Biosensor and method of regenerating recognition molecules

Abstract

To provide a biosensor which can be repetitively and continuously used and enables further miniaturization, and to provide a method of regenerating recognition molecules in the biosensor.SOLUTION: A biosensor 1 is provided, comprising a flow channel 2 for causing a sample solution 120 containing a biomarker 110 to flow, a sensor unit 3 disposed in the flow channel 2 and provided with recognition molecules 32, a first electrode 4 and a second electrode 5 arranged at a distance from each other in the flow channel 2, current application means 6 for applying current to the first electrode 4 and the second electrode 5, and detection means 7 for detecting the biomarker 110 captured by the recognition molecules 32.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to a biosensor and a method for regenerating a recognition molecule in the biosensor.

Background Art

[0002] As a wearable terminal for realizing advanced preventive medicine, the development of a smartwatch or the like incorporating a biosensor has been underway. A biosensor is a sensor that uses a recognition molecule (antibody, enzyme, aptamer, etc.) that specifically binds to a specific biomarker. A biosensor detects a specific protein in blood or the like using the intermolecular interaction between the biomarker and the recognition molecule. In a biosensor, a first recognition molecule is previously immobilized on a substrate. Then, a sample such as a liquid containing a biomarker is flowed through to capture the biomarker with the first recognition molecule. Further, a second recognition molecule modified with a labeling substance is bound to the biomarker to sandwich the biomarker between the first recognition molecule and the second recognition molecule. In this state, the specific biomarker in the sample is quantified by measuring the absorbance, fluorescence intensity, mass change of the second recognition molecule (label), the absorbance of the reaction product of the labeling substance (enzyme) and the substrate, the oxidation-reduction current, or the like.

[0003] Examples of biomarkers detected by a wearable terminal incorporating a biosensor include lifestyle disease markers. Examples of tumor markers suggesting the presence of cancer include CEA, AFP (α-fetoprotein), and the like. Examples of biomarkers suggesting heart disease include oxidized LDL, C-reactive protein (CRP), and the like. Examples of biomarkers suggesting dementia include amyloid-β and the like.

[0004] Currently, in Japan, which has become a super-aged society, the curative medical system has reached its limits. Therefore, the construction of a healthy and long-lived society, the prevention of lifestyle-related diseases, and the shift away from curative medicine are desired. As a means to solve such problems, a wearable terminal incorporating a biosensor that can perform daily health management of the user (wearer) is considered effective.

[0005] Typical biosensors include, for example, sensors using ELISA (a method for quantifying trace biological substances using antigen-antibody reactions), sensors using immunochromatography such as pregnancy test kits, sensors using surface plasmon resonance (SPR) (see, for example, Non-Patent Document 1), sensors using enzymes, aptamers, and the like.

Prior Art Documents

Non-Patent Documents

[0006]

Non-Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0007] Conventional biosensors are basically disposable and cannot be used continuously. This is because after detecting a biomarker, in order to dissociate the biomarker from the recognition molecule and regenerate the biosensor, it is necessary to pump a regeneration solution (such as an acid or an alkali) into the biosensor using a liquid delivery pump and a valve. Such a method for dissociating a biomarker promotes the complication and enlargement of the biosensor and also causes the consumption of reagents (the need for replacement). That is, the conventional regeneration method has inhibited the miniaturization of biosensors.

[0008] The present invention has been made in view of the above circumstances, and an object thereof is to provide a biosensor that can be repeatedly and continuously used and can be further miniaturized, and a method for regenerating a recognition molecule in the biosensor.

Means for Solving the Problems

[0009] The present invention has the following aspects. [1] A flow path through which a sample solution containing a biomarker flows, A sensor unit disposed in the flow path and having a recognition molecule, A first electrode and a second electrode disposed apart from each other in the flow path, Current application means for applying a current to the first electrode and the second electrode, Detection means for detecting a biomarker captured by the recognition molecule, a biosensor comprising the same. [2] The biosensor according to [1], further comprising a pH detection unit for detecting the pH of the sample solution present in the region including the first electrode and its periphery. [3] A method for regenerating a recognition molecule in the biosensor according to [1] or [2], A method for regenerating a recognition molecule, wherein a current is applied to the first electrode and the second electrode by the current application means to control the pH of the sample solution present in the region including the first electrode and its periphery. [4] The method for regenerating a recognition molecule according to [3], wherein an antibody is used as the recognition molecule. [5] The method for regenerating a recognition molecule according to [3] or [4], wherein the pH of the sample solution present in the region including the first electrode and its periphery is controlled to be 1 or more and 3.5 or less. [6] The method for regenerating a recognition molecule according to [3] or [4], wherein the pH of the sample solution present in the region including the first electrode and its periphery is controlled to be 10 or more. [Advantages of the Invention]

[0010] According to the present invention, it is possible to provide a biosensor that can be repeatedly and continuously used and can be further miniaturized, and a method for regenerating a recognition molecule in the biosensor. [Brief Description of the Drawings]

[0011]

Figure 1

Figure 2

[0012] Embodiments of the biosensor of the present invention and a method for regenerating a recognition molecule in the biosensor will be described. Note that this embodiment is specifically described to better understand the gist of the invention, and does not limit the present invention unless otherwise specified.

[0013] [Biosensor] FIG. 1 shows a schematic configuration of a biosensor according to an embodiment of the present invention, and is a schematic diagram showing a cross section along the height direction of the biosensor. As shown in Fig. 1, the biosensor 1 of the present embodiment includes a flow path 2, a sensor unit 3, a first electrode 4, a second electrode 5, a current application means 6, and a detection means 7. The biosensor 1 of the present embodiment may include a pH detection unit 8. The sensor unit 3 has a substrate 31 and a recognition molecule 32 fixed to the surface 31a of the substrate 31. Note that the sensor unit 3 may be composed only of the recognition molecule 32 fixed to the inner surface 2a of the flow path 2. In addition, the biosensor 1 of the present embodiment may include a substrate 9 that supports the flow path 2.

[0014] The flow path 2 is a flow path through which a sample solution 120 containing a biomarker 110 flows.

[0015] The first electrode 4 and the second electrode 5 are arranged to be separated from each other in the flow path 2. The first electrode 4 and the second electrode 5 are arranged, for example, as shown in Fig. 1, in the flow path 2 so as to sandwich the sensor unit 3 along the flow direction of the sample solution 120 (the length direction of the flow path 2). Further, the first electrode 4 and the second electrode 5 may be arranged in the flow path 2 so as to sandwich the sensor unit 3 along the direction perpendicular to the flow direction of the sample solution 120 (the inner circumferential direction of the flow path 2). Further, the substrate 31 of the sensor unit 3 may be the first electrode 4. When the first electrode 4 and the second electrode 5 are arranged so as to sandwich the sensor unit 3 and when the substrate 31 of the sensor unit 3 is the first electrode 4, there is almost no difference in the regeneration ability of the recognition molecule 32 by the biosensor 1.

[0016] The current application means 6 is connected to the first electrode 4 and the second electrode 5, and applies a current with the first electrode 4 as an anode to these two electrodes to oxidize chloride ions and hydroxide ions in the sample solution 120 on the surface 4a of the first electrode 4, generating chlorine and oxygen, and generating a high-concentration hydrogen ion (H + ) region near the surface 4a of the first electrode 4, or applying a current with the first electrode 4 as a cathode to reduce hydrogen ions in the sample solution 120 on the surface 4a of the first electrode 4, generating hydrogen, and generating a high-concentration hydroxide ion (OH - ) region near the surface 4a of the first electrode 4.

[0017] The detection means 7 detects the biomarker 110 captured by the recognition molecule 32.

[0018] The pH detection unit 8 detects the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery. The pH detection unit 8 is connected to the reading unit 10.

[0019] [Flow path] The flow path 2 is not particularly limited as long as it can flow the sample solution 120 containing the biomarker 110. For example, it is preferably a microchannel formed in the flow path substrate 11 by a hydrogel molding method, a microfabrication method such as photolithography, soft lithography, cutting, bonding, or a method combining these microfabrication methods. The microchannel is a channel with an inner diameter (maximum diameter) on the order of nanometers to millimeters. By the flow path 2 being a microchannel, the biomarker 110 can be detected with high sensitivity by the biosensor 1. Also, by the regeneration method described later, the biomarker 110 bound to the recognition molecule 32 can be easily dissociated. Examples of the flow path substrate 11 include silicone rubber, various resins other than silicone rubber, glass, and the like.

[0020] [First electrode] The first electrode 4 is not particularly limited as long as it is not deteriorated by the sample solution 120 and ions such as hydrogen ions and hydroxide ions, and in some cases, the recognition molecule 32 can be immobilized on its surface 4a. Examples of the first electrode 4 include metal electrodes made of metals such as gold and platinum, and carbon-based electrodes. Examples of the carbon-based electrodes include electrodes printed with carbon ink, glassy carbon electrodes, boron-doped diamond electrodes, and the like.

[0021] [Second electrode] The second electrode 5 is not particularly limited as long as it is not deteriorated by the sample solution 120 and ions such as hydrogen ions and hydroxide ions. Examples of the second electrode 5 include metal electrodes made of metals such as gold and platinum, and carbon-based electrodes. Examples of the carbon-based electrodes include electrodes printed with carbon ink, glassy carbon electrodes, boron-doped diamond electrodes, and the like.

[0022] [Current application means] The current application means 6 is not particularly limited as long as it can apply a current to the first electrode 4 and the second electrode 5. Examples of the current application means 6 include a current generator, a voltage-current generator, and the like.

[0023] [Substrate] The substrate 31 is not particularly limited as long as the recognition molecule 32 can be immobilized on its surface 31a and it is not deteriorated by the sample solution 120 and ions such as hydrogen ions and hydroxide ions. Examples of the substrate 31 include metal films made of metals such as gold and platinum, and carbon films.

[0024] [Recognition molecule] The recognition molecule 32 is not particularly limited as long as it can capture the biomarker 110 contained in the sample solution 120, and examples thereof include antibodies, enzymes, aptamers, and the like.

[0025] [Detection means] The detection means 7 is selected according to the use of the biosensor 1, that is, the type of the biomarker 110 to be detected by the biosensor 1. To detect the biomarker 110, a method of binding a second recognition molecule modified with a labeling substance to the biomarker 110 captured by the recognition molecule (first recognition molecule) 32 or a method that does not use the second recognition molecule modified with the labeling substance is used. In the method using the second recognition molecule, as the detection means 7, a means capable of performing a detection method corresponding to the second recognition molecule is used. Examples of the detection means 7 include a device for measuring fluorescence emitted by the second recognition molecule (label), a device for measuring the absorbance of a substance generated by the reaction of the second recognition molecule (label) and a substrate, a device for measuring the redox current of a substance generated by the reaction of the second recognition molecule (label) and a substrate, and the like. As a method that does not use the second recognition molecule, that is, a method for directly measuring the amount of the biomarker 110 bound to the recognition molecule 32, for example, a surface plasmon resonance method, a quartz crystal microbalance method, an electrochemical impedance measurement method, a surface acoustic wave measurement method, and the like can be mentioned. In FIG. 1, the case of directly measuring the amount of the biomarker 110 bound to the recognition molecule 32 by the surface plasmon resonance method is illustrated. In this case, the detection means 7 includes a light source 71 that irradiates the sensor unit 3 with white light, and a detector 72 that detects the amount by which the wavelength of the light attenuated by surface plasmon resonance changes due to the binding of the biomarker 110 and the recognition molecule 32.

[0026] [pH detection unit] Examples of the pH detection unit 8 include a pH meter and the like. By providing the pH detection unit 8, it is not necessary to use a pH indicator to detect the pH of the sample solution 120. Further, by providing the pH detection unit 8, the pH of the sample solution 120 can be detected immediately, and the application of current to the first electrode 4 and the second electrode 5 by the current application means 6 can be adjusted to more easily control the pH of the sample solution 120.

[0027] [Substrate] The substrate 9 is not particularly limited as long as a flow path substrate 11 can be provided on one surface 9a thereof. Examples include a glass substrate, a resin substrate, and the like. When the sensor unit 3 is composed only of the recognition molecule 32 fixed to the inner surface 2a of the flow path 2 (the inner surface 11a of the flow path substrate 11), the flow path substrate 11 is not particularly limited as long as the recognition molecule 32 can be immobilized on the inner surface 11a and it is not deteriorated by the sample solution 120 and ions such as hydrogen ions and hydroxide ions.

[0028] [Biomarker] The biomarker 110 is not particularly limited. For example, it includes tumor markers suggesting the presence of cancer such as CEA and AFP (α-fetoprotein), biomarkers suggesting heart disease such as oxidized LDL and C-reactive protein (CRP), biomarkers suggesting dementia such as amyloid β, and the like.

[0029] [Sample solution] The sample solution 120 is a solution containing the biomarker 110. Specifically, the sample solution 120 includes body fluids such as blood, interstitial fluid, sweat, and saliva, and physiological saline such as phosphate-buffered saline and Tris-buffered saline.

[0030] [Method of using biosensor] The method of using the biosensor 1 of the present embodiment will be described.

[0031] A sample solution 120 containing the biomarker 110 is flowed through the flow path 2. When the sample solution 120 is flowed through the flow path 2, the biomarker 110 contained in the sample solution 120 is captured by the recognition molecule 32 of the sensor unit 3. Next, the biomarker 110 is qualitatively or quantitatively analyzed by the detection means 7. Specifically, white light is irradiated from the light source 71 to the sensor unit 3, and the detector 72 detects the amount by which the wavelength of the light attenuated by surface plasmon resonance changes due to the binding of the biomarker 110 and the recognition molecule 32, thereby qualitatively or quantitatively analyzing the biomarker 110. After the detection of the biomarker 110 is completed, a current is applied by the current application means 6 with the first electrode 4 as the anode and the second electrode 5 as the cathode, and a high-concentration hydrogen ion (H + ) region is generated in the vicinity of the surface 4a of the first electrode 4. As a result, in the flow path 2, the pH of the sample solution 120 existing in the region α including the first electrode 4 and its periphery becomes less than 7 (acidic). Further, due to the flow of the sample solution 120 flowing through the flow path 2, the hydrogen ions (H +) is caused to flow in the 32 directions of the recognition molecule. As a result, the biomarker 110 captured by the recognition molecule 32 dissociates due to the acidic sample solution 120, and the recognition molecule 32 is regenerated to a state where the biomarker 110 is not bound. Thereby, it becomes possible to capture the biomarker 110 contained in the sample solution 120 by the recognition molecule 32 again.

[0032] According to the biosensor 1 of the present embodiment, by the current application means 6, a current is applied with the first electrode 4 as the anode and the second electrode 5 as the cathode, and the pH of the sample solution 120 existing in the region α including the first electrode 4 and its periphery can be made less than 7. As a result, the biomarker 110 captured by the recognition molecule 32 can be dissociated by the acidic sample solution 120, and the recognition molecule 32 can be regenerated to a state where the biomarker 110 is not bound. Further, according to the biosensor 1 of the present embodiment, by the current application means 6, a current is applied with the first electrode 4 as the cathode and the second electrode 5 as the anode, and in the flow path 2, the pH of the sample solution 120 existing in the region α including the first electrode 4 and its periphery can be made greater than 7. As a result, the biomarker 110 captured by the recognition molecule 32 can be dissociated by the alkaline sample solution 120, and the recognition molecule 32 can be regenerated to a state where the biomarker 110 is not bound. Further, unlike the conventional method, in order to regenerate the recognition molecule 32, it is not necessary to feed a regeneration solution such as an acid or an alkali into the flow path 2 where the recognition molecule 32 is disposed using a liquid feed pump or a valve. Therefore, the regeneration of the recognition molecule 32 can be easily performed. Therefore, the biosensor 1 can be used repeatedly and continuously without being disposable. In addition, since the recognition molecule 32 can be regenerated without using large-scale equipment, the biosensor 1 can be miniaturized.

[0033] [Method for regenerating recognition molecule] The method for regenerating a recognition molecule according to an embodiment of the present invention is the method for regenerating a recognition molecule in the biosensor according to an embodiment of the present invention described above.

[0034] "First regeneration method" The method for regenerating the recognition molecule according to this embodiment is, for example, in the biosensor 1, by the current application means 6, applying a current with the first electrode 4 as the anode and the second electrode 5 as the cathode to generate a high-concentration hydrogen ion region in the vicinity of the surface 4a of the first electrode 4, and controlling the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery in the flow path 2 to be less than 7.

[0035] The current applied to the first electrode 4 and the second electrode 5 is not particularly limited, but for example, it is preferably 30 μA or more and 1 mA or less, and more preferably 50 μA or more and 200 μA or less. When the current is less than 30 μA, the reaction amounts of chloride ions and oxygen ions are small, and it becomes difficult to manipulate the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery. When the current exceeds 1 mA, the amount of gas generated from the vicinity of the first electrode 4 increases or the pH becomes too low, increasing the risk of damaging the recognition molecule 32.

[0036] The flow rate of the sample solution 120 flowing in the flow path 2 is not particularly limited, but for example, it is preferably 1 μl / min or more and 1 ml / min or less, and more preferably 10 μl / min or more and 200 μl / min or less.

[0037] In the flow path 2, it is preferable to control the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery to be 1 to 3.5, and more preferably 2 to 3. By controlling the pH of the sample solution 120 to be 1 to 3.5, dissociation of the biomarker 110 from the recognition molecule 32 can be further promoted.

[0038] The sample solution 120 may contain a pH indicator. The pH indicator is not particularly limited as long as it can indicate the range from alkaline (pH 14) to acidic (pH 1). Examples thereof include thymol blue, methyl red, phenolphthalein, etc. Thymol blue shows yellow in the neutral state and changes to red in the acidic state. Thymol blue shows yellow at pH 3, and the closer it is to red, the lower the pH and the higher the acidity. If the sample solution 120 contains a pH indicator, the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery can be confirmed by applying current to the first electrode 4 and the second electrode 5. Based on the pH indicator, it is possible to determine whether to continue applying current to the first electrode 4 and the second electrode 5 or to stop applying current to the first electrode 4 and the second electrode 5.

[0039] Further, the biosensor 1 may include a pH detection unit 8 such as a pH meter that detects the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery. If the pH detection unit 8 is provided, the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery can be confirmed by applying current to the first electrode 4 and the second electrode 5. Based on the pH detection unit 8, it is possible to immediately determine whether to continue applying current to the first electrode 4 and the second electrode 5 or to stop applying current to the first electrode 4 and the second electrode 5.

[0040] According to the method for regenerating a recognition molecule of the present embodiment, by applying a current to the first electrode 4 and the second electrode 5 to control the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery to less than 7, the biomarker 110 captured by the recognition molecule 32 can be dissociated, and the recognition molecule 32 can be regenerated to a state where the biomarker 110 is not bound. Further, unlike the conventional method, in order to regenerate the recognition molecule 32, it is not necessary to feed a regeneration solution such as an acid or an alkali into the flow path 2 in which the recognition molecule 32 is disposed using a liquid feed pump or a valve. Therefore, the recognition molecule 32 can be easily regenerated. Accordingly, the biosensor 1 can be used repeatedly and continuously. In addition, the recognition molecule 32 can be regenerated without using large-scale equipment.

[0041] "Second regeneration method" In the method for regenerating a recognition molecule of the present embodiment, for example, in the biosensor 1, a current is applied by the current application means 6 with the first electrode 4 as the anode and the second electrode 5 as the cathode, and a high-concentration hydrogen ion region is generated in the vicinity of the surface 4a of the first electrode 4, and in the flow path 2, the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery is controlled to less than 7. In the method for regenerating a recognition molecule of the present embodiment, in the biosensor 1, a current is applied by the current application means 6 with the first electrode 4 as the cathode and the second electrode 5 as the anode, and a high-concentration hydroxide ion region is generated in the vicinity of the surface 5a of the second electrode 5, and in the flow path 2, the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery is controlled to more than 7.

[0042] The current applied to the first electrode 4 and the second electrode 5 is not particularly limited, but for example, it is preferably 30 μA or more and 1 mA or less, and more preferably 50 μA or more and 200 μA or less. If the current is less than 30 μA, the reaction amount of hydrogen ions is small, and it becomes difficult to sufficiently control the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery. If the current exceeds 1 mA, the amount of gas generated from the vicinity of the first electrode 4 increases or the pH becomes too high, increasing the risk of damaging the recognition molecule 32.

[0043] The flow rate of the sample solution 120 flowing in the flow path 2 is not particularly limited, but for example, it is preferably 1 μl / min or more and 1 ml / min or less, and more preferably 10 μl / min or more and 200 μl / min or less.

[0044] In the flow path 2, it is preferable to control the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery to 10 or more, more preferably to 10 - 12, and even more preferably to 11 - 12. By controlling the pH of the sample solution 120 to 10 or more and 12 or less, dissociation of the biomarker 110 from the recognition molecule 32 can be further promoted.

[0045] The sample solution 120 may contain a pH indicator. The pH indicator is not particularly limited as long as it can indicate the range from alkaline (pH 14) to acidic (pH 1). For example, alizarin yellow, thymol blue, methyl red, phenolphthalein, etc. can be mentioned. Alizarin yellow shows yellow in neutral and changes to orange in alkaline. Alizarin yellow shows yellow at pH 10, and the closer it is to orange, the higher the pH and the higher the alkalinity. If the sample solution 120 contains a pH indicator, the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery can be confirmed by applying current to the first electrode 4 and the second electrode 5. It is possible to judge whether to continue applying current to the first electrode 4 and the second electrode 5 or to stop applying current to the first electrode 4 and the second electrode 5 based on the pH indicator.

[0046] Also in the second regeneration method, similar to the second regeneration method, the biosensor 1 may be provided with a pH detection unit 8 such as a pH meter that detects the pH of the sample solution 120 present in the region α including the first electrode 4 and its periphery.

[0047] According to the method for regenerating the recognition molecule of the present embodiment, by applying a current to the first electrode 4 and the second electrode 5 to control the pH of the sample solution 120 existing in the region α including the first electrode 4 and its periphery to be greater than 7, the biomarker 110 captured by the recognition molecule 32 can be dissociated, and the recognition molecule 32 can be regenerated to a state where the biomarker 110 is not bound. Also, unlike the conventional method, there is no need to feed a regeneration solution such as an acid or an alkali into the flow path 2 where the recognition molecule 32 is disposed by using a liquid feed pump or a valve in order to regenerate the recognition molecule 32. Therefore, the regeneration of the recognition molecule 32 can be easily performed. Accordingly, the biosensor 1 can be repeatedly and continuously used. Also, the recognition molecule 32 can be regenerated without using large-scale equipment.

Example

[0048] Hereinafter, the present invention will be described more specifically by way of examples, but the present invention is not limited to the following examples.

[0049] [Example] Using the biosensor shown in FIG. 1, while flowing a sample solution in the flow path, a current was applied to the first electrode and the second electrode, and the change in the pH of the sample solution was measured. A pH meter was used to measure the pH of the sample solution. As the sample solution, a 0.9% aqueous sodium chloride solution was used. The flow rate of the sample solution was set to 10 μl / min to 20 μl / min. The current applied to the first electrode and the second electrode was set to 50 μA to 200 μA. The results are shown in FIG. 2. From the results shown in FIG. 2, it was confirmed that the pH of the sample solution can be controlled to 2 to 3, which is effective for the regeneration of the recognition molecule.

Industrial Applicability

[0050] Since the biosensor of the present invention can be repeatedly and continuously used, the biosensor can be miniaturized, and a wearable terminal incorporating the biosensor can be realized.

Explanation of Reference Numerals

[0051] 1 Biosensor 2 Flow path 3 Sensor section 4 First electrode 5 Second electrode 6 Current application means 7 Detection means 8 pH detection section 9 Substrate 10 Reading section 11 Flow path substrate 32 Recognition molecule 110 Biomarker 120 Sample solution

Claims

1. a flow path for flowing a sample solution containing a biomarker; a sensor portion disposed in the flow channel and having a recognition molecule; a first electrode and a second electrode spaced apart from each other within the flow channel; a current applying means for applying a current to the first electrode and the second electrode; A detection means for detecting the biomarker captured by the recognition molecule.

2. The biosensor according to claim 1 , further comprising a pH detection unit that detects a pH of the sample solution present in a region including the first electrode and its periphery.

3. A method for regenerating a recognition molecule in a biosensor according to claim 1 or 2, comprising the steps of: A method for regenerating a recognition molecule, comprising applying a current to the first electrode and the second electrode by the current application means, and controlling the pH of the sample solution present in a region including the first electrode and its periphery within the flow channel.

4. 4. The method for regenerating a recognition molecule according to claim 3, wherein an antibody is used as the recognition molecule.

5. 4. The method for regenerating a recognition molecule according to claim 3, wherein the pH of the sample solution present in the region including the first electrode and its periphery is controlled to be 1 or more and 3.5 or less.

6. 4. The method for regenerating a recognition molecule according to claim 3, wherein the pH of the sample solution present in the region including the first electrode and its periphery is controlled to 10 or higher.

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