JP2016148583A - Manufacturing method for inspection device - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a manufacturing method for an inspection device, which can easily manufacture a target inspection device by processing a microscopic reaction field in a micro fluid device with a processing liquid while properly controlling the inflow and outflow of the processing liquid for the reaction field.SOLUTION: The manufacturing method comprises: with the use of a micro fluid device 10, introducing a liquid including a precursor into a first space S1 and letting some of the liquid flow through a first opening Ta into a microscopic penetration hole T; discharging the liquid out of the first space S1 and letting some liquid temporarily remaining in the microscopic penetration hole T after the discharge flow through the first opening Ta of the microscopic penetration hole T into the first space S1; and forming a film of the precursor, which can directly or indirectly capture the detection target substance that may be included in a liquid sample to be inspected, in the microscopic penetration hole T.SELECTED DRAWING: Figure 1

Description

  The present invention relates to a method of manufacturing a device for inspecting a liquid sample by processing a microfluidic device having a fine through hole.

  In recent years, micro-TAS, microfluidic devices, micromixers, etc. for performing immunological measurement, chemical reaction, cell culture, etc. using antigen-antibody reaction in microscale to nanoscale micro reaction fields have attracted attention. . In order to perform home medical care, point-of-care (POC), clinical examination, cell culture, etc. quickly with a small number of samples, downsizing of these devices is important.

  As a conventional microfluidic device, there has been disclosed a flow channel chip having a space in which cells can be cultured in the middle of a flow channel and having a blocking portion so that cells do not flow out downstream (Patent Document 1). A fine flow path is formed in the upper part of the damming portion, so that cells do not easily pass through and only the culture solution is likely to flow out. Since the culture space and the channel are formed of a transparent plastic substrate, a desired drug can be administered to the cultured cells and the reaction can be optically observed.

JP 2012-185008 A

  A syringe pump is generally used as a liquid feeding method for flowing a liquid containing a target substance into a reaction field provided in a microfluidic device or for discharging a liquid from the reaction field. However, in recent microfluidic devices, not only the flow path but also the reaction field connected to the flow path has been miniaturized (miniaturization). In the case of liquid feeding, an expensive pump capable of precise pressure control is required. Therefore, when manufacturing (manufacturing) the inspection device by processing the reaction field provided in the microfluidic device, an expensive pump is required to appropriately control the inflow and outflow of the processing liquid to the reaction field. It is. For this reason, there exists a problem that the manufacturing cost of the said inspection device rises.

  Furthermore, it is common for air as well as liquid to flow into the flow path that constitutes the microfluidic device. However, liquid feeding in the flow path of such a gas-liquid mixing system is precisely controlled only by a pump. It is difficult to do so, and there is a problem that the manufacturing efficiency of the inspection device is lowered.

  The present invention has been made in view of the above circumstances, and appropriately controls the inflow and outflow of processing liquid with respect to a minute reaction field provided in a microfluidic device, and processes the reaction field with the processing liquid. Thus, an object of the present invention is to provide a method for manufacturing an inspection device that can easily manufacture the target inspection device.

(1) A method for manufacturing a device for inspecting a liquid sample, wherein a first inner wall surface constituting a first space portion, a second inner wall surface constituting a second space portion, and the first inner wall surface are opened. One or more fine through-holes having a first opening and a second opening that opens to the second inner wall surface and spatially connecting the first space and the second space are configured. A liquid containing a precursor is introduced into the first space portion, and the first opening portion is used to enter the one or more fine through holes. After allowing a part of the liquid to flow in, the liquid is formed on the first inner wall surface by discharging the liquid from the first space portion, and subsequently, accompanying the drying process of the liquid film The liquid film and the part of the liquid temporarily remaining in the one or more fine through holes. The part of the liquid is caused to flow out from the first opening of the one or more fine through holes to the first space by force, and further, the one or more fine through holes are configured. A method for producing an inspection device, characterized in that a film made of the precursor is formed on an inner surface, which can directly or indirectly capture a substance to be detected that can be contained in the liquid sample.

According to the inspection device manufacturing method of (1) above, the capillary force (capillary phenomenon) inherently possessed by the fine through-hole is used to allow the liquid to flow into the fine through-hole provided in the microfluidic device. doing. Further, in order to allow the processing liquid to flow out of the fine through hole, the difference in tension between the liquid film and the liquid in the fine through hole, and the difference in tension changes with the drying of the liquid film. Is used. Therefore, since it is not necessary to apply a high pressure generated by the pump to the fine through hole, the processing liquid can be gently and surely flowed into the fine through hole.
The liquid contains the precursor, and the precursor easily flows into the fine through hole. Thereafter, the liquid that is the solvent of the precursor flows out of the fine through hole. At this time, a thin layer made of the liquid is temporarily formed on the inner surface of the fine through hole by the tension of the liquid. Thereafter, the thin layer naturally disappears, and the precursor contained in the liquid remains in the fine through-hole, and a film (capture film) constituted by the precursor is formed. The membrane is useful because it can directly or indirectly capture a substance to be detected contained in a liquid sample to be examined.

  If the inflow and outflow of the liquid to the fine through hole are controlled by a conventionally used pump, the flow rate of the liquid in the fine through hole becomes too large, so that the thin layer is not sufficiently formed. The film formation may be insufficient. However, in the manufacturing method according to the present invention, the liquid is gently introduced by capillarity, and then the fine through-hole is gently utilized by utilizing a flow that spontaneously accompanies discharge of the first space. Therefore, the liquid is allowed to flow out naturally, so that the thin layer is reliably formed. As a result, the film having a sufficient thickness can be formed. That is, according to the manufacturing method of the present invention, a film having a sufficient thickness constituted by the precursor can be formed on the inner surface of the fine through hole more reliably than the conventional method.

(2) A device manufacturing method for inspecting a liquid sample, the first inner wall surface constituting the first space portion, the second inner wall surface constituting the second space portion, ..., and the nth space portion ( n represents an integer of 3 or more.) The n inner wall surfaces including the nth inner wall surface, the first opening portion that opens to the first inner wall surface, and the first opening portion that opens to the second inner wall surface. A first inner surface comprising two or more openings and constituting one or more fine through holes that spatially connect the first space portion and the second space portion; and the first inner wall surface One or more fine through holes that spatially connect the first space portion and the n-th space portion, having a first opening portion that opens to the second wall portion and a second opening portion that opens to the n-th inner wall surface. Using a microfluidic device comprising (n-1) inner surfaces, and (n-1) inner surfaces comprising Liquid is introduced into the first space, and the liquid is introduced into the fine through-holes from the first openings of the fine through-holes formed by the (n-1) inner surfaces. After injecting a part, forming the liquid film made of the liquid on the first inner wall surface by discharging the liquid from the first space, followed by the drying process of the liquid film, Due to the tension between the liquid film and the partial liquid temporarily remaining in the fine through holes, the partial liquid is transferred from the first opening of the fine through holes to the first space. And forming a film composed of the precursor that can directly or indirectly capture a substance to be detected that can be contained in the liquid sample on each inner surface constituting each of the fine through holes. A method of manufacturing an inspection device characterized by the above.

  Also in the manufacturing method of the inspection device of the above (2), the same effect as the manufacturing method of the above (1) is exhibited. That is, in the manufacturing method according to the present invention, the liquid is gently introduced by capillarity, and thereafter, each of the fine penetrations is gently utilized by utilizing a flow that spontaneously accompanies discharge of the first space portion. Since the liquid naturally flows out from the holes, the thin layer is surely formed, and as a result, the film having a sufficient thickness can be formed. That is, according to the manufacturing method of the present invention, a film having a sufficient thickness constituted by the precursor can be formed on the inner surface of the fine through hole more reliably than the conventional method.

(3) A device manufacturing method for inspecting a liquid sample, the first inner wall surface constituting the first space portion, the second inner wall surface constituting the second space portion, ..., and the nth space portion ( n represents an integer of 3 or more.) The n inner wall surfaces including the nth inner wall surface, the first opening portion that opens to the first inner wall surface, and the first opening portion that opens to the second inner wall surface. A first inner surface comprising two or more openings and constituting one or more fine through holes that spatially connect the first space portion and the second space portion; and the first inner wall surface One or more fine through holes that spatially connect the first space portion and the n-th space portion, having a first opening portion that opens to the second wall portion and a second opening portion that opens to the n-th inner wall surface. And (n-1) inner surfaces including the (n-1) inner surface, and the first precursor is contained using a microfluidic device comprising: The introduced first liquid is introduced into the second space portion, and the one or more fine through holes are provided from the second opening portion of the one or more fine through holes constituted by the first inner surface. After flowing a part of the first liquid into the hole, the first liquid is discharged from the second space to form a liquid film made of the first liquid on the second inner wall surface, The part of the first liquid is removed by tension between the liquid film and the part of the first liquid temporarily remaining in the one or more fine through-holes during the drying process of the liquid film. The liquid sample is allowed to flow out from the second opening of the one or more fine through holes to the second space, and further on the first inner surface constituting the one or more fine through holes. Constituted by the first precursor capable of directly or indirectly capturing a target substance that may be contained. The operation of forming the first film, ..., and the (n-1) th liquid containing the (n-1) th precursor are introduced into the nth space, and the (n-1) th After flowing a part of the (n-1) liquid into the one or more fine through holes from the second opening part of the one or more fine through holes constituted by the inner surface. , By discharging the (n−1) th liquid from the nth space, forming a liquid film composed of the (n−1) th liquid on the nth inner wall surface, and subsequently drying the liquid film Due to the tension between the liquid film and the part of the (n-1) th liquid temporarily remaining in the one or more fine through holes, the part of the (n-1) th liquid. From the second opening of the one or more fine through holes to the nth space, and further, the one or more fine through holes are formed. The (n-1) -th inner surface formed by the (n-1) -th precursor that can directly or indirectly capture the target substance that can be included in the liquid sample. And a method of manufacturing an inspection device, comprising: an operation of forming a film.

Also in the manufacturing method of the inspection device of (3), the same effect as the manufacturing method of (1) is exhibited. That is, in the manufacturing method according to the present invention, the liquid is gently introduced by capillarity, and thereafter, using the flow that spontaneously accompanies each discharge of the second space to the nth space, In addition, since the liquid naturally flows out from each fine through-hole, the thin layer is surely formed, and as a result, the film having a sufficient thickness can be formed. That is, according to the manufacturing method of the present invention, a film having a sufficient thickness constituted by the precursor can be formed on the inner surface of the fine through hole more reliably than the conventional method.
Furthermore, in the inspection device, since the inflow and outflow of the liquid in each fine through hole constituted by the first inner surface to the (n-1) inner surface can be controlled independently, A film composed of different precursors can be formed. Therefore, each fine through-hole can be used as an individual reaction field.

(4) The method for manufacturing an inspection device according to (3), wherein the first precursor,..., The (n-1) th precursor are different from each other.

According to the inspection device manufacturing method of (4) above, different substances to be detected are captured on the first inner surface to the (n-1) inner surface constituting each fine through hole provided in the inspection device. Possible first film to (n-1) th film can be formed.
By using only one inspection device manufactured in this way, different detected substances are individually captured in each fine through hole, and a plurality of detected substances that can be included in the liquid sample to be inspected are analyzed. be able to.

(5) At least one set of the same precursor is contained in the precursor group consisting of the first precursor,..., And the (n-1) th precursor. The manufacturing method of the inspection device as described in).

According to the inspection device manufacturing method of (5) above, at least one set of the same detection target is provided on the first inner surface to the (n−1) inner surface constituting each fine through hole provided in the inspection device. A film capable of capturing a substance can be formed.
When a single liquid sample is caused to flow into each fine through-hole using the inspection device manufactured in this way, the same (equivalent (same) in the fine through-hole in which a film capable of capturing the same substance to be detected is formed. Result) should be obtained. That is, since a single liquid sample can be inspected with a plurality of fine through-holes, the inspection accuracy can be improved.

(6) The method for manufacturing an inspection device according to (3), wherein the first precursor,..., The (n-1) th precursor are all the same precursor.

According to the inspection device manufacturing method of (6) above, the same substance to be detected is applied to all of the first inner surface to the (n-1) inner surface constituting each fine through hole provided in the inspection device. A trappable first film to (n-1) th film can be formed.
When a single liquid sample is caused to flow into each fine through-hole using the inspection device manufactured in this way, the same (equivalent (same) in the fine through-hole in which a film capable of binding the same substance to be detected is formed. Result) should be obtained. That is, since a single liquid sample can be inspected with a plurality of fine through-holes, the inspection accuracy can be improved. In addition, when a plurality of different liquid samples are caused to flow into the fine through holes provided in the inspection device, a common substance to be detected that can be contained in each liquid sample can be analyzed with a single inspection device. it can. That is, multi-analyte measurement can be performed efficiently.

  Here, when there are two precursors capable of capturing the same substance to be detected, both precursors may be the same or different from each other.

(7) The film according to (1) above or the first film to the (n-1) film according to (2) or (3) above is specific or nonspecific to the substance to be detected. The method for manufacturing an inspection device according to any one of (1) to (6) above, wherein the inspection device is a film that binds to.
(8) At least one of the precursors described in (1) and the first to (n-1) th precursors described in (2) and (3) is a polymer. The method for manufacturing an inspection device according to any one of (1) to (7), characterized in that:
(9) The method for producing a test device according to (8), wherein the polymer is an antibody, a nucleic acid aptamer, a peptide, a protein or a sugar chain excluding the antibody.

  According to the method for manufacturing an inspection device of the present invention, in order to allow a liquid to flow into a micro through-hole provided in a microfluidic device, a capillary force (capillary phenomenon) inherently possessed by the micro through-hole is used. Yes. Since it is not necessary to apply high pressure generated by the pump to the fine through-holes, it is possible to allow the processing liquid to flow into the fine through-holes gently and reliably.

  The liquid contains the precursor, and the precursor can easily flow into the fine through hole. Thereafter, the liquid that is the solvent of the precursor flows out of the fine through hole. At this time, a thin layer made of the liquid is temporarily formed on the inner side surface of the fine through hole by the tension of the liquid. Thereafter, the thin layer naturally disappears, and the precursor contained in the liquid remains in the fine through hole, so that a film composed of the precursor is formed. The membrane is useful because it can directly or indirectly capture a substance to be detected contained in a liquid sample to be examined.

  If the inflow and outflow of the liquid to the fine through hole are controlled by a conventionally used pump, the flow rate of the liquid in the fine through hole becomes too large, so that a thin layer made of the liquid is not sufficiently formed. As a result, the thickness of the film constituted by the precursor may be insufficient. However, in the manufacturing method according to the present invention, the liquid is gently introduced by capillarity, and then the fine through-hole is gently utilized by utilizing a flow that spontaneously accompanies discharge of the first space. Since the liquid is allowed to flow out naturally, the thin layer is reliably formed, and as a result, the film having a sufficient thickness can be formed. That is, according to the manufacturing method of the present invention, a film having a sufficient thickness constituted by the precursor can be formed on the inner surface of the fine through hole more reliably than the conventional method.

It is typical sectional drawing of the microfluidic device 10 which can be used in the manufacturing method of the test | inspection device of this invention. FIG. 2 is a diagram illustrating a main part of FIG. 1 and a schematic diagram illustrating a state of liquid feeding in a microfluidic device 10. FIG. 2 is a diagram illustrating a main part of FIG. 1 and a schematic diagram illustrating a state of liquid feeding in a microfluidic device 10. FIG. 2 is a diagram illustrating a main part of FIG. 1 and a schematic diagram illustrating a state of liquid feeding in a microfluidic device 10. FIG. 5 is a schematic diagram showing a state in which a thin layer R made of a liquid Q1 containing a precursor is formed on an inner surface Y of a fine through hole T. FIG. 5 is a schematic diagram showing a state in which a film J composed of a precursor is formed on the inner surface Y of the fine through hole T, and the antigen Ag and the contaminant F are captured by the film J. In the fine through-hole T, the antibody Ab binds to the antigen Ag captured by the membrane J made of the precursor by an antigen-antibody reaction, and the complex Comp. It is the schematic diagram which showed a mode that was formed. It is typical sectional drawing showing the 1st space part, the 2nd space part, and several fine through-holes with which the microfluidic device was equipped. It is the schematic diagram which showed an example of the cross-sectional shape of each micro through-hole when the some micro through-hole in the microfluidic device of FIG. 8A is cut | disconnected by A-A 'direction. It is the schematic diagram which showed an example of the cross-sectional shape of each micro through-hole when the some micro through-hole in the microfluidic device of FIG. 8A is cut | disconnected by A-A 'direction. It is the schematic diagram which showed an example of the cross-sectional shape of each micro through-hole when the some micro through-hole in the microfluidic device of FIG. 8A is cut | disconnected by A-A 'direction. It is the schematic diagram which showed an example of the cross-sectional shape of each micro through-hole when the some micro through-hole in the microfluidic device of FIG. 8A is cut | disconnected by A-A 'direction. It is typical sectional drawing of the microfluidic device 20 which can be used in the manufacturing method of the test | inspection device of this invention. It is typical sectional drawing of the microfluidic device 30 which can be used in the manufacturing method of the test | inspection device of this invention. It is typical sectional drawing of the microfluidic device 40 which can be used in the manufacturing method of the test | inspection device of this invention. It is typical sectional drawing of the 1st board | substrate and 2nd board | substrate which comprise a microfluidic device. It is typical sectional drawing of the 1st board | substrate and 2nd board | substrate which comprise a microfluidic device. It is typical sectional drawing of the principal part containing the fine through-hole T of a microfluidic device. It is typical sectional drawing of the principal part containing the fine through-hole T of a microfluidic device. It is typical sectional drawing of the principal part containing the fine through-hole T of a microfluidic device.

<Method for manufacturing inspection device>
A microfluidic device 10 shown in FIG. 1 is an example of a microfluidic device that can be used in the first embodiment of the method for manufacturing an inspection device according to the present invention. FIG. 1 shows a schematic cross-sectional view of a microfluidic device 10.

  The microfluidic device 10 includes a first inner wall surface w1 constituting the first space portion S1, a second inner wall surface w2 constituting the second space portion S2, and a first opening portion Ta opened to the first inner wall surface w1. And an inner side surface Y that forms one or more fine through holes T that spatially connect the first space S1 and the second space S2 with a second opening Tb that opens to the second inner wall surface w2. And at least a main body portion 4 including: In the microfluidic device 10, the first space portion S <b> 1 and the second space portion S <b> 2 included in the main body portion 4 are spatially connected (communicated) with each other by a plurality of fine through holes T.

  In the manufacturing method of the inspection device according to the first embodiment, a processing liquid is introduced into the first space S1, and the first through holes Ta enter the fine through holes T by capillary action (capillary force). After allowing a part of the liquid to flow in, the liquid is discharged from the first space portion S1, and then the part of the liquid temporarily remaining in each fine through hole T when the liquid is discharged. The liquid feeding method (1) for automatically flowing out from the first opening Ta of each fine through hole T to the first space S1 is employed.

<Liquid feeding method (1)>
The description of the liquid feeding method (1) is divided into three steps of an inflow step, a discharge step, and an outflow step.
In the inflow step, an arbitrary liquid is introduced into the first space portion S1, and from the first opening portion Ta of each fine through hole T that opens to the first space portion S1, into each fine through hole T by capillary action. A step of allowing a part of the liquid to flow in;
The discharging step is a step of discharging the liquid from the first space portion S1 while leaving the part of the liquid flowing into each fine through hole T.
The outflow step is a step of automatically flowing out the liquid remaining in each fine through hole T from the first opening Ta of each fine through hole T to the first space S1 after the discharge. .
Hereinafter, each step will be described in order.

(Inflow step)
As shown in FIG. 2, when the liquid Q is introduced into the first space S1 from the opening opened on the surface of the main body 4 constituting the microfluidic device 10, each fine through hole T opened in the first space S1. Part of the liquid Q flows into each fine through hole T from the first opening Ta by capillary action (capillary force). At this time, the air in each fine through hole T is naturally pushed out to the second space S2.

  The hole diameter of the fine through hole T (the diameter or the long diameter of the cross section perpendicular to the longitudinal direction) is not particularly limited as long as the capillary phenomenon occurs, and usually depends on the surface tension, viscosity, etc. of the liquid Q to be used. The range of 1 nm to 1000 μm is preferable.

  The tip of the liquid Q that flows in from the first opening Ta of the fine through hole T may or may not reach the second opening Tb that opens in the second space S2. When the second opening Tb is reached, part of the liquid Q constituting the tip may flow out of the second opening Tb, but basically flows out of the second opening Tb. Instead, the progress of the tip of the liquid Q stops at the second opening Tb. This is because the capillary force keeps the liquid Q in the fine through-hole T and the surface tension acting on the tip of the liquid Q in the second opening Tb keeps the liquid Q in the fine through-hole T. It is guessed.

  When the liquid Q is sufficiently introduced into the first space S1, the inside of each fine through hole T is also filled with a part of the liquid Q (see FIG. 3). At this time, the liquid Q may be continuously introduced into the first space S1, or the liquid Q in the first space S1 is stopped and the first space S1 is filled with the liquid Q. May be kept.

(Discharge step)
Next, as shown in FIG. 4, the liquid Q is discharged from the first space S <b> 1 while leaving the part of the liquid Q flowing into each fine through hole T. The discharge method is not particularly limited, and may be natural discharge using gravity, or forced discharge using a syringe pump, a peristaltic pump, or the like. The discharge speed is not particularly limited, and the first space part S1 may be discharged with a negative pressure. However, it is usually preferable to discharge more gently. That is, the discharge of the liquid Q may cause the first space S1 to have a negative pressure or not to have a negative pressure.

(Outflow step)
When the liquid Q is discharged from the first space portion S1, the liquid Q is discharged and air flows into the first space portion S1, and in order of contact with the air, from the first opening portion Ta of each fine through hole T, The liquid Q remaining in each fine through hole T automatically flows out to the first space S1.

  As one of the factors that cause this automatic outflow, immediately after the liquid Q is discharged from the first space portion S1, the first space portion S1 of the first space portion S1 in which the first opening portion Ta of each fine through-hole T opens. A thin film M made of the liquid Q is formed on the inner wall surface w1. The film M is gradually dried by the air flowing into the first space S1, and the thickness of the film M is gradually reduced, and finally the film M disappears. Along with this drying process, a tension acts between the liquid Q in each fine through hole T and the film M, and the liquid Q in each fine through hole T is drawn in the direction of the film M, and the first opening Ta The mechanism of spilling out is working. By this mechanism, in order from the fine through hole T having the first opening Ta that is fast in contact with air (in FIG. 4, among the multiple fine through holes T arranged from the top to the bottom of the paper, In order from the through-hole T), the liquid Q remaining inside each fine through-hole T automatically flows out into the first space S1 with a certain time difference.

  Through each of the above steps, a part of the liquid Q introduced into the first space portion S1 is caused to flow into each fine through hole T, and the state is maintained as desired. Subsequently, the liquid is discharged from the first space portion S1. By discharging Q, a part of the liquid Q that is temporarily left in each fine through hole T when the liquid Q is discharged can flow out to the first space S1.

  In the above description, the case where the liquid Q is introduced into the first space portion S1 has been described. However, if the liquid Q is introduced into the second space portion S2 in the same manner as this case, each opening to the second space portion S2 is performed. A part of the liquid Q can be caused to flow into each fine through hole T from the second opening Tb of the fine through hole T. Next, by discharging the liquid Q in the second space portion S2, the liquid Q in each fine through hole T can flow out from the second opening portion Tb to the second space portion S2. The structures and configurations of the first space portion S1 and the second space portion S2 may be the same or different from each other.

<First Embodiment of Inspection Device Manufacturing Method>
By using a liquid (processing liquid) for processing the inner surface Y of the fine through-hole T as the liquid Q of the liquid feeding method (1) described above, the inspection device 10 ′ is manufactured using the microfluidic device 10 as a material. be able to.

  In the method for manufacturing an inspection device according to the first embodiment, as a processing liquid, a precursor of the film for forming a film capable of directly or indirectly capturing a target substance that can be included in a liquid sample to be inspected. Use a liquid containing. After this processing liquid is introduced into the first space S1 of the microfluidic device 10 and a part of the processing liquid is caused to flow from the first opening Ta into the fine through hole T by capillary action, The processing liquid is discharged from the space S1, and then the part of the processing liquid temporarily remaining in the fine through-hole T when the processing liquid is discharged is removed from the first through-hole T. It automatically flows out from the opening Ta to the first space S1.

  After the automatic outflow, a thin layer (thin film) R made of a processing liquid is formed on the inner surface of the fine through hole T (see FIG. 5).

  As a mechanism for forming the thin layer R, when most of the processing liquid flows out of the fine through-hole T, friction of the processing liquid and the inner surface Y of the fine through-hole T causes friction of the processing liquid. It is considered that the phenomenon that the remaining part is left on the inner surface Y of the fine through hole T has occurred.

  Subsequent to the formation of the thin layer R, the thin layer R disappears from the inner side surface Y of the fine through hole T, and a film J composed of the precursor is formed on the inner side surface Y.

  As a mechanism in which the thin layer R disappears from the inner surface Y of the fine through-hole T, the processing liquid constituting the thin layer R flows due to the influence of the tension of the processing liquid constituting the thin layer R itself, It is considered that at least one of the first mechanism that flows out of the through hole T and the second mechanism that the solvent that is a component of the processing liquid constituting the thin layer R evaporates occurs. .

  The formation mechanism of the film J when the first mechanism works is that the precursor contained in the processing liquid comes into contact with the inner surface Y, and the intermolecular force and Coulomb force between the precursor and the inner surface Y. It is thought that the phenomenon of adsorbing by physicochemical bonding due to, etc. occurs. The formation mechanism of the film J when the second mechanism works is a phenomenon in which the solubility of the precursor contained in the processing liquid is lowered and the precursor is deposited on the inner surface Y. It is thought that. By any mechanism, it is considered that films J having the same ability to capture a substance to be detected contained in a liquid sample to be inspected are formed.

  The film J formed on the inner side surface Y of the fine through hole T can directly or indirectly capture one or more kinds of detected substances contained in the liquid sample to be inspected.

  When the membrane J directly captures the substance to be detected, the substance to be detected in the liquid sample in contact with the film J may be captured non-specifically by the film J, or specifically captured by the film J. Also good. Specifically, for example, when the precursor is a polymer, low molecule or metal having biocompatibility (affinity) and the membrane J has affinity for many substances derived from organisms, the membrane J Can capture non-specifically biological substances as substances to be detected. As another specific example, when the precursor is an antibody having antigen specificity and the membrane J is composed of the antibody, the membrane J can capture the antigen as a substance to be detected.

  When the membrane J indirectly captures the target substance, the mediator is first adsorbed on the membrane J, and further, the target substance in the liquid sample is trapped non-specifically by the membrane J via the mediator. Alternatively, it may be specifically captured by the membrane J. Specifically, for example, when the precursor is a biocompatible polymer, low molecule, or metal, after the antibody as the mediator is first adsorbed to the membrane J, the membrane J further contains the antibody. Thus, the substance to be detected (in this case, the antigen) in the liquid sample can be indirectly captured. When the antibody has antigen specificity, the substance to be detected can be specifically captured. In addition, by using an antibody with low antigen specificity, it is possible to non-specifically capture many types of antigens as substances to be detected.

  The thickness of the film J is not particularly limited as long as it can capture the substance to be detected directly or indirectly, and depends on the type of the substance to be detected and the diameter of the fine through hole T, but for example, about 1 nm to 1 μm is preferable. It is. If the thickness of the film J is too thin, the capture probability of the substance to be detected may be lowered. If the thickness of the film J is too thick, there is a risk that the inflow and outflow of the liquid sample in the fine through hole T may be hindered, or the film J may be peeled off due to the inflow and outflow.

  The film J may be composed of one kind of precursor, or may be composed of two or more kinds of precursors.

  The precursor is preferably a substance capable of forming a film J on the inner surface Y of the fine through-hole T, and having affinity for a substance to be detected contained in a liquid sample to be inspected. Examples of the precursor include a polymer having a biocompatibility (affinity with a biomolecule), a low molecule, and a metal.

  Examples of the polymer include polystyrene (PS), polymethacrylic acid (PMMA), polyvinyl chloride (PVC), silicone resin, water-soluble collagen, polyacrylamide, and other known thermoplastic elastomers. Examples of the polymer include antibodies, other proteins, peptides (peptide aptamers), DNA aptamers, RNA aptamers, aptamers composed of nucleic acids other than DNA and RNA, sugar chains, and the like. Here, the polymer refers to a molecule having a molecular weight Mw of 10,000 or more.

  Examples of the low molecule include heme (a complex of divalent iron and porphyrin), peptides, various amino acids, nucleic acids, sugars, and the like. Examples of the low molecule include lipophilic substances constituting cell membranes such as phospholipids such as phosphatidylcholine, phosphatidylserine, and phosphatidylethanolamine, other known amphipathic lipids, and cholesterol.

  As said metal, the well-known metal which can form into a film, such as gold | metal | money, silver, copper, titanium, chromium, is mentioned, for example.

The solvent of the processing liquid containing the precursor is not particularly limited as long as the solvent can dissolve or disperse the precursor, and for example, water, an organic solvent, or the like can be used.
The concentration of the precursor contained in the processing liquid is appropriately adjusted in consideration of the viscosity of the processing liquid, the thickness of the film J to be formed, and the like.

  According to the manufacturing method of the inspection device of the first embodiment, in order to allow the processing liquid to flow into the micro through-hole T provided in the microfluidic device 10, the capillary force inherently possessed by the micro through-hole T ( (Capillary phenomenon) is used. Since it is not necessary to apply high pressure generated by the pump to the fine through hole T, the processing liquid can be gently and surely flowed into the fine through hole T. Since the processing liquid contains the precursor, the precursor can easily flow into each fine through hole T provided in the microfluidic device 10.

  In the manufacturing method according to the present invention, each of the processing liquids gently flows in by capillary action, and then gently flows using the flow that spontaneously accompanies the discharge of the processing liquid in the first space S1. The processing liquid naturally flows out from the fine through-holes T. For this reason, it is easy to form the thin layer R made of the processing liquid on the inner surface Y, and subsequently form the film J made of the precursor contained in the processing liquid on the inner surface Y. That is, according to the manufacturing method of the inspection device of the first embodiment, the film J that is configured by the precursor and can capture the target substance is more reliably formed than the conventional method. Can be easily formed.

  By performing the liquid feeding method using the inspection device 10 ′ thus manufactured, each fine through hole T can be used as a reaction field relating to physics, chemistry, and biology (biotechnology). Specifically, for example, the test device 10 ′ can be used as a biochemical test device using an immunochemical reaction.

<Manufacture and use of inspection device 10 '>
As an example of the manufacture and use of the inspection device 10 ′, the manufacture and use of the inspection device 10 ′ in which the film J made of the precursor is formed in the fine through hole T provided in the microfluidic device 10 can be mentioned. Below, it demonstrates with reference to FIGS.

  First, when a first liquid Q1 (an example of a processing liquid) containing polystyrene as a precursor is introduced into the first space portion S1, the liquid Q1 gradually fills the first space portion S1, and each capillary force causes The liquid Q1 flows into the inside from the first opening portion Ta of the fine through hole T (see FIGS. 2 and 3; the sign of the liquid Q1 is Q for convenience).

  After the liquid Q1 is filled in all the fine through holes T, when the liquid Q1 is discharged from the first space S1, air flows into the first space S1. Among the first openings Ta of the fine through holes T that open to the first space S1, the first openings Ta that are in the order of contact with air are sequentially left in the fine through holes T in order. The liquid Q1 flows out into the first space S1 (see FIG. 4; for convenience, the sign of the liquid Q1 is Q). As a result, a thin layer R made of the liquid Q1 is temporarily formed (see FIG. 5), and subsequently, a film J made of the precursor is formed.

  Here, when there is a place where the precursor is not desired to be adsorbed in the flow path, unnecessary adsorption can be prevented by performing in advance a surface treatment such as coating for improving the water repellency of the place. .

  Subsequently, when a second liquid Q2 (an example of a liquid sample) containing an antigen Ag (an example of a substance to be detected) and a foreign substance F (an example of a substance other than the substance to be detected) is introduced into the first space S1. The liquid Q2 gradually fills the first space portion S1, and the liquid Q2 flows into the inside from the first opening portion Ta of each fine through-hole T by a capillary force (not shown). Antigen Ag and contaminant F contained in the liquid Q2 flowing into each fine through hole T are adsorbed non-specifically due to the affinity for the membrane J formed on the inner surface Y of each fine through hole T.

  After the liquid Q2 is filled in all the fine through holes T, when the liquid Q2 is discharged from the first space S1, air flows into the first space S1. Among the first openings Ta of the fine through holes T that open to the first space S1, the first openings Ta that are in the order of contact with air are sequentially left in the fine through holes T in order. The liquid Q2 flows out to the first space S1 (not shown). Even after all the liquid Q2 inside each fine through hole T is discharged, the antigen Ag and the contaminant F in each fine through hole T remain fixed to the membrane J (see FIG. 6).

  Subsequently, when there is a region where the antigen Ag and the contaminant F are not adsorbed and the membrane J is exposed, the region may be subjected to a blocking process. For example, by causing the third liquid Q3 containing skim milk to flow into the fine through-hole T, proteins such as albumin constituting the skim milk are adsorbed on the region. When the blocking process is performed in this manner, it is possible to prevent the antibody Ab contained in the liquid Q4 flowing into the fine through-hole T at a later stage from adsorbing to the membrane J nonspecifically. After the blocking process, the liquid Q3 is caused to flow out from the inside of the fine through hole T, and the next operation is started. The inflow and outflow of the liquid Q3 in the fine through hole T may be performed in the same manner as in the case of the liquid Q2.

  Next, when the fourth liquid Q4 containing the antibody Ab previously given the binding property to the antigen Ag is introduced into the second space part S2, the liquid Q4 gradually fills the second space part S2, and the capillary force As a result, the liquid Q3 flows into the inside from the second opening Tb of each fine through hole T (not shown). The antibody Ab contained in the liquid Q4 flowing into each fine through-hole T recognizes the antigen Ag fixed to the membrane J of each fine through-hole T, binds by an antigen-antibody reaction, and consists of an antigen Ag-antibody Ab. Complex Comp. Form.

  After all the fine through holes T are filled with the liquid Q4, when the liquid Q4 is discharged from the second space S2, air flows into the second space S2. Among the second openings Tb of the fine through holes T that open to the second space S2, the second openings Tb that are in the order of contact with air are sequentially left in the fine through holes T in order. The liquid Q4 flows out to the second space S2 (not shown). Even after all the liquid Q4 in each fine through hole T is discharged, the composite Comp. Maintains the state fixed to the membrane J (see FIG. 7).

  Since the antibody Ab is preliminarily bound (conjugated) with a luminescent labeling substance, for example, when the fine through-hole T is irradiated with excitation light, the complex Comp. The luminescent labeling substance contained in fluoresces. The amount of fluorescence emitted from the complex Comp. Depends on the abundance of Therefore, the content of the antigen Ag contained in the liquid Q2 can be quantified by measuring the fluorescence emission amount.

  Before performing fluorescence measurement by excitation light irradiation, a cleaning solution was flowed into the fine through-hole T for the purpose of washing the antibody Ab non-specifically adsorbed to the membrane J or the antigen Ag in the fine through-hole T. It may be drained later.

In general, it is preferable that the inside of the fine through hole T is cleaned at any one or more of the following stages.
After the membrane J is formed on the inner surface Y of the fine through-hole T After the liquid Q2 containing the antigen Ag and the contaminant F flows in and out of the fine through-hole T The liquid Q3 containing skim milk becomes fine through-hole T After the liquid Q4 containing the antibody Ab flows into and out of the fine through hole T

  The luminescent labeling substance to which the antibody Ab binds may be a fluorescent substance that emits fluorescence when excited by receiving external light irradiation, or a luminescent substance that chemiluminescents another substrate. It may also be a catalytic substance that converts to

  Examples of the fluorescent substance include green fluorescent protein (GFP) and quantum dots. As the catalytic substance, for example, known enzymes used in ELISA can be applied, and specific examples include enzymes such as peroxidase, luciferase, and aequorin. Examples of substrates for this enzyme include 3- (p-hydroxyphenol) propionic acid and its analogs, luciferin and luciferin analogs, coelenterazine and coelenterazine analogs, and the like.

  One kind of luminescent labeling substance may be used alone, or two or more kinds of luminescent substances may be used in combination. Two or more kinds of fluorescent substances may be used in combination, two or more kinds of enzymes and substrates may be used in combination, or one or more kinds of fluorescent substances may be used in combination with one or more kinds of enzymes and substrates. Good. A method for binding various luminescent labeling substances to the antibody Ab is not particularly limited, and a known method can be applied.

  The labeling substance for tracing the substance present in the fine through-hole T may be a luminescent labeling substance as described above or a non-luminescent labeling substance. Examples of non-luminescent labeling substances include radioactive labeling substances as used in known radioimmunoassay methods.

  An example of the use of the inspection device described above is to qualitatively analyze whether or not the antigen Ag is contained in the liquid Q2 to be examined, or to determine the content of the antigen Ag contained in the liquid Q2. For the purpose of quantitative analysis, a format generally called a sandwich immunoassay is adopted. For this reason, it is necessary to prepare in advance an antibody Ab that specifically or non-specifically binds to the antigen Ag. Such antibodies are obtained by known methods.

  In the test using the test device according to the present invention, other known immunoassay formats such as an indirect antibody immunoassay format and a bridging immunoassay format may be employed instead of the sandwich immunoassay format described above. Moreover, you may implement the test | inspection using the interaction between other molecules which does not utilize at least any one of an antibody and an antigen. Since these basic methods for testing using these immunoassay formats and other intermolecular interactions are known, only a brief description will be given here.

  When the indirect antibody immunoassay format is adopted, after the membrane J composed of the antigen Ag (an example of a precursor) is formed on the inner surface Y of the fine through hole T, the primary antibody Ab1 and the secondary antibody Ab2 are sequentially finely penetrated. It is introduced into the hole T. According to this format, when there is a possibility that the primary antibody Ab1 (an example of a substance to be detected) is contained in the liquid sample to be examined, the presence / absence or content of the primary antibody Ab1 is analyzed. be able to.

  When the bridging immunoassay format is adopted, after the membrane J composed of the antigen Ag (an example of the precursor) is formed on the inner surface Y of the fine through-hole T, the primary antibody Ab1 and the antigen Ag having the labeling substance are sequentially formed. It is introduced into the fine through hole T. According to this format, the presence or content of the primary antibody Ab1 (an example of a substance to be detected) in the liquid to be examined can be analyzed.

<Liquid sample>
The substance to be detected contained in the liquid sample to be examined is not particularly limited, for example, an antigen capable of binding specifically or non-specifically to a predetermined antibody, specific or non-specific to a predetermined antigen Antibody, any organic compound, inorganic compound, metal, etc.
The type of the antigen is not particularly limited and is appropriately selected according to the purpose of the test. Specific examples of the antigen include proteins, peptides, nucleic acids, lipids, sugar chains and the like derived from pathogens such as viruses, bacteria, and the like that cause colds, hepatitis, acquired immune deficiencies, and the like.
The detected substance that can be contained in the liquid sample may be one type or two or more types.

<Microfluidic device>
As an example of a microfluidic device that can be used in the method for manufacturing an inspection device according to the present invention, the configuration of the microfluidic device 10, 20, 30, 40 will be described below.

  The plurality of micro through-holes T constituting the microfluidic device 10 shown in FIG. 1 include a first opening Ta and a second space S2 that open to the first inner wall surface w1 constituting the first space S1. And a second opening Tb that opens to the second inner wall surface w2. Each fine through hole T is in the main body 4 constituting the microfluidic device 10 and forms a first flow path S1 and a second flow path in the main body 4 The second space portion S2 to be connected is spatially connected (communicated). That is, the first end of each fine through hole T constitutes a first opening Ta, and the second end of each fine through hole T constitutes a second opening Tb.

  The first space S <b> 1 inherent in the main body 4 is formed by a first inner wall surface w <b> 1 included in the main body 4. The first space S <b> 1 has at least two openings at arbitrary locations on the surface of the main body 4. When any liquid Q is injected from any of the openings, the liquid Q is introduced into the first space S1. Thereafter, the liquid Q in the first space S1 is discharged from any opening at a desired timing. A pump or valve may be attached to the microfluidic device 10 in order to control the introduction and discharge of the liquid Q in the first space S1.

The shape of the first space portion S1 is not particularly limited as long as it has the first inner wall surface w1 in which the first opening portion Ta is opened. For example, the shape of the first space portion S1 is the same shape as the flow path that configures a known fluid device. Alternatively, any solid shape such as a cube, a rectangular parallelepiped, a sphere, and a spheroid may be used. When the shape of the first space S1 is a shape having a longitudinal direction, the shape of the cross section orthogonal to the longitudinal direction may be an arbitrary shape such as a rectangle, a circle, or an ellipse. The cross-sectional area of the cross section is not particularly limited, for ^example, preferably 1mm ² ~400mm 2 about. Within this range, the liquid Q can flow gently and reliably into each fine through hole T opened in the first space S1.

  The shape and size of the second space S2 may be the same as or different from the first space S1. The path of the second flow path formed by the second space part S2 and the path of the first flow path formed by the first space part S1 are different from each other, but some of both paths overlap or intersect. It does not matter. Control of the flow of the liquid Q in each flow path may be performed by a known method, and can be controlled by, for example, a valve or a pump (not shown) provided on the flow path. Since the description regarding other 2nd space part S2 is the same as that of said 1st space part S1, it abbreviate | omits.

  The diameter of the fine through hole T, that is, the diameter or the long diameter (maximum diameter) of the cross section perpendicular to the longitudinal direction of the fine through hole T facilitates the inflow of the liquid Q and the automatic outflow of the liquid Q by the capillary force. For this reason, it is preferable to be uniform from the first opening Ta to the second opening Tb. As described above, the pore diameter is preferably in the range of about 1 nm to 1000 μm. Within this range, the inflow of the liquid Q by the capillary force and the automatic outflow of the liquid Q can be performed gently and reliably.

  The shapes of the first opening Ta and the second opening Tb of the fine through-hole T (the contours of the edges formed by the both ends of the fine through-hole T on the first inner wall surface w1 and the second inner wall surface w2, respectively) are as follows: The shape is not particularly limited as long as it does not prevent the inflow of the liquid Q by the capillary force and the automatic outflow of the liquid Q. Since the inflow and outflow occur more easily, the shape of the first opening Ta and the second opening Tb is a cross section perpendicular to the longitudinal direction of the fine through hole T having the openings Ta and Tb at both ends. The shape is preferably the same.

  The shapes of the first opening Ta and the second opening Tb of the fine through hole T may be the same or different from each other. Further, when a plurality of fine through holes T are provided in the device as in the microfluidic device 10, the shape of the cross section perpendicular to the longitudinal direction of each fine through hole T and the opening of each fine through hole T are provided. The shapes of the parts Ta and Tb may be the same or different from each other.

  A specific example of the cross-sectional shape of each micro through-hole T when the micro through-hole T included in the microfluidic device is cut in the A-A ′ direction of FIG. FIG. 8B is an example in which a plurality of fine through holes T having an elliptical cross section are linearly arranged, and FIG. 8C is an example in which a plurality of fine through holes T having a rectangular cross section are linearly arranged. 8D is an example in which a plurality of fine through holes T having an elliptical cross section are arranged in three straight lines, and FIG. 8E is an elliptical cross section that is flatter than that of FIG. 8B and has a different major axis direction. This is an example in which a plurality of fine through holes T are linearly arranged. The plurality of fine through holes T provided in the microfluidic device may have the same cross-sectional shape as each other, or may have different cross-sectional shapes.

  The length in the longitudinal direction of the fine through-hole T is not particularly limited as long as the liquid Q flows in and the automatic liquid Q flows out due to the capillary force. For example, the length is about 0.5 mm to 15 mm. preferable. Within this range, the inflow and outflow of the liquid Q in the fine through hole T can be caused gently and reliably.

  When there are a plurality of fine through holes T provided in the microfluidic device 10, the distance between the fine through holes T (separation distance) and the distance between the openings Ta and Tb of each fine through hole T (separation distance) are It does not restrict | limit in particular, For example, about 1 micrometer-100 micrometers are preferable. The direction of the central axis along the longitudinal direction of each fine through hole T (the direction indicated by the central axis) may be parallel to each other or non-parallel.

  In the main body 4 of the microfluidic device 10, the first space S <b> 1 and the second space S <b> 2 with which the fine through-hole T communicates form two parallel flow paths. In each flow path, the diameter of the flow path in the direction in which the central axis along the longitudinal direction of the fine through hole T passes through each flow path is not particularly limited, but the viewpoint that the liquid easily flows into the fine through hole T from each flow path. Therefore, for example, about 0.5 mm to 20 mm is preferable.

  The “angle formed” between the central axis along the longitudinal direction of each flow path and the central axis along the longitudinal direction of each fine through hole T is not particularly limited, and may be 90 degrees or an obtuse angle. Or an acute angle. The reason why the angle formed is not particularly limited is that the force dominant to the inflow and outflow of the liquid Q in each fine through hole T is the capillary force and the first space portion around each first opening Ta. This is the tension generated when the film M made of the liquid Q formed on the first inner wall surface w1 constituting S1 moves (drys), and the contribution of the angle made is small.

  In the microfluidic device 10, the first inner wall surface w1 and the second inner wall surface w2 constituting the first space portion S1 and the second space portion S2, and the inner side surface Y constituting the plurality of fine through holes T are the main body portion. 4 included in the base material (substrate).

Next, as a modification of the microfluidic device 10, a microfluidic device 20 (see FIG. 9) and a microfluidic device 30 (see FIG. 10) will be described.
The microfluidic devices 20 and 30 can be used to send liquid by the inflow step, the discharge step, and the outflow step, as in the microfluidic device 10 described above.
Therefore, in the same manner as the method of manufacturing the inspection device 10 ′ using the microfluidic device 10, the inspection devices 20 ′ and 30 ′ are formed using the microfluidic devices 20 and 30 by the liquid feeding method (1) described above. Can be manufactured.

  The microfluidic device 20 shown in FIG. 9 includes a main body portion 4 including a first inner wall surface w1 constituting the first space portion S1 and a second inner wall surface w2 constituting the second space portion S2, and the first space portion S1. The first opening portion Ta that faces and the second opening portion Tb that opens facing the second space portion S2 and spatially connects the first space portion S1 and the second space portion S2. And a sub main body portion 5 (chip) including an inner surface Y constituting one or more fine through holes T (in communication).

  In the microfluidic device 20, the sub body 5 is installed inside the body 4. The first surface 5u of the sub main body 5 is integrated with the first inner wall surface w1 of the main body 4 to constitute the first space S1. The first opening Ta of each fine through-hole T included in the sub-main body 5 is open to the first surface 5u so as to face the first space S1. Similarly, the second surface 5v of the sub main body 5 is integrated with the second inner wall surface w2 of the main body 4 to constitute a second space S2. The second opening Tb of each fine through-hole T included in the sub-main body 5 is open to the second surface 5v so as to face the second space S2.

  A microfluidic device 30 shown in FIG. 10 includes a main body portion 4 including a first inner wall surface w1 constituting the first space portion S1 and a second inner wall surface w2 constituting the second space portion S2, and the first space portion S1. The first auxiliary inner wall surface ww1 and the second auxiliary inner wall surface ww2 forming the second space S2 are included, and the first opening Ta and the second auxiliary inner wall surface ww2 that open to the first auxiliary inner wall surface ww1 are included. And includes an inner surface Y that constitutes one or more fine through holes T that spatially connect (communicate) the first space S1 and the second space S2. A sub main body 5 (chip).

  In the microfluidic device 30, the sub main body 5 is installed inside the main body 4. The first sub inner wall surface ww1 of the sub main body 5 is connected to the first inner wall surface w1 of the main body 4 to form one first space S1 as a whole. The first opening portion Ta of each fine through hole T included in the sub body portion 5 opens to the first sub inner wall surface ww1 so as to face the first space portion S1. Similarly, the second sub inner wall surface ww2 of the sub main body portion 5 is connected to the first inner wall surface w2 of the main body portion 4 to form one second space portion S2 as a whole. The second opening Tb of each fine through-hole T included in the sub main body 5 opens to the second sub inner wall surface ww2 so as to face the second space S2.

  The sub main body 5 (chip) constituting the microfluidic devices 20 and 30 may be installed so that it can be detached from the main body 4 and attached to the main body 4 or removed from the main body 4. It may be installed in a state where it is bonded or bonded so that it is not possible.

  The shape of the base constituting the sub main body 5 is not particularly limited as long as it can be installed on the main body 4. Examples of the shape of the substrate include a box shape (chip shape) such as a rectangular parallelepiped and a cube. The material of the substrate is not particularly limited, and the same material as the material of the substrate constituting the main body 4 can be applied.

  A method for forming the fine through hole T, the first space portion S1, and the second space portion S2 in the base body constituting the main body portion 4 or the sub main body portion 5 is not particularly limited, and a known fine processing technique can be applied. A manufacturing method of the microfluidic device 10 will be described in detail later as a representative example.

The microfluidic devices 10, 20, and 30 described above are devices including two space portions S1 and S2.
Hereinafter, a microfluidic device 40 having three or more spaces will be exemplified with reference to FIG.

  The microfluidic device 40 includes a first inner wall surface w1 constituting the first space portion S1, a second inner wall surface w2 constituting the second space portion S2, and an nth inner portion constituting the nth space portion Sn. A total of n inner wall surfaces including the wall surface wn are included. The “n” represents an integer (ordinal number) of 3 or more.

  Furthermore, the microfluidic device 40 has a first opening Ta that opens to the first inner wall surface w1 and a second opening Tb that opens to the second inner wall surface w2, and includes the first space S1 and the second space. A first inner surface Y1 constituting one or more fine through-holes T that spatially connect the portion S2, and a first opening Ta and an nth inner wall surface wn that open to the first inner wall surface w1. The (n−1) th (n−1) th one or more fine through holes T having a second opening Tb that opens and spatially connect (communicate) the first space S1 and the nth space Sn. A total of (n−1) inner surfaces including the inner surface Y (n−1) are included. One inner surface constitutes one or more fine through holes. The “n” represents an integer (ordinal number) of 3 or more.

  Here, “a first inner wall surface w1 constituting the first space portion S1, a second inner wall surface w2 constituting the second space portion S2, and an nth inner wall surface wn constituting the nth space portion Sn. "..." in the notation of "is the third inner wall surface w3 constituting the third space portion S3, the fourth inner wall surface w4 constituting the fourth space portion S4, and the fifth inner portion constituting the fifth space portion S5. It represents that any n number of space portions are repeated in the order of the wall surfaces w5,. The “n” represents an integer (ordinal number) of 3 or more, and FIG. 11 illustrates a case where n = 5.

Similarly, “... Has a first opening Ta that opens to the first inner wall surface w1 and a second opening Tb that opens to the nth inner wall surface, and the first space portion S1 and the nth space portion Sn. "..." in the notation of the (n-1) th inner surface Y (n-1) constituting one or more fine through-holes T that spatially connect (communicate) with each other,
It has a first opening Ta that opens to the first inner wall surface w1 and a second opening Tb that opens to the third inner wall surface, and spatially connects the first space portion S1 and the third space portion S3 ( A second inner surface Y2 constituting one or more fine through-holes T (in communication);
The first opening portion Ta has a first opening portion Ta that opens on the first inner wall surface w1 and a second opening portion Tb that opens on the fourth inner wall surface, and spatially connects the first space portion S1 and the fourth space portion S4 ( A third inner surface Y3 constituting one or more fine through-holes T (in communication);
In this order, any (n-1) inner surfaces are repeated. The “n” represents an integer (ordinal number) of 3 or more, and is the same integer as “n” in the nth space portion Sn. Therefore, FIG. 9 illustrates the case where n = 5.

  The microfluidic device 40 communicates the first micro space group G1 composed of one or more micro through holes T communicating the first space portion S1 and the second space portion S2, and the first space portion S1 and the third space portion S3. A second micro space group G2 composed of one or more micro through holes T, (n-1) of one or more micro through holes T communicating the first space portion S1 and the n th space portion Sn. It has a minute space group G (n-1). Here, the notation “...” Represents that any (n−1) minute space groups are repeated in the same order as described above. The “n” represents an integer (ordinal number) of 3 or more, and is the same integer as “n” in the nth space portion Sn. Therefore, FIG. 11 illustrates the case where n = 5.

<Liquid feeding method (2)>
The first liquid feeding method (liquid feeding method (2)) in the microfluidic device 40 can be performed by the inflow step, the discharge step, and the outflow step, as in the above-described microfluidic device 10 and the like.

  In the first liquid feeding method, the inflow step introduces the liquid Q into the first space S1, and opens to the first space S1, and the first through holes T of the fine through holes T constituting the first minute space group G1. From the first opening Ta of each fine through hole T that opens into the first opening Ta,... And the first space S1, and constitutes the (n−1) th minute space group G (n−1). In this step, a part of the liquid Q is caused to flow into each fine through hole T by capillary action. The discharging step is a step of discharging the liquid Q from the first space portion S1 while leaving the part of the liquid Q flowing into each fine through hole T. The outflow step is a step of automatically flowing out the liquid Q in each fine through hole T from the first opening Ta of each fine through hole T to the first space S1 after the discharge. Note that n represents an integer (ordinal number) of 3 or more.

  In the first liquid feeding method, the first openings Ta of the micro through holes T constituting the first micro space group to the (n-1) micro space group of the microfluidic device 40 are commonly opened. By introducing the liquid Q into the first space portion S1, the same liquid Q flows into each micro through-hole T of the first micro space group to the (n−1) th micro space group, and then flows out. ing.

<Second Embodiment of Inspection Device Manufacturing Method>
By using the liquid (processing liquid) for processing the inner surface of the fine through-hole as the liquid Q of the liquid feeding method (2) described above, the inspection device 40 ′ can be manufactured using the microfluidic device 40 as a material. it can.

  In the method for manufacturing an inspection device according to the second embodiment, as a processing liquid, a precursor of the film for forming a film capable of directly or indirectly capturing a target substance that can be included in a liquid sample to be inspected Use a liquid containing. This processing liquid is introduced into the first space S1 of the microfluidic device 40, and one of the processing liquid is introduced into each micro through-hole T by capillarity from the first opening S1 of each micro through-hole T. The processing liquid is discharged from the first space portion S1 after the portion is introduced, and then the part of the processing liquid temporarily remaining in each fine through hole T when the processing liquid is discharged. Are automatically allowed to flow out from the first opening Ta of each fine through hole T to the first space S1.

  After the automatic outflow, a thin layer (thin film) R made of a processing liquid is formed on each inner surface constituting the fine through hole T. Subsequent to the formation of the thin layer R, the thin layer R disappears from each inner surface constituting the fine through hole T, and a film J composed of the precursor is formed on each inner surface along with the disappearance. The mechanism by which the thin layer R and the film J are formed is considered to be the same as the manufacturing method of the first embodiment described above.

  According to the inspection device manufacturing method of the second embodiment, the same effects as the inspection device manufacturing method of the first embodiment are exhibited. That is, in the manufacturing method of the inspection device according to the second embodiment, the processing liquid is gently introduced by capillary action, and then the flow spontaneously generated along with the discharge of the first space S1 is used to gently In addition, the processing liquid can naturally flow out from each fine through hole T. For this reason, the thin layer R can be formed, and the film J composed of the precursor can be easily formed. That is, according to the inspection device manufacturing method of the second embodiment, the film J made of the precursor can be more reliably formed on the inner surface Y of each fine through hole T than the conventional method.

  In the inspection device 40 ′ manufactured by the manufacturing method according to the second embodiment, various different processing liquids are allowed to flow in and out independently of each of the micro space groups G1 to G (n−1). You can make it. Therefore, the fine through hole T in each minute space group provided in the inspection device 40 ′ can be used as an independent reaction field.

<Liquid feeding method (3)>
As the second liquid feeding method (liquid feeding method (3)) different from the above-mentioned first liquid feeding method (liquid feeding method (2)), the first to (n-1) minute space groups are used. It is also possible for different liquids to flow into and out of each group.

  The second liquid feeding method (liquid feeding method (3)) in the microfluidic device 40 is performed by changing a part of the inflow step, the discharge step, and the outflow step described above. In the second liquid feeding method, the liquid Q flows from the second opening Tb of each fine through hole T.

The second opening Tb of each micro through-hole T in the microfluidic device 40 is open to any one of the second space to the nth space. When the first liquid to the (n-1) th liquid are respectively introduced into each of these space portions, each of these liquids has each fine through hole T opened to the second space portion to the nth space portion, that is, the first minute space group. To the inside of the (n-1) minute space group by capillary force. Since introduction and discharge of each liquid in each space part can be controlled independently, an individual liquid can be supplied to each fine through hole T opened in the second space part to the nth space part. Depending on the timing, they can be made to flow independently or flow out independently.
The second liquid feeding method (liquid feeding method (3)) will be specifically described below.

  In the second liquid feeding method using the microfluidic device 40, each of the first microspace group to the (n-1) th microspace group independently performs an inflow step, an exhaust step, and an outflow step. In the inflow step, the second opening Tb of each fine through hole T constituting each group is individually opened for each group, and each of the second space part S2 to the nth space part Sn is desired. Introduce liquid.

  When the predetermined liquid A is allowed to flow into and out of each fine through-hole T constituting the first minute space group G1, the liquid A may be introduced and discharged in the second space portion S2. Independently of the inflow and outflow of the liquid A in the first microspace group G1, the nth space portion is used when the liquid B flows in and out of the (n-1) th microspace group G (n-1). The liquid B may be introduced into Sn and then discharged.

A part of the liquid A introduced into the second space S2 flows into the fine through holes T from the second openings Tb of the fine through holes T constituting the first minute space group G1. Next, when the liquid A is discharged from the second space portion S2, a part of the liquid A temporarily remaining in each minute through hole T constituting the first minute space group G1 is second from the second opening portion Tb. It flows out to space part S2.
The inflow and outflow of the liquid B introduced into the nth space portion Sn into the (n-1) th minute space group G (n-1) are the same as in the case of the liquid A introduced into the second space portion S2. It is.

  That is, in the second liquid feeding method, the main body portion 4, the first space portion S <b> 1, the second space portion S <b> 2,... And the first micro space group G1, which is formed in the main body portion 4 and includes one or more fine through holes T communicating with the first space portion S1 and the second space portion S2, and the first space. Using the microfluidic device 40 including the (n-1) th micro space group G (n-1) including one or more fine through holes T communicating with the part S1 and the nth space part Sn, As will be described below, this is a liquid feeding method in which the liquid feeding in the first micro space group G1,..., The liquid feeding in the (n-1) th micro space group G (n-1) is performed independently.

  Here, the notation “...” Represents that any (n−1) minute space groups are repeated in the same order as described above. The “n” represents an integer (ordinal number) of 3 or more, and is the same integer as “n” in the nth space portion Sn. Therefore, FIG. 11 illustrates the case where n = 5.

  The liquid feeding in the first micro space group G1 introduces the first liquid into the second space portion S2, and at the same time the first through holes T constituting the first micro space group G1 opening in the second space portion S2. From the second opening Tb, an inflow step for allowing a part of the liquid to flow into each fine through hole T by capillary action, and the one that has flowed into each fine through hole T constituting the first micro space group G1. A discharge step of discharging the liquid from the second space portion S2 while leaving the liquid of the portion, and after the discharge, from the second opening Tb of each fine through hole T constituting the first minute space group G1, An outflow step for automatically outflowing the liquid in each fine through hole T to the second space S2.

  In the (n−1) th minute space group G (n−1), liquid feeding introduces the (n−1) th liquid into the nth space portion Sn and opens the (n−1) th space portion Sn. -1) an inflow step for allowing a part of the liquid to flow into each micro through hole T by capillary action from the second opening Tb of each micro through hole T constituting the micro space group G (n-1); The liquid is discharged from the nth space portion Sn while leaving the part of the liquid flowing into each fine through hole T constituting the (n-1) th minute space group G (n-1). After the discharge step, and after the discharge, the liquid in each fine through hole T is supplied from the second opening Tb of each fine through hole T constituting the (n-1) th minute space group G (n-1). an outflow step for automatically flowing out into the n space portion Sn.

  In the second liquid feeding method using the microfluidic device 40, the same liquid may be fed to each group independently, or different liquids may be fed.

<Third Embodiment of Manufacturing Method of Inspection Device>
By using the liquid (processing liquid) for processing the inner surface of the fine through hole as the liquid Q of the liquid feeding method (3) described above, the inspection device 40 ″ can be manufactured using the microfluidic device 40 as a material. it can.

  In the method for manufacturing an inspection device according to the third embodiment, as a processing liquid, a precursor of the film for forming a film capable of directly or indirectly capturing a target substance that can be included in a liquid sample to be inspected Are used from the first liquid to the (n-1) th liquid.

  The method for manufacturing an inspection device according to the third embodiment includes a first operation to an (n-1) th operation. Here, n is an integer of 3 or more, and is the same integer as “n” in the “nth space” of the microfluidic device 40.

  In the first operation, a first liquid containing a first precursor for forming a first film capable of directly or indirectly capturing a substance to be detected that can be contained in a liquid sample to be inspected is converted into a second space. Introduced into the part S2, and enters each fine through hole T by capillary action from each second opening Tb of one or more fine through holes T (micro space group G1) constituted by the first inner surface Y1. After flowing a part of the first liquid, the first liquid is discharged from the second space portion S2, and then, the one remaining temporarily in each fine through hole T when the first liquid is discharged. Part of the first liquid is automatically caused to flow out from the second opening of each fine through-hole to the second space S2, and further, the first inner side surface Y1 constituting the one or more fine through-holes T is formed. And a first precursor that can directly or indirectly capture a substance to be detected that can be contained in the liquid sample. An operation of forming the first film.

  The (n-1) th operation is a (n-1) precursor for forming a (n-1) film capable of directly or indirectly capturing a substance to be detected that can be contained in a liquid sample to be examined. The (n-1) th liquid containing is introduced into the nth space portion Sn, and one or more fine through-holes T (microspaces) constituted by the (n-1) th inner surface (n-1) From the second opening Tb of the group G (n-1)), a part of the (n-1) liquid is caused to flow into each fine through hole T by capillary action, and then from the nth space Sn. The (n-1) th liquid is discharged, and then the part of the (n-1) liquid temporarily remaining in each fine through-hole T when the (n-1) th liquid is discharged. The (n-1) th inner surface that automatically flows out from the second opening Tb of each fine through hole T to the nth space Sn and further constitutes the one or more fine through holes T. , Which can be captured substance to be detected which may be contained in the liquid sample directly or indirectly, an operation of forming the (n-1) (n-1) th layer, which is constituted by the precursor.

In the inspection device manufacturing method of the third embodiment, (n-1) operations are performed. That is, the first operation, the second operation, the third operation, the fourth operation,..., And the (n−1) th operation are performed.
Here, n is an integer (ordinary number) of 3 or more that is the same, and is an integer that coincides with n representing the “nth space portion” of the microfluidic device 40 to be used.

Since each of the above operations can be controlled independently, each operation may be performed simultaneously or individually at a desired timing.

  The first precursor contained in the first liquid and the (n-1) th precursor contained in the (n-1) th liquid may be different from each other. In this case, the first film to the (n−1) film formed in the respective micro through holes T constituting the first to (n−1) th minute space groups are composed of different precursors. Therefore, the capture performance of each film with respect to the substance to be detected is different. In the inspection device manufactured in this way, different substances to be detected can be captured in each film.

  The first precursor contained in the first liquid, ..., and the precursor group consisting of the (n-1) th precursor contained in the (n-1) th liquid, at least one set of the same precursors May be included. In this case, at least one set of the first film to the (n−1) film formed in each minute through hole T constituting the first to (n−1) th minute space group. Since the films are made of the same precursor, at least one set of films capable of capturing the same substance to be detected can be formed. When a single liquid sample is caused to flow into each fine through-hole using the inspection device manufactured in this way, the same (equivalent (same) in the fine through-hole in which a film capable of capturing the same substance to be detected is formed. Result) should be obtained. That is, since a single liquid sample can be inspected with a plurality of fine through-holes, the inspection accuracy can be improved.

  The first precursor contained in the first liquid and the (n-1) th precursor contained in the (n-1) th liquid may all be the same precursor. In this case, all of the first film to the (n−1) film formed in each minute through hole T constituting the first to (n−1) th minute space group are made of the same precursor. Since it is comprised, the film | membrane which can capture | acquire the same to-be-detected substance in all the fine through-holes T can be formed. When a single liquid sample is caused to flow into each fine through-hole using the inspection device manufactured in this way, the same (equivalent (same) in the fine through-hole in which a film capable of binding the same substance to be detected is formed. Result) should be obtained. That is, since a single liquid sample can be inspected with a plurality of fine through-holes, the inspection accuracy can be improved. In addition, when a plurality of different liquid samples are caused to flow into the fine through holes provided in the inspection device, a common substance to be detected that can be contained in each liquid sample can be analyzed with a single inspection device. it can. That is, multi-analyte measurement can be performed efficiently.

  Here, when there are two precursors capable of capturing the same substance to be detected, both precursors may be the same or different from each other.

  The first film to the (n-1) film may specifically bind to the substance to be detected, or may bind nonspecifically. The first precursor to the (n-1) th precursor may be precursors that can specifically bind the detected substance, or substances that can bind the detected substance nonspecifically. It may be. Further, among the first precursor to the (n-1) th precursor, at least one precursor may be the polymer, the low molecule, or the metal. Good.

  According to the inspection device manufacturing method of the third embodiment, the same effects as the inspection device manufacturing method of the first embodiment are exhibited. That is, in the inspection device manufacturing method according to the third embodiment, the introduction of the processing liquid into the second space part to the nth space part is independently controlled, and each microscopic penetration of the processing liquid is gently performed by capillary action. From the fine through-holes T gently, using the flow that spontaneously flows along with the discharge of the processing liquid that is independently controlled in the second space part to the n-th space part. Processing liquid can flow out naturally. As described above, since the processing liquid can be gently flowed in and out of each fine through hole T, the thin layer R can be formed and the film J composed of the precursor can be easily formed. it can. That is, according to the inspection device manufacturing method of the third embodiment, the film J made of the precursor can be more reliably formed on the inner side surface Y of each fine through hole T than the conventional method.

  In the inspection device 40 ″ manufactured by the manufacturing method of the third embodiment, various different liquids are allowed to flow in and out independently of each of the micro space groups G1 to G (n−1). Therefore, the fine through hole T in each minute space group provided in the inspection device 40 ″ can be used as an independent reaction field.

<Manufacture and usage example of inspection device 40 ''>
In the second liquid feeding method (liquid feeding method (3)) using the inspection device 40 ″, the same liquid is fed independently to each of the micro space groups G1 to G (n−1). For example, ELISA for detecting different antigens in each micro space group G1 to G (n-1) can be performed. Since it can be performed independently, a plurality of ELISAs can be performed in parallel.

  In the manufacture and use of the inspection device 40 ″, a liquid feeding method that combines the second liquid feeding method and the first liquid feeding method may be performed. Utilizing this, it is possible to realize efficient liquid feeding. As a specific example, an example in which the inspection device 40 ″ is continuously manufactured and used is given below.

  First, in the microfluidic device 40, a solution containing protein A is sent to the micro through-holes T constituting all the micro space groups G1 to G (n-1) by the first liquid feeding method. Liquid. By forming the film J composed of protein A on the inner side surfaces Y1 to Y (n-1) of the micro through holes T constituting each micro space group by this liquid feeding, by the first liquid feeding method, A solution containing skim milk is fed to block the inner side surfaces Y1 to Y (n-1) of each fine through-hole T constituting all the minute space groups. The protein A corresponds to the precursor, and the microchannel device 40 in which a film made of protein A is formed on the inner surface of the fine through hole T is an inspection device 40 ″.

  In the inspection device 40 ″ manufactured (produced) as described above, a solution containing an individual primary antibody having a desired antigen specificity is independently obtained for each microspace group by the second liquid feeding method. By this liquid feeding, the protein A and the primary antibody bind to each other, and individual primary antibodies are fixed to the inner side surfaces Y1 to Y (n-1) of the respective micro through holes T constituting each micro space group. Is done.

  Next, after the cleaning liquid is supplied to the micro through holes T constituting all the micro space groups G1 to G (n-1) by the first liquid supply method, all the micro space groups are supplied. The liquid sample to be inspected is sent in a batch. By this liquid feeding, the substance to be detected contained in the liquid sample can bind to the primary antibody fixed in each micro through hole T of each micro space group. Subsequently, a solution containing a secondary antibody to which a labeling substance is bound is fed to all the microspace groups in a lump by the first liquid feeding method. By this liquid feeding, a ternary complex composed of a primary antibody, a substance to be detected and a secondary antibody can be formed in each micro through hole T of each micro space group. Thereafter, independent ELISA can be performed in each microspace group by detecting or measuring the labeling substance bound to the secondary antibody.

<< Method for Manufacturing Microfluidic Device >>
The above-described microfluidic device can be manufactured by applying a known microfabrication technique.
The material in particular of the main-body part 4 of a microfluidic device is not restrict | limited, For example, glass, a plastic (resin), a semiconductor, a metal, ceramics etc. are mentioned. The shape of the main body 4 is not particularly limited, and it is easy to connect and install the main body 4 to other devices (for example, pumps, reagent bottles, waste liquid reservoirs, etc.). A box shape is preferable. The main body 4 having such a shape is referred to as a “substrate”. Hereinafter, although the manufacturing method of the microfluidic device when the main-body part 4 is comprised with the board | substrate is demonstrated, even if the main-body part 4 is shapes other than a board | substrate, it can manufacture similarly.

  As a method for forming the fine through hole T inherent in the substrate (main body portion 4), for example, two formation methods, a first formation method and a second formation method, can be exemplified.

(First forming method)
In the first forming method, as shown in FIGS. 12 and 13, a groove Ya (concave portion) is formed on the surface of the first substrate 4A, the second substrate 4B is joined to the surface, and the ceiling is formed in the groove Ya. In this method, fine through holes T are formed.

  12 and 13 are schematic cross-sectional views in which main parts including the fine through hole T of the main body 4 are enlarged. The main body 4 is formed by joining the first substrate 4A and the second substrate 4B. The fine through hole T is formed at the interface between the two substrates. In the case of FIG. 12, one fine through hole T having a rectangular cross section is formed. In the case of FIG. 13, a plurality of fine through holes T having a rectangular cross section are formed.

  As shown in FIG. 13, when the height H1 of the groove Ya is larger than the separation distance L2 between the adjacent fine through holes T (when H1> L2), the surface area of each fine through hole T (the fine through holes The total of the area of the inner side surface to be configured becomes larger than the surface area of one large fine through hole T shown in FIG. 12 (the area of the inner side surface forming the fine through hole). Therefore, when it is desired to increase the surface area of the fine through hole T in the main body 4, that is, the contact area between the liquid Q and the inner surface constituting the fine through hole T, the fine through hole T as shown in FIG. A plurality of layers may be formed. In order to further increase the surface area (contact area), the aspect ratio (height H1 / base L1) of each fine through hole T may be made larger than 1. The surface area can be increased as the aspect ratio is increased.

  As a method of forming the groove Ya in the first substrate 4A, various known methods can be cited depending on the material of the first substrate 4A.

  In the case of using a resin substrate as the first substrate 4A, for example, a forming method in which known methods such as molding, nanoimprinting, injection molding, and the like for transferring and forming a fine pattern produced by a soft lithography technique are appropriately combined.

  When a glass substrate is used as the first substrate 4A, for example, a method of appropriately combining known methods such as photolithography, laser processing, machining, dry etching, and wet etching can be used. In the case of forming the nanoscale groove Ya, a combination of substrate modification by short pulse laser processing and wet etching is preferable.

  Examples of a method for bonding the first substrate 4A and the second substrate 4B include known methods such as a method of bonding with an adhesive, an anodic bonding method, a self-welding method, and a low-temperature pressure bonding method using surface modification. .

  The materials of the first substrate 4A and the second substrate 4B may be the same or different from each other. Specifically, both substrates may be glass substrates, a combination of a glass substrate and a silicon substrate, a combination of a glass substrate and a plastic substrate, or both substrates. It may be a resin substrate.

  The kind of plastic substrate is not particularly limited. For example, PET (polyethylene terephthalate), PS (polystyrene), PMMA (polymethyl methacrylate), PDMS (polydimethylsiloxane), AS (acrylonitrile styrene) resin, PC (polycarbonate), PLA (Polylactic acid) and the like. The kind of glass which comprises a glass substrate is not specifically limited, For example, quartz glass, borosilicate glass, soda-lime glass, etc. are mentioned.

(Second forming method)
The second forming method is a method of directly forming the fine through hole T in the first substrate 4C as shown in FIGS. By connecting the focal point (condensing part) of short pulse laser light inside the substrate and scanning the part where the fine through hole T is formed in the substrate, the etching resistance of the scanned part (modified part) is weakened. To reform. Thereafter, the modified through hole T can be formed in the substrate by removing the modified portion from the substrate by wet etching.

  14, 15, and 16 are schematic cross-sectional views of a main part including a plurality of fine through holes T formed in the main body 4. The main body 4 is constituted by a single substrate 4C. The fine through hole T is formed inside the substrate 4C.

  In the case of FIG. 14, the fine through holes T having a plurality of elliptical cross sections are arranged in a line in a direction perpendicular to the substrate thickness direction K (plane direction of the substrate). Since the major axis of the ellipse is along the substrate thickness direction K, the integration density of the fine through holes 4 in the plane direction of the substrate can be increased as compared with the case where the cross section is a perfect circle. By increasing the integration density, the number of measurement samples per unit volume increases, so that measurement accuracy can be improved.

  In the case of FIG. 15, the fine through holes T having a plurality of elliptical cross sections are arranged in two rows in a direction orthogonal to the substrate thickness direction K (plane direction of the substrate). In the drawing, the fine through holes T arranged in the upper stage and the fine through holes T arranged in the lower stage are arranged so as not to overlap each other when viewed in the thickness direction K of the substrate. By arranging in this way, the integration density of the plurality of fine through holes T can be increased without impairing the ease of observing each fine through hole T in the substrate thickness direction K.

  In the case of FIG. 16, the fine through holes T having a plurality of elliptical cross sections are arranged in a line in the substrate thickness direction K. When arranged in this way, since the major axis of the fine through hole T is along the substrate plane direction (the minor axis of the minute through hole T is along the substrate thickness direction K), light ( For example, when the fine through-hole T is irradiated with excitation light, a backlight for observing the inside, or the like, the irradiation light is hardly refracted and the irradiation light is easily transmitted. Therefore, it is easy to irradiate light inside the fine through hole T, and light emission inside the fine through hole T can be easily observed from the thickness direction K of the substrate.

(Specific example of the second forming method)
In this specific example, an apparatus that generates a titanium sapphire laser beam, which is a femtosecond laser, is used, but an apparatus that generates another type of laser beam may be used.
First, laser light is incident on the first surface of a quartz glass substrate placed on a precision stage, the laser light is focused on a predetermined position inside the substrate, and perpendicular to the propagation direction (optical axis) of the laser light. The focal point of the laser beam is scanned in the direction of. At this time, if the laser polarization is perpendicular to the scanning direction, it is possible to easily form the fine through hole T having a minor diameter of nano order (for example, about 10 nm to 500 nm). Moreover, it is preferable that the irradiation intensity of a laser beam is set to be equal to or higher than the processing lower limit threshold and lower than the processing upper limit threshold. The optimum irradiation intensity is preferably examined in advance using the same type of quartz glass substrate.
As an example, the following irradiation conditions are mentioned, for example.

Wavelength (center wavelength) = 800 nm, spectrum width = 10 nm (± 5 nm), pulse time width = ˜250 fs, numerical aperture (NA) of objective lens = 0.5, polarization = linear polarization, optical axis and scanning direction Angle of about 90 degrees / peak intensity (laser fluence per pulse / pulse time width) = 9 TW / cm ²
・ Scanning speed (μm / sec) = 1,000 μm / sec, repetition frequency (kHz) = 200 kHz
・ Scan at a constant speed while shifting so that the focus of each pulse overlaps.

  By irradiating the quartz substrate with laser light as described above, a modified portion with reduced etching resistance is formed in the region scanned by the light collecting portion including the focal point of the laser light and its periphery. For example, it is possible to form a modified portion that extends substantially parallel to the substrate surface and has an elliptical (substantially rectangular) cross-sectional shape perpendicular to the extending direction. As an example, a modified portion having a long diameter (vertical length) (length in the substrate thickness direction K) of about 5 μm and a short diameter (horizontal length) (length in the substrate plane direction) of about 30 nm. Can be formed. By such laser processing, it is possible to form a plurality of modified portions arranged in parallel to each other.

  Next, the quartz substrate with both ends of each modified portion exposed on the surface is immersed in hydrofluoric acid or an aqueous potassium hydroxide solution for etching. In this etching, the etching solution permeates into each modified portion from both ends of each modified portion, and each modified portion is removed from the quartz substrate. As a result, a plurality of fine through holes T penetrating the quartz substrate can be formed. As an example, both end portions are open on the surface of the quartz substrate, the cross-sectional shape in a direction orthogonal to the longitudinal direction is an ellipse (substantially rectangular), and a long diameter (vertical length) (length in the substrate thickness direction K) ) Is about 5.5 μm, and a fine through hole having a short diameter (horizontal length) (length in the substrate plane direction) of about 300 nm can be formed.

  When the modified portion is formed on the quartz substrate, if both ends of the modified portion are not exposed on the surface of the substrate, the modified portion can be formed by a method such as photolithography, grinding, polishing, etc. before etching. What is necessary is just to pre-process so that the both ends of may be exposed to the substrate surface.

  The configurations and combinations thereof in the embodiments described above are examples, and the addition, omission, replacement, and other modifications of the configurations can be made without departing from the spirit of the present invention. Further, the present invention is not limited by each embodiment, and is limited only by the scope of the claims.

  The method for manufacturing an inspection device according to the present invention can be widely used in the field of medical inspection and the like.

10, 20, 30, 40 ... microfluidic device, S1 ... first space, S2 ... second space, S3 ... third space, S4 ... fourth space, S5 ... fifth space, w1 ... first 1 inner wall surface, w2 ... second inner wall surface, w3 ... third inner wall surface, w4 ... fourth inner wall surface, w5 ... fifth inner wall surface, Ta ... first opening, Tb ... second opening, T ... Fine through-hole, Y ... inside surface, 4 ... main body part, 5 ... sub-main body part, M ... film made of liquid, G1 ... first minute space group, G2 ... second minute space group, G3 ... third minute space group G4 ... fourth micro space group, Q ... liquid, Q1 ... first liquid, Q2 ... second liquid, Q3 ... third liquid, R ... thin layer made of liquid containing precursor, J ... detected Membrane capable of capturing substances, Ab ... antibody, Ag ... antigen, Comp. ... complex

Claims (9)

A device manufacturing method for inspecting a liquid sample, comprising:
A first inner wall surface constituting the first space portion, a second inner wall surface constituting the second space portion,
One having a first opening opening in the first inner wall surface and a second opening opening in the second inner wall surface, and spatially connecting the first space portion and the second space portion. Using a microfluidic device comprising an inner surface that constitutes the above fine through hole,
After introducing a liquid containing a precursor into the first space, and flowing a part of the liquid into the one or more fine through holes from the first opening,
By discharging the liquid from the first space portion, a liquid film made of the liquid is formed on the first inner wall surface, and subsequently, the liquid film and the one or more ones accompanying a drying process of the liquid film Due to the tension with the part of the liquid temporarily remaining in the fine through hole, the part of the liquid flows out from the first opening of the one or more fine through holes to the first space part. Let me
Forming a film made of the precursor, capable of directly or indirectly capturing a substance to be detected that can be contained in the liquid sample, on the inner surface constituting the one or more fine through-holes; Method for manufacturing inspection device. A device manufacturing method for inspecting a liquid sample, comprising:
The first inner wall surface constituting the first space portion, the second inner wall surface constituting the second space portion,..., And the nth space portion (n represents an integer of 3 or more). N inner wall surfaces including a wall surface;
One having a first opening opening in the first inner wall surface and a second opening opening in the second inner wall surface, and spatially connecting the first space portion and the second space portion. The first inner surface constituting the fine through hole, ..., the first opening that opens in the first inner wall surface, and the second opening that opens in the nth inner wall surface, (N-1) inner surfaces including the (n-1) th inner surface that constitutes one or more fine through holes that spatially connect the first space portion and the nth space portion,
Using a microfluidic device comprising
A liquid containing a precursor is introduced into the first space, and each fine through-hole is formed from a first opening of each fine through-hole formed by the (n-1) inner surfaces. After flowing a part of the liquid into the hole,
By discharging the liquid from the first space portion, a liquid film made of the liquid is formed on the first inner wall surface, and subsequently, the liquid film and each fine through-hole that accompany the drying process of the liquid film The partial liquid is caused to flow out from the first opening of each of the fine through holes to the first space due to the tension with the partial liquid temporarily remaining therein, and
An inspection device characterized in that a film made of the precursor material capable of directly or indirectly capturing a substance to be detected that can be contained in the liquid sample is formed on each inner surface constituting each fine through-hole. Manufacturing method. A device manufacturing method for inspecting a liquid sample, comprising:
The first inner wall surface constituting the first space portion, the second inner wall surface constituting the second space portion,..., And the nth space portion (n represents an integer of 3 or more). N inner wall surfaces including a wall surface;
One having a first opening opening in the first inner wall surface and a second opening opening in the second inner wall surface, and spatially connecting the first space portion and the second space portion. The first inner surface constituting the fine through hole, ..., the first opening that opens in the first inner wall surface, and the second opening that opens in the nth inner wall surface, (N-1) inner surfaces including the (n-1) th inner surface that constitutes one or more fine through holes that spatially connect the first space portion and the nth space portion,
Using a microfluidic device comprising
The first liquid containing the first precursor is introduced into the second space portion, and the second opening portion of the one or more fine through holes formed by the first inner surface includes After a part of the first liquid flows into one or more fine through-holes, the first liquid is discharged from the second space portion, whereby the liquid made of the first liquid on the second inner wall surface. A film is formed, and subsequently, the one of the liquid films and the part of the first liquid temporarily remaining in the one or more fine through-holes in accordance with the drying process of the liquid film The first inner surface that causes the first liquid of a portion to flow out from the second opening of the one or more fine through-holes to the second space and further constitutes the one or more fine through-holes The first precursor is capable of directly or indirectly capturing a substance to be detected that can be contained in the liquid sample. Thus the operation of forming the first film made, ..., and,,
The (n-1) liquid containing the (n-1) th precursor is introduced into the nth space, and the one or more fine penetrations constituted by the (n-1) inner surface After a part of the (n−1) th liquid flows into the one or more fine through holes from the second opening of the hole, the (n−1) th through the nth space. ) Forming a liquid film made of the (n-1) th liquid on the nth inner wall surface by discharging the liquid; and subsequently, the liquid film and the one or more liquids accompanying the drying process of the liquid film Due to the tension with the partial (n-1) liquid temporarily remaining in the fine through-holes, the partial (n-1) liquid is caused to pass through the one or more fine through-holes. The second (n-1) inner surface constituting the one or more fine through-holes is further discharged to the n-th space from the second opening. An operation for forming a (n-1) film composed of the (n-1) precursor, which can directly or indirectly capture a substance to be detected that can be contained in a liquid sample. Device manufacturing method.   The method for manufacturing an inspection device according to claim 3, wherein the first precursor,..., The (n−1) th precursor are different from each other.   The at least one set of the same precursor is contained in the precursor group consisting of the first precursor, ..., and the (n-1) th precursor. Manufacturing method of inspection device.   4. The method of manufacturing an inspection device according to claim 3, wherein the first precursor, and the (n−1) th precursor are all the same precursor. 5.   The film according to claim 1 or the first film to the (n-1) film according to claim 2 or 3 is a film that specifically or nonspecifically binds to the substance to be detected. The manufacturing method of the test | inspection device as described in any one of Claims 1-3 characterized by the above-mentioned.   The precursor according to claim 1, and at least one of the first precursor to the (n-1) th precursor according to claims 2 and 3, is a polymer. The manufacturing method of the test | inspection device as described in any one of -3.   The method for producing a test device according to claim 8, wherein the polymer is an antibody, a nucleic acid aptamer, a peptide, a protein or a sugar chain excluding the antibody.

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