Surface plasmon resonance biosensor chip comprising corrugated graphene channel, method of manufacturing same, and method of detecting target material by using same

Abstract

The technical idea of the present invention provides a surface plasmon resonance biosensor chip, a method of manufacturing same, and a method of detecting a target material by using same, the surface plasmon resonance biosensor chip comprising: a substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction; a graphene channel which induces surface plasmon resonance and includes a peak part and a valley part with height difference in the vertical direction on the upper surface of the substrate; a probe disposed on the graphene channel and capable of chemically binding to a target substance to be detected; and a linker molecule for coupling the probe and the graphene channel, wherein the peak part and the valley part are alternately arranged in a zigzag form along the first horizontal direction or a diagonal direction between the first horizontal direction and the second horizontal direction.

Description

Surface plasmon resonance biosensor chip including wrinkled graphene channel and method for manufacturing the same, and method for detecting target substance using the same

[0001] The technical idea of ​​the present invention relates to a surface plasmon resonance (SPR) biosensor chip including a wrinkled graphene channel.

[0002] A biosensor is a system that converts information from a target object into recognizable signals, such as color, fluorescence, or electrical signals, by utilizing biological elements or mimicking biological systems. Biosensors are a future-oriented convergence technology with diverse applications and significant economic value, and interest in them has been growing recently.

[0003] Biosensors are fundamentally composed of biological elements, physical components, and interfaces. Common biosensors include piezoelectric biosensors, optical biosensors, electrochemical biosensors, and thermistor-based biosensors.

[0004] Piezoelectric biosensors can detect even subtle changes within a substance by altering the resonant frequency of a vibrating quartz crystal. Optical biosensors utilize the principle that biological elements interact with the target substance to attenuate, absorb, or emit light. Examples include transient wave detectors and surface plasmon resonance biosensors. Electrochemical biosensors directly convert signals into electrical signals through a reaction between the biosensor and analyte. Thermistor-based biosensors immobilize biorecognition compounds on a temperature-sensitive element and detect minute temperature changes.

[0005] Biosensors have a wide range of applications, including medical diagnosis, direct monitoring of critically ill patients, drug delivery systems, fermentation monitoring and control, food and pharmaceutical processing, drug action and misuse detection, environmental pollution monitoring, fish farming, and livestock care. Among these, the healthcare sector is the most demanding for biosensors. A common need across all these applications is real-time, on-site chemical information on rapidly evolving issues.

[0006] In this respect, surface plasmon resonance biosensors offer several advantages as biosensors. Specifically, surface plasmon resonance biosensors utilize optical principles to measure molecular interactions without the need for separate labels, such as fluorescent substances, and have the advantage of being able to measure reaction progress in real time.

[0007] Surface plasmon refers to the collective charge density oscillation of electrons that occurs on the surface of a metal thin film, and the surface plasmon wave generated by it is a surface electromagnetic wave that propagates along the interface between the metal thin film and the adjacent dielectric. When an electric field or light is applied to the interface between the metal thin film and the dielectric from the outside, surface charges are induced due to the discontinuity of the vertical component of the electric field at the interface between the two media, and the oscillation of these surface charges appears as a surface plasmon wave. Unlike electromagnetic waves in free space, surface plasmon waves are waves that oscillate parallel to the incident plane and have a p-polarized polarization component. Therefore, surface plasmon excitation by optical methods is only possible with vertically polarized electromagnetic waves.

[0008] Surface plasmon resonance biosensors utilize evanescent fields to examine samples. Evanescent fields are the exponential decrease in electric field strength over a distance approximately the wavelength of the incident light when total internal reflection occurs at the interface between two media. Normally, there is no energy transfer across the interface, so the reflectivity after reflection remains 100%. However, under special conditions, when the evanescent field interacts with matter near the interface, energy transfer occurs, resulting in a decrease in reflectivity—a phenomenon known as attenuated total reflection.

[0009] For attenuated total reflection to occur, a material capable of interacting with incident light must be deposited on one surface of the prism. However, unlike the reflectivity of general total reflection, the reflectivity of a metal film placed on one surface of the prism decreases sharply at a certain angle of incidence. This is because the incident light excites surface plasmons, the oscillations of the surface electron charge density present at the interface between the metal film and the dielectric, resulting in energy transfer. This is called surface plasmon resonance, and the angle of incidence at this time is called the resonance angle.

[0010] Raman scattering is a phenomenon in which molecules emit red-shifted scattered light according to their unique vibrational and rotational energy states when exposed to short-wavelength light. The Raman scattering signal is a unique property of the molecule, and while Raman scattering signal measurement is highly suitable for non-destructive and label-free optical detection of biomaterials, it has the disadvantages of weak signals, low reproducibility, and long measurement times. One method to overcome these problems is surface-enhanced Raman spectroscopy, which amplifies the signal to enable detection in seconds or minutes, and occurs on rough metal surfaces. Surface-enhanced Raman scattering is a phenomenon in which the Raman scattering signal is amplified by a factor of 10^9 to 15 due to the surface plasmon resonance phenomenon.

[0011] Surface plasmon resonance biosensors can be broadly divided into an optical section, a sample fixing section, and a signal processing section. The optical section is divided into a surface plasmon resonance excitation light source, a surface plasmon resonance conversion section, and a light detection section. The sample fixing section consists of a metal thin film, a sample fixing section, etc., and the signal processing section consists of a circuit and operating software that converts the measured reflectivity into an electrical signal in real time.

[0012] Additionally, surface plasmon resonance generated from a zero-dimensional metal nanoparticle structure formed on a metal thin film is called localized surface plasmon resonance (LSPR). Localized surface plasmon resonance has also recently attracted considerable attention in the scientific community.

[0013] Currently, surface plasmon resonance biosensors and localized surface plasmon resonance biosensors with Kretschmann structures are widely used, but with the development of nanotechnology, there is a demand for the development of sensors with higher sensitivity.

[0014] Furthermore, lithography is typically used to form metal nanoparticles on the metal film of a surface plasmon resonance biosensor. Therefore, conventional methods require the fabrication of a mask with a nanoparticle pattern drawn on the metal film and the additional process of shaving the metal film. This necessitates a complex, costly, and time-consuming manufacturing process.

[0015] Graphene, a two-dimensional hexagonal carbon (C) structure, is a new material that has been actively researched worldwide as a replacement for semiconductors. Furthermore, graphene is known to possess electrically semi-metallic properties, yet possess extremely high electrical conductivity (intrinsic electron mobility of 20,000 cm2 / Vs) because charges within it act as zero-effective mass particles. In particular, graphene, which is made by mechanically exfoliating graphite and forming a two-dimensional hexagonal structure of carbon (C) atoms, has been reported to exhibit field effect characteristics when used in transistors. Since then, graphene has been attracting attention as a material that can replace conventional semiconductor materials such as silicon. Accordingly, much research is being conducted on technologies that utilize the inherently excellent electrical conductivity of graphene to replace the metal film of surface plasmon resonance biosensors with graphene, or to rapidly modify the surface of graphene without additional chemical reaction steps instead of forming metal nanoparticles on the metal film through a lithographic process.

[0016] The present invention has been designed to solve such problems, and the technical idea of ​​the present invention is to provide a surface plasmon resonance biosensor chip including a wrinkled graphene channel and a method for manufacturing the same, and a method for detecting a target substance using a surface plasmon resonance biosensor chip including a wrinkled graphene channel.

[0017] In order to solve the above-described problem, the technical idea of ​​the present invention provides a surface plasmon resonance biosensor chip including a substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction; a graphene channel that generates surface plasmon resonance and includes peaks and valleys having a height difference in a vertical direction on an upper surface of the substrate; a probe disposed on the graphene channel and capable of chemically bonding to a target substance to be detected; and a linker molecule that connects the probe and the graphene channel, wherein the peaks and the valleys are alternately arranged in a zigzag shape along the first horizontal direction or a diagonal direction between the first horizontal direction and the second horizontal direction.

[0018] According to one embodiment, a surface plasmon resonance biosensor chip is provided, characterized in that the substrate comprises a transparent thermoplastic polymer or silicon dioxide (SiO2), titanium oxide (TiO2), tantalum oxide (Ta2O5), or aluminum oxide (Al2O5).

[0019] According to one embodiment, a surface plasmon resonance biosensor chip is provided, characterized in that the peak portions are arranged at regular intervals within a range of 1 nanometer to 10 nanometers along the first horizontal direction.

[0020] According to one embodiment, a surface plasmon resonance biosensor chip is provided, characterized in that the height difference between the peak portion and the valley portion is in the range of 0.2 nanometers to 20 nanometers.

[0021] According to one embodiment, a surface plasmon resonance biosensor chip is provided, characterized in that the peak portion and the valley portion are alternately arranged in a zigzag shape along the first horizontal direction, and the peak portion and the valley portion extend in a line shape.

[0022] In one embodiment, when the probe is DNA having a nucleotide sequence complementary to the nucleotide sequence of the target material, the linker molecule provides a surface plasmon resonance biosensor chip including a PASE (Pyrenebutanoic acid succinimidyl ester) molecule.

[0023] In order to solve the above-described problem, the technical idea of ​​the present invention provides a method for manufacturing a surface plasmon resonance biosensor chip, comprising the steps of: preparing a substrate including a transparent thermoplastic polymer; forming a graphene channel including peaks and valleys having a height difference on the substrate; and connecting a linker molecule to the graphene channel and connecting a probe capable of chemically bonding to a target substance to be detected to the linker molecule, wherein the step of forming the graphene channel comprises: forming a preliminary graphene channel on the substrate; and heating the substrate to cause the preliminary graphene channel to undergo shrinkage deformation.

[0024] According to one embodiment, a method for manufacturing a surface plasmon resonance biosensor chip is provided, characterized in that, in the step of forming the graphene channel, the length in the direction in which the graphene channel extends is formed to be 45% of the length in the direction in which the preliminary graphene channel extends.

[0025] According to one embodiment, a method for manufacturing a surface plasmon resonance biosensor chip is provided, characterized in that, in the step of forming the graphene channel, the preliminary graphene channel is heated within 110°C to 120°C for 10 to 120 minutes, so that the length in the direction in which the graphene channel extends is formed to be 30% to 90% of the length in the direction in which the preliminary graphene channel extends.

[0026] In order to solve the above-described problem, the technical idea of ​​the present invention provides a method for manufacturing a surface plasmon resonance biosensor chip, comprising the steps of: preparing a substrate including silicon dioxide (SiO2), titanium oxide (TiO2), tantalum oxide (Ta2O5), or aluminum oxide (Al2O5); etching the substrate to form a preliminary peak portion and a preliminary valley portion having a height difference; forming a graphene channel on the substrate including a peak portion and a valley portion corresponding to each of the preliminary peak portion and the preliminary valley portion; and connecting a linker molecule to the graphene channel and connecting a probe capable of chemically bonding to a target substance to be detected to the linker molecule.

[0027] According to one embodiment, a method for manufacturing a surface plasmon resonance biosensor chip is provided, characterized in that, in the step of forming the graphene channel, the graphene channel is formed by depositing a material including carbon (C) on the substrate using a chemical vapor deposition process.

[0028] In order to solve the above-described problem, the technical idea of ​​the present invention is a step of preparing a surface plasmon resonance biosensor, which comprises: a substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction on one surface of a prism; a graphene channel which generates surface plasmon resonance on one surface of the prism and includes peaks and valleys having a height difference in a direction perpendicular to an upper surface of the substrate; a probe which is disposed on the graphene channel and is capable of chemically bonding with a target substance to be detected; a linker molecule which connects the probe and the graphene channel; a light source which irradiates incident light into the graphene channel; a detector which detects reflected light reflected from the graphene channel; and a step of preparing a surface plasmon resonance biosensor in which the peaks and the valleys are alternately arranged in a zigzag shape along the first horizontal direction or a diagonal direction between the first horizontal direction and the second horizontal direction; a step of the light source irradiating the incident light into the graphene channel; And a step of monitoring the reflected light reflected from the graphene channel is included, and a detection method of a surface plasmon resonance biosensor is provided, characterized in that, in the resonance angle and resonance wavelength of the incident light that causes surface plasmon resonance in the graphene channel, a change in the resonance angle and the resonance wavelength corresponds to the presence of the target material.

[0029] The present invention provides a detection method of a surface plasmon resonance biosensor, comprising: a substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction; a graphene channel that generates surface plasmon resonance and includes peaks and valleys having a height difference in a direction perpendicular to an upper surface of the substrate; a probe disposed on the graphene channel and capable of chemically bonding to a target substance to be detected; a linker molecule that connects the probe and the graphene channel; a light source that irradiates incident light into the graphene channel; a detector that detects a Raman signal emitted from the graphene channel; and a step of preparing a surface plasmon resonance biosensor in which the peaks and the valleys are alternately arranged in a zigzag shape along the first horizontal direction or a diagonal direction between the first horizontal direction and the second horizontal direction; a step of irradiating the incident light from the light source into the graphene channel; and a step of monitoring the Raman signal reflected from the graphene channel.

[0030] According to exemplary embodiments of the present invention, a surface plasmon resonance biosensor chip including a wrinkled graphene channel according to the present invention can provide a biosensor with improved stability, since the binding force between the wrinkled graphene channel and the linker molecule is greater than the binding force between the flat graphene channel and the linker molecule.

[0031] Furthermore, since the surface area of ​​the wrinkled graphene channel is larger than that of the flat graphene channel, the wrinkled graphene channel can bind to more probes, thereby providing a biosensor with improved sensitivity to target substances.

[0032] In addition, compared to a surface plasmon resonance biosensor chip including a conventional metal thin film, the surface plasmon resonance biosensor chip including a wrinkled graphene channel according to the present invention quickly modifies the surface of graphene to form wrinkled graphene without additional mask production and etching processes required to form a metal thin film of a certain pattern, ultimately enabling the rapid production of a surface plasmon resonance biosensor chip.

[0033] FIG. 1 is a schematic diagram illustrating the structure of a surface plasmon resonance biosensor including a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0034] FIG. 2 is a drawing illustrating a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0035] Figure 3 is an enlarged view of the R1 region of the wrinkled graphene channel of Figure 2.

[0036] FIG. 4 is a drawing for explaining a method for manufacturing a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0037] FIGS. 5A to 5C are graphs illustrating the number of linker molecules bound to a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0038] FIG. 6 is a drawing for explaining a method for manufacturing a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to another embodiment of the present invention.

[0039] Hereinafter, embodiments of the technical concept of the present invention will be described in detail with reference to the attached drawings. Identical components in the drawings are designated by the same reference numerals, and redundant descriptions thereof will be omitted.

[0040] The surface plasmon resonance biosensor according to embodiments of the present invention is a sensor that detects a substance using a change in resonance angle. As an example, the surface plasmon resonance biosensor according to embodiments of the present invention can be used for analyzing biomolecules contained in a biological sample, thereby performing gene expression profiling, genotyping, detecting mutations and polymorphisms such as SNPs (Single nucleotide polymorphisms), analyzing proteins and peptides, screening for potential drugs, developing and manufacturing new drugs, etc.

[0041] Surface plasmon resonance biosensors employ appropriate probes depending on the target of the biosample to be analyzed. Examples of probes that can be employed in surface plasmon resonance biosensors include DNA probes, enzymes or antibodies / antigens, protein probes such as bacteriorhodopsin, microbial probes, and neuronal probes. Biosensors manufactured in the form of chips are also referred to as biochips. For example, depending on the type of probe employed, they may be referred to as DNA chips, protein chips, cell chips, neuron chips, etc. Surface plasmon resonance biosensors according to some embodiments of the present invention may include oligomeric probes as probes. The oligomeric probes imply that the number of monomers of the employed probe is at the oligomeric level. Here, the term "oligomer" may be used to mean a polymer composed of two or more covalently bonded monomers having a molecular weight of about 1000 or less. Specifically, it may contain about 2 to 500 monomers, preferably 5 to 30 monomers. However, the meaning of the oligomer probe is not limited to the above number.

[0042] The monomers that make up the oligomeric probe can be modified depending on the type of biological sample to be analyzed, and can be, for example, nucleosides, nucleotides, amino acids, peptides, etc. Nucleosides and nucleotides can include not only the known purine and pyrimidine bases, but also methylated purines or pyrimidines, acylated purines or pyrimidines, etc. Furthermore, nucleosides and nucleotides can include not only the conventional ribose and deoxyribose sugars, but also modified sugars in which one or more hydroxyl groups are substituted with halogen atoms or aliphatics, or functional groups such as ethers and amines are bonded. Amino acids can be L-, D-, and achiral forms of amino acids found in nature, as well as modified amino acids, or amino acid analogs. A peptide refers to a compound formed by an amide bond between the carboxyl group of an amino acid and the amino group of another amino acid.

[0043] Unless otherwise specifically stated, the probes exemplified in the following examples are DNA probes, which are oligomeric probes covalently bonded with monomers of about 5 to 30 nucleotides. However, the present invention is not limited thereto, and it goes without saying that various probes described above can be applied.

[0044] FIG. 1 is a schematic diagram illustrating the structure of a surface plasmon resonance biosensor including a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0045] Referring to FIG. 1, a surface plasmon resonance biosensor (100) may include a prism (110), a surface plasmon resonance biosensor chip (120) disposed on the prism (110), a light source (130) for irradiating light onto one surface of the surface plasmon resonance biosensor chip (120) through the prism (110), and a detector (140) for detecting light reflected from the surface plasmon resonance biosensor chip (120) or detecting a Raman signal. The surface plasmon resonance biosensor chip (120) is for detecting a target substance and may include a substrate (122), a graphene channel (124) disposed on the substrate (122), a probe (126) on the graphene channel (124), and a linker molecule (128).

[0046] The prism (110) serves to increase the wave vector of incident light for excitation of surface plasmons, and causes light provided from a light source (130) to be incident on the graphene channel (124) to excite surface plasmons of the graphene channel (124), and emits light reflected from the graphene channel (124). In the present specification, the light provided from the light source (130) may be referred to as incident light, and the light reflected from the graphene channel (124) may be referred to as reflected light. The prism (110) may be made of various optical glasses or optical plastics. In addition, the cross-sectional shape of the prism (110) may be changed to a hemisphere, a parallelogram, an inverted trapezoid, a semi-cylinder, etc. other than the illustrated triangle. Alternatively, the prism (110) may be omitted from the surface plasmon resonance biosensor (100).

[0047] As described above, the surface plasmon resonance biosensor chip (120) may include a substrate (122), a graphene channel (124) disposed on the substrate (122), a probe (126) on the graphene channel (124), and a linker molecule (128).

[0048] As an example, the substrate (122) may include a material that can cause shrinkage deformation such that the graphene channels (124) disposed on the substrate (122) are wrinkled. The substrate (122) may include a material that causes shrinkage deformation, particularly at high temperatures. For example, the substrate (122) may include a thermoplastic polymer that causes shrinkage deformation at about 100° C. to 200° C. However, for use in a surface plasmon resonance biosensor, it is preferable that the substrate (122) include one or a combination of colorless and transparent polystyrene, polyetherimide, polyethersulfone, polyetheretherketone, polytetrafluoroethylene, and liquid crystal polymer so that incident light can easily reach the graphene channels (124). For example, when a substrate (122) including polystyrene is annealed at about 110°C to 115°C for about 10 to 120 minutes, wrinkles may be formed in the range of several micrometers to several hundred nanometers on the substrate (122). The degree of shrinkage deformation of the substrate (122) may vary depending on the annealing temperature. For example, the degree of shrinkage deformation of the substrate (122) may increase as the annealing temperature increases. In addition, the degree of shrinkage deformation of the substrate (122) may vary depending on the annealing time. For example, the degree of shrinkage deformation of the substrate (122) may increase as the annealing time increases.

[0049] As another example, the substrate (122) may include an insulator that does not affect the electronic structure of the graphene channel (124), such as silicon dioxide or aluminum oxide, or a conductor that has little interaction with the graphene channel (124), such as platinum (Pt), copper (Cu), silver (Ag), or gold (Au). However, the present invention is not limited thereto, and the substrate (122) may include a conductor, a semiconductor, or an insulator. However, in terms of use of a surface plasmon resonance biosensor, it is preferable that the substrate (122) include a transparent material, such as silicon dioxide, titanium oxide, tantalum oxide, or aluminum oxide, so that incident light can easily reach the graphene channel (124).

[0050] A graphene channel (124) including graphene having a two-dimensional hexagonal carbon (C) structure may be interposed on the substrate (122). The graphene channel (124) may extend in a first horizontal direction (X) on the substrate (122). However, the graphene channel (124) does not have a flat structure extending along the upper surface of the substrate (122), but may include peaks and valleys having a height difference in a direction perpendicular to the upper surface of the substrate (e.g., the vertical direction (Z)). In the present specification, a graphene channel (124) having peaks and valleys may be referred to as a “wrinkled graphene channel.” Conversely, a graphene channel that does not include peaks and valleys may be referred to as a “flat graphene channel.” A specific description of the structure of the wrinkled graphene channel will be described later in the description of FIG. 2. The degree of wrinkles in the graphene channel (124) may vary depending on the temperature and time of the annealing process described above. Considering the yield and sensitivity of the graphene channel (124), the degree of wrinkles from a flat graphene channel to a wrinkled graphene channel is preferably about 55%.

[0051] In a surface plasmon resonance biosensor (100) according to one embodiment of the present invention, incident light incident on a prism (110) has an incident angle (θ) in ) can be completely absorbed into the graphene channel (124) at a specific angle exceeding the total reflection angle. That is, the incident angle (θ) of light incident on the prism (110) in ) can be excited at a specific angle exceeding the total reflection angle on the graphene channel (124), and surface plasmon resonance can occur. In the present specification, the "incident angle (θ) of light incident on the prism in ) A specific angle exceeding the total reflection angle is referred to as the resonance angle, and the wavelength of the surface plasmon wave at the resonance angle is referred to as the resonance wavelength.

[0052] The surface plasmon resonance biosensor (100) may further include a probe (126) capable of chemically binding to a target substance molecule. The probe (126) may be disposed on the graphene channel (124). For example, the probe (126) may be one of a polynucleotide, a peptide nucleic acid (PNA) probe, an aptamer (DNA having a specific binding ability to a specific substance), a protein, an antibody, or a capture agent. The probe (126) may be any one selected from the polynucleotide, peptide nucleic acid probe, aptamer, protein, antibody, and capture agent for chemical binding to the target substance. For example, when the target substance is DNA, the probe (126) may have a nucleotide sequence complementary to the nucleotide sequence of the DNA, which is the target substance, for chemical binding to the target substance.

[0053] The surface plasmon resonance biosensor (100) may further include a linker molecule (128) to facilitate chemical bonding between the probe (126) and the graphene channel (124). The linker molecule (128) may be disposed on the graphene channel (124). In one embodiment, the graphene channel (124) may be bonded to the linker molecule (128), and the probe (126) may be bonded to the graphene channel (124) via the linker molecule (128). In one embodiment, when the target material is DNA, a PASE (Pyrenebutanoic acid succinimidyl ester) molecule of the following chemical formula 1 may be selected as the linker molecule (128).

[0054] [Chemical Formula 1]

[0055]

[0056] The pyrene group in the PASE molecule can bind to the graphene channel (124), and the succinimidyl ester group in the PASE molecule can bind to the probe (126). In this case, the probe to which the succinimidyl ester group binds may be DNA having a nucleotide sequence complementary to the nucleotide sequence of the target substance, DNA. The binding of the pyrene group in the PASE molecule to the graphene channel (124) may be through a π-π bond among van der Waals bonds.

[0057] The resonance angle and resonance wavelength of the surface plasmon resonance biosensor (100) are very sensitive to changes in the refractive index of a material close to the graphene channel (124). When a target material molecule binds to a probe (126) bonded on the surface plasmon resonance biosensor (100), the resonance angle and resonance wavelength of the surface plasmon resonance biosensor (100) change, so that by measuring the amount of change in the resonance angle and the amount of change in the resonance wavelength of the surface plasmon resonance biosensor (100), it is possible to determine whether a target material molecule is bound on the graphene channel (124). Accordingly, when a light source (130) irradiates incident light onto the graphene channel (124) and a detector (140) monitors the reflected light reflected from the graphene channel (124), the change in the resonance angle and the resonance wavelength of the incident light can correspond to the presence of a target material.

[0058] The surface plasmon resonance biosensor (100) according to the technical idea of ​​the present invention can determine the presence of a target substance by monitoring changes in the resonance angle and resonance wavelength as described above, but as another example, the target substance can also be determined using a Raman scattering signal. Since the intensity of the Raman scattering signal can be amplified by the peak and valley portions having a height difference of the graphene channel (124), the target substance can be detected using the surface-enhanced Raman spectroscopy signal emitted from the graphene channel (124).

[0059] A coherent laser or LED, etc. can be used as the light source (130). Since the surface plasmon resonance depends on the wavelength of the light source (130), a wide range of wavelengths can be used as needed. When a wrinkled graphene channel (124) with a constant period is used, a light source with a wavelength larger than the period of the graphene channel (124) that can excite the surface plasmon suitable for sensing can be used, or a white light source for utilizing multiple wavelengths can be used. In addition, the power of the incident light can be adjusted by adding an optical component such as a pinhole.

[0060] The detector (140) can detect surface plasmon resonance generated due to the wrinkled structure of the graphene channel (124). The detector (140) collects light reflected from the prism (110) and the surface plasmon resonance biosensor chip (120), and is a general term for a component that can be used as a photodetector. Typically, a photodiode can be used for the detector (140), and the power of the reflected light can be controlled by adding an optical component such as a pinhole.

[0061] A bonding layer (150) may be interposed between the prism (110) and the surface plasmon resonance biosensor chip (120) to couple them to each other. The bonding layer (150) may include a metal, for example, chromium (Cr) or titanium (Ti). The bonding layer (150) may be omitted, in which case the prism (110) and the surface plasmon resonance biosensor chip (120) may be directly bonded by various methods, such as an optical method or a deposition method.

[0062] FIG. 2 is a drawing illustrating a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0063] Referring to FIG. 2, the graphene channel (124) may include a peak portion (P) and a valley portion (V) having a height difference in the vertical direction (Z). The wrinkled shape of the graphene channel (124) may vary. As an example, referring to (a) of FIG. 2, the peak portions (P) and the valley portions (V) of the wrinkled graphene channel (124) may be alternately arranged in a zigzag shape along the first horizontal direction (X). As another example, referring to (b) of FIG. 2, the peak portions (P) and the valley portions (V) of the wrinkled graphene channel (124) may be alternately arranged in a zigzag shape in the diagonal direction between the first horizontal direction (X) and the second horizontal direction (Y). The wrinkled shape of the graphene channel (124) is not limited to that illustrated, and may be wrinkled in a shape other than that illustrated.

[0064] The interval between the peaks (P) and the valleys (V) of the graphene channel (124) may vary. In one embodiment, the peaks (P) may be arranged to be spaced apart at a constant first interval (W1) in a range of about 1 nanometer to 10 nanometers along the first horizontal direction (X). The valleys (V) may be arranged to be spaced apart at a constant second interval (W2) in a range of about 1 nanometer to 10 nanometers along the first horizontal direction (X). In another embodiment, the peaks (P) and the valleys (V) may be arranged to be spaced apart at a non-constant interval along the first horizontal direction (X). As the degree of wrinkles in the graphene channel (124) increases, the interval between the peaks (P) and the valleys (V) along the first horizontal direction (X) may decrease.

[0065] As one embodiment, the height difference in the vertical direction (Z) between adjacent peaks (P) and valleys (V) of the graphene channel (124) may be constant within a range of about 0.2 nanometers to 20 nanometers. As another embodiment, the height difference in the vertical direction (Z) between adjacent peaks (P) and valleys (V) of the graphene channel (124) may not be constant.

[0066] A valley portion (V) may be arranged between adjacent peak portions (P) of the graphene channel (124), and a peak portion (P) may be arranged between adjacent valley portions (V). The shapes of the peak portions (P) and the valley portions (V) of the graphene channel (124) are not limited to those illustrated, and may be formed in various shapes. In one embodiment, for the first valley portion (V1) and the second valley portion (V2), which are valley portions (V) adjacent to the peak portion (P) of the graphene channel (124), the slope of the first intermediate region (COL1) from the first valley portion (V1) to the peak portion (P) and the slope of the second intermediate region (COL2) from the second valley portion (V2) to the peak portion (P) may be different from each other. In another embodiment, for the first valley portion (V1) and the second valley portion (V2), which are valley portions (V) adjacent to the peak portion (P) of the graphene channel (124), the slope of the first intermediate region (COL1) from the first valley portion (V1) to the peak portion (P) and the slope of the second intermediate region (COL2) from the second valley portion (V2) to the peak portion (P) may be the same. In this case, the first intermediate region (COL1) from the first valley portion (V1) to the peak portion (P) and the second intermediate region (COL2) from the second valley portion (V2) to the peak portion (P) may be symmetrical with respect to the peak portion (P). Depending on the shape of the graphene channel (124), the wavelength range of the surface plasmon absorption band may vary, and the measurement sensitivity may also vary. For example, if a graphene channel (124) with a long wavelength range of the surface plasmon band absorption band is provided, the measurement sensitivity of the surface plasmon resonance biosensor (100) can be increased.

[0067] The graphene channel (124) may have a thickness of about 1 nanometer to 2 nanometers and a surface area of ​​about 1 square millimeter to 100 square millimeters.

[0068] In addition, the number of graphene layers constituting the graphene channel (124) is not limited to that illustrated. The graphene channel (124) may be composed of a plurality of graphene layers. The graphene channel (124) may be composed of first graphene and second graphene disposed on the first graphene. The first graphene and the second graphene may have the same or different shapes. For example, the first graphene may have a smaller separation distance between adjacent peaks (P) than the second graphene. In addition, a plurality of carbon (C) atoms included in the plurality of graphene layers constituting the graphene channel (124) may be arranged in a row in the vertical direction (Z) or may be arranged in an offset manner. In one embodiment, the carbon (C) atoms constituting the first graphene and the carbon (C) atoms constituting the second graphene may be arranged in a row in the vertical direction (Z). As another example, the carbon (C) atoms constituting the first graphene and the carbon (C) atoms constituting the second graphene may be arranged to be misaligned in the vertical direction (Z).

[0069] Typically, surface plasmon resonance biosensors include metal films and probes combined with the metal films. Graphene is known to possess electrically semi-metallic properties, yet possesses extremely high electrical conductivity (intrinsic electron mobility of 20,000 cm2 / Vs) because its internal charge acts as a zero-effective mass particle. Therefore, introducing graphene channels instead of metal films into surface plasmon resonance biosensors can be advantageous in terms of cost, with no loss in electrical conductivity compared to surface plasmon resonance biosensors utilizing metal films.

[0070] In addition, the surface plasmon resonance biosensor (100) according to the present invention may include a wrinkled graphene channel (124). At this time, the equilibrium state energy (unit: eV) of the wrinkled graphene channel (124) is much lower than the equilibrium state energy of the flat graphene channel, so that the stability of the carbon (C) atoms constituting the graphene can be further increased. Table 1 below compares the equilibrium state energies between the flat graphene channel and the wrinkled graphene channel. Therefore, when the wrinkled graphene channel (124) according to the present invention is introduced into the surface plasmon resonance biosensor (100), the binding force between the graphene channel (124) and the linker molecule increases, so that a surface plasmon resonance biosensor (100) with improved stability can be provided.

[0071] [Table 1]

[0072]

[0073] Figure 3 is an enlarged view of the R1 region of the wrinkled graphene channel of Figure 2.

[0074] Referring to FIG. 3, an intermediate region (COL) between a peak portion (P) and a valley portion (V) of a graphene channel (124) is illustrated. A distance (d1) in the vertical direction (Z) from a line (A-A') crossing the intermediate region (COL) of the graphene channel (124) to the peak portion (P) may be about 0.54 nanometers. A distance (d2) in the vertical direction (Z) from a line (A-A') crossing the intermediate region (COL) of the graphene channel (124) to the valley portion (V) may be about 0.54 nanometers.

[0075]

[0076] FIG. 4 is a diagram illustrating a method for manufacturing a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to one embodiment of the present invention. In particular, the method for manufacturing a wrinkled graphene channel described in FIG. 4 may be for manufacturing a wrinkled graphene channel using a substrate (122) comprising a thermoplastic polymer. The present invention is not limited to what is illustrated, and wrinkled graphene channels can be formed through various processes.

[0077] Referring to FIG. 4, a preliminary graphene channel (124') can be formed on a preliminary substrate (122'). The preliminary graphene channel (124') can be formed by depositing a carbon compound including carbon (C) on the preliminary substrate (122') using a chemical vapor deposition (CVD) process. For example, the carbon compound may be natural graphite, synthetic graphite, highly ordered pyrolytic graphite, activated graphite, carbon monoxide, carbon dioxide, methane, ethane, ethylene, methanol, ethanol, acetylene, propane, propylene, butane, butadiene, pentane, pentene, cyclopentadiene, hexane, cyclohexane, benzene, pyridine, hexaphenylborazine, poly(4-vinylpyridine) (P4VP), 1,3,5-triazine, piperidine, toluene, polymethyl methacrylate, polystyrene, acrylonitrile, polyacrylonitrile, Melamine, polycyclic aromatic hydrocarbon structure methylnaphthalene, hexabromobenzene, naphthalene, pentachloropyridine, terphenyl, tetrabromothiophene, benzopyrene, azulene, trimethylnaphthalene, acenaphthene, acenaphthylene, anthracene, fluorene, phenalene, phenanthrene, benz(a)anthracene,Benzofluorene, Benzo(c)phenanthrene, Chrysene, Fluoranthene, Pyrene, Tetracene, Triphenylene, Benz(e)acephenanthrylene, Benzofluoranthene, Dibenzanthracene, Olympicene, Pentacene, Perylene, Picene, Tetraphenylene, Zethrene, Ovalene, Kekulene, Hexacene, Heptacene, Diindenoperylene, Dicoronylene, It may include Coronene, Corannulene, Benzo(ghi)perylene, Anthanthrene, Hexamethyldihydro-4H-benzoquinolizinoacridine, 4Hbenzoquinolizinoacridinetrione or Hexaazatriphenylenehexacarbonitrile.

[0078] The preliminary substrate (122') may include a metal such as copper (Cu), nickel (Ni), or platinum (Pt). When the preliminary substrate (122') includes copper (Cu), it may be preferable for forming a single layer of graphene. A raw material gas (e.g., methane (CH4)) containing carbon (C) may be injected onto the preliminary substrate (122') to form a preliminary graphene channel (124').

[0079] Next, a protective film (134) may be formed on the preliminary graphene channel (124'). As an example, the protective film (134) may include a polymer such as an acrylic resin (PMMA; Polymethyl methacrylate). As an example, the protective film (134) may be formed by a spin coating method. Specifically, a small amount of liquid for forming the protective film (134) may be dropped on the preliminary graphene channel (124'), and then the preliminary substrate (122') may be rotated. The rotation of the preliminary substrate (122') may cause the small amount of liquid to spread to the edge of the preliminary graphene channel (124'), thereby forming a protective film (134) that coats the preliminary graphene channel (124').

[0080] Next, the preliminary substrate (122') under the preliminary graphene channel (124') can be removed. In one embodiment, the preliminary substrate (122') can be removed using a chemical etching method. The etchant of the chemical etching method can include a substance that can dissolve metal. In one embodiment, the etchant can include sodium persulfate (Na2S2O8). Thereafter, the substrate (122) can be formed under the preliminary graphene channel (124') using a direct transfer method. The substrate (122) and the preliminary graphene channel (124') on which the protective film (134) is formed can be combined by performing a plasma treatment on the upper portion of the substrate (122). In one embodiment, the plasma treatment can include oxygen (O) as a reaction gas. After the preliminary graphene channel (124') is bonded to the substrate (122), the protective film (134) on the preliminary graphene channel (124') can be removed. In one embodiment, when the protective film (134) includes acrylic resin (PMMA), acetic acid (CH3COO H ) and DI water can be used to remove the protective film (134).

[0081] Next, the substrate (122) to which the preliminary graphene channel (124') from which the protective film (134) is removed is bonded can be annealed. For example, the annealing process can be performed at about 110°C to 120°C and can be performed for about 10 minutes to 120 minutes. At this time, the length in the direction in which the graphene channel (124) extends can be formed to be 30% to 90% of the length in the direction in which the preliminary graphene channel (124') extends. Through shrinkage deformation of the substrate (122), the preliminary graphene channel (124') can be formed into a wrinkled graphene channel (124).

[0082] The degree of wrinkles in the graphene channel (124) may vary depending on the time or temperature of the annealing process. In one embodiment, when the length of the preliminary graphene channel (124') in the first horizontal direction (X) is defined as the first length and the length of the graphene channel (124) in the first horizontal direction (X) is defined as the second length, when the substrate (122) to which the preliminary graphene channel (124') is bonded is annealed at about 120° C. for about 60 minutes, the second length may be 1 / 4 of the first length. Since the greater the degree of wrinkles in the wrinkled graphene channel (124), the longer the time consumed in the process for bonding the graphene channel (124) and the linker molecule, it is preferable that the degree of wrinkles in the wrinkled graphene channel (124) from the preliminary graphene channel (124') be about 55% when considering yield and sensitivity. That is, it is preferable that the length in the direction in which the graphene channel (124) extends is about 45% of the length in the direction in which the preliminary graphene channel (124') extends.

[0083] Hereinafter, a method for binding a PASE molecule as a linker molecule to a graphene channel will be described in detail with reference to an experimental example of the present invention. This experimental example is only intended to more clearly understand the present invention and is not intended to limit the scope of the present invention. Referring to an experimental example of the present invention, a PASE solution was prepared from DMSO (Dimethyl sulfoxide). The PASE solution was prepared at a concentration of 30 μM. The prepared PASE solution was dropped onto the surface of the graphene channel so that the graphene channel and the prepared PASE solution were in contact with each other. The graphene channel treated with the PASE solution was washed three times with a DMSO solution and three times with ethanol. The washed graphene channel was dried for approximately 1 hour or more. The dried graphene channel was washed with DI water. As a result, a graphene channel to which a PASE molecule is bound can be obtained.

[0084]

[0085] Hereinafter, a method for binding DNA as a probe to a graphene channel is specifically described with reference to an experimental example of the present invention. In the present specification, DNA bound as a probe is referred to as probe DNA. Referring to an experimental example of the present invention, a graphene channel that had been treated with a PASE solution was incubated in 1X PBS (1X phosphate-buffered saline) for more than one day. After the incubation, a diluted solution containing the probe DNA was dropped onto the graphene channel, and after about 3 hours, it was washed using a 1X PBS solution. The probe DNA was diluted to a concentration of about 264 μM. The washed graphene channel was dried at about 25°C for about 1 hour or more. As a result, a graphene channel to which the probe DNA was bound via a PASE molecule can be obtained.

[0086]

[0087] Next, we describe two experimental examples of a method for detecting a target substance through a graphene channel bound to a probe DNA via a PASE molecule.

[0088] [Experimental Example 1]

[0089] After the PASE solution treatment was completed, the graphene channel was incubated in 1XPBS, washed with 1XPBS, and then incubated with 50 μL of 100 μM pDNA-NH2 for about 3 hours or more. After the incubation with 100 μM pDNA-NH2 was completed, it was washed with 1XPBS three or more times. A 50 μL tDNA solution containing x μM (where x is a number greater than or equal to 0) tDNA was brought into contact with the second end of the graphene channel, and then left to stand for about 20 minutes. After the waiting period, the electrode was washed with 1XPBS, and then brought into contact with a sample containing a mixture of 5 mM methylene blue, 5 mM [Fe(CN)6]-4, and the target substance. Afterwards, the CV (Cyclic voltammetry) graph was observed at a scan frequency of 50 mV, and the current was observed while applying the gate voltage from approximately 0 V to -0.6 V. The above-described experimental process was repeated while gradually increasing the concentration of tDNA in the tDNA solution.

[0090]

[0091] [Experimental Example 2]

[0092] After PASE solution treatment, the graphene channel was incubated in 1XPBS, washed with 1XPBS, and then 50 μL of 100 μM MB-pDNA-NH2 was applied onto the graphene channel and incubated for more than 3 hours. After incubation with 50 μL of 100 μM MB-pDNA-NH2, the channel was washed with 1XPBS three or more times. The subsequent process is similar to the first experimental example.

[0093] However, there is a difference in that the first experimental example incubates the graphene channel with pDNA-NH2 after incubation in 1XPBS, and the second experimental example incubates with MB-pDNA-NH2 containing methylene blue.

[0094]

[0095] FIGS. 5A to 5C are graphs illustrating the number of linker molecules bound to a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to one embodiment of the present invention.

[0096] FIGS. 5A to 5C are CV graphs that observe the current density using a wrinkled graphene channel or a flat graphene channel created through the experimental example of the present invention described above. FIG. 5C is a graph that records the anodic current peak (Ipa) of FIGS. 5A and 5B. Line (a) of FIG. 5A shows the change in current density when voltage is applied to a flat graphene channel, and line (b) shows the change in current density when voltage is applied to a flat graphene channel that has undergone PASE processing. Line (a) of FIG. 5B shows the change in current density when voltage is applied to a wrinkled graphene channel, and line (b) shows the change in current density when voltage is applied to a wrinkled graphene channel that has undergone PASE processing.

[0097] Referring to Fig. 5c, the cathode current peak is -0.907 Acm when voltage is applied to the wrinkled graphene channel. -2 When a voltage of -8.185 Acm- was applied to the wrinkled graphene channel after PASE processing was completed, 2 , while when voltage was applied to the flat graphene channel, it was -2.671 Acm -2 When a voltage was applied to the flat graphene channel after PASE processing, -5.503 Acm -2The increase in the cathode current peak can be attributed to the PASE electron-mediated transport reaction, which confirms that the PASE electron-mediated transport reaction occurs more actively on the wrinkled graphene channel.

[0098] The reason why the change in the cathode current peak from the wrinkled graphene channel to the PASE-treated wrinkled graphene channel is greater than the change in the cathode current peak from the flat graphene channel to the PASE-treated wrinkled graphene channel is that the wrinkled graphene channel has a larger surface area than the flat graphene channel, allowing more PASE molecules to be adsorbed onto the graphene surface. Since more PASE molecules are adsorbed onto the surface of the wrinkled graphene channel, more electron transfer is induced, allowing the change in the cathode current peak to be greater than that of the flat graphene channel. Therefore, the surface plasmon resonance biosensor including the wrinkled graphene channel according to the present invention can provide a surface plasmon resonance biosensor with enhanced sensitivity to a target substance, since the wrinkled graphene channel binds to more linker molecules than the flat graphene channel.

[0099]

[0100] FIG. 6 is a drawing for explaining a method for manufacturing a wrinkled graphene channel included in a surface plasmon resonance biosensor chip according to another embodiment of the present invention. The present invention is not limited to what is illustrated, and the wrinkled graphene channel can be formed through various processes. In particular, the method for manufacturing the wrinkled graphene channel described in FIG. 6 may be a method for forming a wrinkled graphene channel on a substrate including a transparent material such as silicon dioxide (SiO2), titanium oxide (TiO2), tantalum oxide (Ta2O5), or aluminum oxide (Al2O5).

[0101] Referring to FIG. 6, the substrate (122) may include a preliminary peak portion (PP) and a preliminary valley portion (PV) having a height difference on one surface. The preliminary peak portion (PP) and the preliminary valley portion (PV) may include those formed through an etching process on one surface of the substrate (122). A carbon compound including carbon (C) may be deposited on the substrate (122) including the preliminary peak portion (PP) and the preliminary valley portion (PV) through a chemical vapor deposition (CVD) process to form a graphene channel (124). At this time, the carbon compound may include materials described as an example in FIG. 4.

[0102] The graphene channel (124) is formed on a substrate (122) including a preliminary peak portion (PP) and a preliminary valley portion (PV), and thus can be formed to include a peak portion (P) and a valley portion (V) having a height difference. The peak portion (P) and the valley portion (V) of the graphene channel (124) can be formed to correspond to the preliminary peak portion (PP) and the preliminary valley portion (PV), respectively. For example, the peak portion (P) of the graphene channel (124) can be formed to overlap with the preliminary peak portion (PP) in the vertical direction (Z). The valley portion (V) of the graphene channel (124) can be formed to overlap with the preliminary valley portion (PV) in the vertical direction (Z). As a result, a graphene channel (124) having a wrinkled shape can be obtained.

[0103] Typically, an optical lithography process can be used to form metal nanoparticles on a metal thin film included in a surface plasmon resonance sensor. To form metal nanoparticles using the optical lithography process, a mask with a pattern of nanoparticles drawn on the metal thin film must be created, and a process of etching the portion of the metal thin film other than the pattern of the nanoparticles must be performed. Therefore, the process of creating a mask with a pattern of metal nanoparticles drawn on it and the process of precisely etching the portion of the metal thin film other than the pattern of the nanoparticles are required separately, so there was a problem that the manufacturing process was complicated and took a long time. Furthermore, even when graphene was used instead of a metal thin film, separate processes were required to form metal nanoparticles, so there was still a problem that the manufacturing process of the surface plasmon resonance sensor was complicated and took a long time.

[0104] In the case of the wrinkled graphene channel (124) included in the surface plasmon resonance biosensor (100) according to one embodiment of the present invention, a preliminary graphene channel (124') having a flat shape may be formed on a substrate (122), and then the substrate (122) may be heated to cause the preliminary graphene channel (124') to contract and deform, thereby forming the wrinkled graphene channel (124). Alternatively, the substrate (122) may be etched to form a preliminary peak portion (PP) and a preliminary valley portion (PV) having a height difference on the substrate (122), and a material constituting the graphene channel (124) may be deposited on the substrate (122) by a chemical vapor deposition method, thereby forming a wrinkled graphene channel (124) having a peak portion (P) and a valley portion (V) corresponding to the preliminary peak portion (PP) and the preliminary valley portion (PV), respectively. Therefore, the wrinkled graphene channel (124) included in the surface plasmon resonance biosensor (100) according to one embodiment of the present invention does not require a separate mask manufacturing process and a precise etching process, thereby simplifying the manufacturing process of the surface plasmon resonance biosensor (100) and shortening the manufacturing time.

[0105] As described above, exemplary embodiments have been disclosed in the drawings and specifications. While specific terminology has been used to describe embodiments herein, it is intended solely to illustrate the technical concept of the present disclosure and is not intended to limit the scope of the present disclosure as defined in the claims. Therefore, those skilled in the art will appreciate that various modifications and equivalent embodiments are possible. Therefore, the true technical protection scope of the present disclosure should be determined by the technical concept of the appended claims.

Claims

1. A substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction; A graphene channel that generates surface plasmon resonance and includes peaks and valleys having a height difference in a direction perpendicular to the upper surface of the substrate; A probe disposed on the graphene channel and capable of chemically bonding with a target material to be detected; and comprising a linker molecule that connects the probe and the graphene channel; A surface plasmon resonance biosensor chip in which the peak portion and the valley portion are alternately arranged in a zigzag shape along the first horizontal direction or the diagonal direction between the first horizontal direction and the second horizontal direction.

2. In paragraph 1, A surface plasmon resonance biosensor chip characterized in that the substrate comprises a transparent thermoplastic polymer or silicon dioxide (SiO2), titanium oxide (TiO2), tantalum oxide (Ta2O5), or aluminum oxide (Al2O5).

3. In paragraph 1, A surface plasmon resonance biosensor chip, characterized in that the peak portions are arranged at regular intervals within a range of 1 nanometer to 10 nanometers along the first horizontal direction.

4. In paragraph 1, A surface plasmon resonance biosensor chip, characterized in that the height difference between the peak and the valley is in the range of 0.2 nanometers to 20 nanometers.

5. In paragraph 1, A surface plasmon resonance biosensor chip characterized in that the peak portion and the valley portion are alternately arranged in a zigzag shape along the first horizontal direction, and the peak portion and the valley portion extend in a line shape.

6. In paragraph 5, A surface plasmon resonance biosensor chip in which the linker molecule comprises a PASE (Pyrenebutanoic acid succinimidyl ester) molecule, when the probe is DNA having a nucleotide sequence complementary to the nucleotide sequence of the target material.

7. A step of preparing a substrate containing a transparent thermoplastic polymer; A step of forming a graphene channel including a peak portion and a valley portion having a height difference on the substrate; and A step of connecting a linker molecule to the graphene channel and connecting a probe capable of chemically bonding to a target substance to be detected to the linker molecule, The step of forming the above graphene channel is: a step of forming a preliminary graphene channel on the substrate; and A method for manufacturing a surface plasmon resonance biosensor chip, characterized in that it comprises a step of heating the substrate so that the preliminary graphene channel undergoes shrinkage deformation.

8. In paragraph 7, In the step of forming the above graphene channel, A method for manufacturing a surface plasmon resonance biosensor chip, characterized in that the length in the direction in which the graphene channel extends is formed to be 45% of the length in the direction in which the preliminary graphene channel extends.

9. In paragraph 7, In the step of forming the graphene channel, the preliminary graphene channel is heated at 110°C to 120°C for 10 to 120 minutes, A method for manufacturing a surface plasmon resonance biosensor chip, characterized in that the length in the direction in which the graphene channel extends is formed to be 30% to 90% of the length in the direction in which the preliminary graphene channel extends.

10. A step of preparing a substrate including silicon dioxide, titanium oxide, tantalum oxide or aluminum oxide; A step of etching the above substrate to form a preliminary peak portion and a preliminary valley portion having a height difference; A step of forming a graphene channel including a peak portion and a valley portion corresponding to each of the preliminary peak portion and the preliminary valley portion on the substrate; and A method for manufacturing a surface plasmon resonance biosensor chip, comprising the steps of connecting a linker molecule to the graphene channel and connecting a probe capable of chemically bonding to a target substance to be detected to the linker molecule.

11. In paragraph 10, A method for manufacturing a surface plasmon resonance biosensor chip, characterized in that, in the step of forming the graphene channel, the graphene channel is formed by depositing a material containing carbon (C) on the substrate using a chemical vapor deposition process.

12. A step for preparing a surface plasmon resonance biosensor, comprising: a substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction on one side of a prism; a graphene channel having a peak portion and a valley portion that generate surface plasmon resonance on one side of the prism and have a height difference in a direction perpendicular to the upper surface of the substrate; a probe disposed on the graphene channel and capable of chemically bonding with a target material to be detected; a linker molecule that connects the probe and the graphene channel; a light source that irradiates incident light into the graphene channel; a detector that detects reflected light reflected from the graphene channel; and a step for preparing a surface plasmon resonance biosensor in which the peak portion and the valley portion are alternately arranged in a zigzag shape along the first horizontal direction or a diagonal direction between the first horizontal direction and the second horizontal direction; a step of the light source irradiating the incident light into the graphene channel; and Comprising a step of monitoring the reflected light reflected from the graphene channel, A detection method of a surface plasmon resonance biosensor, characterized in that, in the resonance angle and resonance wavelength of the incident light that causes surface plasmon resonance in the graphene channel, a change in the resonance angle and the resonance wavelength corresponds to the presence of the target material.

13. A step for preparing a surface plasmon resonance biosensor, comprising: a substrate extending in a first horizontal direction and a second horizontal direction intersecting the first horizontal direction; a graphene channel including a peak portion and a valley portion having a height difference in a direction perpendicular to an upper surface of the substrate and generating surface plasmon resonance; a probe disposed on the graphene channel and capable of chemically bonding with a target material to be detected; a linker molecule connecting the probe and the graphene channel; a light source irradiating incident light into the graphene channel; a detector detecting a Raman signal emitted from the graphene channel; and a step for preparing a surface plasmon resonance biosensor in which the peak portion and the valley portion are alternately arranged in a zigzag shape along the first horizontal direction or a diagonal direction between the first horizontal direction and the second horizontal direction; a step of the light source irradiating the incident light into the graphene channel; and A detection method of a surface plasmon resonance biosensor, comprising a step of monitoring the Raman signal reflected from the graphene channel.

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