Multi-reflection liquid-immersion silicon-based microfluidic measurement device and measurement method

Abstract

A multi-reflection liquid-immersion silicon-based microfluidic measurement device and measurement method provided by one embodiment of the present invention completely separates the first reflected light reflected from the sample detection layer from the second reflected light reflected from the prism-buffer solution interface through multiple reflections, and amplifies the measurement sensitivity by achieving multiple incidences through multiple reflections. The multi-reflection liquid-immersion silicon-based microfluidic measurement device of an embodiment of the present invention includes: a microfluidic structure, consisting of a support and at least one microfluidic path, the microfluidic path being formed on the support, a sample detection layer being formed in the microfluidic path, and a bioconjugate for detecting the sample being fixed in the sample detection layer; a sample injection portion for injecting a buffer solution containing the sample into the microfluidic path; a prism unit, provided with a prism and a reflector, the reflector being formed by coating a reflective mirror surface on the bottom surface of the prism; a polarized light generating portion for generating polarized light; and a polarized light detecting portion for detecting polarization changes of the reflected light.

Description

Technical Field

[0001] The present invention relates to a multiple-reflection liquid-immersion silicon-based microfluidic measurement device and a measurement method. More specifically, the present invention relates to the following multiple-reflection liquid-immersion silicon-based microfluidic measurement device and a measurement method, namely, through multiple reflections, the first reflected light reflected from the sample detection layer and the second reflected light reflected from the prism-buffer solution interface are completely separated and multiple incidences are achieved through multiple reflections to amplify the measurement sensitivity. Background Art

[0002] Reflectometry and ellipsometry are optical analysis techniques that measure changes in reflectivity or polarization of light reflected from a sample's surface and analyze the resulting values ​​to detect sample thickness or optical properties.

[0003] Measuring equipment utilizing this technology includes reflectometers and ellipsometers. In the semiconductor field, these devices are used to evaluate the thickness and physical properties of various nanoscale thin films during nanofilm manufacturing processes. Furthermore, in the biological field, due to its expanding application range, it continues to be used for surface analysis of biological substances such as DNA, viruses, and new pharmaceutical substances.

[0004] While existing reflectometers are sufficient for assessing the thickness and properties of nanofilms larger than nanometers (nm), they suffer from reduced reliability due to reduced measurement sensitivity when analyzing low-molecular-weight biological materials within the required sensitivity range of approximately 1 nm to 0.001 nm. Ellipsometers, on the other hand, have a measurement sensitivity of approximately 0.01 mm or less, and are particularly sensitive when measuring the thickness of oxide films on semiconductor substrates with a relatively low refractive index compared to high-refractive-index semiconductors, where the refractive index contrast is relatively large.

[0005] However, in order to analyze low-molecular-weight biological substances using ellipsometers, a measurement method with enhanced sensitivity is required.

[0006] When analyzing biological substances, a conventional technique for improving measurement sensitivity is a surface plasmon resonance sensor (hereinafter referred to as a "surface plasmon resonance sensor") that utilizes a combination of reflectometry and surface plasmon resonance (SPR) technology.

[0007] Surface plasmon resonance (SPR) is a phenomenon in which electrons on a metal surface are excited by light waves and collectively vibrate along the surface's vertical direction, resulting in the absorption of light energy. Because SPR is sensitive to the polarization characteristics of light, SPR sensors can not only measure changes in the thickness and refractive index of nanofilms contacting metal surfaces, but also measure changes in the adsorbed concentration of biomolecules in real time using a non-labeling method that does not use fluorescent substances.

[0008] At the structural level, a surface plasmon resonance sensor consists of a nanoscale metal film coated on a material such as glass, onto which a biomaterial-attached sensor is formed. When a sample dissolved in a buffer solution is attached to the sensor, the resonance angle is measured based on the change in reflectivity. When light enters the surface plasmon resonance sensor, the glass serves as the incident medium. As light passes through the biomaterial-attached thin film layer, the buffer solution ultimately becomes the substrate.

[0009] In the above configuration, the refractive index of the buffer solution, which acts as the substrate, directly affects the shift in the resonance angle, similar to the changes in the biofilm layer caused by the binding of the sample to be measured. Therefore, in order to measure only the pure binding dynamics, it is necessary to separately measure the refractive index of the buffer solution for calibration.

[0010] Therefore, to correct for changes in the refractive index of the buffer solution and prevent errors caused by diffusion between the sample and the buffer solution, methods currently used involve sophisticated valve mechanisms, air injection devices, and the correction of two or more channels using a single reference channel. However, this method not only makes it difficult to distinguish between changes in the surface plasmon resonance angle caused by changes in the refractive index of the buffer solution and those caused by pure adsorption and dissociation properties, but also consistently contributes to measurement errors. As a result, existing surface plasmon resonance sensors face fundamental problems when measuring the adsorption and dissociation properties of low-molecular-weight substances, due to these limitations.

[0011] Furthermore, existing surface plasmon resonance sensors require the use of thin metal films made of precious metals such as gold (Au) and silver (Ag) to achieve surface plasmon resonance, resulting in relatively high manufacturing costs. Furthermore, depending on the manufacturing process, the metal film can suffer from uneven surface roughness and significant refractive index variations. This not only hinders accurate measurement of biological substances due to unstable optical properties, but also introduces errors due to varying sensitivity characteristics at different locations when comparing the sensor against a reference channel.

[0012] To address the shortcomings of surface plasmon resonance sensors, a biomaterial binding sensor layer is formed on a substrate material such as silicon. Ellipsometry is then used to measure the amplitude and phase of light reflected from the substrate material by a buffer solution in an immersion microfluidic environment using p-polarized light with no reflection. This yields a signal that is insensitive to changes in the refractive index of the buffer solution and is sensitive to the binding dynamics of the biomaterial. In contrast to surface plasmon resonance measurements, when measuring the binding properties of biomaterials adsorbed on a substrate material in an immersion microfluidic environment, the buffer solution serves as the incident medium, and light passing through the biomaterial adsorption layer is reflected by the substrate material.

[0013] Under these measurement conditions, the ellipsometric angle Ψ, which represents the measurement amplitude, is sensitive only to changes in the biofilm and substrate material, and is insensitive to changes in the refractive index of the incident medium, such as the buffer solution. For substrates with a stable refractive index, such as silicon, the measured ellipsometric angle Ψ can produce a signal that is sensitive only to changes in the biofilm. When using Figure 1 In the case of the prism incident structure shown, the ellipsometric angle Δ representing the phase can simultaneously measure the thickness of the biofilm and the refractive index of the buffer solution by expressing only a signal sensitive to the refractive index of the buffer solution. However, when using a substrate such as a prism parallel to the plane incident structure, the light reflected by the interface between the prism and the buffer solution should be eliminated and only the light reflected by the substrate should be used. In order to minimize the amount of sample used, it is necessary to reduce the distance between the prism surface and the substrate material. In this case, the two reflected lights are difficult to separate because they are located at a very close distance, and this becomes a factor in measurement error. Therefore, in a planar three-dimensional structure such as a prism, a new structural measurement method is required to distinguish between the light reflected by the interface between the prism and the buffer solution and the light reflected by the substrate material including the sensor.

[0014] Figure 1 FIG1 is a cross-sectional view showing a biomaterial binding property measurement sensor of the prior art. Figure 1 As shown, a conventional biomaterial binding property sensor generally comprises a prism 100, a microfluidic structure 200, a polarized light generating unit 300, and a polarized light detecting unit 400. In this case, the microfluidic structure 200 of the conventional biomaterial binding property sensor forms a liquid-immersed microfluidic channel 210 environment by placing an adsorption layer 530 on a substrate 510 or a dielectric film 520. When a buffer solution 50 containing a dissolved biomaterial sample 1 is injected into the microfluidic channel 210, the ligand 2 formed on the surface of the adsorption layer 530 adsorbs the biomaterial, forming an adsorption layer of a predetermined thickness.

[0015] Furthermore, the polarized incident light generated by the polarized light generating unit 300 passes through the incident surface 110 of the prism and is incident on the interface between the buffer solution 50 and the substrate 510 at an angle that creates a non-reflective condition for p-polarized light waves. In this case, the reflected light reflected by the substrate 510 includes optical data related to the adsorption layer of the sample 1 and the refractive index of the buffer solution. That is, during the process of adsorption and dissociation of the sample 1 by the ligand 2, the molecular adsorption and dissociation kinetics (binding and dissociation kinetics) such as the adsorption concentration, the thickness or refractive index of the adsorption layer, and the refractive index of the buffer solution change, thereby causing the measured ellipsometry angle to change. Furthermore, the reflected light including the optical data is detected by the polarized light detection unit 400. At this time, according to the changes caused by the polarized light component of the reflected light, that is, the polarized light detection unit 400 can grasp the molecular adsorption and dissociation kinetics of the sample 1 and the refractive index of the buffer solution by measuring the ellipsometry angle.

[0016] Figure 2 The adsorption curve showing the adsorption process of the sample 1 by the metal film 30 and the dissociation curve showing the dissociation process are shown. A larger association rate constant (ka) means faster absorption of the biosubstance, while a smaller dissociation rate constant (kd) means slower dissociation.

[0017] That is, the normal dissociation constant (KD = kd / ka) can be calculated by measuring the association rate constant and the dissociation rate constant. For example, the adsorption or dissociation characteristics of a low-molecular-weight new drug candidate that can be used as a carcinogen inhibitor on a protein containing a carcinogen can be measured to determine whether it can be used as a new drug.

[0018] Below, refer to Figure 3 and Figure 4 , the characteristics and limitations of existing biomaterial analysis sensors are explained. Figure 3 In the case of the prism-based incident structure shown, the incident light will be incident on the interface at an angle of approximately θ2 = 72.14°. When the incident light enters the buffer solution from the prism, the angle will change by approximately -0.026° due to the change in the refractive index of the buffer solution (0.0002). The no-reflection condition for p-polarized light waves is close to θ2 = 72.14°. Due to the change in the refractive index of the buffer solution, the current angle becomes 72.114°, which is less than 0.026°. Therefore, as shown in Figure 4 As shown, the curves of Ψ and Δ are formed, and the non-reflection angle of the p-polarized light wave does not change with the change of the refractive index. Therefore, the values ​​of Ψ and Δ are measured at an angle of 72.114°, which is less than 0.026°.

[0019] exist Figure 4In the case where the refractive index of the buffer solution 50 is different from each other, the solid line curve indicates that the refractive index of the buffer solution 50 is equivalent to 1.3330, and the dashed line curve indicates that the refractive index of the buffer solution 50 is equivalent to 1.3332. When a prism structure is used, as shown in FIG. Figure 4 As shown, the Ψ value does not change due to changes in the refractive index of the buffer solution; instead, the Δ value increases. In other words, the ellipsometric constant Δ, which relates to phase difference, is sensitive only to changes in the refractive index of the buffer solution and is virtually unaffected by the bonding characteristics. Therefore, changes in the refractive index of the buffer solution alone can be measured with high sensitivity. The change in the ellipsometric constant Δ significantly changes with decreasing film thickness. When used in applied research to analyze material properties or changes in bonding characteristics by measuring refractive index changes, this method enables refractive index measurement with ultra-high sensitivity compared to existing surface plasmon resonance measurement methods.

[0020] When a buffer solution whose refractive index changes due to the continuous supply of a buffer solution or a solvent for a sample is supplied to the sensor through a microchannel, the pure binding kinetics and the refractive index change of the buffer solution can be measured simultaneously.

[0021] However, if Figure 3 As shown, when the distance between the prism base and the substrate is relatively small, it becomes difficult to separate the light reflected from the prism-buffer interface from the light reflected from the substrate. When measuring with p-polarized light without reflection, the intensity of light reflected from the substrate is relatively weak compared to the light reflected from the prism-buffer interface, which can lead to measurement errors. Furthermore, not only does ultra-high sensitivity require high sensitivity, but when measuring extremely low concentrations of substances contained in air, increasing measurement sensitivity can introduce additional challenges.

[0022] Prior art literature

[0023] Korean Patent Gazette No. 10-1105328

[0024] Korean Patent Gazette No. 10-1383652 Summary of the Invention

[0025] Technical problems to be solved

[0026] In order to solve the problems mentioned above, the object of the present invention is to provide the following multiple-reflection liquid-immersion silicon-based microfluidic measurement device and measurement method, that is, high-sensitivity measurement can be achieved by completely separating the first reflected light reflected from the sample detection layer and the second reflected light reflected from the prism-buffer solution interface through multiple reflections.

[0027] Another object of the present invention is to provide a multi-reflection liquid-immersion silicon-based micro-channel measuring device and measuring method that amplify measurement sensitivity by causing light to be incident on a sample multiple times through multiple reflections.

[0028] Furthermore, the object of the present invention is to provide the following multiple-reflection liquid-immersion silicon-based microfluidic measurement device and measurement method, namely, in order to minimize sample consumption, the flow path height is minimized and a multi-channel microfluidic path is set up, thereby providing experimental conditions that can change the concentration of the sample or the adsorption degree of the self-assembled monolayer film in various ways.

[0029] Furthermore, the purpose of the present invention is to provide the following multiple-reflection liquid-immersion silicon-based microfluidic measurement device and measurement method, that is, in a liquid-immersion microfluidic environment, high-sensitivity measurement of biological binding substances can be achieved in a non-labeling manner, and therefore can be widely used in various fields such as biology, medicine, food, and environment.

[0030] The technical objectives to be achieved by the present invention are not limited to the technical objectives mentioned above. Ordinary technicians in the technical field to which the present invention belongs can clearly understand other technical objectives not mentioned through the following description.

[0031] Means used to solve problems

[0032] In order to achieve the above-mentioned purpose, the multi-reflection liquid-immersion silicon-based microfluidic assay device of the present invention is characterized in that it includes: a microfluidic structure, which is composed of a bracket and at least one microfluidic path, wherein the microfluidic path is formed on the bracket, a sample detection layer is formed on the microfluidic path, and a bio-binding substance for detecting the sample is fixed on the sample detection layer; a sample injection part, which is used to inject a buffer solution containing the sample into the microfluidic path; a prism unit, which is provided with a prism and a reflector, wherein the reflector is formed by coating a reflective mirror surface on the bottom surface of the prism; a polarized light generating part, which is used to generate polarized light; and a polarized light generating part. The polarization detection unit is used to detect the polarization change of the reflected light. The above polarized light forms incident light that passes through the above prism and is incident on the prism-buffer solution interface where the above prism contacts the above buffer solution. After a portion of the above incident light is reflected by the above prism-buffer solution interface, it forms a first reflected light that passes through the above prism. After another portion of the above incident light passes through the above prism-buffer solution interface, it performs repeated multiple reflections and multiple reflections of the incident light through the above sample detection layer and the above reflector, forming a second reflected light that passes through the above prism. The above first reflected light and the above second reflected light are completely separated at the spatial level through the above multiple reflections.

[0033] According to an embodiment of the present invention, another part of the above-mentioned incident light passes through the above-mentioned buffer solution and forms transmitted light that is incident on the above-mentioned sample detection layer at an incident angle that meets the no-reflection condition of the p-polarized light wave. After the above-mentioned transmitted light is reflected by the above-mentioned sample detection layer and is mirror-reflected by the above-mentioned reflector at the above-mentioned prism-buffer solution interface to perform the above-mentioned multiple reflections, a second reflected light that passes through the above-mentioned prism can be formed.

[0034] According to an embodiment of the present invention, the sample detection layer includes: a substrate; a dielectric film formed on the upper portion of the substrate; and an adsorption layer formed on the upper portion of the dielectric film, wherein the bioconjugate for detecting the sample may be fixed on the adsorption layer.

[0035] According to an embodiment of the present invention, when light is reflected multiple times by the sample due to the multiple reflections and the non-reflection reflectivity of the p-polarized light wave decreases, the signal intensity of the transmitted light can be prevented from decreasing by increasing the thickness of the dielectric film.

[0036] According to an embodiment of the present invention, when light is reflected multiple times by the sample due to the above-mentioned multiple reflections and causes the non-reflection reflectivity of the above-mentioned p-polarized light wave to decrease, the signal intensity of the above-mentioned transmitted light can be prevented from decreasing by forming the above-mentioned transmitted light to be incident on the above-mentioned sample detection layer at an incident angle that satisfies the non-reflection condition of the s-polarized light wave.

[0037] According to an embodiment of the present invention, the substrate may be made of one or more materials selected from silicon, dielectrics, or semiconductors.

[0038] According to an embodiment of the present invention, the polarization detection unit may calculate the thickness or concentration of the sample adsorbed on the adsorption layer based on the polarization change of the second reflected light.

[0039] According to an embodiment of the present invention, the polarized light generating unit can adjust the brightness of the incident light incident through the prism and control the shape of a beam spot formed by the incident light on the interface between the prism and the buffer solution.

[0040] According to an embodiment of the present invention, the sample injection unit may inject gas into the microchannel instead of the buffer solution in order to measure biomarkers contained in air or gas.

[0041] According to an embodiment of the present invention, when the gas is injected into the microchannel, the polarized light passes through the prism and forms incident light that is incident on the prism-gas interface where the prism contacts the gas. After a portion of the incident light is reflected by the prism-gas interface, it can pass through the prism and form the first reflected light.

[0042] In order to achieve the above-mentioned purpose, the multi-reflection liquid-immersion silicon-based microfluidic measurement method of the present invention includes: a first step, injecting a buffer solution into the above-mentioned microfluidic structure through a sample injection part, the above-mentioned microfluidic structure includes at least one microfluidic path, the above-mentioned sample detection layer is formed in the above-mentioned microfluidic path, and the above-mentioned sample detection layer is fixed with a bio-binding substance for detecting the sample; a second step, allowing the above-mentioned sample contained in the above-mentioned buffer solution to be adsorbed by the antibody of the above-mentioned sample detection layer; a third step, allowing the above-mentioned polarized light generating part to generate polarized light; a fourth step, allowing the above-mentioned polarized light to pass through the above-mentioned prism to form an incident light incident on the above-mentioned prism-buffer solution interface where the above-mentioned prism contacts the above-mentioned buffer solution; a fifth step, allowing a part of the above-mentioned incident light to be reflected by the above-mentioned prism-buffer solution interface to form a transmission light. The above-mentioned first reflected light of the prism causes another part of the above-mentioned incident light to pass through the above-mentioned buffer solution to form transmitted light incident on the above-mentioned sample detection layer at an incident angle that meets the no-reflection condition of polarized light waves; the sixth step causes the above-mentioned transmitted light to be reflected by the above-mentioned sample detection layer and to be mirror-reflected by the above-mentioned reflector at the above-mentioned prism-buffer solution interface to perform repeated multiple reflections and multiple reflections of the incident light, and then pass through the above-mentioned prism to form the second reflected light; the seventh step detects the polarization change of the above-mentioned second reflected light by the above-mentioned polarization detection unit; and the eighth step detects the concentration of the sample adsorbed on the above-mentioned sample detection layer based on the polarization change of the above-mentioned second reflected light, and the above-mentioned first reflected light and the above-mentioned second reflected light released to the outside of the above-mentioned prism unit are completely separated on the spatial level through the above-mentioned multiple reflections.

[0043] Effects of the Invention

[0044] In existing measurement methods, the light energy reflected from the interface between the prism and the measurement medium is greater than the light energy reflected from the substrate material, and measurement errors may occur due to difficulty in separation. In contrast, the present invention having the above-mentioned structure can apply a multiple reflection structure to completely separate the light reflected from the interface between the prism and the measurement medium and the light reflected from the substrate material to obtain a signal amplification effect based on multiple reflections.

[0045] Furthermore, the present invention has the following effect: in order to minimize sample consumption, the flow path height is minimized and a multi-channel microflow path is provided, thereby providing experimental conditions that can change the concentration of the sample or the adsorption degree of the self-assembled monolayer film in various ways.

[0046] Moreover, in a liquid-immersion microfluidic environment, the present invention can achieve high-sensitivity determination of bioconjugates in a non-labeling manner, and therefore can be widely used in various fields such as biology, medicine, food, and environment.

[0047] The effects of the present invention are not limited to the above-mentioned effects, but should be understood to include all effects that can be derived from the inventive structure described in the detailed description of the present invention or the scope of the claims. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 FIG1 is a cross-sectional view showing a biomaterial binding property measuring sensor of the prior art.

[0049] Figure 2 Schematic diagram showing the change in adsorption concentration during the adsorption and dissociation process of the sample on the metal film.

[0050] Figure 3 Schematic diagram of a prism-incident silicon-based liquid-immersion micro-flow measurement sensor for explaining the problems of the prior art.

[0051] Figure 4 This graph shows the ellipsometry constants Ψ and Δ, which represent the adsorption of biomaterials and the change in the refractive index of buffer solutions, measured using a conventional biomaterial binding property measurement sensor.

[0052] Figure 5 This is a simplified diagram of a microfluidic measurement device without a reflector according to an embodiment of the present invention.

[0053] Figure 6 and Figure 7 FIG. 1 is a schematic diagram of a micro-fluidic measurement device having a reflector formed thereon according to an embodiment of the present invention.

[0054] Figure 8 FIG. 1 is a perspective view of a microfluidic assay device according to an embodiment of the present invention.

[0055] Figure 9 This is an exploded perspective view of a microfluidic assay device according to an embodiment of the present invention.

[0056] Figure 10 FIG. 4 is a perspective view of a prism unit and a first structure according to an embodiment of the present invention.

[0057] Figure 11 FIG. 4 is a cross-sectional view of a second structure according to an embodiment of the present invention.

[0058] Figure 12 FIG. 1 is a graph showing how the amplitude of the second reflected light changes with the number of reflections of the transmitted light due to the p-polarized light wave according to an embodiment of the present invention.

[0059] Figure 13 FIG. 1 is a graph showing how the amplitude of the second reflected light changes with the number of reflections of the transmitted light due to the s-polarized light wave according to an embodiment of the present invention. DETAILED DESCRIPTION

[0060] According to the most preferred embodiment of the present invention, it is characterized in that it includes: a microfluidic structure, which is composed of a bracket and at least one microfluidic path, the microfluidic path is formed on the bracket, a sample detection layer is formed on the microfluidic path, and a bio-binding substance for detecting the sample is fixed on the sample detection layer; a sample injection part, which is used to inject a buffer solution containing the sample into the microfluidic path; a prism unit, which is provided with a prism and a reflector, and the reflector is formed by coating a reflective mirror surface on the bottom surface of the prism; a polarized light generating part, which is used to generate polarized light; and a polarized light detecting part, which is used to detect the sample. In detecting the polarization change of the reflected light, the polarized light forms incident light that passes through the prism and is incident on the prism-buffer solution interface where the prism contacts the buffer solution. A portion of the incident light is reflected by the prism-buffer solution interface to form a first reflected light that passes through the prism. Another portion of the incident light passes through the prism-buffer solution interface and then, after repeated multiple reflections and multiple reflections of the sample detection layer and the reflector, forms a second reflected light that passes through the prism. The first reflected light and the second reflected light are completely separated on a spatial level through the multiple reflections.

[0061] The present invention is described below with reference to the accompanying drawings. However, the present invention can be implemented in a variety of different embodiments and is not limited to the embodiments described herein. Furthermore, to clarify the present invention, portions not relevant to the description will be omitted, and similar reference numerals will be used throughout the specification to denote similar portions.

[0062] Throughout this specification, when a component is described as being "connected (coupled, in contact with, or combined with)" to another component, this includes not only "direct connection" but also "indirect connection" with other components interposed therebetween. Furthermore, when a component "includes" another component, this implies that the component also includes the other component and does not exclude the other component, unless otherwise specified.

[0063] The terms used in this specification are only used to illustrate specific embodiments and are not intended to limit the present invention. Unless the context clearly indicates otherwise, expressions in the singular include expressions in the plural. Terms such as "including" or "having" in this specification are only used to specify the presence of features, numbers, steps, tasks, structural elements, components, or combinations thereof described in this specification, and do not preclude the presence or additional possibility of one or more other features, numbers, steps, tasks, structural elements, components, or combinations thereof.

[0064] Structure of the First Embodiment

[0065] Hereinafter, the present invention will be described in detail with reference to the accompanying drawings.

[0066] Figure 5 FIG. 1 is a schematic diagram of a microfluidic measurement device without the reflector 120 according to an embodiment of the present invention. Figure 6 and Figure 7 FIG. 1 is a schematic diagram of a microfluidic measurement device having a reflector 120 according to an embodiment of the present invention. Figure 5 is a diagram showing a situation where the transmitted light 20 is not reflected by the reflector 120. Figure 6 is a diagram showing the situation in which the transmitted light 20 is reflected once by the reflector 120. Figure 7 FIG2 is a diagram showing the situation in which the transmitted light 20 is reflected twice by the reflector 120. Figure 6 and Figure 7 Although the polarized light generating unit 300 and the polarized light detecting unit 400 are omitted, it should be understood that Figure 6 and Figure 7 Also like Figure 5 , a polarized light generating unit 300 and a polarized light detecting unit 400 are provided.

[0067] and, Figure 8 is a perspective view of a microfluidic assay device according to an embodiment of the present invention. Figure 9 This is an exploded perspective view of a microfluidic measurement device according to an embodiment of the present invention. Figure 10 FIG. 1 is a perspective view of a prism unit 100 and a first structure 200 a according to an embodiment of the present invention. Figure 11 FIG. 2 is a cross-sectional view of a second structure 200 b according to an embodiment of the present invention.

[0068] like Figures 5 to 11 As shown, the microfluidic channel 210 measuring device of the present invention includes: a microfluidic channel structure 200, which is composed of a bracket 220 and at least one microfluidic channel 210, the above-mentioned microfluidic channel 210 is formed on the bracket 220, and a sample detection layer 500 is formed on the above-mentioned microfluidic channel 210, and a bioconjugate for detecting sample 1 is fixed on the above-mentioned sample detection layer 500; a sample 1 injection part, which is used to inject a buffer solution 50 including sample 1 into the microfluidic channel 210; a prism unit 100, which is provided with a prism 110 and a reflector 120, and the above-mentioned reflector 120 is formed by coating a reflecting mirror surface on the bottom surface of the prism 110; a polarized light generating part 300, which is used to generate polarized light; and a polarized light detecting part 400, which is used to detect the polarization change of the reflected light.

[0069] Among them, polarized light forms incident light 10 that transmits through the prism 110 and is incident on the prism-buffer solution interface 111 where the prism 110 contacts the buffer solution 50. After a portion of the incident light 10 is reflected by the prism-buffer solution interface 111, it forms first reflected light that transmits through the prism 110. After another portion of the incident light 10 transmits through the buffer solution 50, it forms transmitted light 20 that is incident on the sample detection layer 500 at an incident angle that satisfies the no-reflection condition for polarized light waves. The transmitted light 20 is reflected by the sample detection layer 500 and is mirror-reflected by the reflector 120 at the prism-buffer solution interface 111 to perform repeated multiple reflections and multiple reflections of the incident light, thereby forming second reflected light 40 that transmits through the prism 110.

[0070] Furthermore, the first reflected light 30 and the second reflected light 40 released to the outside of the prism unit 100 can be completely separated spatially by multiple reflections.

[0071] One embodiment of the present invention utilizes ellipsometry to measure the adsorption and dissociation kinetics of bioconjugates, including low-molecular-weight substances, and has a structure in which a buffer solution 50 containing a bioconjugate sample (not shown) is injected into a microfluidic structure 200. In this case, the microfluidic structure 200 may include a multi-channel microfluidic path 210.

[0072] The prism 110 can be mainly made of optical glass, for example, BK7 or SF10, but is not limited thereto. Furthermore, the prism 110 can also be assembled from a plurality of unit prisms.

[0073] The reflector 120 may be formed by applying a mirror coating to the bottom surface of the prism 110, thereby reflecting the transmitted light 20 toward the reflector 120. Furthermore, the length of the reflector 120 may be changed according to the number of reflections of the transmitted light.

[0074] When the number of reflections of the transmitted light increases due to an increase in the length of the reflector 120 (the length in the direction of travel of the transmitted light 20), the distance between the first reflected light 30 and the second reflected light 40 increases, thereby improving the spatial separation performance of the first reflected light 30 and the second reflected light 40, making it easier to measure the second reflected light 40. However, when the number of reflections of the transmitted light 20 increases, the reflectivity decreases as the light is reflected multiple times by the sample 1. Therefore, the length of the reflector 120 can be set to reflect the change in the reflectivity of the transmitted light 20.

[0075] As described above, when light is reflected multiple times by sample 1 due to multiple reflections, resulting in a decrease in the non-reflection reflectivity of the p-polarized light wave, a decrease in the signal intensity of transmitted light 20 can be prevented by increasing the thickness of dielectric film 520. When the thickness of dielectric film 520 is increased, that is, when the film thickness is changed from several nanometers to several hundred nanometers, the reflectivity increases dramatically, and thus, even if transmitted light 20 is reflected multiple times, a decrease in the signal intensity of transmitted light 20 can be prevented.

[0076] Furthermore, even when light is reflected multiple times by sample 1 due to multiple reflections, resulting in a decrease in the non-reflection reflectivity of p-polarized light, a decrease in the signal intensity of transmitted light 20 can be prevented by forming transmitted light 20 incident on sample detection layer 500 at an angle of incidence that satisfies the non-reflection condition for s-polarized light. The non-reflection condition for s-polarized light is the opposite of the non-reflection condition for p-polarized light, resulting in a very large signal. Therefore, even when light is reflected multiple times by sample 1 due to multiple reflections, a large signal can be detected.

[0077] The microchannel structure 200 includes one or more microchannels 210. The microchannels 210 are disposed below the prism unit 100 and form a sample detection layer 500. A bio-binding substance for detecting a sample may be immobilized on the sample detection layer 500.

[0078] Specifically, in one embodiment of the present invention, the microfluidic structure 200 may include a plurality of microfluidic paths 210 serving as passages for inflow or outflow of a buffer solution 50 containing a sample 1. In this case, the width of the microfluidic paths 210 may be a small size of several millimeters or less than 1 millimeter. Furthermore, each of the plurality of microfluidic paths 210 may include an inflow path 210a, a microfluidic channel 210c, and an outflow path 210b. In other words, the microfluidic paths 210 may be formed by connecting the inflow path 210a, the microfluidic channel 210c, and the outflow path 210b.

[0079] On the other hand, the sample injection unit 600 can inject or discharge the buffer solution 50 including the sample 1 into the microchannel 210 .

[0080] The polarized light generating unit 300 can adjust the brightness of the incident light 10 incident through the prism 110 and control the shape of a beam spot formed by the incident light 10 on the prism-buffer solution interface.

[0081] Specifically, the polarized light generating unit 300 is used to generate polarized light, and may include a light source 310 and a polarizer 320 , and may also include a collimator 330 , a condenser 340 , or a first compensator 350 .

[0082] The polarizer 320 and the first compensator 350 are rotatable structures, or other polarization light modulation units may be included. The incident light 10 may include all polarization components of p-polarized light waves and s-polarized light waves. In order to improve the signal-to-noise ratio (SNR), the polarizer 320 can be arranged at an angle close to the p-polarized light wave. Preferably, the ratio of the p-polarized light wave and the s-polarized light wave included in the second reflected light 40 received by the polarized light detection unit 400 described below can be similar. In this way, the shape of the beam spot formed by the incident light 10 on the prism-buffer solution interface 111 can be controlled.

[0083] In an embodiment of the present invention, incident light 10 may be incident on the sample detection layer 500 at an incident angle θ1 that satisfies the no-reflection condition for p-polarized light. In the ellipsometry equation, the complex reflection coefficient ratio p is the ratio of the reflection coefficient ratio Rs of the s-polarized light wave to the reflection coefficient Rp of the p-polarized light wave, that is, it can be expressed as p = Rp / Rs. The no-reflection condition for p-polarized light wave means that the reflection coefficient ratio Rp of the p-polarized light wave is close to 0. The no-reflection condition for p-polarized light wave is similar to the surface plasmon resonance condition of existing surface plasmon resonance sensors and can maximize the measurement sensitivity of the present invention.

[0084] Light source 310 can be various lamps emitting monochromatic light or white light in the infrared, visible, or ultraviolet regions; light-emitting diodes (LEDs); solid-state, liquid, or gas lasers; and semiconductor laser diodes (LDs) including laser diodes. Furthermore, light source 310 can have a structure in which the wavelength varies depending on the optical system structure. On the other hand, when p-polarized light waves approach a non-reflective condition, the optical signal of the reflected light is relatively small. Therefore, in this case, high-intensity laser irradiation with coherence can be used to improve the signal-to-noise ratio and achieve high-sensitivity measurement.

[0085] The collimator 330 receives light from the light source 310 and provides parallel light to the polarizer 320. Furthermore, the condenser 340 converges the parallel light passing through the polarizer 320 to increase the brightness of the incident light 10. Furthermore, the first compensator 350 serves to phase-delay the polarization component of the incident light 10.

[0086] The polarization detection unit 400 can calculate the thickness or concentration of the sample 1 adsorbed on the adsorption layer 530 based on the change in polarization of the reflected light. Specifically, the polarization detection unit 400 receives the second reflected light 40 and detects the change in polarization. As the second reflected light 40 is received and reflected by the adsorption layer 530, the change in its polarization state can be detected. The polarization detection unit 400 may include an analyzer 410, a light detector 420, and a processor 430. Additionally, it may include a second compensator 440 and a spectrometer 450.

[0087] Analyzer 410 corresponds to polarizer 320 and may be provided with a polarizing plate. It controls the polarization degree or polarization plane of the reflected light by further polarizing the second reflected light 40. Depending on the structure of the optical system, analyzer 410 may be rotatable or may further include a polarization modulation unit that can perform functions such as phase shifting and elimination of polarized light components.

[0088] The photodetector 420 obtains optical data by detecting the polarized second reflected light 40 and converting it into an electrical signal. In this case, the optical data may include information related to changes in the polarization state of the second reflected light 40. A CCD-type solid-state imaging device, a photomultiplier tube (PMT), or a silicon photodiode may be used as the photodetector 420.

[0089] The arithmetic processor 430 can obtain electrical signals from the photodetector 420 and derive measurement values. The arithmetic processor 430 contains a predetermined analysis program utilizing reflectometry and ellipsometry. Therefore, the arithmetic processor 430 can extract and analyze the optical data converted into electrical signals to derive measurement values ​​such as the sample adsorption concentration, the thickness of the adsorption layer 160, the adsorption constant, the dissociation constant, and the refractive index. In this case, to improve measurement sensitivity, the arithmetic processor 430 preferably derives the measurement values ​​by calculating the ellipsometry constants Ψ and Δ, which are related to the phase difference of the ellipsometry method.

[0090] By using the memory analysis program described above, the micro-flow channel measurement device of the present invention can not only perform ellipsometry but also perform the above-mentioned measurement based on reflectometry under the condition that there is no reflection of p-wave or s-wave.

[0091] The second compensator 440 is used to adjust the phase of the polarized light component of the reflected light by implementing phase delay. The second compensator 440 is a rotatable structure, or may further include other polarization light modulation units.

[0092] Beam splitter 450 can be used when light source 310 is white light. This is because the reflected light must be split to separate the reflected light within a narrow wavelength range in order to transmit it to photodetector 420. In this case, photodetector 420 can obtain optical data on the distribution of the reflected light using a two-dimensional image sensor such as a CCD solid-state imaging device.

[0093] Hereinafter, the micro-flow channel structure 200 will be described in further detail.

[0094] like Figure 8 and Figure 9 As shown, the micro-channel structure 200 of the present invention may include a first structure 200a and a second structure 200b.

[0095] In this case, the first structure 200a can be formed at the bottom of the prism unit 100. In particular, in the embodiment of the present invention, the prism unit 100 and the first structure 200a can be formed as an integral body, but the present invention is not limited thereto. Furthermore, the first structure 200a and the second structure 200b can be separated from each other, and the second structure 200b can include the microfluidic channel layer 200c.

[0096] On the other hand, the first structure 200 can be made of a transparent material such as glass or a transparent synthetic resin material. In this case, an acrylic resin such as polymethyl methacrylate (PMMA) can be used as the synthetic resin material. Alternatively, a silicon-based material such as polydimethylsiloxane (PDMS) can be used.

[0097] Specifically, the first structure 200a may include: a plurality of inflow paths 210a formed on one side of the first structure 200a; and a plurality of outflow paths 210b formed on the other side of the first structure. The inflow paths 210a are formed by connecting a first inflow port 212 formed on one side of the first structure 200a with a second inflow port 214 formed at the bottom of the first structure 200a. The outflow paths 210b are formed by connecting a first outflow port 216 formed at the bottom of the first structure 200a with a second outflow port 218 formed on the other side of the first structure 200a.

[0098] On the other hand, the plurality of inflow paths 210a and the exhaust paths 210b can be respectively connected to the plurality of microfluidic channels 210c formed in the microfluidic channel layer 200c of the second structure 200b. Specifically, the second inlet 214 can be coupled to one side of the microfluidic channel 210c, and the first inflow path 210a can be connected to the microfluidic channel 210c.

[0099] Furthermore, the first outlet 216 contacts the other side of the micro-channel 210 c , and the outlet path 210 b may be connected to the micro-channel 210 c .

[0100] That is, the inflow path 210a, the microfluidic channel 210c, and the discharge path 210b may be connected so that the buffer solution 50 including the sample injected through the inflow path 210a may be discharged along the discharge path 210b through the microfluidic channel 210c.

[0101] In other words, the second structure 200b includes a microchannel layer 200c, which may include a plurality of microchannels 210c. The microchannel layer 210c may be made of acrylic resin such as polymethylmethacrylate (PMMA), but is not limited thereto.

[0102] Second structure 200b may include a sample detection layer 500 formed on the bottom surface of the groove formed by the plurality of microfluidic channels 210c. Sample detection layer 500 comprises a substrate 510, a dielectric film 520 formed on top of substrate 510, and an adsorption layer 530 formed on top of dielectric film 520. A bio-binding substance used to detect sample 1 may be immobilized on adsorption layer 530.

[0103] Substrate 510 can be made of one or more materials selected from silicon, dielectrics, or semiconductors. Silicon has the advantage of providing stable, predetermined physical properties at a low cost. Furthermore, semiconductors or dielectrics offer a significant difference in refractive index from biological materials, resulting in higher measurement sensitivity compared to silicon under non-reflective conditions for p-waves or s-waves. Furthermore, at 655 nm, substrate 510 can have a complex refractive index of approximately 3.8391 + 10.018186.

[0104] The dielectric film 520 formed on the substrate 510 can be made of a transparent semiconductor oxide film or a glass film. The thickness of the dielectric film 520 can be 0 mm to 10 mm.

[0105] On the other hand, the most readily available dielectric film 520 can be a silicon oxide film (SiO2) grown to a thickness of several nanometers by naturally oxidizing silicon. The refractive index of the silicon oxide film is approximately 1456 at 655 nm. Since this refractive index is significantly different from that of the silicon substrate 510, it is beneficial for improving the measurement sensitivity of the present invention.

[0106] Furthermore, a glass film made of optical glass can be used as the dielectric film 520. Compared to metal films made of gold, silver, etc., the refractive index of the dielectric film 520 made of silicon, silicon oxide film, or glass film is relatively stable, thereby providing stable optical characteristics and having the advantage of reducing manufacturing costs.

[0107] In one embodiment of the present invention, adsorption layer 530 can be made of either a self-assembled thin film or a biofilm. Furthermore, a biobinding substance used to detect a specific sample can be immobilized on adsorption layer 530. In this case, adsorption layer 530 can adsorb and dissociate the low-molecular-weight biobinding substance sample 1 and reflect the transmitted light 20.

[0108] In other words, the sample contained in the buffer solution flowing in through the inflow path 210 a may be adsorbed by the adsorption layer 530 or desorbed from the adsorption layer 530 .

[0109] The sample injection unit 600 can inject or discharge a buffer solution containing a sample into the microchannel 210. Therefore, not only the buffer solution can be used, but also gas can be used as a medium in the microchannel 210 to measure biomarkers contained in the gas. While air can be used as the gas, this is not limiting, and other gases can also be used. In this case, the air containing the biomarkers can be injected into the microchannel 210 from the sample injection unit 600.

[0110] When gas is injected into microchannel 210, polarized light passes through prism 110 and forms incident light 10 that is incident on the prism-gas interface where prism 110 and gas are in contact. A portion of incident light 10 is reflected by the prism-gas interface and then passes through prism 110 to form first reflected light 30. Thus, even in the above-described situation, a highly sensitive signal can be obtained under conditions where there is no reflection of p-waves or s-waves.

[0111] Operation of the First Embodiment

[0112] Hereinafter, the operation of the preferred embodiment will be described in detail with reference to the accompanying drawings.

[0113] like Figures 5 to 7As shown, the prism 110 can be a single prism structure. Incident light 10 can pass through the prism 110 and be incident on the prism-buffer solution interface 111. At the prism-buffer solution interface 111, the incident light 10 can be separated into first reflected light 30 and transmitted light 20. After being reflected by the sample detection layer 500, the transmitted light 20 can be incident on the reflector 120 at the prism-buffer solution interface 111. Furthermore, after being reflected by the reflector 120, the transmitted light 20 can be incident on the sample detection layer 500 again and reflected. Furthermore, after performing the above-mentioned multiple reflections and multiple reflections, the transmitted light 20 can pass through the prism-buffer solution interface 111 and the prism 110 without the reflector 120, and form second reflected light 40. The spatial separation between the first reflected light 30 and the second reflected light 40 (the distance between the respective light paths) can increase with the number of reflections in the multiple reflections.

[0114] As described above, the polarization detection unit 400 can obtain physical property information about the thickness or optical characteristics of the sample 1 by receiving the second reflected light 40 and analyzing its polarization state. Figure 5 As shown, when the reflector 120 is not used, the optical path interval between the first reflected light 30 and the second reflected light 40 is relatively small. Since it is difficult to separate the first reflected light 30 and the second reflected light 40, each reflected light may be incident on the polarized light detection unit 400.

[0115] Specifically, in the conventional art, as light reflected from the interface between the prism 110 and the medium is refracted, it becomes difficult to separate it from the light incident on the adsorption layer 530. Consequently, the light reflected from the interface between the prism 110 and the medium is refracted and detected by the polarized light detection unit 400 together with the light incident on the adsorption layer 530. Consequently, the light reflected from the interface between the prism 110 and the medium, which has relatively high energy, causes measurement errors, resulting in reduced measurement sensitivity.

[0116] Furthermore, in order to allow the polarization detection unit 400 to receive only the second reflected light 40 , there arises a need to configure an additional optical system in front of the polarization detection unit 400 or minimize the beam spot size of the incident light 10 .

[0117] In order to solve the above problems, Figure 6 and Figure 7As shown, by forming a reflector 120 on the bottom surface of the prism 110 and guiding the multiple reflections of the transmitted light 20, the optical path distance between the first reflected light 30 and the second reflected light 40 can be significantly increased, thereby spatially separating the first reflected light 30 and the second reflected light 40. This allows the polarization detection unit 400 to receive the second reflected light 40. Thus, the present invention can improve the measurement sensitivity of the microfluidic measurement device by preventing measurement errors.

[0118] That is, if the polarized light detection unit 400 is positioned at a specific distance from the prism unit 100, the distance separating the second reflected light 40 from the first reflected light 30 will become longer. Therefore, the brightness of the first reflected light 30 gathered in the polarized light detection unit 400 can be minimized. Moreover, if the optical axis of the polarized light detection unit 400 is aligned with the optical path of the second reflected light 40, the effect of a small amount of the first reflected light 30 unnecessarily gathered in the polarized light detection unit 400 can be minimized. That is, in the process of traveling toward the inside of the polarized light detection unit 400, the propagation loss of the first reflected light 30 is greater than the propagation loss of the second reflected light 40. Therefore, compared with the brightness of the second reflected light 40, the brightness of the first reflected light 30 reaching the polarized light detection unit 400 can be limited to a relatively small amount.

[0119] As a result, the micro-flow measurement device of the present invention can easily separate the second reflected light 40 containing characteristic information of the sample 1 from the first reflected light 30 , thereby preventing measurement errors and easily achieving high-sensitivity analysis of the sample 1 .

[0120] Figure 12 FIG. 4 is a graph showing how the amplitude of the second reflected light 40 changes with the number of reflections of the transmitted light 20 due to the p-polarized light wave according to an embodiment of the present invention. Figure 13 FIG. 4 is a graph showing how the amplitude of the second reflected light 40 changes with the number of reflections of the transmitted light 20 according to an embodiment of the present invention. Figure 12 and Figure 13 The curve shown is the ellipsometric constant Ψ curve.

[0121] in, Figure 12 Part (a) is because Figure 5 The graph is a graph showing a case where the number of reflections of the transmitted light 20 is 0 due to the p-polarized light wave of the reflector 120. Figure 12 Part (b) is because for Figure 6 The graph is a case where the number of reflections of the transmitted light 20 is 1 due to the p-polarized light wave of the reflector 120. Figure 12 Part (c) is because Figure 7 The graph shows a case where the number of reflections of the transmitted light 20 is 2 due to the p-polarized light wave shown.

[0122] and, Figure 13 Part (a) is because Figure 5 The graph is a graph showing a case where the number of reflections of the transmitted light 20 is 0 due to the s-polarized light wave of the reflector 120. Figure 13 Part (b) is because for Figure 6 The graph is a graph showing a case where the number of reflections of the transmitted light 20 is 1 due to the s-polarized light wave of the reflector 120. Figure 13 Part (c) is because Figure 7 The graph is a case where the number of reflections of the transmitted light 20 is 2 due to the s-polarized light wave shown.

[0123] exist Figure 12 Part (a), (b), (c) and Figure 13 In the graphs shown in parts (a), (b), and (c), the indicator curve is a thick line (red).

[0124] like Figure 12 As shown in parts (a), (b), and (c), as the number of reflections of the transmitted light 20 increases due to the p-polarized light wave on the reflector 120, it can be confirmed that the amplitude of the second reflected light 40 increases (the amplitude increases from 1 nm to 3 nm). Figure 13 As shown in parts (a), (b), and (c), as the number of reflections of the transmitted light 20 increases due to the s-polarized light wave of the reflector 120, it can be confirmed that the amplitude of the second reflected light 40 increases (the amplitude increases from 1 nm to 3 nm).

[0125] As described above, it can be confirmed that the amplitude of the second reflected light 40 increases and is amplified as the number of reflections of the transmitted light 20 by the reflector 120 increases.

[0126] Hereinafter, a microflow measurement method using the microflow measurement device of the present invention will be described.

[0127] First, in a first step, the sample injection unit 600 injects a buffer solution 50 into the microfluidic structure 200. The microfluidic structure 200 includes at least one microfluidic channel 210. A sample detection layer 500 is formed in the microfluidic channel 210, and a bio-binding substance for detecting a sample is immobilized in the sample detection layer 500. The buffer solution 50 is then injected into each microfluidic channel 210 with a time difference. Furthermore, the buffer solution 50 can be injected into only a portion of the microfluidic channels 210, eliminating the need to use the remaining microfluidic channels 210.

[0128] Then, in the second step, the sample 1 included in the buffer solution 50 can be adsorbed by the antibody of the sample detection layer 500. Alternatively, the sample can be Figure 11As shown, multiple adsorption layers formed on multiple different self-assembled monolayers or the same self-assembled monolayer of the single micro-channel 210c are adsorbed to form adsorption layers with different bonding properties. In the third step, the polarized light generating unit 300 can generate polarized light.

[0129] Subsequently, in the fourth step, the polarized light may pass through the prism 110 and form incident light 10 that is incident on the prism-buffer solution interface 111 where the prism 110 contacts the buffer solution 50. Furthermore, in the fifth step, a portion of the incident light 10 is reflected by the prism-buffer solution interface 111 and forms reflected light that passes through the prism 110, while another portion of the incident light 10 may pass through the buffer solution 50 and form transmitted light 20 that is incident on the sample detection layer 500 at an incident angle that satisfies the no-reflection condition for polarized light waves.

[0130] Then, in the sixth step, after the transmitted light 20 is reflected by the sample detection layer 500 and specularly reflected by the reflector 120 at the prism-buffer solution interface 111, it undergoes repeated multiple reflections and multiple reflections of the incident light, and then passes through the prism 110 to form the second reflected light 40. The fifth and sixth steps can be performed so that the first reflected light 30 and the second reflected light 40 released to the outside of the prism unit 100 are completely separated in space due to the multiple reflections.

[0131] Next, in the seventh step, the polarization detection unit 400 detects the polarization change of the second reflected light 40. Furthermore, in the eighth step, the concentration of the sample 1 adsorbed on the sample detection layer 500 is detected based on the polarization change of the second reflected light 40.

[0132] Specifically, analyzer 410 receives the elliptically polarized second reflected light 40 from adsorption layer 530, allowing only light that matches the polarization characteristics to pass through. Next, photodetector 420 detects changes in the polarization component of second reflected light 40 to obtain specified optical data, converting it into an electrical signal and transmitting it to processor 430.

[0133] Subsequently, the processor 430 storing programs related to reflectometry and ellipsometry can extract and analyze the optical data converted into electrical signals to derive the adsorption concentration, adsorption constant and dissociation constant, refractive index, refractive index of the buffer solution and other measured values ​​of the sample.

[0134] In this case, the processor 430 of the present invention can measure the refractive index of the buffer solution 50 by calculating the ellipsometric constant Δ related to the phase difference in ellipsometry, and calculate the bonding dynamics by measuring the ellipsometric constant Ψ related to the amplitude ratio. This is because, under conditions where p-polarized light is not reflected, the ellipsometric constant Δ related to the phase difference is sensitive only to changes in the refractive index of the buffer solution and is largely unaffected by the bonding dynamics. Therefore, the advantage is that only changes in the refractive index of the buffer solution 50 can be measured, while the ellipsometric constant Ψ related to the amplitude ratio is highly sensitive primarily to the bonding dynamics of the material.

[0135] Therefore, the bonding characteristics of a sample flowing into a buffer solution can be measured by Ψ, and when dissolved in a buffer solution, the pure bonding characteristics can be calculated by simultaneously measuring the change in refractive index by △ or the change in refractive index of the buffer solution including a solvent such as dimethyl sulfoxide (DMSO) used to dissolve the sample.

[0136] The above description of the present invention is merely illustrative. It should be understood that those skilled in the art can readily adapt the present invention to other embodiments without altering the technical concept or essential features of the present invention. Therefore, the embodiments described above are merely illustrative and should not be construed as limiting. For example, the various structural elements of a single structure may be implemented separately, and similarly, separate structural elements may be implemented in combination.

[0137] The scope of the present invention should be defined based on the scope of protection claimed in the invention. It should be understood that all modified or deformed embodiments derived from the meaning, scope and equivalent concepts of the scope of protection claimed in the invention belong to the scope of the present invention.

[0138] Description of Reference Signs

[0139] 1: Sample

[0140] 2: Ligand

[0141] 10: Incident light

[0142] 20: Transmitted light

[0143] 30: First reflected light

[0144] 40: Second reflected light

[0145] 50: buffer solution

[0146] 100: Prism unit

[0147] 110: Prism

[0148] 111: Prism-buffer solution interface

[0149] 120: Reflector

[0150] 200: Microfluidic structure

[0151] 200a: First structure

[0152] 200b: Second structure

[0153] 200c: Microfluidic channel layer

[0154] 210: Microfluidics

[0155] 210a: Inflow path

[0156] 210b: Exhaust path

[0157] 210c: Microfluidic channel

[0158] 212: First flow entrance

[0159] 214: Second inlet

[0160] 216: First row exit

[0161] 218: Second row exit

[0162] 220: Bracket

[0163] 300: Polarized light generating unit

[0164] 310: Light Source

[0165] 320: Polarized filter

[0166] 330: Collimating mirror

[0167] 340: Condenser

[0168] 350: First compensator

[0169] 400: Polarization detection unit

[0170] 410: Analyzer

[0171] 420: Photodetector

[0172] 430: Computational processor

[0173] 440: Second compensator

[0174] 450: Optical Splitter

[0175] 500: Sample detection layer

[0176] 510: Substrate

[0177] 520: Dielectric film

[0178] 530: Adsorption layer

[0179] 600: Sample injection part

Claims

1. A multi-reflection liquid-immersion silicon-based microfluidic measurement device, characterized in that: include: A microfluidic channel structure is composed of a support and at least one microfluidic channel, wherein the microfluidic channel is formed on the support, a sample detection layer is formed on the microfluidic channel, and a bio-binding substance for detecting a sample is fixed on the sample detection layer; a sample injection portion, for injecting a buffer solution containing the sample into the microchannel; A prism unit is provided with a prism and a reflector, wherein the reflector is formed by coating a reflective mirror surface on the bottom surface of the prism; a polarized light generating unit, configured to generate polarized light; and The polarization detection unit is used to detect the polarization change of the reflected light. The polarized light forms incident light that transmits the prism and is incident on the prism-buffer solution interface where the prism contacts the buffer solution. A portion of the incident light is reflected by the prism-buffer solution interface to form a first reflected light that passes through the prism. Another portion of the incident light passes through the prism-buffer solution interface and then undergoes repeated multiple reflections and incident multiple reflections through the sample detection layer and the reflector to form a second reflected light that passes through the prism. The polarization detection unit detects a change in polarization of the second reflected light. The first reflected light and the second reflected light are completely separated in space through the multiple reflections. Another portion of the incident light passes through the buffer solution and forms transmitted light incident on the sample detection layer at an incident angle that satisfies the no-reflection condition for p-polarized light waves. The transmitted light is reflected by the sample detection layer and then specularly reflected by the reflector at the prism-buffer solution interface to perform the multiple reflections, thereby forming second reflected light that passes through the prism. The transmitted light does not pass through the reflector and is reflected. The length of the reflector is changed according to the number of reflections of the transmitted light.

2. The multi-reflection liquid immersion silicon-based microfluidic measurement device according to claim 1, characterized in that: The sample detection layer includes: substrate; a dielectric thin film formed on an upper portion of the substrate; and The adsorption layer is formed on the upper part of the dielectric film. The biobinding substance used to detect the sample is immobilized on the adsorption layer.

3. The multi-reflection liquid immersion silicon-based microfluidic measurement device according to claim 2, characterized in that: When light is reflected multiple times by the sample due to the multiple reflections and the reflectivity of the transmitted light incident at an incident angle that satisfies the p-polarized light wave no-reflection condition is reduced, the thickness of the dielectric film is increased to prevent the signal intensity of the transmitted light from decreasing.

4. The multi-reflection liquid immersion silicon-based microfluidic measurement device according to claim 2, characterized in that: When light is reflected multiple times by the sample due to the multiple reflections and the reflectivity of the transmitted light incident at an angle that satisfies the no-reflection condition for the p-polarized light wave is reduced, the signal intensity of the transmitted light is prevented from decreasing by forming transmitted light incident on the sample detection layer at an angle that satisfies the no-reflection condition for the s-polarized light wave.

5. The multi-reflection liquid immersion silicon-based microfluidic measurement device according to claim 2, characterized in that: The substrate is made of one or more materials selected from silicon and dielectrics.

6. The multi-reflection liquid immersion silicon-based microfluidic measurement device according to claim 2, characterized in that: The polarization detection unit calculates the thickness or concentration of the sample adsorbed on the adsorption layer based on the polarization change of the second reflected light.

7. The multi-reflection liquid immersion silicon-based microfluidic measurement device according to claim 1, characterized in that: The polarized light generating unit adjusts the brightness of the incident light entering through the prism and controls the shape of a beam spot formed by the incident light on the interface between the prism and the buffer solution.

8. The multi-reflection liquid-immersion silicon-based microfluidic measurement device according to claim 1, characterized in that: The sample injection unit injects gas into the microchannel instead of the buffer solution in order to measure the biomarker contained in the air or gas.

9. The multi-reflection liquid-immersion silicon-based microfluidic measurement device according to claim 8, characterized in that: When the gas is injected into the microchannel, the polarized light passes through the prism and becomes incident light on a prism-gas interface where the prism contacts the gas. A portion of the incident light is reflected by the prism-gas interface, then passes through the prism and forms the first reflected light.

10. A measurement method using the multi-reflection liquid-immersion silicon-based microfluidic measurement device according to claim 1, characterized in that: include: In a first step, a buffer solution is injected into the microfluidic structure through a sample injection portion, wherein the microfluidic structure includes at least one microfluidic channel, the sample detection layer is formed in the microfluidic channel, and a bio-binding substance for detecting a sample is immobilized on the sample detection layer; The second step is to allow the sample contained in the buffer solution to be adsorbed by the antibody of the sample detection layer; The third step is to make the polarized light generating unit generate polarized light; The fourth step is to allow the polarized light to pass through the prism to form incident light incident on the prism-buffer solution interface where the prism contacts the buffer solution; A fifth step is to cause a portion of the incident light to be reflected by the prism-buffer solution interface to form the first reflected light that passes through the prism, and to cause another portion of the incident light to pass through the buffer solution to form transmitted light that is incident on the sample detection layer at an incident angle that satisfies the no-reflection condition for polarized light waves; In a sixth step, the transmitted light is reflected by the sample detection layer and then reflected by the reflector at the prism-buffer solution interface to perform repeated multiple reflections and multiple reflections of the incident light, and then passes through the prism to form a second reflected light; In a seventh step, the polarization detection unit is used to detect the polarization change of the second reflected light; and In the eighth step, the concentration of the sample adsorbed on the sample detection layer is detected based on the polarization change of the second reflected light. The first reflected light and the second reflected light released to the outside of the prism unit are completely separated on a spatial level through the multiple reflections.

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