属层102之上形成介质层101；分离金属层102与模板。采用该制备方法能够使金属层与模板具有同样的光滑表面，能够有效提高金属层的光滑性，这样可以进一步降低谐振峰线宽、提高品质因子。同时采用上述制备方法能够有效避免金属层厚度变化和金属层不对称而影响生物传感器的光学响应等问题。优选地，模板为硅片。硅片具有更优的光滑性能，能够进一步提高金属层的光滑性。
具体地，当本实施例中的方法制备生物传感器时，金属凹槽的光学响应主要由硅片和金属层之间的界面决定，当金属层厚度表现为如图15中(a)所示的对称、如图15中(b)所示的非对称或者如图15中(c)所示的厚度变薄时，硅片和金属层之间的界面是一致的，因此不会影响生物传感器的光学响应。
进一步地，采用本实施例中的制备方法能够得到金属层的表面粗糙度的均方根为0.2nm～1.9nm的金属层，这样能够大大提高金属层的光滑性。进一步减小FP-WA耦合模式的谐振峰线宽。而传统的直接蒸镀或者溅射的金属膜表面的粗糙度的均方根大于2nm。比如，传统的直接蒸镀或者溅射的金膜表面的粗糙度的均方根大于2nm，传统的直接蒸镀或者溅射的银膜表面的粗糙度的均方根大于4nm。即本实施例中提供了一种能够有效降 低金属层表面粗糙度均方根的金属层的制备方法，可以将其应用到低表面粗糙度金属层的制备中。
在一个具体的示例中，在模板上形成周期性的辅助凸起201包括如下步骤：在模板表面覆盖掩膜材料；对掩膜材料进行曝光、显影处理，在模板表面形成与辅助凸起201相匹配的掩膜材料300，得到掩膜模板(掩膜模板如图3所示)；对掩膜模板进行蚀刻处理，形成辅助凸起201；去除掩膜模板上的掩膜材料。
可以理解的是，掩膜材料可以是但不限定为聚甲基丙烯酸甲酯，在实际操作过程中，在硅模板的表面旋涂一层聚甲基丙烯酸甲酯薄膜。优选地，控制聚甲基丙烯酸甲酯薄膜的厚度为850nm～950nm，进一步优选为900nm。
覆盖掩膜材料之后采用曝光、显影对掩膜材料进行处理，得到图3所示的掩膜模板。对掩膜模板进行蚀刻处理，然后去除掩膜模板上剩余的掩膜材料，得到如图4所示的具有周期性辅助凸起201的模板。得到如图4所示的模板之后，在该模板上形成金属层，得到具有如图5所示的具有金属层的模板，然后再在具有金属层的模板上形成介质层，还可以继续在介质层表面形成衬底，进而得到具有如图6所示的结构。可以理解的是，曝光、显影、蚀刻可以选用标准的电子束光刻流程。在模板上形成金属层可以采用金属沉积的方式，优选地，控制金属层的厚度为200nm～500nm。在得到具有如图5所示的具有金属层的模板之后，分离金属层和模板，可以得到表面光滑且具有周期性凹槽的金属层。
进一步地，在金属层的表面形成介质层包括如下步骤：将热固化环氧树脂或热固化环氧树脂转移到金属层的表面，然后固化成型。更进一步地，固化成型之前在介质层表面覆盖衬底以促进介质层成型。可选地，衬底为玻璃。
在一个具体的示例中，分离金属层与模板之后还包括如下步骤：对模板进行清洗处理。对模板进行清洗处理除去模板表面残余的介质层材料和金属层材料，可以实现模板的重复利用。具体地，对模板进行清洗处理包括如下步骤：对模板依次采用丙酮、碘/碘化钾混合溶液以及乙醇进行清洗处理。进一步地，碘/碘化钾混合溶液中碘、碘化钾以及水的质量比为3:(8～12):(135～145)。更进一步地，碘/碘化钾混合溶液中碘、碘化钾以及水的质量比为3:10:140。
本发明还有一实施例提供了一种上述生物传感器在目标物感兴趣参数检测中的应用。具体地，该应用包括如下步骤：
将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
在金属层表面分别滴加不同已知感兴趣参数的目标物溶液，并测量该溶液对应的反射光的标准光学参数；
分别计算各不同已知感兴趣参数的溶液对应的标准光学参数与初始光学参数的差值，并基于该差值与对应的已知感兴趣参数获取光学参数随溶液感兴趣参数的变化关系；
在金属层表面滴加待测溶液，测试待测溶液对应的反射光的样品光学参数，并计算该样品光学参数与初始光学参数的差值，进而根据该差值与变化关系获取待测溶液的感兴趣参数。
进一步地，本发明还有一实施例提供了一种上述生物传感器以非疾病诊断目的在目标物感兴趣参数检测中的应用。具体地，该应用包括如下步骤：
将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
在金属层表面分别滴加不同已知感兴趣参数的目标物溶液，并测量该溶液对应的反射光的标准光学参数；
分别计算各不同已知感兴趣参数的溶液对应的标准光学参数与初始光学参数的差值，并基于该差值与对应的已知感兴趣参数获取光学参数随溶液感兴趣参数的变化关系；
在金属层表面滴加待测溶液，测试待测溶液对应的反射光的样品光学参数，并计算该样品光学参数与初始光学参数的差值，进而根据该差值与变化关系获取待测溶液的感兴趣参数。
更具体地，目标物为盐、有机溶剂、抗原、抗体、DNA或RNA。感兴趣参数可以是浓度或折射率等能够通过光学参数反映的参数。
在一个具体的示例中，上述生物传感器以非疾病诊断目的在生物体目标物浓度检测中的应用。具体地，该应用包括如下步骤：
将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
在金属层表面分别滴加不同已知浓度的生物体目标物溶液，并测量不同已知浓度的溶液对应的反射光的标准光学参数；
分别计算各不同已知浓度的溶液对应的标准光学参数与初始光学参数的差值，并基于该差值与对应的浓度获取光学参数随浓度的变化关系；
在金属层表面滴加待测溶液，测试待测溶液对应的反射光的光学参数，并计算该光学参数与初始光学参数的差值，进而根据该差值与变化曲线获取待测溶液的浓度。
进一步地，生物体目标物为抗原、抗体、DNA或RNA。
可以理解的是，光学参数为谐振波长、反射率以及反射光强中的至少一种。其中，谐振波长为生物传感器的反射谷最低点所在的波长。反射率是指生物传感器反射光强和银镜(100％反射校准)反射光强之比。还可以理解的是，在计算光学参数之间的差值时，计算同一种光学参数之间的差值。
作为上述应用的一个具体例子，该应用为生物传感器在抗原浓度检测中的应用。具体地，该应用包括如下步骤：
在生物传感的金属层表面滴加抗体溶液，并进行抗体固定处理；
将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
在金属层表面分别滴加不同已知浓度的抗原溶液，使抗原和抗体进行特异性结合，并分别测量滴加各已知浓度的抗原溶液后对应的反射光的试验光学参数；
分别计算各不同已知浓度的抗原溶液对应的试验光学参数与初始光学参数的差值，并基于该差值与对应的抗原溶液的浓度获取光学参数差值随抗原溶液溶度的变化曲线；
在金属层表面滴加待测抗原溶液，测试待测抗原溶液对应的反射光的光学参数，并计算该光学参数与所述初始光学参数的差值，进而根据该差值与所述变化曲线获取待测抗原溶液的抗原浓度。
在一个具体的示例中，抗体固定包括如下步骤：滴加抗体溶液之后在37℃下孵化，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。使抗原和抗体进行特异性结合包括如下步骤：滴加抗原溶液之后在37℃下孵化，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
在另一个具体的示例中，在生物传感的金属层表面滴加抗体溶液之前还包括如下步骤：对金属层的表面进行羧基化处理，然后进行羧基活化处理。具体地，羧基化处理包括如下步骤：将金属层与巯基丙酸的磷酸盐缓冲液溶液接触，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。羧基活化处理包括如下步骤将金属层与1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDC)和N-羟基琥珀酰亚胺(NHS)的混合溶液接触，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。EDC与NHS的摩尔比为4:1。
可以理解的是，对金属层的表面进行羧基化处理之前还包括如下步骤：对金属层的表面进行亲水化处理。可选地，亲水化处理包括对金属层进行等离子蚀刻的步骤。
可以理解的是，将入射光垂直照射到金属层的表面之前还包括如下步骤：对进行抗体固定处理之后的金属层的表面进行自由羧基封闭处理。具体地，自由羧基封闭处理包括如下步骤：将金属层与牛血清蛋白(BSA)溶液接触，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
本发明还有一实施例提供了一种上述生物传感器在测量溶液折射率的应用。具体地，该应用包括如下步骤：
将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
在金属层表面分别滴加不同已知折射率的溶液，并测量不同已知折射率溶液对应的反射光的标准光学参数；
分别计算各不同已知折射率溶液对应的标准光学参数与初始光学参数的差值，并基于该差值与对应的折射率获取光学参数随折射率的变化关系；
在金属层表面滴加待测溶液，测试待测溶液对应的反射光的光学参数，并计算该光学参数与初始光学参数的差值，进而根据该差值与变化关系获取待测溶液的折射率。
优选地，多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直；入射光的偏振方向与行方向垂直，或者入射光的偏振方向与列方向垂直。比如，当金属凹槽为如图20中(b)所示的二维阵列分布时，入射光的偏振方向与行方向垂直，或者入射光的偏振方向与列方向垂直。
进一步地，各行的介质凹槽相连通以形成宽度均匀的连通凹槽；或者，各列的介质凹槽相连通以形成宽度均匀的连通凹槽；入射光的偏振方向与连通凹槽的延伸方向垂直。比如，当金属凹槽为如图20中(a)所示的一维阵列分布时，入射光的偏振方向与金属凹槽的延伸方向垂直。
以下为具体实施例。
实施例1
本实施例中生物传感器的金属凹槽为如图20中(a)所示的一维阵列分布。
本实施例中生物传感器的制备方法包括如下步骤：
S101：在硅片上旋涂一层聚甲基丙烯酸甲酯薄膜。控制聚甲基丙烯酸甲酯薄膜的厚度为900nm。
S102：对聚甲基丙烯酸甲酯薄膜进行电子束曝光和显影，使得聚甲基丙烯酸甲酯薄膜具有光栅结构，得到掩膜硅片。其中，光栅周期是700nm，光栅条宽度是350nm。
S103：以聚甲基丙烯酸甲酯光栅结构作为掩模，对掩膜硅片进行反应离子束蚀刻，在硅片上形成辅助凸起，相邻的两个辅助凸起之间表现为凹槽结构，其中，凹槽的槽口宽度和槽底宽度分别为450nm和260nm，凹槽深度为350nm，阵列周期为700nm。
S104：用氧等离子蚀刻S103中得到的硅片，清除掉残余的聚甲基丙烯酸甲酯，形成具有辅助凸起的硅模板。
S105：利用磁控溅射仪在S104得到的硅模板上沉积垂直厚度为200nm的金，得到金层。其中，溅射电流为35mA，时间为500s。
S106：在金层上滴液态的紫外固化环氧树脂(型号为NOA61，Norland公司)，用干净的载玻片盖在环氧树脂上，并向下压紧，使得载玻片和硅片之间的环氧树脂层厚度均匀，并覆盖满整个金层区域。所用载玻片的尺寸大于硅模板的尺寸。然后，用紫外灯从载玻片一端照射环氧树脂，使其固化。紫外灯的功率和固化时间分别为48W和20分钟。
S107：用干净的小刀划开金层和硅模板的界面，使两者分离，形成金层表面光滑、具有周期性金凹槽、金凹槽呈一维阵列分布的生物传感器。该周期性金凹槽的结构参数为：周期为700nm，槽深为350nm，槽口宽度为440nm，槽底宽度为250nm，金层垂直厚度为200nm。该金层的原子力显微镜图如图10所示，由图10可知，金层的表面粗糙度的均方根为0.36nm，表明该方法能够得到超光滑的金凹槽。
S108：依次用丙酮、乙醇和碘/碘化钾水溶液去除分离之后的硅模板上残余的环氧树脂和金，得到再生的硅模板。其中碘/碘化钾水溶液的配比为碘：碘化钾：去离子水＝3：10：140。
本实施例中生物传感器具有很窄的谐振线宽，谐振线宽仅为4.7nm，具有优异的传感性能。
作为本实施例中生物传感器的一种性能验证，在本实施例中，固定w 0＝440nm，w 1＝250nm，a＝700nm，ε d＝1(空气)，ε m取自Johnson-Christy的金介电常数的实验数据，根据式(2)，画出不同阶数的FP模式的谐振波长和金凹槽深度的关系曲线，如图14中(a)中的虚线所示。根据式(1)，画出WA模式的谐振波长和金凹槽深度的关系曲线，如图14中(a)中的点划线所示。利用有限时域差分法计算出不同金凹槽深度的金层的反射谱扫描图，如图14中(a)中的背景灰度图所示，其中，不同灰度代表不同的反射率。寻找图14中(a)中两种模式色散曲线的交点，发现交点处周期性金凹槽的谐振模式的色散曲线发生反常弯曲，出现线宽极窄的反射谷。具体地，在凹槽深度为h＝350nm时，如图14中(b)所示，反射谷的强度降到0，说明达到临界耦合条件，也就是系统的辐射衰减率等于其内部衰减率。在该凹槽深度值下，极窄的谐振线宽和极大的谐振强度(即反射谷的深度，定义为谐振处光强的极大值和极小值之差)同时达到，因此是十分理想的结构参数。
本实施例中生物传感器在抗原浓度检测中的应用包括如下步骤：
S201：对金层的表面进行亲水化处理：将S107中得到的生物传感器用氧等离子体刻蚀30秒，功率为200W。
S202：对金层的表面进行羧基化处理：在室温下，将S201处理后的生物传感器浸泡在10mM的巯基丙酸的磷酸盐缓冲液中12小时，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
S203：对金层的表面进行羧基活化处理：在室温下，将S202处理后的生物传感器在400mM的1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDC)和100mM的N-羟基琥珀酰亚胺(NHS)混合水溶液中浸泡2小时，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
S204：在生物传感的金属层表面滴加抗体溶液并进行抗体固定：在S203处理后的生物传感器的金层表面滴加100μg/ml甲胎蛋白(AFP)抗体溶液，在37℃培养箱中孵化1小时，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
S205：将S204处理后的生物传感器在牛血清蛋白(BSA)溶液中浸泡40分钟，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
S206：采用微区光学测量系统将入射光沿垂直于金凹槽的槽底的方向入射到金属层的表面，且入射光的偏振方向与金凹槽的延伸方向垂直，记录下此时生物传感器的反射谱，如图16中(a)～(e)中的实线所示。
S207：取浓度分别为0.01ng/ml、0.1ng/ml、5ng/ml、60ng/ml和200ng/ml的甲胎蛋白(AFP)抗原溶液分别滴在5个结构参数相同的生物传感器上，在37℃培养箱中孵化1小时，然后依次用磷酸盐缓冲液和去离子水清洗3次，并用氮气吹干。
S208：采用微区光学测量系统将入射光垂直照射到S207处理后的生物传感器的金层上，入射光的入射方向与金层槽底垂直，入射光的偏振方向与入射方向垂直，记录下此时生物传感器的反射谱，如图16中(a)～(e)中的虚线所示。
S209：对比加AFP抗原前后的光谱，整理出传感器的谐振波长移动随AFP抗原浓度的变化曲线，如图17中(a)所示。同时，整理出波长为704.645nm处传感器的相对反射率改变随AFP抗原浓度的变化曲线，如图17中(b)所示。由图16和图17可知，生物传感器对AFP抗原的检测限为0.01ng/ml，线性范围为0.01ng/ml-200ng/ml。基于图17中(a)和(b)中的变化曲线，可以确定在线性范围内相同结构参数的生物传感器的谐振波长移动或相对反射率改变所对应的抗原溶液浓度。
实施例2
本实施例中验证了实施例1中制备方法过程中硅模板的重复利用性。重复实施例1中S105～S108，得到另外5个由同一硅模板制备的生物传感器，分别记为复制1(实施例1中的生物传感器)、复制2、复制3、复制4、复制5和复制6。分别用显微光谱仪测量6个生物传感器的反射谱，结果如图13所示。由图13可知，6个传感器具有几乎相同的光谱，其谐振波长的标准误差仅0.06nm，其波长为702.5nm处的反射光强的标准误差仅为0.54％，说明实施例1中的制备方法得到的生物传感器具有很好的光学响应一致性。
实施例3
本实施例中生物传感器的金属凹槽为如图20中(a)所示的一维阵列分布。本实施例中周期性金凹槽的结构参数为：周期为700nm，槽深为350nm，槽口宽度为450nm，槽底宽度为260nm，金层垂直厚度为300nm。
请参阅图12，本实施例中生物传感器的耦合模式下的半高全宽值仅为2nm，其中半高全宽值的定义为反射谷最高点与最低点光强之和的一半处对应的谱线宽度。
本实施例测量丙三醇的折射率的方法包括如下步骤：
S301：配置浓度为0％、10％、20％、30％、40％、50％和60％的丙三醇水溶液，其对应的折射率为1.33303、1.34481、1.35749、1.3707、1.38413、1.39809和1.41299。
S302：将入射光沿垂直于金凹槽的槽底的方向入射到金属层的表面，且入射光的偏振方向与金凹槽的延伸方向垂直，测量反射光的初始谐振波长。
S303：在金属层表面分别滴加S301中不同浓度的丙三醇水溶液，并测量对应的反射光的谐振波长。可以理解的是，在每次测量之后，用去离子水清洗传感器。
S304：分别计算各不同折射率溶液对应的反射光的谐振波长与初始谐振波长的差值，并基于该差值与对应的折射率获取谐振波长差值与折射率的变化曲线，如图18中(b)所示。
S305：在金属层表面滴加待测溶液，测试待测溶液对应的反射光的谐振波长，并计算该谐振波长与初始谐振波长的差值，进而根据该差值与图18中(b)所示的变化曲线获取待测溶液的折射率。在S303中还可以测量得到不同浓度的丙三醇水溶液情况下，反射光的反射率和谐振波长的关系，如图18中(a)所示。结合图18中(a)和(b)，可以计算得出生物传感器具有681nm/RIU的灵敏度(S)和高达285的波长品质因子(FOM λ)。其中S＝Δλ/Δn，Δλ为波长的改变量，Δn为折射率的改变量；FOM λ＝S/FWHM，FWHM为谐振的半高全宽。
实施例4
本实施例中对不同工作波长下的优选金凹槽尺寸进行了设计。本实施例中生物传感器的金属凹槽为如图20中(a)所示的一维阵列分布。
S401：固定金凹槽的周期为600nm，槽口宽度和槽底宽度分别为420nm和220nm，根据式(1)确定其一阶WA的谐振波长为600nm。再根据式(2)估计二阶FP模式的谐振波长在600nm附近的槽深范围为200-400nm。利用时域有限差分法模拟不同槽深(200-400nm)的金凹槽的反射谱，从中搜寻线宽最窄、反射谷最深的FP-WA耦合模式，如图19中的#1所示。
S402：重复S401，分别固定金凹槽的周期(从600nm到1500nm)，然后利用时域有限差分法分别扫描不同周期所对应的槽深范围内的反射谱，从中搜寻线宽最窄的FP-WA耦合模式。最终得到若干个不同工作波长的金纳米槽阵列的最佳结构参数，如下表所示。不同工作波长的金凹槽的反射谱如图19所示。
Figure PCTCN2021099061-appb-000009
以上所述实施例的各技术特征可以进行任意的组合，为使描述简洁，未对上述实施例中的各个技术特征所有可能的组合都进行描述，然而，只要这些技术特征的组合不存在矛盾，都应当认为是本说明书记载的范围。
以上所述实施例仅表达了本发明的几种实施方式，其描述较为具体和详细，但并不能因此而理解为对发明专利范围的限制。应当指出的是，对于本领域的普通技术人员来说，在不脱离本发明构思的前提下，还可以做出若干变形和改进，这些都属于本发明的保护范围。因此，本发明专利的保护范围应以所附权利要求为准，说明书及附图可以用于解释权利要求的内容。

Claims (10)

  1. 一种实现FP-WA耦合模式的生物传感器，其特征在于，包括介质层和金属层；所述介质层具有多个介质凹槽，多个所述介质凹槽周期性等间距分布，所述介质凹槽在自其槽口到槽底的方向上的开口宽度逐渐减小；所述金属层设在所述介质层之上并对应各所述介质凹槽形成金属凹槽；
    所述金属凹槽的周期满足式(1)：
    Figure PCTCN2021099061-appb-100001
    所述金属凹槽的深度、所述金属凹槽的槽口宽度以及所述金属凹槽的槽底宽度满足式(2)：
    Figure PCTCN2021099061-appb-100002
    其中，
    Figure PCTCN2021099061-appb-100003
    a代表所述金属凹槽的周期，h代表所述金属凹槽的深度，w 0代表所述金属凹槽的槽口宽度，w 1代表所述金属凹槽的槽底宽度，i代表WA模式的阶数，λ WA代表WA模式的谐振波长，m代表FP模式的阶数，λ WG代表FP模式的谐振波长，k代表FP模式的谐振波数，
    Figure PCTCN2021099061-appb-100004
    代表FP模式在金属凹槽的槽口和金属凹槽的槽底的反射相位之和，ε d代表所述生物传感器所处环境的介电常数，ε m代表所述金属层的介电常数。
  2. 如权利要求1所述的生物传感器，其特征在于，所述金属凹槽的周期为600nm～1500nm，所述金属凹槽的深度为300nm～800nm，所述金属凹槽的槽口宽度为400nm～600nm，所述金属凹槽的槽底宽度为200nm～400nm。
  3. 如权利要求1所述的生物传感器，其特征在于，所述生物传感器的谐振线宽为3nm～9nm；和/或，
    所述金属层的厚度为200nm～500nm；和/或，
    所述金属层的表面粗糙度的均方根为0.2nm～1.9nm。
  4. 如权利要求1～3中任一项所述的生物传感器，其特征在于，多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直。
  5. 如权利要求4所述的生物传感器，其特征在于，各行的介质凹槽相连通以形成宽度均匀的连通凹槽；或者，各列的介质凹槽相连通以形成宽度均匀的连通凹槽。
  6. 如权利要求1～5中任一项所述的生物传感器的制备方法，其特征在于，包括如下步骤：
    在模板上形成周期性等间距分布的辅助凸起；
    在所述模板的表面形成金属层，所述金属层对应各所述辅助凸起形成所述金属凹槽；
    在所述金属层之上形成所述介质层；
    分离所述金属层与所述模板。
  7. 如权利要求1～5中任一项所述的生物传感器以非疾病诊断目的在目标物感兴趣参数检测中的应用。
  8. 如权利要求7所述的应用，其特征在于，包括如下步骤：
    将入射光沿垂直于金属凹槽的槽底的方向入射到金属层的表面，测量反射光的初始光学参数；
    在金属层表面分别滴加不同已知感兴趣参数的目标物溶液，并测量该溶液对应的反射光的标准光学参数；
    分别计算各不同已知感兴趣参数的溶液对应的标准光学参数与所述初始光学参数的差值，并基于该差值与对应的已知感兴趣参数获取光学参数随溶液感兴趣参数的变化关系；
    在金属层表面滴加待测溶液，测试所述待测溶液对应的反射光的样品光学参数，并计算该样品光学参数与所述初始光学参数的差值，进而根据该差值与所述变化关系获取所述待测溶液的感兴趣参数。
  9. 如权利要求8所述的应用，其特征在于，多个介质凹槽呈多行多列分布，各行相互平行，各列相互平行，且各行与各列相互垂直；所述入射光的偏振方向与行方向垂直，或者所述入射光的偏振方向与列方向垂直。
  10. 如权利要求9所述的应用，其特征在于，各行的介质凹槽相连通以形成宽度均匀的连通凹槽；或者，各 列的介质凹槽相连通以形成宽度均匀的连通凹槽；所述入射光的偏振方向与所述连通凹槽的延伸方向垂直。
PCT/CN2021/099061 2021-05-11 2021-06-09 实现fp-wa耦合模式的生物传感器及其制备方法和应用 WO2022236905A1 (zh)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN202110510078.8 2021-05-11
CN202110510078.8A CN113433094B (zh) 2021-05-11 2021-05-11 实现fp-wa耦合模式的生物传感器及其制备方法和应用

Publications (1)

Publication Number Publication Date
WO2022236905A1 true WO2022236905A1 (zh) 2022-11-17

Family

ID=77753392

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2021/099061 WO2022236905A1 (zh) 2021-05-11 2021-06-09 实现fp-wa耦合模式的生物传感器及其制备方法和应用

Country Status (2)

Country Link
CN (1) CN113433094B (zh)
WO (1) WO2022236905A1 (zh)

Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4502756A (en) * 1982-09-30 1985-03-05 The United States Of America As Represented By The Secretary Of The Navy Bandpass filter formed by serial gratings operating in a Wood's anomaly region
US6093536A (en) * 1987-01-21 2000-07-25 Ares-Serono Research & Development Limited Partnership Wood's anomaly assay technique and kit
CN103196867A (zh) * 2013-04-01 2013-07-10 中山大学 局域等离子体谐振折射率传感器及其制造方法
CN103811580A (zh) * 2014-03-05 2014-05-21 中国科学院半导体研究所 InGaAs红外光探测器
CN103808691A (zh) * 2014-02-19 2014-05-21 中国科学院半导体研究所 非对称Au粒子阵列和FP腔耦合的折射率传感器
CN207300884U (zh) * 2017-05-15 2018-05-01 安徽大学 嵌入式纳米金阵列表面等离子共振传感器基底
CN110360935A (zh) * 2019-07-31 2019-10-22 西北工业大学 一种基于简化光学纳米谐振腔的面内位移传感单元及方法
CN111398217A (zh) * 2019-06-05 2020-07-10 江西师范大学 一种高品质等离激元光学传感器及其制备方法

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CA2416177A1 (en) * 2000-07-31 2002-02-07 Spectalis Corp. Optical waveguide filters
CN102353652B (zh) * 2011-07-12 2013-09-18 中山大学 一种化学折射率传感器及其设计方法
GB201221330D0 (en) * 2012-11-27 2013-01-09 Univ Glasgow Terahertz radiation detector, focal plane array incorporating terahertz detector, and combined optical filter and terahertz absorber
JP2014232098A (ja) * 2013-04-30 2014-12-11 日本精工株式会社 標的物質捕捉装置及び標的物質検出装置
CN108027313B (zh) * 2015-08-26 2021-01-29 光子系统有限责任公司 谐振周期性结构以及使用它们作为滤光器和传感器的方法
CN108519352B (zh) * 2018-04-09 2021-06-08 南京邮电大学 一种基于金属-介质-金属波导布拉格光栅的折射率传感器
CN109856087B (zh) * 2018-12-29 2021-01-29 复旦大学 传感芯片及其制备方法、检测系统、检测方法

Patent Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4502756A (en) * 1982-09-30 1985-03-05 The United States Of America As Represented By The Secretary Of The Navy Bandpass filter formed by serial gratings operating in a Wood's anomaly region
US6093536A (en) * 1987-01-21 2000-07-25 Ares-Serono Research & Development Limited Partnership Wood's anomaly assay technique and kit
CN103196867A (zh) * 2013-04-01 2013-07-10 中山大学 局域等离子体谐振折射率传感器及其制造方法
CN103808691A (zh) * 2014-02-19 2014-05-21 中国科学院半导体研究所 非对称Au粒子阵列和FP腔耦合的折射率传感器
CN103811580A (zh) * 2014-03-05 2014-05-21 中国科学院半导体研究所 InGaAs红外光探测器
CN207300884U (zh) * 2017-05-15 2018-05-01 安徽大学 嵌入式纳米金阵列表面等离子共振传感器基底
CN111398217A (zh) * 2019-06-05 2020-07-10 江西师范大学 一种高品质等离激元光学传感器及其制备方法
CN110360935A (zh) * 2019-07-31 2019-10-22 西北工业大学 一种基于简化光学纳米谐振腔的面内位移传感单元及方法

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
SHEN YANG, SHE XIAOYI, JIN CHONGJUN: "Mechanically Reconfigurable Pd Nanogroove Array: An Ultrasensitive Optical Hydrogen Detector", ACS PHOTONICS, vol. 5, no. 4, 18 April 2018 (2018-04-18), pages 1334 - 1342, XP093004061, ISSN: 2330-4022, DOI: 10.1021/acsphotonics.7b01323 *
TAPSANIT PIYAWATH, YAMASHITA MASATSUGU, ISHIHARA TERUYA, OTANI CHIKO: "Quasi-analytical solutions of hybrid platform and the optimization of highly sensitive thin-film sensors for terahertz radiation", JOURNAL OF THE OPTICAL SOCIETY OF AMERICA - B., OPTICAL SOCIETY OF AMERICA, WASHINGTON., US, vol. 33, no. 12, 1 December 2016 (2016-12-01), US , pages 2535, XP093004063, ISSN: 0740-3224, DOI: 10.1364/JOSAB.33.002535 *

Also Published As

Publication number Publication date
CN113433094A (zh) 2021-09-24
CN113433094B (zh) 2023-01-17

Similar Documents

Publication Publication Date Title
Ahmadivand et al. Photonic and plasmonic metasensors
Sreekanth et al. Enhancing the angular sensitivity of plasmonic sensors using hyperbolic metamaterials
US6421128B1 (en) Coupled plasmon-waveguide resonance spectroscopic device and method for measuring film properties in the ultraviolet and infrared special ranges
Huang et al. Application of photonic crystal enhanced fluorescence to cancer biomarker microarrays
KR101006793B1 (ko) 라벨 없는 바인딩 검출 및 형광 증폭과 조합된 격자-기반센서 및 센서를 위한 판독 시스템
US6330387B1 (en) Coupled plasmon-waveguide resonance spectroscopic device and method for measuring film properties in the ultraviolet and infrared spectral ranges
CN109856087B (zh) 传感芯片及其制备方法、检测系统、检测方法
CN110398479B (zh) 一种基于光学芯片基底的显微光谱测量装置及方法
CN112461787B (zh) 一种基于布洛赫表面波的铌酸锂光学传感器及方法
WO2019039551A1 (ja) メタマテリアル構造体および屈折率センサ
Jin et al. A stable and high resolution optical waveguide biosensor based on dense TiO2/Ag multilayer film
Saadatmand et al. Graphene-based integrated plasmonic sensor with application in biomolecule detection
Wang et al. Surface plasmon resonance waveguide biosensor by bipolarization wavelength interrogation
Guselnikova et al. Coupling of plasmonic hot spots with shurikens for superchiral SERS-based enantiomer recognition
Gryga et al. Narrow Tamm resonances in one-dimensional photonic crystals employed in sensor applications
JP4987737B2 (ja) 蛍光検出デバイス
Liu et al. Flexible hyperspectral surface plasmon resonance microscopy
WO2022236905A1 (zh) 实现fp-wa耦合模式的生物传感器及其制备方法和应用
Wang et al. Plasmonic metasurface enhanced by nanobumps for label-free biosensing of lung tumor markers in serum
Jian et al. Enhancement of the volume refractive index sensing by ROTE and its application on cancer and normal cells discrimination
Chen et al. A low cost surface plasmon resonance biosensor using a laser line generator
Pokhriyal et al. Coupled external cavity photonic crystal enhanced fluorescence
TWI770836B (zh) 生物感測裝置、生物感測系統及其使用方法
US20240241052A1 (en) Biosensor implementing fp-wa coupling mode, preparation method therefor, and use thereof
Abdulhalim II et al. Resonant and scatterometric grating-based nanophotonic structures for biosensing

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 21941474

Country of ref document: EP

Kind code of ref document: A1

WWE Wipo information: entry into national phase

Ref document number: 18560188

Country of ref document: US

NENP Non-entry into the national phase

Ref country code: DE