Biosensor and module for detecting target substance in body fluid

Abstract

The present invention relates to a biosensor and a biomodule, for detecting a target substance in a body fluid, which are capable of collecting a body fluid such as saliva, urine, or blood from an examinee to detect a target substance (for example, a hormone) in the body fluid, and performing diagnosis based on a level thereof. The biosensor for detecting a target substance in a body fluid, according to the present invention, is for detecting a target substance in a body fluid of an examinee, and comprises: a substrate; an electrode formed on the substrate; a carbon thin film formed on the electrode; and an immobilized material immobilized on the carbon thin film. The carbon thin film is composed of a hydrogen (H)-free amorphous structure, and has both a carbon-carbon double bond (C=C) and a carbon-carbon single bond (C-C). The immobilized material includes the same substance as the target substance. In the carbon thin film, on the basis of XPS peak intensities determined by XPS analysis, the XPS peak intensity related to the carbon-carbon double bond may be higher than the XPS peak intensity related to the carbon-carbon single bond.

Description

Biosensor and module for detecting target substances in body fluids

[0001] The present invention relates to a biosensor, and more specifically, to a biosensor and bio module for detecting target substances in body fluids, capable of collecting body fluids such as saliva, urine, and blood from a subject, detecting target substances (e.g., hormones) within the body fluids, and diagnosing their levels.

[0002] Biosensors are sensors that convert signals generated from biological substances or systems into electrical or optical signals for measurement. By utilizing the unique characteristics of biomaterials, they can detect specific hormones, diseases, or drugs, and are thus utilized in various fields such as medical diagnosis and new drug development.

[0003] Meanwhile, mental stress is significantly increasing in modern society due to complex and diverse social and environmental factors, and this is emerging as a serious social problem as it can lead to diseases directly related to life.

[0004] Depression is a representative example of a modern disease that can be caused by increased stress. Depression is known to be directly related to the levels of (anti)stress hormones, such as serotonin and cortisol. Therefore, there is an urgent need for the development of technology capable of rapidly and accurately measuring these (anti)stress hormones for the early diagnosis and prevention of depression.

[0005] Conventional biosensors or measurement methods capable of detecting such biological hormones are as follows.

[0006] Conventional methods for detecting and measuring biological hormones include High Performance Liquid Chromatography (HPLC), fluorometric assay, and reverse phase chromatography.

[0007] However, high-performance liquid chromatography (HPLC), fluorescence analysis, and reverse-phase chromatography all had the disadvantage of requiring a long analysis time and being expensive.

[0008] To overcome these drawbacks, a biosensor has been proposed that detects biological hormones such as serotonin and cortisol by utilizing changes in electrical signals resulting from antigen-antibody immune responses.

[0009] However, conventional biosensors utilizing electrical signals in this manner often exhibit high noise levels compared to the effective signal, making it difficult to accurately distinguish between signals from the actual target substance and signals from other substances (i.e., noise). This acted as a factor that significantly reduced the measurement accuracy, reproducibility, and sensitivity of the biosensor.

[0010] Furthermore, conventional biosensors utilizing electrical signals had limitations in simultaneously achieving high levels of measurement reproducibility and measurement resolution. In other words, biosensors with high measurement resolution exhibited very low measurement reproducibility, while biosensors with high measurement reproducibility had low measurement resolution.

[0011] Specifically, when looking at conventional biosensors that use electrical signals, for biosensors with high measurement resolution of 1 pg / ml to 1 ng / ml, the measurement reproducibility is evaluated to be around 30 to 40%, and is reported to be practically at a low level of 20% or less.

[0012] Meanwhile, biosensors utilizing electrical signals must be equipped with electrodes, and these electrodes must be formed from a material that possesses high electrical conductivity while simultaneously exhibiting excellent biocompatibility, corrosion resistance, and biostability.

[0013] If the electrode is formed from a material with low biocompatibility or stability, the target substance may react directly with the electrode, acting as a noise component that ultimately leads to a significant decrease in the measurement accuracy, reproducibility, and resolution of the biosensor.

[0014] For this reason, conventional biosensors typically use gold (Au) to form electrodes. Gold (Au) is known as a representative material that possesses all the properties required for biosensor electrodes, including electrical conductivity, biocompatibility, corrosion resistance, and biostability.

[0015] However, since gold (Au) is very expensive, there was a disadvantage that the price of the biosensor would increase significantly if gold were used to form the electrode.

[0016] The present invention aims to solve the aforementioned problems, and the objective of the present invention is to provide a biosensor and module for detecting target substances in body fluids that can simultaneously improve measurement accuracy, measurement range, measurement resolution, and measurement reproducibility compared to conventional biosensors, in a biosensor utilizing electrical signals.

[0017] Another objective of the present invention is to provide a biosensor and module for detecting target substances in body fluids that can maximize the immobilization rate of target substances (i.e., substances to be detected) when target substances such as antigens and hormones, rather than target binding substances such as antibodies, receptors, markers, and probes, are immobilized on an electrode.

[0018] Another objective of the present invention is to provide a biosensor and module for detecting target substances in body fluids that can satisfy all the requirements for electrical signal sensing performance, biocompatibility, corrosion resistance, and biostability for biosensor electrodes, even without using gold (Au) when forming the electrodes of the biosensor.

[0019] Another objective of the present invention is to provide a biosensor and module for detecting target substances in body fluids that can prevent damage such as cracks, peeling, and pinholes from occurring in the carbon film during sterilization of the biosensor, even when a carbon-based thin film (i.e., a carbon film) is formed on an electrode.

[0020] Another objective of the present invention is to provide a biosensor and module for detecting target substances in body fluids that can prevent damage, particularly pinholes, from occurring during the process of removing immobilized material and conductive polymer on the electrode for the reuse of the electrode.

[0021] A biosensor for detecting a target substance in body fluid according to the present invention for achieving the above objective is a biosensor for detecting a target substance in the body fluid of a subject, comprising a substrate, an electrode formed on the substrate, a carbon thin film formed on the electrode, and a fixing material immobilized on the carbon thin film.

[0022] The above carbon thin film is composed of an amorphous structure that does not contain hydrogen (H) and has both carbon double bonds (C=C) and carbon single bonds (CC).

[0023] The above fixed material contains the same material as the above target material.

[0024] The carbon thin film may have a carbon double bond-related XPS peak intensity that is higher than the carbon single bond-related XPS peak intensity based on XPS analysis.

[0025] According to one example of the present invention, the fixing substance and the target substance may be hormones.

[0026] A bio module for detecting a target substance in body fluid according to the present invention comprises a biosensor for detecting a target substance in body fluid and a buffer solution into which the subject's body fluid is mixed.

[0027] The above buffer solution includes a target binding substance that specifically binds to the target substance, and when the target substance in the subject's body fluid is detected, the target binding substance and the subject's body fluid are mixed and administered onto the carbon thin film.

[0028] The above bio module is configured to detect the target substance by utilizing the result of a competitive reaction between the target binding substance in the buffer solution and the target substance in the body fluid when the buffer solution is administered onto the carbon thin film.

[0029] According to the biosensor and module for detecting target substances in body fluids according to the present invention, by applying a carbon thin film having specific physical properties, a structure opposite to that of conventional biosensors is formed, that is, a structure in which a target substance (i.e., a substance to be detected) such as an antigen or hormone is immobilized on an electrode, rather than a target binding substance such as an antibody, receptor, marker, or probe. By detecting the target substance using a method or principle entirely different from that of conventional biosensors based on this, it is possible to simultaneously improve measurement accuracy, measurement range, as well as measurement resolution and measurement reproducibility compared to conventional biosensors.

[0030] In addition, even if target substances such as antigens or hormones (i.e., substances to be detected) are immobilized on the electrode instead of target binding substances such as antibodies, receptors, markers, or probes, there is an effect of significantly increasing the immobilization rate of such target substances.

[0031] In addition, even if a thin film of carbon main components (i.e., a carbon thin film) is formed on the electrode, damage such as peeling, cracking, or pinholes can be prevented from occurring in the carbon thin film during the sterilization process of the biosensor, so there is an advantage of being able to apply various and stable sterilization treatments suitable for the target substance and diagnostic environment.

[0032] In addition, by applying a carbon thin film with specific physical properties, it is possible to satisfy all the requirements for electrical signal sensing performance, biocompatibility, corrosion resistance, and biostability for biosensor electrodes. This allows for the construction of electrodes without using gold (Au), thereby offering the advantage of significantly reducing the manufacturing cost of biosensors.

[0033] In addition, for the reuse of the electrode, it is possible to prevent damage, such as pinholes, from occurring during the process of removing the immobilized material and conductive polymer on the electrode, thereby having the advantage of efficiently recycling the electrode as is.

[0034] FIG. 1 is a cross-sectional view of a biosensor for detecting a target substance in body fluid according to the present invention.

[0035] FIG. 2 is a plan view showing a biosensor electrode according to one embodiment of the present invention.

[0036] FIG. 3 is a plan view showing a biosensor electrode having a carbon thin film formed thereon according to the present invention.

[0037] FIG. 4 is a schematic diagram of a buffer solution mixed with body fluid containing a target substance according to the present invention.

[0038] FIG. 5 is a field emission scanning electron microscope (FESEM) analysis image of a carbon thin film according to one embodiment of the present invention.

[0039] FIG. 6 is experimental data showing graphs of XPS peak intensities related to carbon single bonds (CC) and carbon double bonds (C=C) of a carbon thin film according to the present invention, measured according to plasma density when forming a carbon thin film by physical vapor deposition (PVD).

[0040] Figure 7 is experimental data showing a graph of plasma density measured according to process pressure during the physical vapor deposition (PVD) process.

[0041] FIG. 8 is a photograph of the first result of detecting a target substance using the competitive reaction result of the bio module according to the present invention.

[0042] FIG. 9 is a photograph of a second result in which a target substance was detected using the competitive reaction result of a bio module according to the present invention.

[0043] Figure 10 is a photograph of the carbon thin film state of Comparative Example 1 after sterilization treatment.

[0044] Figure 11 is a photograph showing the state of the carbon thin film of Example 1 after sterilization treatment.

[0045] The terms used in this specification are used merely to describe specific embodiments and are not intended to limit the invention. Singular expressions include plural expressions unless the context clearly indicates otherwise. In this specification, terms such as “comprising” or “having” are intended to indicate the presence of the features, numbers, steps, actions, components, parts, or combinations thereof described in the specification, and should be understood as not precluding the existence or addition of one or more other features, numbers, steps, actions, components, parts, or combinations thereof.

[0046] In addition, in this specification, "on or above" means being located above or below the target part, but this does not necessarily mean being located on the upper side with respect to the direction of gravity. That is, the term "on or above" as used in this specification includes not only cases where it is located above or below the target part, but also cases where it is located in front or behind the target part.

[0047] Furthermore, when it is stated that a part, such as a region or plate, is "on or above" another part, this includes not only cases where it is in contact with or spaced apart from "immediately on or above" another part, but also cases where there is another part in between.

[0048] In addition, when a component is described in this specification as being "connected" or "connected" to another component, it should be understood that the component may be directly connected to or directly connected to the other component, but unless otherwise specifically stated, it may also be connected or connected through another component in between.

[0049] Additionally, in this specification, terms such as "first," "second," etc., may be used to describe various components, but said components should not be limited by said terms. These terms are used solely for the purpose of distinguishing one component from another.

[0050] Hereinafter, various embodiments, advantages, and features of the present invention will be described in detail with reference to the attached drawings.

[0051] Prior to describing the present invention, terms related to the performance of the biosensor are defined as follows.

[0052] Measurement accuracy is an indicator of how close a measured value is to the actual value. In other words, the higher the measurement accuracy, the closer the measured value is to the actual value.

[0053] Measurement resolution refers to the ability of a biosensor to detect minute numerical values, signifying the smallest unit of change that the sensor can measure. In other words, the higher the measurement resolution, the finer the numerical values ​​can be detected.

[0054] The measurement range refers to the difference between the maximum and minimum values ​​that a biosensor can measure. For example, if the measurement range of a blood glucose sensor is 0 to 600 mg / dL, the sensor can only measure blood glucose when it is between 0 mg / dL and 600 mg / dL.

[0055] Measurement reproducibility is an indicator of how consistently measurement values ​​are obtained when the same subject is measured multiple times using the same biosensor under identical conditions. Therefore, the higher the measurement reproducibility, the more similar the measurements are to one another, and the higher the reliability of the measurement results.

[0056] FIG. 1 is a cross-sectional view of a biosensor for detecting a target substance in body fluid according to the present invention, FIG. 2 is a plan view showing a biosensor electrode according to one embodiment of the present invention, and FIG. 3 is a plan view showing a biosensor electrode having a carbon thin film formed thereon according to the present invention.

[0057] Referring to FIGS. 1 to 3, the biosensor according to the present invention is a biosensor for detecting a target substance in a body fluid, capable of collecting a subject's body fluid, detecting a target substance contained in the body fluid, and diagnosing the value thereof, comprising a substrate (10), an electrode (20) formed on the substrate (10), a carbon thin film (30) formed on the electrode (20), and a fixing material (40) immobilized on the carbon thin film (30). Here, the subject's 'body fluid' is a liquid fluid flowing inside the subject's body, and may be, for example, saliva, urine, or blood.

[0058] The substrate (10) of the present invention may be formed from an insulating material such as synthetic resin, natural resin, or glass as a substrate for forming an electrode (20).

[0059] The electrode (20) of the present invention is formed of an electrically conductive material such as a metal or a conductive polymer, as a means for detecting an electrical signal resulting from a specific binding reaction between a target binding material (50) contained in a buffer solution (60) and a fixing material (40) immobilized on a carbon thin film (30).

[0060] According to one example of the present invention, the electrode (20) may include at least one selected from the group consisting of Cu, Co, Bi, Be, Ag, Al, Au, Hf, Cr, In, Mn, Mo, Mg, Ni, Nb, Pb, Pd, Pt, Re, Rh, Sb, Ta, Te, Ti, W, V, Zr, Zn and combinations thereof, but is not limited thereto.

[0061] The electrode (20) can be formed by a known method. For example, the electrode (20) can be formed by a deposition method such as photolithography, thermal deposition, E-beam deposition, PECVD (Plasma Enhanced Chemical Vapor Deposition), LPCVD (Low Pressure Chemical Vapor Deposition), PVD (Physical Vapor Deposition), sputtering, and ALD (Atomic Layer Deposition).

[0062] According to another example of the present invention, the electrode (20) may include at least one selected from the group consisting of polypyrrole, polythiophene, polyaniline and polyacetylene, but is not limited thereto.

[0063] According to one example of the present invention, the electrode (20) may be composed of multiple electrodes formed at a distance from each other, and in the above case, the multiple electrodes may be three electrodes consisting of a counter electrode (21), a working electrode (23), and a reference electrode (25).

[0064] The working electrode (23) is an electrode where a reaction in which the target binding substance (50) and the immobilizing substance (40) are oxidized or reduced occurs. The electrical signal generated at the working electrode (23) is analyzed through the signal processing circuit of the biosensor, thereby enabling the determination of the concentration or characteristics of the target substance (i.e., the target substance).

[0065] The counter electrode (21) is an electrode where the opposite reaction to the oxidation or reduction reaction occurring at the working electrode (23) takes place. The counter electrode (21) can facilitate the electrochemical reaction by allowing current to flow together with the working electrode (23).

[0066] The reference electrode (25) is an electrode that serves as a reference for measuring potential in electrochemical analysis. The reference electrode (25) maintains a constant potential and can be used to measure the potential of the working electrode (23) and the counter electrode (21).

[0067] According to one example of the present invention, the total area of ​​the two-dimensional plane of the working electrode (23) can be formed to be smaller than the total area of ​​the two-dimensional plane of the counter electrode (21) and also smaller than the total area of ​​the two-dimensional plane of the reference electrode (25).

[0068] The carbon thin film (30) of the present invention is a layer upon which a fixing material (40) is placed and fixed, and when a specific binding reaction occurs between the fixing material (40) and the target binding material (50), it detects the electrical signal and transmits it to the electrode (20), and in particular, plays a role in ensuring that the fixing material (40) is not damaged, deformed, or destroyed by the metal electrode (20) and can maintain a normal active state for a long period of time.

[0069] That is, the biosensor of the present invention has a carbon thin film (30) formed on an electrode (20), and a substance identical to the target substance (i.e., a fixing substance (40)) is formed on the carbon thin film (30), so that the fixing substance (40) is provided in a structure in which it is in direct contact with the carbon thin film (30) rather than the electrode (20), and is formed in a structure that is immobilized on the electrode (20) through the carbon thin film (30).

[0070] For reference, in order to detect a target substance, a target binding substance (50) that specifically binds to the target substance is formed on the electrode (20). Therefore, the conventional biosensor is structured such that the target binding substance (50), rather than the target substance, is fixed in contact with the electrode (20).

[0071] Here, the target substance may be, for example, an antigen, which is a substance produced by pathogens such as bacteria, viruses, or fungi, or abnormal cells such as cancer cells, or a biohormone that performs a specific function. And, the target binding substance (50) may be a biomarker or bioprobe, such as an antibody, aptamer, or receptor, that specifically binds to such antigen or hormone.

[0072] On the other hand, the biosensor of the present invention is different in that, in order to detect a target substance, a substance identical to the target substance (i.e., a fixing substance (40)) is formed on the electrode (20) rather than the aforementioned target binding substance (50), and the fixing substance (40) is characterized by being formed on a carbon thin film (30) formed on the electrode (20).

[0073] However, unlike conventional biosensors, if a target substance such as an antigen is formed to come into direct contact with the electrode (20) instead of a target binding substance (50) such as an antibody, the following problems may occur.

[0074] That is, the electrode (20) is generally formed of a metallic material, and if antigens or biological hormones such as bacteria, viruses, or cancer cells are exposed to the metallic component of the electrode (20) for a long period of time, the function may be impaired or deformation, death, etc. may be induced due to the reaction with the electrode (20) or the toxicity of the metallic component.

[0075] For this reason, conventional biosensors provide a method for detecting a target substance based on a structure in which a target binding substance (50) other than the target substance is formed on an electrode (20).

[0076] In addition, when target substances (i.e., substances to be detected) such as antigens and hormones are directly immobilized on the metal electrodes of conventional biosensors, rather than target binding substances such as antibodies, receptors, markers, and probes, there was a problem in that the immobilization rate of such target substances was difficult to exceed about 20 to 30 percent relative to the amount of target substances input.

[0077] According to these conventional methods, there was a limitation in that it was difficult to simultaneously achieve a high level of measurement reproducibility and measurement resolution. In other words, there was a limitation where biosensors with high measurement resolution had very low measurement reproducibility, and biosensors with high measurement reproducibility had low measurement resolution.

[0078] On the other hand, the biosensor of the present invention is configured with a structure opposite to that of conventional biosensors (i.e., a structure in which a target substance, rather than a target binding substance, is immobilized on an electrode). Based on this, it is configured to detect a target substance using a detection method or principle entirely different from that of conventional biosensors. Through this, it has the advantage of simultaneously improving measurement accuracy, measurement range, as well as measurement resolution and measurement reproducibility compared to conventional biosensors. Detailed information regarding this will be described later.

[0079] Meanwhile, when the carbon thin film (30) of the present invention is applied, damage such as pinholes can be prevented during the process of removing the fixed material (40) and the conductive polymer fixed on the electrode for the reuse of the electrode (20), thereby providing the advantage of efficiently recycling the electrode as is.

[0080] For reference, when a fixing material (40) is fixed using a conductive polymer on a conventional metal electrode without applying a carbon thin film (30) as in the present invention, there was a disadvantage in that when the fixing material (40) and the conductive polymer were removed for the reuse of the electrode, a pinhole occurred in the metal electrode, making it impossible to recycle the electrode as is.

[0081] The carbon thin film (30) of the present invention is a key component that enables the opposite structure to the above-mentioned conventional biosensor, and can provide an environment that satisfies all biocompatibility, corrosion resistance, and biostability for target substances such as antigens or hormones, so that even if such target substances are formed on the electrode (20), problems such as damage, deformation, or death of the above-mentioned substances can be prevented, and it is also configured to detect an electrical signal resulting from a specific binding reaction between the target substance (i.e., the immobilizing substance (40)) and the target binding substance (50).

[0082] In addition, even if target substances such as antigens or hormones (i.e., substances to be detected) are immobilized on the electrode instead of target binding substances such as antibodies, receptors, markers, or probes, there is an effect of significantly increasing the immobilization rate of such target substances.

[0083] The carbon thin film (30) of the present invention is formed on the electrode (20), is formed of a carbon main component that does not contain hydrogen (H), has an amorphous structure, and has both carbon double bonds (C=C) and carbon single bonds (CC).

[0084] According to one example of the present invention, the electrode (20) may be composed of a counter electrode (21), a working electrode (23), and a reference electrode (25), and in the above case, a carbon thin film (30) may be formed on the working electrode (23).

[0085] The fixing material (40) of the present invention is a material that is immobilized on a carbon thin film (30) and includes the same material as the target substance (i.e., target substance).

[0086] For example, if the first biosensor is intended to detect a first target substance present in the subject's body fluid, the fixation substance (40) of the first biosensor includes the same substance as the first target substance.

[0087] As another example, if the second biosensor is intended to detect a second target substance present in the subject's body fluid, the fixation substance (40) of the second biosensor includes the same substance as the second target substance.

[0088] As another example, if the first biosensor is intended to detect a first target substance and a second target substance present in the subject's body fluid, the fixation material (40) of the first biosensor includes both the same substance as the first target substance and the same substance as the second target substance.

[0089] According to one example of the present invention, the target substance and the fixation substance (40) may be hormones, for example, (anti)stress hormones. In the above case, the (anti)stress hormone may be at least one selected from serotonin and cortisol.

[0090] For reference, (anti)stress hormones such as serotonin and cortisol are known to be directly related to depression. Therefore, depression can be diagnosed by measuring the levels of these (anti)stress hormones.

[0091] According to one example of the present invention, the fixing material (40) can be immobilized on the carbon thin film (30) by a conductive polymer.

[0092] In the above case, the conductive polymer may include at least one selected from polypyrrole, polythiophene, polyaniline, and polyacetylene.

[0093] According to one embodiment, the fixing material (40) may be formed by electrical polymerization, and the fixing material (40) may be immobilized simultaneously.

[0094] The bio module according to the present invention further includes a buffer solution (60) in addition to the bio sensor as described above.

[0095] FIG. 4 is a schematic diagram of a buffer solution mixed with body fluid containing a target substance according to the present invention.

[0096] The buffer solution (60) is a solution containing a target binding substance (50) and serves to help the target binding substance (50) maintain its activity for a long period of time within a specific pH range. That is, the buffer solution (60) contains a target binding substance (50) that reacts with a substance to be detected (i.e., a target substance), and serves to optimize the activity of the target binding substance (50).

[0097] When detecting a target substance in the subject's body fluid, the buffer solution (60) is mixed with the body fluid collected from the subject.

[0098] The buffer solution (60) includes a target binding substance (50) that specifically binds to a target substance, that is, a target substance present in the subject's body fluid, and when the target substance in the subject's body fluid is detected, the target binding substance (50) in the buffer solution (60) and the subject's body fluid are mixed and then administered onto at least a carbon thin film (30).

[0099] Here, the administration of the buffer solution (60) may mean an application method in which the buffer solution (60) is dropped onto the carbon film (30) in the form of a droplet, or the buffer solution (60) is applied onto the carbon film (30) so that the fixed material (40) immobilized on the carbon film (30) can come into contact with the buffer solution (60).

[0100] According to one example of the present invention, the target substance and the fixation substance (40) may be hormones, and in the above case, the target binding substance (50) mixed in the buffer solution (60) may be at least one selected from antibodies, aptamers, and receptors that specifically bind to the hormone.

[0101] For example, the target substance and the fixation substance (40) may be cortisol, and in the above case, the target binding substance (50) may be an antibody that specifically binds to cortisol and may include at least one of N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide and N-hydroxysuccinimide. Alternatively, the target binding substance (50) may be a structure in which N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide and N-hydroxysuccinimide are coupled.

[0102]

[0103] Below, the principle of detecting target substances of the bio module as described above will be explained first, followed by a detailed explanation of the carbon thin film (30).

[0104] Another bio module of the present invention is configured to detect a target substance in the subject's body fluid by utilizing the result of a competitive reaction between the target binding substance (50) in the buffer solution (60) and the target substance in the subject's body fluid when the aforementioned buffer solution (60) is administered onto the carbon thin film (30).

[0105] According to one example of the present invention, the competitive reaction result for detecting a target substance may include the following. That is, the competitive reaction result may include a result in which a portion of the target binding substance (50) in the buffer solution (60) preferentially reacts with the target substance in the body fluid mixed in the buffer solution (60), and the remaining portion reacts with the immobilized substance (40) immobilized on the carbon thin film (30).

[0106] Specifically, the competitive reaction results can be divided into a first case in which the concentration of the target substance in the subject’s body fluid mixed in the buffer solution (60) is above a threshold and a second case in which it is below a threshold, and different competitive reaction results are shown depending on the first and second cases.

[0107] Here, the 'threshold' is a preset value, for example, let us assume that a target binding substance (50) of concentration 1a is mixed in a buffer solution (60), and that this concentration 1a of the target binding substance (50) is a concentration capable of reacting with all of the target substances of concentration 2a, and that the threshold is set to concentration 2a.

[0108] In the above case, if the concentration of the target substance in the body fluid mixed in the buffer solution (60) is below the threshold (i.e., the 2a concentration), the following first competitive reaction result is produced.

[0109] Specifically, the result of the first competitive reaction includes the following: that is, only some of the target binding substances (50) contained in the buffer solution (60) preferentially react with the target substance in the body fluid (i.e., specific binding reaction), and the remaining portion of the target binding substances (50) is unable to react with the target substance in the body fluid. Then, the remaining portion that did not react with the target substance in the body fluid reacts with the immobilized material (40) immobilized on the carbon thin film (30).

[0110] When the concentration of the target substance in the body fluid mixed in the buffer solution (60) is greater than or equal to the threshold (i.e., the 2a concentration), the following second competitive reaction result is produced.

[0111] Specifically, the result of the second competitive reaction includes the following: that is, because a sufficiently large amount of target substance is present in the body fluid mixed in the buffer solution (60), all of the target binding substance (50) contained in the buffer solution (60) preferentially reacts (i.e., specific binding reaction) with the target substance in the body fluid, so that there is no remaining target binding substance (50) that did not react with the target substance in the body fluid. Accordingly, the specific binding reaction of the target binding substance (50) in the buffer solution (60) with respect to the immobilized substance (40) on the carbon thin film (30) does not occur.

[0112] According to one example of the present invention, the aforementioned threshold of the target substance concentration can be set to various values ​​by adjusting the concentration of the target binding substance (50) mixed in the buffer solution (60).

[0113] For example, in the case of the example described above, the concentration of the target binding substance (50) mixed in the buffer solution (60) is the first concentration, and in this case, the threshold of the target substance concentration is set to the second concentration. Here, let us assume a case where the concentration of the target binding substance (50) is mixed at the first concentration, which is lower than the first concentration.

[0114] In the above case, this 1b concentration of the target binding substance (50) is a concentration capable of reacting with all of the target substances of the maximum 2b concentration, and the 2b concentration corresponds to a value lower than the previously mentioned 2a concentration.

[0115] Accordingly, when the concentration of the target binding substance (50) in the buffer solution (60) is adjusted to the 1b concentration, the threshold of the target substance concentration described above can be set to the 2b concentration, which is a value lower than the 2a concentration.

[0116] The bio module of the present invention generates the following electrical signal according to the results of the aforementioned competitive reaction, and can use this to detect a target substance present in the subject's body fluid and also calculate the concentration value.

[0117] That is, when the target binding material (50) in the buffer solution (60) specifically binds to the fixing material (40) on the carbon thin film (30), an oxidation-reduction reaction occurs and a transfer of charge occurs, which can be measured as an electrical signal.

[0118] Furthermore, the magnitude or change of these electrical signals may appear as different values ​​depending on the level of the target substance contained in the subject's body fluids; therefore, by measuring the magnitude or change of the electrical signals, the target substance in the subject's body fluids can be detected and its level diagnosed.

[0119] According to one example of the present invention, when a buffer solution (60) is administered onto a carbon thin film (30), the bio module of the present invention generates an electrical signal greater than a threshold (hereinafter referred to as the "first threshold") when the concentration of the target substance in the body fluid mixed in the buffer solution (60) is within a normal range, and generates an electrical signal less than the first threshold when the concentration of the target substance in the body fluid mixed in the buffer solution (60) is within an abnormal range. Here, the "abnormal range" may refer to a concentration range higher than the maximum concentration of the target substance (e.g., hormone) that can be contained in the body fluid of a normal person.

[0120] Accordingly, as a result of a test using the bio module of the present invention, if an electrical signal greater than or equal to a first threshold is detected, the target substance level (e.g., hormone level) of the subject can be determined to be within the normal range, and if an electrical signal less than the first threshold is detected, the target substance level of the subject can be determined to be within the abnormal range.

[0121] According to one example of the present invention, when a buffer solution (60) is administered onto a carboxyl film, the bio module of the present invention generates an electrical signal of smaller size as the concentration of the target substance in the body fluid mixed in the buffer solution (60) is higher, and generates an electrical signal of larger size as the concentration of the target substance in the body fluid mixed in the buffer solution (60) is lower.

[0122] Accordingly, according to the bio module of the present invention, the concentration level of a target substance contained in the body fluid of the subject can be calculated by utilizing the magnitude or change of the measured electrical signal.

[0123] In other words, the smaller the magnitude (or change) of the measured electrical signal, the higher the concentration of the target substance (e.g., hormone level) contained in the subject's body fluid is determined to be, and the larger the magnitude (or change) of the measured electrical signal, the lower the concentration of the target substance contained in the subject's body fluid is determined to be.

[0124] As can be seen from the explanation above, the bio-module of the present invention generates electrochemical signal results opposite to those of conventional bio-sensors that utilize electrical signals. That is, conventional bio-sensors that utilize electrical signals generate a larger electrical signal in proportion to the higher the level of the target substance in the subject's body fluid.

[0125] On the other hand, the bio module of the present invention may be configured such that a smaller electrical signal is generated as the level of the target substance in the subject's body fluid increases, and furthermore, no electrical signal is generated when the level of the subject's target substance exceeds a set threshold (e.g., an abnormal level).

[0126] According to one example of the present invention, the 'electrical signal' may be an electrochemical signal generated by an oxidation-reduction reaction between a fixing material (40) immobilized on a carbon thin film (30) and a target binding material (50) in a buffer solution (60).

[0127] According to one example of the present invention, the 'electrical signal' may be at least one selected from current, voltage, resistance, and impedance.

[0128] In the above case, the bio module of the present invention can detect a target substance in body fluid and calculate the value by using the magnitude / strength of current, voltage, resistance, or impedance, or by using a change in magnitude / strength of current, voltage, resistance, or impedance.

[0129] According to one example of the present invention, the bio module of the present invention may be configured so that when a buffer solution (60) is administered onto a carbon film (30), if the concentration of a target substance in the body fluid mixed in the buffer solution (60) is within an abnormal range, a specific binding reaction between a fixation substance (40) immobilized on the carbon film (30) and a target binding substance (50) contained in the buffer solution (60) does not occur.

[0130] In the above case, since no specific binding reaction occurs between the fixation substance (40) and the target binding substance (50), no change in electrical signal occurs. Therefore, if no change in electrical signal is detected as a result of testing using the bio module of the present invention, the target substance level (e.g., hormone level) of the subject can be determined to be within an abnormal range.

[0131] If the concentration of the target substance in the body fluid mixed in the buffer solution (60) is within the normal range, a specific binding reaction occurs between the immobilized substance (40) on the carbon thin film (30) and the target binding substance (50) contained in the buffer solution (60), and an electrical signal is generated accordingly. Therefore, if a change in the electrical signal is detected as a result of an examination using the bio module of the present invention, the target substance level (e.g., hormone level) of the subject can be determined to be within the normal range.

[0132] A diagnostic method using a diagnostic kit for detecting a target substance in body fluid according to the present invention comprises a step of mixing the subject's body fluid, a step of administering a mixing buffer solution, a step of detecting the target substance, and a step of displaying the detection result.

[0133] The subject body fluid mixing step is a step of mixing the subject's body fluid with the buffer solution (60) of the diagnostic kit for detecting target substances in the aforementioned body fluid.

[0134] The mixed buffer solution administration step is a step of applying a buffer solution (60) mixed with the subject's body fluid onto a carbon film (30). At this time, the buffer solution (60) is dropped onto the carbon film (30) in the form of droplets, or the buffer solution (60) is applied onto the carbon film (30) so that the fixing material (40) immobilized on the carbon film (30) can come into contact with the buffer solution (60).

[0135] The target substance detection step is a step of detecting a target substance in a body fluid by utilizing the aforementioned competitive reaction result of the target binding substance (50) in the buffer solution with respect to the target substance in the body fluid mixed in the buffer solution (60).

[0136] The method for detecting target substances in body fluids and measuring their concentrations using competitive reaction results is the same as previously explained, so a detailed explanation thereof will be omitted.

[0137] According to one example of the present invention, the detection result display step may be a display step that visually displays the detection result of a target substance.

[0138] In the above case, the detection result display step may include the step of displaying a first label when the concentration of the target substance in the body fluid is below a threshold, and displaying a second label distinct from the first label when the concentration of the target substance in the body fluid is above the threshold.

[0139] According to one example of the present invention, the detection result display step may include the step of calculating and quantitatively displaying the numerical value of the concentration of a target substance in the body fluid of a determined subject.

[0140]

[0141] Hereinafter, the carbon thin film according to the present invention will be described in detail.

[0142] As explained above, the biosensor and module according to the present invention are configured with a structure opposite to that of a conventional biosensor (i.e., a structure in which a target substance, rather than a target binding substance, is immobilized on an electrode), and by detecting a target substance using a method or principle completely different from that of a conventional biosensor based on this, these differentiating features can be realized by the following carbon thin film (30).

[0143] For reference, when detecting a target substance based on a structure in which a carbon thin film (30) is formed on an electrode (20) and a target substance (i.e., a fixing substance (40)) is immobilized thereon, it must be possible to provide an environment that satisfies all biocompatibility, corrosion resistance, and biostability for the target substance, such as an antigen or hormone (hereinafter, required performance 1), and it must be possible to detect an electrochemical signal resulting from a specific binding reaction between the fixing substance (40) and the target binding substance (50) (hereinafter, required performance 2), and furthermore, it must be possible to satisfy the following required performance related to sterilization treatment (hereinafter, required performance 3).

[0144] That is, when immobilizing a fixing material (40) on a carbon film (30), sterilization treatment of the surface of the carbon film (30) may be required. Through this sterilization treatment, microorganisms such as bacteria and fungi unrelated to the fixing material (40) are removed to prevent contamination of the fixing material (40) and to minimize noise generation during the reaction of the target binding material (50).

[0145] Various sterilization methods may be used, such as high temperature / high pressure sterilization, chemical sterilization, ultraviolet (UV) sterilization, ultrasonic sterilization, and electric sterilization, and an appropriate sterilization method may be selected depending on the type of fixed material (40).

[0146] For example, high-temperature sterilization is a method of removing all microorganisms attached to the carbon film (30) using high temperature and high pressure steam of 121°C or higher, and undergoes a cooling process after high-temperature sterilization.

[0147] Chemical sterilization is a method of removing microorganisms on a carbon film (30) using alcohol or hydrogen peroxide, and ultraviolet (UV) sterilization is a method of removing microorganisms by irradiating the surface of a carbon film (30) with ultraviolet light.

[0148] However, when the carbon film (30) is formed with carbon as the main component, cracks, pinholes, peeling (hereinafter collectively referred to as ‘carbon film damage’) may occur in the carbon film (30) due to the high temperature / high pressure atmosphere or chemical substances of the sterilization process, and in the above case, it cannot function as a biosensor for detecting target substances in body fluids.

[0149] Therefore, when forming a carbon thin film (30) of carbon main component on an electrode (20) in the biosensor, it is important to satisfy the aforementioned required performance 1 and required performance 2, but it must also satisfy the required performance 3 related to sterilization treatment.

[0150] To this end, the inventors of the present invention have found that if a carbon thin film (30) is formed by physical vapor deposition (PVD) and the carbon thin film (30) is formed from a carbon-based material having hardness, carbon bonding structure (XPS peak intensity ratio), and pore volume characteristics within a specific range, all of the aforementioned requirements 1, 2, and 3 can be satisfied.

[0151] (1) Deposition method of carbon thin film

[0152] The carbon thin film (30) is formed by physical vapor deposition (PVD) and may be made of amorphous carbon.

[0153] Here, the term 'amorphous' refers to a state in which the arrangement of multiple atoms or molecules lacks periodic regularity, does not have a specific shape, or is formless. Conversely, a material that maintains its atomic arrangement regularity over long distances corresponds to a crystal.

[0154] Therefore, the term 'amorphous carbon' used in the present invention refers to carbon that exists in an amorphous form without a crystal structure.

[0155] According to one example of the present invention, the carbon thin film (30) can be formed by physical vapor deposition (PVD) using a plasma source.

[0156] Physical Vapor Deposition (PVD) methods utilizing a plasma source include ion plating and sputtering.

[0157] The carbon thin film (30) can be deposited on the upper surface of the electrode (20) and can be formed in the form of a thin film or a membrane.

[0158] According to one example of the present invention, the carbon thin film (30) may have a thickness of 5 to 1000 nm, but is not necessarily limited thereto. For example, the thickness of the carbon thin film (30) may be 10 to 1000 nm; 50 to 1000 nm; 100 to 1000 nm; 300 to 1000 nm; 500 to 1000 nm; 5 to 900 nm; 5 to 700 nm; 5 to 500 nm; or 5 to 300 nm.

[0159]

[0160] (2) Carbon bonding structure of the carbon thin film (XPS peak intensity ratio (I))

[0161] The carbon thin film (30) of the present invention is formed on the electrode (20), is formed of a carbon main component that does not contain hydrogen (H), has an amorphous structure, and has both carbon double bonds (C=C) and carbon single bonds (CC).

[0162] According to one example of the present invention, the carbon thin film (30) is formed such that, based on the XPS peak intensity according to XPS analysis, the XPS peak intensity related to the carbon double bond (C=C) is higher than the XPS peak intensity related to the carbon single bond (CC).

[0163] This is because if the XPS peak intensity related to the carbon double bond (C=C) of the carbon film (30) is smaller than the XPS peak intensity related to the carbon single bond (CC), the detection performance for the electrochemical signal generated during the specific binding reaction between the immobilizing material (40) and the target binding material (50) is significantly reduced, and thus the aforementioned requirement 2 cannot be satisfied.

[0164] In this specification, XPS analysis refers to X-ray photoelectron spectroscopy, and the XPS peak intensity related to carbon double bonds is the sp of carbon. 2 It may be the bond-related XPS peak intensity, and the carbon single bond-related XPS peak intensity is the carbon's sp 3 It may be the XPS peak intensity related to the bond.

[0165] According to one example of the present invention, the ratio of carbon single bonds in the carbon thin film (30) is formed to have a higher value than the ratio of carbon single bonds in graphite and a lower value than the ratio of carbon single bonds in PVD-DLC as described above, based on the XPS peak intensity according to XPS analysis.

[0166] That is, the XPS peak intensity related to carbon single bonds of the carbon thin film (30) has a higher value than the XPS peak intensity related to carbon single bonds of graphite and can have a lower value than the XPS peak intensity related to carbon single bonds of PVD-DLC.

[0167] According to one example of the present invention, the ratio of carbon double bonds in the carbon film (30) is formed to have a value lower than the ratio of carbon double bonds in graphite and a value higher than the ratio of carbon double bonds in PVD-DLC, based on the XPS peak intensity according to XPS analysis.

[0168] That is, the XPS peak intensity related to carbon double bonds of the carbon film (30) has a lower value than the XPS peak intensity related to carbon double bonds of graphite and can have a higher value than the XPS peak intensity related to carbon double bonds of PVD-DLC.

[0169] Here, the graphite used as a comparison standard for the carbon bonding structure (XPS peak intensity ratio (I)) characteristics of the carbon thin film (30) may be pure graphite that is not doped with other elements or other materials.

[0170] And, PVD-DLC, which is another comparison standard for the carbon bonding structure (XPS peak intensity ratio (I)) characteristics of the carbon thin film (30), may be PVD-DLC that is not doped with other elements or other materials, and may be, for example, Ta-DLC (Tetrahedral Amorphous Diamond Like Carbon).

[0171] Meanwhile, the size comparison of the carbon single / double bond ratio of the carbon thin film (30) with the carbon single / double bond ratio of graphite and PVD-DLC can be determined through XPS (X-ray photoelectron spectroscopy) analysis.

[0172] More specifically, the carbon thin film (30) can be formed such that the intensity ratio (I) of the XPS (X-ray photoelectron spectroscopy) peaks calculated according to the following mathematical formula 1 satisfies the following conditional formula 1.

[0173] [Mathematical Formula 1]

[0174] XPS peak intensity ratio (I) = I1 / I2

[0175] (In Equation 1, I1: XPS peak intensity related to carbon single bonds (CC) of the carbon thin film, I2: XPS peak intensity related to carbon double bonds (C=C) of the carbon thin film)

[0176]

[0177] [Condition 1]

[0178] 0.3 < XPS peak intensity ratio (I) < 1.0

[0179]

[0180] If the XPS peak intensity ratio (I) according to mathematical formula 1 of the carbon film (30) is 0.3 or less, peeling, cracking, pinholes, etc. of the carbon film (30) occur during the aforementioned sterilization treatment, and the aforementioned required performance 3 cannot be satisfied.

[0181] And, if the XPS peak intensity ratio (I) according to mathematical formula 1 of the carbon thin film (30) is 1.0 or higher, the electrical signal detection performance due to the specific binding reaction between the fixed material (40) and the target binding material (50) is significantly reduced, and the aforementioned required performance 2 cannot be satisfied.

[0182] Preferably, the carbon thin film (30) has single bonds (CC) and double bonds (C=C) between carbons, and can be formed so that the intensity ratio (I) of the XPS peaks calculated according to the above mathematical formula 1 further satisfies the following condition formula 2 or the following condition formula 3.

[0183] [Condition 2]

[0184] 0.4 < XPS peak intensity ratio (I) < 0.9

[0185]

[0186] [Condition 3]

[0187] 0.5 < XPS peak intensity ratio (I) < 0.8

[0188]

[0189] Meanwhile, the carbon thin film (30) can be formed to have the aforementioned XPS peak intensity ratio (I) characteristics by controlling the process conditions for its deposition.

[0190] For example, when forming a carbon thin film (30) using sputtering, the plasma density (cm) -3 By controlling process conditions including ) and process pressure (mTorr), a carbon thin film (30) satisfying the aforementioned XPS peak intensity ratio (I) characteristic can be formed.

[0191]

[0192] (3) Hardness and resistivity of the carbon film

[0193] The carbon film (30) is an electrically conductive material having a hardness within a specific range as follows.

[0194] The carbon film (30) is formed to have a higher hardness compared to the hardness of graphite, which is another carbon allotrope.

[0195] The carbon film (30) is formed to have a lower hardness compared to the hardness of another carbon allotrope, diamond-like carbon (DLC: Diamond-Like Carbon, hereinafter referred to as 'PVD-DLC').

[0196] And, the carbon thin film (30) is formed to have a higher electrical conductivity compared to the electrical conductivity of the PVD-DLC.

[0197] That is, the carbon thin film (30) is formed to have a lower resistivity compared to the resistivity of the PVD-DLC.

[0198] The carbon film (30) can have a lower electrical conductivity compared to the electrical conductivity of graphite.

[0199] That is, the carbon film (30) can have a higher resistivity compared to the resistivity of graphite.

[0200] Here, the hardness and resistivity values ​​of graphite, which are the physical property comparison standards of the carbon thin film (30), can be based on the maximum hardness and maximum resistivity values ​​that graphite can have.

[0201] Therefore, the carbon film (30) may be a material having a hardness higher than the maximum hardness of graphite and a resistivity higher than the maximum resistivity of graphite.

[0202] Here, PVD-DLC, which is another physical property comparison standard for the carbon thin film (30), is formed by physical vapor deposition (PVD), is composed of amorphous carbon, and is intended to have characteristics similar to diamond (i.e., carbon bonding structure and physical properties).

[0203] The physical vapor deposition (PVD) method for forming PVD-DLC can be sputtering or ion plating.

[0204] In addition, when PVD-DLC is formed by sputtering, an inert gas such as argon (Ar) can be used as the process gas.

[0205] The hardness and resistivity values ​​of PVD-DLC, which are the physical property comparison standards of the carbon thin film (30), can be based on the minimum hardness and minimum resistivity values ​​that PVD-DLC can have.

[0206] Accordingly, the carbon thin film (30) may be a material having a lower hardness than the minimum hardness of PVD-DLC and a lower resistivity than the minimum resistivity of PVD-DLC.

[0207] Specifically, when diamond-like carbon (DLC) is formed by physical vapor deposition (PVD), it is amorphous and exhibits a hardness of at least 35 GPa, and generally exhibits a high hardness of 40 to 80 GPa.

[0208] And, PVD-DLC is 1×10 14 ~ 1×10 16 It exhibits a resistivity of Ωcm.

[0209] A representative example of such PVD-DLC is Ta-DLC (Tetrahedral Amorphous Diamond Like Carbon). This Ta-DLC has a very high hardness of 60 to 80 GPa. For reference, diamond has a hardness of 70 to 150 GPa.

[0210] According to one example of the present invention, the PVD-DLC, which serves as a comparison standard for the hardness and resistivity characteristics of the carbon thin film (30), may be a PVD-DLC that is not doped with other elements or other materials, and may be, for example, Ta-DLC (Tetrahedral Amorphous Diamond Like Carbon).

[0211] And, graphite, which is another comparison criterion for the hardness and resistivity characteristics of the carbon film (30), may be pure graphite that is not doped with other elements or other materials.

[0212] Graphite, one of the carbon allotropes, is known to have a hardness of 1.5 GPa or less.

[0213] Graphite is a conductor with very high electrical conductivity, 1×10⁻⁶ -4 ~ 1×10 -3 It has a resistivity of Ωcm.

[0214] Accordingly, the carbon thin film (30) is formed to have a hardness greater than the hardness range of graphite, which is 1.5 GPa or less, and a hardness lower than the hardness range of PVD-DLC, which is 35 to 80 GPa.

[0215] That is, the carbon thin film (30) is formed to have a hardness value higher than 1.5 GPa, which corresponds to the maximum hardness value of graphite, and a hardness value lower than 35 GPa, which corresponds to the minimum hardness value of PVD-DLC.

[0216] Meanwhile, in order to prevent damage to the carbon film (30) during sterilization treatment and, in particular, to maximize electrical signal detection performance, conditions related to the XPS peak intensity ratio (I) and pore volume described later can be considered together, and in the above case, the carbon film (30) can be formed to have the following hardness.

[0217] According to one example of the present invention, the hardness of the carbon thin film (30) may be 8 GPa to 28 GPa, preferably 9 GPa to 27 GPa; or 10 GPa to 26 GPa, but is not limited thereto.

[0218] Here, the hardness of the carbon thin film (30) may be a value measured by a nanoindenter, which is one of the hardness measuring devices. Here, the nanoindenter may be a nanoindentation tester that measures in compliance with standard test methods ASTM E 2546 to ISO 14577.

[0219] And, the carbon thin film (30) has a resistivity range of 1×10⁻⁶, which is the resistivity range of graphite. -4 ~ 1×10 -3 It can have a resistivity greater than Ωcm, and the resistivity range of PVD-DLC is 1×10⁻⁶ 14 ~ 1×10 16 It has a resistivity lower than Ωcm.

[0220] That is, the carbon thin film (30) is 1×10⁻¹⁰, which corresponds to the maximum resistivity value of graphite.-3 It can have a resistivity value higher than Ωcm, and 1×10⁻⁶, which corresponds to the minimum resistivity value of PVD-DLC. 15 It is a conductive material having a resistivity value lower than Ωcm.

[0221] According to one example of the present invention, the resistivity (Ωcm) of the carbon thin film (30) may be 1 to 1500 Ωcm, but is not limited thereto. For example, the resistivity (Ωcm) of the carbon thin film (30) may be 1 to 1800 Ωcm; 1 to 1600 Ωcm; 1 to 1400 Ωcm; 1 to 1200 Ωcm; 10 to 2000 Ωcm; 50 to 2000 Ωcm; 100 to 2000 Ωcm; or 200 to 2000 Ωcm.

[0222] That is, the carbon thin film (30) is made of an electrically conductive material having a lower resistivity than PVD-DLC, at least 10 times the maximum resistivity of graphite; at least 10 2 Times; or at least 10 3 It can have twice as high resistivity.

[0223] Meanwhile, the carbon thin film (30) can be formed to have the aforementioned hardness and resistivity characteristics by controlling the process conditions for its deposition.

[0224] For example, when forming a carbon thin film (30) using sputtering, the current density (W / cm²) and plasma density (cm²) -3 A carbon thin film (30) satisfying the aforementioned hardness and resistivity characteristics can be formed by controlling at least a plurality of process conditions selected from process pressure (mTorr), deposition rate (nm / min), electron temperature (eV), and ion flux (mA / cm²).

[0225]

[0226] (4) Porosity of the carbon film

[0227] The carbon film (30) has a number of pores, and these pores have the following characteristics.

[0228] That is, the porosity of the carbon film (30) can be formed to have a value greater than the maximum porosity that graphite can have.

[0229] Here, the porosity of the carbon film (30) refers to the value obtained by dividing the total volume occupied by the pores within the carbon film (30) (i.e., the total volume of pores) by the total volume of the carbon film (30).

[0230] Furthermore, the pore ratio of graphite refers to the value obtained by dividing the total volume occupied by the pores within the graphite (i.e., the total pore volume) by the total volume of the graphite.

[0231] Accordingly, the carbon film (30) can be formed such that, when compared to the carbon film (30) and graphite having the same volume, the total pore volume of the carbon film (30) has a value greater than the total pore volume of the graphite.

[0232] When the carbon thin film (30) is formed to have the aforementioned pore volume characteristics, the carbon thin film (30) may be formed to have a lower density than graphite. For reference, the theoretical density of graphite is 2.267 g / cm³.

[0233] Accordingly, according to one example of the present invention, the carbon thin film (30) can be formed to have a density of less than 2.267 g / cm³.

[0234] According to one example of the present invention, the density of the carbon thin film (30) may be at most 2.2 g / cm³ and at least 1.05 g / cm³, but is not limited thereto. For example, the density of the carbon thin film (30) may be 1.05 g / cm³ or less and less than 2.15 g / cm³; 1.05 g / cm³ or less and more than 2.05 g / cm³; 1.05 g / cm³ or less and more than 2.0 g / cm³; 1.05 g / cm³ or less and more than 2.0 g / cm³; 1.1 g / cm³ or less and more than 2.2 g / cm³; 1.15 g / cm³ or less and more than 2.2 g / cm³; 1.2 g / cm³ or less and more than 2.2 g / cm³; or 1.25 g / cm³ or less and more than 2.2 g / cm³.

[0235] According to one example of the present invention, the porosity of the carbon thin film (30) can be formed to have a value greater than the porosity of the PVD-DLC.

[0236] Here, the porosity of PVD-DLC refers to the value obtained by dividing the total volume occupied by the pores within the PVD-DLC (i.e., the total pore volume) by the total volume of the PVD-DLC.

[0237] Accordingly, the carbon thin film (30) is formed such that, when compared to the carbon thin film (30) and PVD-DLC having mutually equal volumes, the total pore volume of the carbon thin film (30) has a value greater than the total pore volume of the PVD-DLC.

[0238] In addition, when the carbon thin film (30) is formed to have the aforementioned porosity characteristics, the carbon thin film (30) may be formed to have a lower density than PVD-DLC.

[0239] Here, the PVD-DLC, which is a comparison standard for the porosity characteristics of the carbon thin film (30), may be a PVD-DLC that is not doped with other elements or other materials, and may be, for example, Ta-DLC (Tetrahedral Amorphous Diamond Like Carbon).

[0240] Ta-DLC can have a density similar to that of diamond, specifically a density of about 2.39 to 3.26 g / cm³. For reference, the theoretical density of diamond is 3.515 g / cm³.

[0241] And, graphite, which is another comparison criterion for the porosity characteristics of the carbon film (30), may be pure graphite that is not doped with other elements or other materials.

[0242] That is, the carbon thin film (30) according to one example of the present invention can be formed to have a lower density than PVD-DLC, and furthermore, can have a lower density than graphite, which has a lower density than PVD-DLC.

[0243] FIG. 5 is a field emission scanning electron microscope (FESEM) analysis image of a carbon thin film according to one embodiment of the present invention. As can be seen in FIG. 5, the carbon thin film (30) according to the present invention has a large number of pores formed with a very large size, and the porosity is found to have a value greater than that of PVD-DLC as well as graphite.

[0244] In addition, when the carbon thin film (30) is formed to have porosity characteristics as described above, the carbon thin film is shown to have a lower density than PVD-DLC as well as graphite.

[0245] Meanwhile, the carbon thin film can be formed to have the aforementioned pore volume characteristics by controlling the process conditions for its deposition.

[0246] For example, when forming a carbon thin film (30) using sputtering, the current density (W / cm²) and plasma density (cm²) -3 A carbon thin film (30) satisfying the aforementioned porosity characteristics can be formed by controlling at least a plurality of process conditions selected from process pressure (mTorr), deposition rate (nm / min), electron temperature (eV), and ion flux (mA / cm²).

[0247]

[0248] (5) Process conditions (plasma density and process pressure)

[0249] According to one example of the present invention, a carbon thin film (30) may be formed by sputtering deposition, and in the case, the carbon thin film (30) may be formed by controlling the plasma density and working pressure to specific process conditions.

[0250] That is, when a carbon thin film (30) is formed by sputtering in a process atmosphere having plasma density and process pressure within the following ranges, a carbon-based material having the aforementioned hardness, carbon bond structure (XPS peak intensity ratio), and pore volume characteristics can be formed.

[0251] According to one example of the present invention, when a carbon thin film (30) is formed by sputtering, the working pressure may be greater than 1 mTorr and less than 130 mTorr, and preferably 3 mTorr to 100 mTorr; 3 mTorr to 70 mTorr; 3 mTorr to 50 mTorr; or 3 mTorr to 20 mTorr.

[0252] According to one example of the present invention, when a carbon thin film (30) is formed by sputtering, the plasma density is at least 1.0 × 10 10 cm -3 It may be more than, preferably at least 5.0×10 10 cm -3 It could be more than that.

[0253] According to one example of the present invention, when a carbon thin film (30) is formed by sputtering, the plasma density is 1.0 × 10 10 cm -3 Up to 5.0×10 11 cm -3 It can be, preferably 5.0×10 10 cm -3 Up to 5.0×10 11cm -3 It could be.

[0254] Here, plasma density can refer to the density of ionized particles in the sputtering process.

[0255] According to one example of the present invention, the carbon thin film (30) may be formed by magnetron sputtering, and in this case, the magnetron sputtering may be unbalanced magnetron sputtering (UBMS).

[0256] Figure 6 is experimental data showing the XPS peak intensities related to carbon single bonds (CC) and carbon double bonds (C=C) of a carbon thin film according to the present invention, measured according to plasma density when the carbon thin film is formed by physical vapor deposition (PVD).

[0257] Figure 7 is experimental data showing the plasma density measured according to process pressure during the physical vapor deposition (PVD) process and plotted as a graph.

[0258] Referring to FIGS. 6 and 7, when a carbon thin film (30) according to the present invention is formed by physical vapor deposition (PVD) using a plasma source, the plasma density is 5.0 × 10 9 cm -3 When controlled as described above, it can be confirmed that a carbon thin film (30) satisfying the "carbon bonding structure (XPS peak intensity ratio (I) of the carbon thin film (30)" described above, i.e., condition 1, is formed.

[0259] According to one example of the present invention, physical vapor deposition (PVD) using a plasma source may be sputtering.

[0260] In addition, by controlling the current density (W / cm²), process pressure (mTorr), deposition rate (nm / min), electron temperature (eV), ion flux (mA / cm²), etc., under specific conditions, a carbon thin film (30) having hardness, resistivity, and pore volume characteristics within the aforementioned specific range can be formed.

[0261]

[0262] (6) Extended Example

[0263] According to an extended embodiment of the present invention, the carbon thin film (30) may be a carbon material doped with nitrogen (N).

[0264] When nitrogen (N) is doped into the aforementioned carbon thin film (30), nitrogen gas can be injected at a level below a specific threshold relative to the total process pressure during the sputtering process for forming the carbon thin film (30) to form a nitrogen-doped carbon thin film (30).

[0265] According to one example of the present invention, the partial pressure of nitrogen during the sputtering process for forming a carbon thin film (30) may be up to 10% (i.e., 10% or less) relative to the total process pressure, and preferably 9% or less; 8% or less; 7% or less; 6% or less; or 5% or less.

[0266] This is because if the partial pressure of nitrogen exceeds 10%, the stress increases, making it difficult to form the carbon thin film (30), and the film properties deteriorate, and the adhesion and bonding strength (i.e., bonding strength) of the carbon thin film (30) to the electrode (20) decreases rapidly.

[0267] For reference, if there is a region where the adhesion between the electrode (20) and the carbon thin film (30) is not uniform or the bonding force is weak, the carbon thin film (30) may peel off from the electrode (20) or crack in that region, and this may act as a factor that reduces the measurement accuracy and reproducibility of the biosensor.

[0268] According to one example of the present invention, the nitrogen content of the carbon thin film (30) may be up to 10 weight% (i.e., 10 weight% or less), and preferably 9 weight% or less; 8 weight% or less; 7 weight% or less; 6 weight% or less; 5 weight% or less; or 4 weight% or less.

[0269] This is because if the nitrogen content of the carbon thin film (30) exceeds 10 weight%, it becomes difficult to form the carbon thin film (30), and the adhesion and bonding strength (i.e., bonding strength) of the carbon thin film (30) to the electrode (20) decreases rapidly, which ultimately leads to a problem where the measurement accuracy and reproducibility of the biosensor decrease.

[0270] Accordingly, in order to ensure the measurement accuracy and reproducibility of the biosensor, the carbon thin film (30) must be formed on the electrode (20) with uniform adhesion and strong bonding strength throughout, and for this purpose, it can be formed with a nitrogen partial pressure or nitrogen content within the aforementioned range.

[0271] In this way, when nitrogen is doped into the carbon film (30) within a specific range of nitrogen partial pressure or nitrogen content, it is possible to effectively prevent damage to the carbon film (30) during sterilization treatment, while maximizing the aforementioned required performance 2 (i.e., detection performance for electrochemical signals).

[0272] According to an extended embodiment of the present invention, the carbon thin film (30) may be formed to have a gradient portion in which the ratio of carbon single bonds gradually increases from the upper portion to the lower portion.

[0273] And, the carbon film (30) can be formed to have a gradient region in which the ratio of carbon double bonds gradually decreases from the top to the bottom.

[0274] Here, the upper portion of the carbon film (30) may be a portion that includes the surface of the carbon film (30) or a portion that is closer to the surface than the lower portion, and the lower portion may be a portion that includes the opposite side of the surface or a portion that is closer to the opposite side than the upper portion.

[0275]

[0276] Hereinafter, the biosensor and module for detecting target substances in body fluids according to the present invention as described above will be further explained based on examples and comparative examples. However, the following examples are merely illustrative of the content of the present invention and the scope of the present invention is not limited by the following examples.

[0277] [Example 1]

[0278] The biosensor and module according to Example 1 were fabricated so that the XPS peak intensity ratio, hardness, and porosity of the carbon thin film all fall within the range of physical properties presented in the present invention.

[0279] The hardness of the carbon thin film according to Example 1 was measured using a nanoindenter compliant with ISO 14577, and the XPS peak intensity ratio (I) was measured using a MultiLab 2000 spectrometer (Thermo Electron Corporation, UK). Using the XPS data analyzed in this way, the XPS peak intensity ratio (I) was calculated according to the aforementioned Equation 1.

[0280] (1) Electrode

[0281] - Three electrodes consisting of a working electrode, a counter electrode, and a reference electrode

[0282] (2) Target substances and immobilized substances

[0283] - Target substance: cortisol

[0284] - Fixed substance: cortisol

[0285] (3) Target binding substance

[0286] - Cortisol Antibody: N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide (Purchased from R&D Systems (Minneapolis, MN, USA))

[0287] (3) Carbon thin film process conditions

[0288] - Deposition method: Sputtering

[0289] - Process pressure: 3×10 -3 torr

[0290] - Plasma density: 8×10 10 cm -3

[0291] - Sputtering Target: 99.99% purity graphite target

[0292] (4) Carbon thin film properties

[0293] - XPS Peak Intensity Ratio (I): 0.62

[0294] - Hardness: 8.3 Gpa

[0295] - Porosity: Greater than graphite

[0296]

[0297] [test]

[0298] FIG. 8 is a photograph of the first result of detecting a target substance (cortisol) using the competitive reaction result of the bio module according to the present invention, corresponding to the competitive reaction result that appears when the level of the target substance (cortisol) contained in the subject's body fluid falls within the normal range (i.e., negative reaction).

[0299] That is, the first result, as shown in Figure 8, confirms that the cortisol concentration in the subject's body fluid is below the threshold, and accordingly, only some of the target binding substances (cortisol antibodies) contained in the buffer solution preferentially undergo a specific binding reaction with the target substance in the body fluid, while the remaining parts that did not undergo a specific binding reaction with the target substance in the body fluid undergo a specific binding reaction with the immobilized substance (i.e., cortisol) immobilized on the carbon foil.

[0300] FIG. 9 is a photograph of a second result in which a target substance (cortisol) is detected using the competitive reaction result of the bio module according to the present invention, and corresponds to the competitive reaction result that appears when the level of the target substance (cortisol) contained in the subject's body fluid falls within an abnormal range (i.e., a positive reaction).

[0301] That is, the second result, as shown in Fig. 9, confirms that the cortisol concentration in the subject's body fluid is above a threshold, and accordingly, all of the target binding substances (cortisol antibodies) contained in the buffer solution preferentially undergo a specific binding reaction with the cortisol in the body fluid, and thus no specific binding reaction occurs with the immobilized substance (i.e., cortisol) immobilized on the carbon thin film.

[0302] Figure 11 is a photograph of the carbon thin film state of Example 1 after sterilization treatment.

[0303] Referring to FIG. 11, according to Embodiment 1 of the present invention, as can be seen in FIG. 8 and FIG. 9, the target substance can be accurately detected, and it can also be confirmed that no damage such as peeling or cracking occurs even when sterilization is performed.

[0304]

[0305] [Comparative Example 1]

[0306] Comparative Example 1 involved forming a carbon thin film on an electrode that did not satisfy the XPS peak intensity ratio (I) and hardness characteristics among the carbon thin film properties presented in the present invention, and then performing sterilization treatment. Specifically, the properties of the carbon thin film according to Comparative Example 1 are as follows.

[0307] - XPS Peak Intensity Ratio (I): 0.26

[0308] - Hardness: 1.6 Gpa

[0309] - Porosity: Greater than graphite

[0310] Figure 10 is a photograph of the carbon thin film state of Comparative Example 1 after sterilization treatment.

[0311] Referring to Fig. 10, in the case of Comparative Example 1, it can be seen that damage to the carbon film, including peeling and cracking as shown in Fig. 10, occurs when sterilization is performed, and in this case, it cannot function as a biosensor for detecting target substances in body fluids.

[0312]

[0313] Although preferred embodiments of the present invention have been described and illustrated above using specific terms, such terms are intended solely to clarify the invention, and it is obvious that various modifications and changes may be made to the embodiments and described terms without departing from the technical spirit and scope of the following claims. Such modified embodiments should not be understood separately from the spirit and scope of the invention, but should be considered to fall within the scope of the claims of the invention.

[0314]

[0315] Explanation of the symbols

[0316] 10: Material 20: Electrode

[0317] 21: Counter electrode 23: Working electrode

[0318] 25: Reference electrode 30: Carbon thin film

[0319] 40: Immobilizing material 50: Target binding material

[0320] 60: Buffer solution

Claims

1. As a biosensor for detecting target substances in a subject's body fluids, A substrate; an electrode formed on the substrate; a carbon thin film formed on the electrode; and a fixing material immobilized on the carbon thin film; comprising The above carbon thin film is, It is composed of an amorphous structure that does not contain hydrogen (H), and It has both carbon double bonds (C=C) and carbon single bonds (CC), and The above-mentioned fixed material is, A biosensor for detecting a target substance in body fluids, characterized by containing a substance identical to the target substance.

2. In Paragraph 1, The above target substance and the above immobilized substance are, A biosensor for detecting target substances in body fluids characterized by being hormones.

3. In Paragraph 1, The above carbon thin film is, Based on XPS peak intensity according to XPS analysis, A biosensor for detecting target substances in body fluids, characterized in that the XPS peak intensity related to the carbon double bond is higher than the XPS peak intensity related to the carbon single bond.

4. In Paragraph 1, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized in that the XPS peak intensity ratio (I) calculated according to the following mathematical formula 1 satisfies the following condition formula 1. Mathematical formula 1 XPS peak intensity ratio (I) = I1 / I2 (In Equation 1, I1: XPS peak intensity related to the carbon single bond, I2: XPS peak intensity related to the carbon double bond) Conditional expression 1 0.3 < XPS peak intensity ratio (I) < 1.0 5. In Paragraph 4, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized in that the intensity ratio (I) of XPS peaks calculated according to the above mathematical formula 1 further satisfies the following condition formula 2. Conditional expression 2 0.4 < XPS peak intensity ratio (I) < 0.9 6. In Paragraph 1, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by having a hardness greater than the maximum hardness that graphite, a carbon allotrope, can have.

7. In Paragraph 6, The hardness of the above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by having a value of 8 GPa to 28 GPa.

8. In Paragraph 1, The above carbon thin film is, It has multiple voids, and The porosity of the above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by having a porosity greater than the maximum porosity that graphite, a carbon allotrope, can have.

9. In Paragraph 1, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by being a material having a resistivity greater than the maximum resistivity that carbon allotropes graphene and graphite can have.

10. In Paragraph 1, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by having a gradient region in which the ratio of carbon single bonds gradually increases from the upper part to the lower part of the carbon thin film.

11. In Paragraph 1, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by having a gradient region in which the ratio of carbon double bonds gradually decreases from the upper part to the lower part of the carbon thin film.

12. In Paragraph 1, The above carbon thin film is, A biosensor for detecting target substances in body fluids characterized by being doped with nitrogen (N).

13. In Paragraph 12, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by having a nitrogen content of 10 weight% or less.

14. In Paragraph 12, The above carbon thin film is, It is formed by physical vapor deposition (PVD), and The nitrogen is doped by injecting nitrogen gas during the physical vapor deposition (PVD) process for forming the carbon thin film. A biosensor for detecting target substances in body fluids, characterized in that the partial pressure of nitrogen during the physical vapor deposition (PVD) process is 10% or less of the total process pressure.

15. In Paragraph 1, The above carbon thin film is, At least 1.0×10 10 cm -3 A biosensor for detecting target substances in body fluids, characterized by being formed by a sputtering process having a plasma density greater than or equal to the above.

16. In Paragraph 1, The above carbon thin film is, 5.0×10 10 cm -3 Up to 5.0×10 11 cm -3 A biosensor for detecting target substances in body fluids, characterized by being formed by a sputtering process having a plasma density.

17. In Paragraph 1, The above electrode is, It includes a working electrode, a counter electrode, and a reference electrode, The above carbon thin film is, A biosensor for detecting target substances in body fluids, characterized by being formed on the above-mentioned working electrode.

18. In Paragraph 17, The total area of ​​the two-dimensional plane of the above working electrode is, A biosensor for detecting a target substance in body fluid, characterized by having a total area of ​​the two-dimensional plane of the counter electrode and a total area of ​​the two-dimensional plane of the reference electrode that is smaller than the total area of ​​the two-dimensional plane of the reference electrode.

19. A biosensor for detecting a target substance in body fluid according to any one of claims 1 to 18; and a biomodule for detecting a target substance in body fluid, comprising a buffer solution mixed with the subject's body fluid, The above buffer solution is, It includes a target binding substance that specifically binds to the above target substance, and When a target substance is detected in the body fluid of the subject, the target binding substance and the subject's body fluid are mixed and administered onto the carbon thin film. The above bio module is, A bio module for detecting a target substance in a body fluid, characterized by detecting the target substance by utilizing the result of a competitive reaction between the target binding substance in the buffer solution and the target substance in the body fluid when the buffer solution is administered onto the carbon thin film.

20. In Paragraph 19, The result of the above competitive response is, A portion of the target binding substance in the above buffer solution reacts preferentially with the target substance in the above body fluid, and A bio-module for detecting target substances in body fluids, wherein the remainder includes a result of reacting with the immobilized material immobilized on the carbon thin film.

21. In Paragraph 19, The result of the above competitive response is, If the concentration of the target substance in the body fluids of the above-mentioned subject is below the threshold, Only some of the target binding substances included in the above buffer solution preferentially undergo a specific binding reaction with the target substance in the body fluid, and A bio-module for detecting a target substance in a body fluid, wherein the remaining portion that did not undergo a specific binding reaction with the target substance in the body fluid undergoes a specific binding reaction with the immobilized substance immobilized on the carbon thin film.

22. In Paragraph 19, The above bio module is, When the above buffer solution is administered onto the carbon thin film, When the concentration of the target substance in the body fluid mixed in the above buffer solution is within the normal range, an electrical signal exceeding the threshold is generated, and A bio module for detecting a target substance in a body fluid, characterized by generating an electrical signal below the threshold when the concentration of the target substance in the body fluid mixed in the buffer solution is within an abnormal range.

23. In Paragraph 19, The above bio module is, When the above buffer solution is administered onto the carbon thin film, The higher the concentration of the target substance in the body fluid mixed in the buffer solution, the smaller the size of the electrical signal generated, and A bio-module for detecting a target substance in a body fluid, characterized in that the lower the concentration of the target substance in the body fluid mixed in the buffer solution, the larger the electrical signal generated.

24. In Paragraph 19, The above target substance and the above fixation substance are hormones, and A biomodule for detecting a target substance in body fluids, wherein the target binding substance is at least one selected from antibodies, aptamers, and receptors that specifically bind to the hormone.

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