JP2012211819A - Biosensor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a biosensor which has a compact structure so that a user can always carry the biosensor, can largely reduce probability that false positive occurs due to impurities, and can shorten inspection time when measurement of a low concentration sample is performed.SOLUTION: Fine particles 107 each having a capture carrier which is bonded to a surface of the particle and specifically adsorbs a specific target molecule are dispersed in a solution. There is formed, on a substrate, a concave groove having a fixed width, length, and depth where the solution is located on a surface. The capture carriers are bound to surfaces of at least a pair of facing electrodes exposed inside the concave groove. Molecule measurement means measures target molecular weight in a liquid sample according to the change of an electrical connection state of the pair of the electrodes.

Description

  The present invention relates to a biosensor capable of detecting the presence of a substance group containing a specific target molecule using a capture carrier having a property of specifically adsorbing the specific target molecule.

  In recent years, there have been various activities such as terrorism using explosives and firearms, distribution of prohibited drugs such as stimulants and narcotics, outbreaks of new influenza and unknown infectious agents, food contamination, air pollution, water pollution, etc. The existence of threats threatens safe and secure social life.

  Sensing technology targeting substances and molecules that are the source of these threats is indispensable for the formation of a safe and secure society. In particular, there is a demand for the development of inexpensive and simple methods and sensing devices that anyone can use easily.

  Currently, X-ray fluorescence analysis, mass spectrometer (MS: MASSSPECTROMETRY), ion mobility spectrometry (IMS: IONMOBILITY SPECTROMETRY), gas chromatography (GC: GASCHROMATOGRAPHY), liquid chromatography (GC) There are various gas sensors such as LC (LIQUIDCHROMATOGRAPHY), sintered semiconductor type, constant potential electrolytic type, galvanic type, infrared absorption type.

  In addition, chemiluminescence method using chemical reaction, chemical staining method, immunochromatography using bioreactive substances such as antibody, enzyme, aptamer, enzyme immunoassay (ELISA) method, surface plasmon There are various methods such as resonance (SPR: SURF ACEPLASMONRESONANCE) method, quartz crystal microbalance (QCM: QUARTZCRYSTALM ICROBALANCE) method, surface acoustic wave (SAW) sensor, and bioluminescence method.

  Therefore, it is not difficult to adopt the most suitable method according to the target substance to be detected. However, when using it as a sensor, in order to respond to various application aspects, the viewpoint of portability that can be carried at all times and can be sensed by a device as compact as possible is also a very important factor.

  From this point of view, techniques generally called physical detection methods such as X-ray fluorescence, MS, GC, and LC generally have high molecular recognition ability and high sensitivity. However, the range of applications is limited in that the apparatus is of a desktop size, has low portability, is expensive, and takes time for analysis.

  On the other hand, various gas sensors, immunochromatography, chemical staining / luminescence methods, bioluminescence methods, and the like can produce highly portable sensing devices and have been used in various places.

  For example, various gas sensors, such as gas leak detectors and oxygen concentration meters, are widely used. In chemical substance detection methods using chemical reactions, for example, a mechanism that detects a target molecular substance using a drug that reacts with a specific functional group and emits light, such as a luminol reaction, can be used to simplify the apparatus. is there. Some devices using a bioluminescence mechanism using a specific reaction between luciferin and luciferase are compact and have high sensitivity.

  Immunochromatography can detect the target substance with a very simple procedure by specializing target molecules such as influenza virus and preparing the corresponding antibodies for detection in advance. It is.

  Specifically, two types of target substance antibodies are prepared, one of which is fixed to a specific region of the substrate, and the other is modified with a coloring label such as gold fine particles. And when the mixed solution of the sample solution that seems to contain the target substance and the colloidal gold labeled antibody is spread on the substrate, the target substance is sandwiched in a specific area of the substrate on which the antibody is fixed, Gold colloid is fixed.

  As a result, a change in color accumulates on the substrate due to the light absorption of the colloidal gold, and the content of the target substance is estimated based on the change. In addition, by changing the antibody used, there is an advantage that it can cope with various target molecules.

  There is also a proposal of a method for detecting a giant biopolymer using an electrode structure. The technique includes an electrode structure having first and second electrodes, and a first electrical measurement is performed at the electrodes.

  In other steps, the solution to be investigated is brought into contact with the electrode structure. At this time, the solution may contain a giant biopolymer to be detected. In yet another step, the macrobiopolymer to be detected contained in the solution under investigation binds to the capture molecules on the first and second electrodes, and the electrode structure binds to the macrobiopolymer and A reagent for increasing the conductivity of a macromolecular biomolecule that imparts mechanical conductivity is contacted. Thereafter, the second electrical measurement is performed on the electrode, and the result of the two electrical measurements is compared on the electrode to detect the giant biopolymer (Patent Document 1).

  There is also a proposal of a biosensor cell in which each biosensor cell includes a sensing zone and a biosensor array including a plurality of biosensor cells. In that technique, a first sensing electrode, a second sensing electrode, and a gap separating the sensing electrodes are arranged in the sensing zone.

  The first sensing electrode is electrically isolated from the second sensing electrode using a gap. The capture molecule is immobilized within the sensing zone, the field effect transistor has a gate electrode, a source electrode, and a drain electrode, the first sensing electrode is electrically connected to the gate electrode of the field effect transistor, The second sensing electrode can be electrically connected to the gate voltage (Patent Document 2).

JP-T-2004-524534 Special table 2009-524046 gazette

  Immunochromatography is highly portable and has great advantages in terms of sensing methods that can handle a variety of target molecules, but on the other hand, it is inferior in terms of sensitivity and quantitativeness compared to ELISA and SPR methods, and has high detection. There are methods for obtaining sensitivity, but there are problems such as an increase in inspection time and operation procedures, and a lack of knowledge other than limited target substances.

  In addition, the test time is sufficiently short compared to ELISA and the like, which is one of the advantages of immunochromatography, but detection in a shorter time may be required, so there are still problems in this respect. It can be said that.

  To describe each problem in detail, the sensitivity is usually inferior to that of an ELISA having a signal amplification function and an SPR using a resonance effect because the color development (light J absorption) by a gold colloid is usually observed visually. . Further, there is a limit to the quantitativeness in visual observation.

  Therefore, as a method for improving the sensitivity, a method for improving the sensitivity by reacting a sensitizer with a metal colloid as a catalyst has been proposed. A method for quantifying the amount of colloidal gold adsorbed optically using a laser or the like has also been proposed.

  However, in any method, in addition to the conventional immunochromatography measurement kit, in addition to the operation procedure, an error factor increases and a new measuring instrument is required. Since the advantage of simplicity is greatly impaired, it is not widely used.

  Next, regarding the feeling at the time of measurement, in immunochromatography, a strip-shaped fibrous material having a width of about 5 MM is usually used as a substrate, and a liquid sample is transported through the gap inside the fibrous material using capillary action. It takes a longer period of 15 to 30 minutes to make an important decision in actual operations, such as whether it takes at least about 5 minutes, or whether the test object is negative or positive because of its low concentration.

  One of the major problems of immunochromatography is to further shorten the examination time. As a method for improving measurement sensitivity and measurement time, immunochromatography using an electrical detection method has been studied.

  There has also been proposed a method for detecting a molecule by a change in conductivity between electrodes caused by crosslinking between two electrodes separated by a certain gap using a biological macromolecule such as DNA. There has also been proposed a method of quantifying target molecules by preparing two comb electrodes facing each other and measuring a change in conductivity therebetween.

  An advantage of the method using these electrodes is an improvement in quantitativeness. The sensitivity is basically the same as that of conventional immunochromatography, but it can be expected that the substantial minimum detection limit is lowered, that is, the sensitivity is improved by improving the quantitativeness.

  However, one of the problems to be solved in these methods is that they are easily affected by impurities. In both of the above methods, the distance between the electrodes is short and the surface is exposed, so even a very small particle of about 1 UM contained in the sample solution can cause a short circuit between the electrodes and easily cause false positives. End up. Therefore, as a result, no reliability can be obtained at the practical inspection stage.

  Another problem is the problem of downsizing the sensor chip and measuring time. In electrochromatography, which is electrically evaluated, a flat substrate with patterned electrodes is usually used, but in terms of surface area around the volume of the substrate when mounted as a kit, it is compared with the fibrous substrate used in the conventional method for visual evaluation. And greatly inferior.

  Therefore, when an equivalent surface area is prepared for performing an immunoassay, the size of the flat substrate becomes very large. If the size of the substrate is limited, sensitivity and measurement time must be sacrificed.

  The present invention has been made in view of the above-mentioned problems, and has a compact structure that can be carried at all times, can greatly reduce the probability of false positives caused by contaminants, and can be used for low-concentration sample measurement. A biosensor capable of reducing time is provided.

  The biosensor of the present invention includes a solution in which fine particles in which a capture carrier that specifically adsorbs a specific target molecule is bound to a surface are dispersed, and a groove having a certain width, depth, and depth at which the solution is located on the surface. A liquid sample in accordance with a change in the electrical coupling state of the pair of electrodes, a base that is formed inside the groove, at least a pair of opposing electrodes that are exposed inside the concave groove and the capture carrier is coupled to the surface And a molecular measuring means for measuring the target molecular weight in the medium.

  In the biosensor of the present invention, fine particles in which a capture carrier that specifically adsorbs a specific target molecule is bound to the surface are dispersed in a solution. A concave groove having a certain width, depth and depth at which the solution is located on the surface is formed in the substrate. A capture carrier is coupled to the surfaces of at least a pair of opposing electrodes exposed inside the concave groove. The molecular measurement means measures the target molecular weight in the liquid sample according to the change in the electrical binding state of the pair of electrodes. For this reason, with a compact structure that can be carried at all times, the probability of false positives due to contaminants can be greatly reduced, and the inspection time when measuring a low-concentration sample can be shortened.

FIG. 2 shows a part of the manufacturing process of the biosensor according to the first embodiment of the present invention, in which (A) is a schematic front view and (B) is a schematic plan view. A part of manufacturing process of a biosensor is shown, (A) is a schematic front view, and (B) is a schematic plan view. The structure of the completed biosensor is shown, (A) is a schematic front view, and (B) is a schematic plan view. It is a typical front view which shows the state which adsorb | sucks a target molecule with a biosensor. It is a typical front view which shows the microscopic state which adsorb | sucks a target molecule with a biosensor. It is a typical top view which shows the structure of the biosensor of the 2nd form of implementation of this invention. It is a typical top view which shows the mounting method of the biosensor of the 2nd form of implementation of this invention.

  An embodiment of the present invention will be described below with reference to the drawings. As shown in FIGS. 3 and 4, the biosensor of the present embodiment includes a solution (not shown) in which fine particles 107 having a capture carrier that specifically adsorbs a specific target molecule bound to the surface are dispersed. A substrate 102 to 104 forming a groove having a constant width, depth and depth, and at least a pair of opposing comb pattern electrodes 101 exposed inside the groove and having a capture carrier bonded to the surface thereof. And a molecular measuring means for measuring the target molecular weight in the liquid sample in accordance with a change in the electrical coupling state of the pair of comb pattern electrodes 101.

  The biosensor of the present embodiment is a molecule (capture molecule) serving as a capture carrier having a property of specifically adsorbing a specific target molecule (antigen specificity), particularly a protein, hapten, nucleic acid aptamer, receptor, etc. The biologically derived capture molecule is used.

  In particular, the presence of target molecules in a liquid sample using fine particles 107 dispersed in a solution with capture molecules bound to the surface and two opposed comb-shaped pattern electrodes 101 with capture molecules bound to the surface. The presence / absence, content, and the like are detected by a change in the electrical coupling state between the two opposing comb pattern electrodes 101.

  As the capture molecule, a molecule having a molecular recognition function that is used in gene information transmission performed inside a living body, signal transmission inside and outside a cell, nerve information transmission, immune system, and the like can be applied. For example, a G protein-coupled receptor or an immunoglobulin can be used, and an amino acid sequence such as a molecular recognition site can be modified as necessary.

  It is also possible to use a specially selected capture molecule by using an existing method for extracting only molecules having a property of specifically adsorbing to a specific molecule from nucleic acids and peptides. These capture molecules can be additionally modified with a functional group, amino acid sequence, base sequence, or the like for the purpose of effectively adsorbing or binding to the surface of the comb pattern electrode 101 or the surface of the fine particle 107.

  The material of the comb pattern electrode 101 can be a metal. Preferably, noble metals such as gold, platinum, silver, and palladium can be used. In particular, an alloy containing at least one of these noble metals can be used in order to increase the affinity with the thiol molecule used to facilitate the fixation of the capture molecule.

  In addition, it is possible to reduce the manufacturing cost of the structure of the comb pattern electrode 101 by using an organic material instead of a metal as the material of the comb pattern electrode 101. As the material of the comb pattern electrode 101, it is possible to use a carbon conductive material such as a carbon nanotube, polyacetylene, polythiophene, or the like that conducts conduction by a pi-conjugated electron network. Furthermore, it is preferable that the surface of the comb pattern electrode 101 made of an organic material is modified with a functional group used for facilitating fixation of the capture molecule.

  Each comb pattern electrode 101 is provided with a coating layer made of an insulating material so as to cover a part of the comb pattern electrode 101 in order to prevent a contaminant current caused by non-specific contact and short-circuiting of the impurities. As the insulating material, an oxide insulating film such as a silicon oxide film, an organic insulating film such as polyimide or parylene, an insulating self-alignment film, or the like can be used.

  Between the comb-shaped pattern electrodes 101 facing each other, a groove having a depth obtained by combining at least the thickness of the electrode layer and the thickness of the coating layer is generated. The lateral width of the concave groove is set to be equal to or larger than the diameter of the fine particles 107 having capture molecules bound to the surface, which will be described below. Further, the optimum adsorption efficiency of the fine particles 107 and the optimum effect of suppressing the contaminant current are obtained. You can choose the size you want.

  The depth of the groove is at least equal to the lateral width of the groove, preferably twice or more. It is also possible to etch the substrate in order to deepen the groove. As a result, particles larger than the gap size cannot physically enter the concave grooves between the opposing comb pattern electrodes 101.

  Therefore, as a result, the contaminant current can be suppressed, and the probability that a false positive occurs can be greatly reduced. In order to process the structure of the comb pattern electrode 101, semiconductor lithography can be used. On the other hand, the fine particles 107 are also used by binding capture molecules.

  As the capture molecules to be bound to the fine particles 107, those having the same properties as the capture molecules bound to the comb pattern electrode 101 and the specific substances and molecules to be measured are used. That is, the same capture molecule can be used, or two completely different types of capture molecules having specific binding forces at different sites of the molecule to be measured can be used.

  The fine particles 107 to which the capture molecules are bound are used after being adjusted to an optimum concentration and dispersed in the solution. At the time of measurement, first, it is confirmed that the comb-shaped pattern electrodes 101 facing each other are not electrically short-circuited before measurement, and are immersed in a standard solution for measurement. In this state, first, a constant voltage VDC is applied between the opposing comb pattern electrodes 101, and the initial value IDC1 of the current is recorded.

  Next, the sample to be actually measured and the solution in which the fine particles 107 are dispersed are added to the standard solution, and an adsorption reaction is performed. Stirring can be appropriately performed during the adsorption reaction. In some cases, a constant potential is applied to both of the opposing comb pattern electrodes 101 or a high frequency of several hundred KHZ or more is applied between the opposing comb pattern electrodes 101 to promote the adsorption reaction. Is possible.

  Conversely, the adsorption of the fine particles 107 can be suppressed, or the fine particles 107 that do not exhibit specific adsorption can be moved away from the comb pattern electrode 101. Then, by applying a constant voltage VDC again between the opposing comb-shaped pattern electrodes 101 and recording the current value IDC2 after processing, if the binding due to the antigen-antibody reaction occurs preferentially, the current change rate (IDC2− As IDC1) / IDC1, a significantly large value can be obtained.

  In addition, voltage application and current measurement are not limited to this method, but AC voltage and pulse voltage. Various measurement methods such as frequency sweep can be used.

  Next, the best mode for carrying out the present invention will be described in detail with reference to FIGS. FIG. 1 to FIG. 3 are schematic diagrams for explaining a manufacturing process of the comb-shaped pattern electrode 101 which is a basic configuration of a sensor for testing human chorionic gonadotropin (HCG) in urine used in the present embodiment.

  FIG. 1 schematically shows a cross-sectional view and a top view of a substrate in which a comb-shaped pattern electrode 101 is first prepared by a lift-off process or a milling process using a resist pattern as a mask. The cross-sectional view of the substrate corresponds to the cross section of the portion indicated by AB in the top view.

  The substrate is an SOI (SILICON ON INSULATOR) substrate having a silicon oxide layer 102 with a thickness of 20 NM on the surface, a silicon layer 103 with a thickness of 300 NM below it, and a buried silicon oxide layer 104 with a thickness of 50 NM below it. Was used.

  The comb pattern electrode 101 is obtained by patterning gold having a thickness of 20 NM produced by an electron beam evaporation method by milling using a resist pattern as a mask. The lateral width is 80 NM and the gap between the electrodes is 80 NM.

  Further, a positive resist is used for the electrode, and the region 105 on the comb pattern electrode shown in the top view is exposed and developed to open a window. Subsequently, the silicon oxide layer 102 in a region not covered with the resist defined in the region 105 is removed by using anisotropic reactive ion etching, and then the resist is removed.

  Then, using the comb pattern electrode 101 and the silicon oxide layer 102 as a mask, the exposed silicon layer is removed using anisotropic reactive ion etching, resulting in the structure shown in FIG.

  Finally, a 50 NM silicon oxide layer 106 is sputter-deposited as a coating film, leaving the contact portion of the comb-shaped pattern electrode 101, thereby obtaining a buried electrode structure exposed in the gap as shown in FIG.

  In this electrode structure, a plurality of opposing comb pattern electrodes 101 are arranged in parallel, and one of the opposing comb pattern electrodes 101 is electrically connected and grounded. The other of the opposing comb pattern electrodes 101 is electrically independent and has a structure connected to a circuit that performs arithmetic processing such as signal synthesis and difference.

  Next, a method for immobilizing capture molecules on the surface of the comb pattern electrode 101 will be described. First, the comb-shaped electrode surface is cleaned by oxygen plasma treatment, then immersed in 1MM carboxydecanethiol using ethanol as a solvent for 1 hour, and further immersed in 1MM hydroxyundecanethiol for 1 hour. Thus, the surface of the comb pattern electrode 101 is terminated with a carboxyl group.

  Next, after treatment with 0.1 M N-hydroxysuccinic acid aqueous solution for 15 minutes, anti-HCG-Α-mouse IGG adjusted to a concentration of 10 UG / ML in 10MM PH7 phosphate buffered saline (PBS) Was treated for 1 hour to immobilize anti-HCG-IV antibody on the surface of the comb pattern electrode 101. As a result of immobilizing the antibody, the thickness of the film containing the antibody or the like was about 10 NM, so that the gap between the electrodes was reduced from 80 NM to 20 NM to 60 NM.

  On the other hand, as the fine particles 107 with a capture molecule, a commercially available product in which an anti-HCG-IV antibody is similarly immobilized on a gold colloid having an average particle size of 40 NM is used. Similarly, a 10 MM PH7 PBS, a nonionic surfactant Was adjusted so that OD520 would be 0.1.

  At this time, since the thickness of the film containing the antibody on the gold colloid surface was about 10 NM, the average size of the fine particles 107 with a capture molecule was about 60 NM. This size is exactly the same as the effective gap 60 NM between electrodes, but since the actual structure of the capture molecule film is fluid, HCG is a protein having a size of about 4 NM as a target molecule. Is fixed in the gap without any problem.

  In addition, the size of the trapped molecule-attached fine particle 107 is D, the thickness of the capture molecule film 109 on the fine particle surface is D1, the thickness of the capture molecule film 110 on the electrode surface is D2, the size of the target molecule 111 is A, a comb shape When the distance between the electrodes of the pattern electrode 101 is L, when the size of each component is adjusted so that the relationship of L = 2 × (D1 + D2 + A) + D is established, the fine particles 107 face each other as shown in FIG. It is possible to realize a situation where the two electrode surfaces are fixed at two locations.

  When such a condition is satisfied, if the divergence multiplier between the electrode and the fine particle is defined, it is effectively smaller than the divergence multiplier when the fixation occurs at one place, that is, the divergence multiplier of the capture molecule with respect to the target molecule. Therefore, in the case of the same concentration, more fine particles are fixed and a large signal can be obtained rather than fixing at one place.

  In the biosensor of this embodiment, as described above, fine particles 107 in which a capture carrier that specifically adsorbs a specific target molecule is bound to the surface are dispersed in a solution. Concave grooves having a certain width, depth and depth at which the solution is located on the surface are formed by the substrates 102 to 104.

  A capture carrier is coupled to the surfaces of at least a pair of opposing electrodes exposed inside the concave groove. When a target molecule is present in the liquid sample, a change in the electrical binding state of the pair of electrodes occurs, and the molecular measurement means measures the target molecular weight in the liquid sample according to the amount of change and the response time. Alternatively, the molecular measurement means measures the target molecular weight in the liquid sample in accordance with the change in the number of electrodes in which a change in the electrical coupling state occurs or the response time.

  For this reason, with a compact structure that can be carried at all times, the probability of false positives due to contaminants can be greatly reduced, and the inspection time when measuring a low-concentration sample can be shortened.

  Subsequently, a biosensor according to a second embodiment of the present invention will be described below with reference to FIG. In the biosensor of the present embodiment, two sets of comb pattern electrodes 101 for HCG inspection are used as shown in the figure.

  One is an electrode set for evaluation 201 in which the anti-HCG-IV antibody is immobilized on the surface of the comb-shaped pattern electrode 101 by the above-described method, and the other is the bovine serum albumin (BSA) immobilized by the same method as described above. This is a reference electrode set 202.

  In each electrode set, one of the opposing comb pattern electrodes 101 is all grounded, and the other comb pattern electrode 101 is independently connected to the signal selection circuits 203 and 204. In the signal selection circuits 203 and 204, before the measurement solution is dropped onto each electrode set, first, the comb pattern electrodes 101 whose resistance to the ground side is equal to or less than a certain standard are selected, and the comb pattern electrodes 101 are selected. Is judged as a defective line, and the electrical connection is cut off using the signal selection circuits 203 and 204.

  Next, the gold colloid solution 1UL with the anti-HCG-ＨＣ antibody described above is dropped onto each of the electrode sets 201 and 202 as a measurement solution. Further, with the solution dropped, the ground resistance of each comb pattern electrode 101 is evaluated again every 10 seconds and recorded in the arithmetic recording circuit 205 as reference data.

  Subsequently, 1 UL of PBS of 10MM PH7 containing HCG at a concentration of 10NG / ML is added as a standard sample. As a result, the HCG is fixed so as to be sandwiched between the antibody on the comb pattern electrode 101 and the antibody on the gold colloid.

  At this time, as shown in the cross-sectional view of the comb pattern electrode 101 shown in FIG. 6, even if a conductive substance such as 108 larger than the gap between the 80 NM comb pattern electrodes 101 is mixed in the measurement solution. Since it cannot physically enter between the comb-shaped pattern electrodes 101, the influence of impurities is greatly reduced.

  On the other hand, when the colloid is added and the solution is added, the pH and electrolyte concentration of the entire measurement solution on the electrode set change, so that the entire comb pattern electrode 101 of the electrode set 201 for evaluation and the electrode set 202 for reference The ground resistance varies uniformly. A circuit for judging the shock due to the addition as the addition timing of the measurement sample is provided, and thereafter, the ground resistance of each comb pattern electrode 101 is measured every second for 2 minutes.

  When each comb-shaped pattern electrode 101 is evaluated with the change rate of the ground resistance before and after the addition, the ground resistance is less than a certain standard, the change rate is close to zero at the addition timing, and the ground resistance is a constant reference before the addition. As described above, although the absolute value of the rate of change gradually increases after the addition timing, it can be roughly classified into two groups.

  The arithmetic recording circuit 205 can obtain four groups of data, a low change rate group and a high change rate group, from the evaluation electrode set 201 and the reference electrode set 202, respectively. Thus, it is determined whether or not the number of signals from the electrode set 201 for evaluation that belong to the high change rate group and the change rate are significant, and the sample concentration is estimated from the average change rate. The estimation is performed in light of calibration data that has been measured in advance.

  FIG. 7 illustrates a cross section of the biosensor according to the second embodiment of the present invention mounted on a liquid sample measurement chip.

  The substrate 301 on which the electrode set of the second form was created was equipped with a connector 302 for electrical input / output after dicing, substrate cleaning, and antibody modification to the electrode.

  Further, a hydrophilic nonwoven fabric 303 on the sample introduction side and a hydrophilic nonwoven fabric 304 on the sample reservoir side processed so that one end is tapered and thinned are prepared, and the nonwoven fabric 303 on the sample introduction side is provided at the position illustrated in 305. The same probe fixing region 305 was prepared by dropping and drying the fine particles with capture molecules, which was the same as that used in the second embodiment of the present invention.

  The created electrode set 301 with a connector was formed with electrodes on the gap side so that a gap of about 15 mm was formed between the resin package 306 by sandwiching the nonwoven fabrics 302 and 303 as shown in FIG. It was fixed so that the surface was facing.

  Furthermore, by arranging the longitudinal direction of the groove of the electrode set 301 so as to be substantially parallel to the direction along the flow of the sample solution, the contact frequency between the electrode surface and the target molecule can be increased. Similarly, the frequency of contact between the electrode surface and the target molecule can be further increased by arranging the longitudinal direction of the concave groove at an angle of 45 degrees or less with respect to the direction of the solution flow.

  At the time of measurement, the prepared liquid sample measuring chip is connected to an electric circuit for measurement, and in the same manner as in the second embodiment of the present invention, after the defective line is cut off, the opening in the resin package 306 is opened. From the part 307, a liquid sample of about 30 L was introduced.

  The introduced liquid sample traveled through the introduction-side nonwoven fabric 303 by a capillary phenomenon, passed through the probe fixing position 305 on the way, and reached the substrate 301 in a state where the fine particles with capture molecules were suspended in the liquid. The liquid sample was further introduced into the void portion by capillary action, and finally reached the reservoir-side nonwoven fabric 304 in about 3 minutes.

  After 3 minutes from the introduction of the sample, the resistance change rate was investigated through the measurement electric circuit that was reconnected, and the analysis was performed in the same manner as in the second embodiment of the present invention. The HCG concentration in the liquid sample could be examined.

  The present invention is not limited to the present embodiment, and various modifications are allowed without departing from the scope of the present invention. In the above-described embodiment, the structure of each part has been specifically described. However, the structure and the like can be variously changed within a range that satisfies the present invention.

101 Comb pattern electrode 102 Silicon oxide layer 103 Silicon layer 104 Silicon oxide layer 105 Region 106 Silicon oxide layer 107 Fine particles 108 Contaminant 109 Capture molecule film 110 Capture molecule film 111 Target molecule 201 Electrode set 202 Electrode set 203, 204 Signal selection circuit 205 Operation Recording Circuit 301 Electrode Set 302 Connector 303 Sample Introduction Side Nonwoven Fabric 304 Reservoir Side Nonwoven Fabric 305 Probe Fixing Area 306 Resin Package 307 Opening

Claims (9)

A solution in which microparticles bound to the surface of a capture carrier that specifically adsorbs a specific target molecule are dispersed;
A substrate on which concave grooves of a certain width, depth and depth at which the solution is located are formed;
At least a pair of opposing electrodes that are exposed inside the concave groove and the capture carrier is bonded to the surface;
A molecular measuring means for measuring a target molecular weight in the liquid sample in accordance with a change in an electrical binding state of the pair of electrodes;
A biosensor.   The average particle diameter of the fine particles is adjusted so that the fine particles having the capture carrier bound to the surface are captured by at least one target molecule on each of the opposing electrodes. The biosensor according to claim 1.   2. The biosensor according to claim 1, wherein an average particle diameter of the fine particles having the capture carrier bound to the surface thereof is equal to a length between the opposing electrodes.   The target molecular weight in the liquid sample is measured by generating a laminar flow of the solution mixed with the solution and the liquid sample on the surface of the electrode. Biosensor.   5. The biosensor according to claim 4, wherein a longer axis among the constant lateral width and depth of the concave groove is substantially parallel to a direction in which the laminar flow is generated.   5. The biosensor according to claim 4, wherein a longer axis among the constant lateral width and depth of the concave groove is in contact with an angle of 45 degrees or less with respect to a direction in which the laminar flow is generated.   The biosensor according to any one of claims 1 to 6, comprising a pair of electrically independent electrodes.   The biosensor according to claim 7, further comprising conduction disconnecting means for individually disconnecting conduction between each of the plurality of electrically independent electrodes and the molecular measurement means.   The biosensor according to claim 7, wherein each of the electrically independent electrodes has a plurality of types of capture carriers that adsorb different target molecules.

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