JPH03223674A - Reaction vessel - Google Patents

Abstract

PURPOSE:To simultaneously measure many items with respect to many specimens by providing a reaction unit having a passage provided with at least one fluid inlet and having at least one reagent fixing part on the way of the passage on the downstream side of the fluid inlet. CONSTITUTION:A specimen is introduced from a fluid inlet 10 and three kinds of the antigens in the specimen are respectively bonded to the antibodies corresponding to the antigens immobilized on reagent immobilized parts 30, 31, 32 and mutually having no cross reactivity. A solution mixture of three kinds of labelled antibodies is introduced from the fluid inlet 10 to bond the respective antigens bonded to the respective antibodies immobilized on the reagent immobilized parts 30, 31, 32 to the corresponding labelled antibodies and subsequently discharged from an outlet 20. If necessary, when the antibody corresponding to the substance to be measured in the specimen and the antibody corresponding to a reference substance are immobilized on the reagent immobilized parts 30, 31, 32 measuring the signals showing labels after cleaning, the substance to be measured and the reference substance can be simultaneously measured.

Description

DETAILED DESCRIPTION OF THE INVENTION <Industrial Application Field> The present invention relates to a reaction vessel for easily measuring trace substances in a living body by enzyme immunoassay, DNA detection method, or the like.

<Prior art> Measurement of trace amounts of substances in living organisms is frequently carried out for the purpose of diagnosing various diseases and determining therapeutic effects, etc. Depending on the purpose, simple measurement methods that can be easily operated, Measurement methods with high sensitivity and high measurement accuracy are being developed one after another. Among these, the simple measurement method is a very convenient method and can be easily carried out without using measuring instruments or reaction devices, so it is suitable for applications where qualitative or semi-quantitative measurement alone is enough to make a diagnosis. Widely used. For example, it is used for measuring urine glucose, other biochemical tests, pregnancy diagnosis, etc., but it is also used for detecting pathogenic viruses by detecting DNA using nucleic acid hybridization methods, and for other reactions. Applications are becoming widespread.

By the way, current simple measurement methods that utilize immune reactions (antigen-antibody reactions) as a measurement principle include agglutination reactions or agglutination inhibition reactions (hereinafter referred to as agglutination reactions) using red blood cells or latex as carriers; and enzyme immunoassay using an enzyme as a labeling agent (E I
A) etc.

In the agglutination reaction, red blood cells or other similar compounds are used as carriers, and the reaction is carried out in an ampoule with a spherical bottom, and the size of sedimentation rings or spots formed on the spherical surface or their The method of determining the presence or absence of red blood cells (agglutination sedimentation reaction method) is easy to operate and has relatively high measurement sensitivity, but the determination of the result depends on the sedimentation of red blood cells or other similar compounds. It takes a long time to get results. On the other hand, the method (latex agglutination reaction method), in which latex is used as a carrier and the degree of agglutination is determined based on the agglutination image while stirring on a slide, has slightly lower sensitivity, but is simple to operate and can produce results in a short time. can get. Therefore, the latex agglutination reaction method is currently most widely used for applications that do not require relatively high sensitivity, such as pregnancy diagnosis. However, since this method requires skill to judge the results, accurate judgments are mainly limited to doctors and laboratory technicians at medical institutions such as hospitals and clinics.

Although EIA as a simple measurement method has higher measurement sensitivity than other methods, a relatively long reaction time may be required to obtain the high sensitivity. Furthermore, in the B/F separation operation (in the antigen-antibody reaction, the bound form (Bound: B) produced by the binding of antigen and antibody and the unbound free form (Bound: B)
Free: Physically separating F) is complicated, and if this B/F separation is not performed accurately,
In the enzymatic reaction, which is the next step, non-specific reactions occur, resulting in erroneous results.

As mentioned above, although the simple measurement method based on the agglutination reaction method is capable of detecting the presence or absence of a substance, it is not suitable for measuring the amount of a substance present, that is, for quantitative measurement. On the other hand, EIA has the disadvantages that measurement takes a long time and the B/F separation operation is complicated. Since this is a quantitative measurement method based on the degree of judgment or color development, it is possible to perform not only qualitative judgment but also quantitative measurement, and anyone can easily and accurately judge the results. Therefore, regarding EIA which has such advantages,
Although many studies have been made to shorten reaction time and simplify B/F separation operations, the reality is that no satisfactory measurement method has yet been put to practical use.

In addition, recently, simple measurement methods are being applied, but tests that use nucleic acid hybridization to detect specific DNA or RNA (hereinafter referred to as DNA, etc.) have a reaction mode that is DNA etc. is a specific DN
It is similar to immunoassay methods using antigen-antibody reactions, especially EIA, in that it utilizes the property of selectively binding to A and the like. Therefore, a process similar to EIA is required, and conventional measurement methods require a long reaction time.
It has the same problems as EIA, such as the complicated F separation operation.

Therefore, as a means to solve this problem, Japanese Patent Application Laid-Open No. 63-20063 and Japanese Patent Application No. 62-2159
No. 92 proposes a dish-shaped container.

In fact, by using this container, EI for qualitative reactions can be
The operation of A has been significantly simplified.

However, even if this container is used, it is said that it has excellent quantitative performance.
The benefits of IA are not fully exploited.

Furthermore, implementing EIA requires multiple operations such as sample dispensing, addition of washing solution, addition of enzyme-labeled antibody solution, addition of coloring agent or enzyme substrate, and these multiple operations must be performed. No solution has yet been found to shorten the time required for measurement due to the complexity of having to do this and the large number of operations.

In other words, in simplifying the measurement method that allows quantitative measurements, the challenges are to simplify the B/F separation operation and the addition operation of samples and reagents. However, there is still room for improvement regarding these issues.

<Problems to be Solved by the Invention> As mentioned above, a simple measuring method that is highly sensitive, easy to operate, and performs accurate measurements has not yet been developed.

The present invention has been made in view of the above circumstances, and enables highly sensitive measurements to be performed accurately and easily, particularly in the B/F separation operation, and in addition, in addition to the addition operation of the sample and reagent. The purpose of the present invention is to provide a reaction container that can easily perform the following steps.

Another object of the present invention is to provide a container that is widely applicable to various reactions, such as detection methods using EIA and nucleic acid hybridization mechanisms.

A further object of the present invention is to provide a reaction device that can perform simultaneous measurements of multiple items with simple operations.

Means for Solving the Problems> The present invention provides a method for comparing a series of operations such as antigen-antibody reaction or pipelidization reaction, B/F separation operation, enzyme reaction, and result determination in EIA and nucleic acid hybridization methods. This invention relates to a reaction vessel that can be used continuously in a short period of time.

The reaction vessel of the present invention allows ordinary measurements to be carried out without requiring any special equipment. Moreover, it can also be adapted to automatic measuring equipment for continuous processing of a large number of samples or for the purpose of quantitative measurement.

The reaction container of the present invention embodies the above-mentioned characteristics, and has a passageway having at least one fluid inlet in the structure, and has a passageway having at least one fluid inlet in the middle of the passageway, and has a passageway having at least one fluid inlet. The reactor also has at least one reagent fixing portion on the downstream side, and at least one reaction unit having an exhaust mechanism communicating with the passage.

The present invention will be explained in detail below.

The reaction vessel of the invention has at least one reaction unit. Therefore, first, the configuration of a reaction vessel having one reaction unit will be explained based on the drawings. Since the present invention has various aspects, the drawings will first be explained.

FIG. 1a is a perspective view of an embodiment of the present invention, and FIG. 1b is a perspective view of an embodiment of the present invention.
It is a sectional view of the passage part.

FIG. 2a is a perspective view of one embodiment of the present invention, and FIGS. 2b and 2c are views taken along the line A-A and line B-B.

Figures 3a, 3b, and 3c are plan views of each component of an embodiment of the present invention, Figure 3d is a side view thereof, and Figure 3e
The figure and FIG. 3f are arrow views along the line CC and line DD.

FIG. 4 is a perspective view of one embodiment of the present invention.

FIG. 5 is a plan view of one embodiment of the present invention.

FIGS. 6a and 6b are plan views of each component of an embodiment of the present invention, and FIG. 6c is a side view thereof.

7a and 7b are plan views of each component of an embodiment of the present invention, and FIGS. 7c and 7d are side views thereof.

Figures 8a, 8b, and 8c are plan views of each component of an embodiment of the present invention, Figure 8d is a side view thereof, and Figure 8e is an enlarged longitudinal sectional view of part A in Figure 8b. It is.

9a and 9b are plan views of each component of an embodiment of the present invention, and FIG. 9c is a cross-sectional view taken along line X-X49.

FIG. 10, FIG. 11, and FIG. 12 are each a plan view of an embodiment of the present invention.

One reaction unit of the reaction vessel of the present invention described here is:
The components include a structure, a passageway having at least one fluid inlet in the structure, at least one reagent fixing part located in the middle of the passageway and downstream of all the fluid inlets, and communicating with the passageway. It has a continuous exhaust mechanism.

The specimen 2 may be composed of a single member, as shown in FIG. 1a, or may be composed of two parts, as shown in FIGS.
Alternatively, as shown in FIGS. 3d, 3e, and 3f, the lid 3 and the split mold 4.5 of the structure may be composed of a separate mold 4.5. Alternatively, it may be composed of a larger number of split molds.

Furthermore, as shown in FIG. 6C, it may be composed of the lid 3 and split mold 4 of the structure, and the legs 9a, which are also specimens, or as shown in FIG. 7c, the split mold 4 and It is composed of a lid body 3 made of sheet 8 that covers the split mold 4 and serves as a lid, and the lower side of the split mold 4 may be shaped like a ship's bottom. Alternatively, as shown in Fig. 8d, the lid body 3 of the structure, the split mold 4.5
It may also be composed of a stand 9b which is also a structure.

In the manufacturing process of the reaction container of the present invention, considering the ease of manufacturing the reagent fixing part and the reagent adhering part in the passage, which will be described later, the structure is composed of a plurality of split molds each having at least one passage, which will be described later. It is preferable that Alternatively, it is preferable to include a structure having a passage (to be described later) in an open state, and a lid which is a structure that opens only a necessary part of the passage.

Furthermore, if the configurations shown in FIGS. 6c, 7C, and 8d are used, when the reaction is substantially completed, the reaction vessel will tilt in the direction shown by the arrow in the figure, indicating that the reaction has ended, in other words. Such a configuration is also preferable because it informs that measurement or determination is now possible. Specifically, the reaction vessel changes from the state shown in FIG. 7C to the state shown in FIG. 7D when the reaction is completed.

The material of the specimen includes various plastics such as glass, epoxy resin, polyacrylic resin, polyester resin, polystyrene resin, and polyvinyl chloride resin. It is a plastic that can be treated to make it hydrophilic.

Further, when a part of the structure, for example, a lid body, is a sheet, the material of the sheet includes those mentioned above as the material of the specimen, and metals such as aluminum.
The sheet may be thermocompression bondable to other parts of the structure or may have an adhesive laminated to one side of the sheet.

The color of the structure (including the sheet) is not particularly limited, but if the reaction results are shown in color, it is preferably transparent, or the upper sheet or split mold is transparent and the lower split mold is white. In this case, transparent is preferable.

A passage is formed within the structure or in the split mold that constitutes the specimen. Various specimens (for example, urine, serum, etc.), liquids such as washing liquids, reaction solutions, and air pass through this passage. The passageway then has at least one fluid inlet leading to an evacuation mechanism, such as an evacuable outlet.

The fluid inlet 10 (11) is one as shown in FIGS. 1a, 2a, 4, 5, 6b, 8a, 98, 10 and 12. You can use just one, or the third one.
As shown in Figures a and 3b and 7a, the knees,
Or there may be more.

If there are multiple fluid inlets, e.g. sample and reaction solution,
It is also possible to add several liquids simultaneously or in a predetermined sequence.

If there are multiple fluid inlets, please refer to Figure 3a, Figure 3b,
As shown in the examples shown in Fig. 3C, Fig. 3d, Fig. 3e, and Fig. 3f, by devising the position U of the fluid inlet 10.21, the fluid can flow in from the fluid inlet 10 located downstream from the most upstream. It is preferable that the liquid flowing into the fluid inlet 11 travels in substantially the same direction as the liquid flowing in from the most upstream fluid inlet 11.

Further, one of the fluid inlets is often provided at the most upstream side, but the fluid inlet IO may be provided in the middle portion of the passage 50, for example, as in the example shown in FIG. 9b. In the case of the example shown in FIG. 9b, the liquid that has flowed into the upstream side (the right side in the figure) of the fluid inlet 10 also ultimately flows downstream.

An example of an evacuation mechanism that communicates with the passageway is an evacuable outlet provided in a portion of the passageway.

To describe a specific example, as shown in FIG. 1a, FIG. 2a, FIG. 4, FIG. 5, and FIG. 10, one end of the passage is
Liquids added such as specimens and gases such as air can flow out.
The open end opens on the side of the specimen, and the open end serves as the outlet 20. As shown in Figure 3C, one end of the passage opens toward the F direction of body #4 and serves as the outlet 20. There are things like that.

Alternatively, as shown in Figures 6a and 6b, Figures 7a and 7b, or Figure 8a, there are two outlets (exit 20 and outlet 21) that open upward, downward, or toward the side of the specimen. There are things that I have. however,
6a and 6b, 7a and 7b, or 8a, 8b, and 8c, which show exploded plan views of each component, the liquid storage section described below 90 and in which the water-absorbing material 81 is housed in at least a portion of the liquid storage section 90, the outlet 20
Very little liquid flows out from the outlet 20, which is provided for evacuation. Further, the outlet 21 is provided to allow liquid to flow into a reagent adhesion region T having a reagent adhesion portion 40, which will be described later, and also serves as an exhaust port.

The example shown in Fig. 11 is also an example in which there are multiple outlets, but in the example shown in Fig. 11, the downstream side of the passage described later is branched, and the ends of each of the branched passages (capillary portions 52°, 53, 54) are connected to each other. are the exits 20, 21, and 22.

In this way, the outlet can be designed so that the added liquid, such as the sample, is discharged, or the added liquid remains in the passage without being discharged, and only the gas that was in the passage is discharged. It can also be designed to

In order to configure the reaction vessel of the present invention to have an exhaust mechanism communicating with the passage, it is not necessarily necessary to provide an outlet in the passage.

For example, as in the example shown in FIGS. 9a, 9b, and 9c, only the four circumferences of the upper surface of the split mold 4 having the passage are connected to the lid 3.
The upper surface of the passage portion may be later closed with the lid 3 through a space. With such a configuration, when liquid is added from the fluid inlet 10, the gas (air) that was in the passage moves to the space between the lid 3 and the split mold 4, so there is no special effect on the passage. Although no outlet is provided, liquid can flow within the passageway.

Various variations are possible in the configuration of the passage itself, as explained below.

First, regarding the directionality of the passage, for example, the passage 50 may be horizontal with respect to the specimen 2 as shown in Figs. 1a and 1b, or the passage 50 may be horizontal to the specimen 2 as shown in Figs. 3e and 3f. Additionally, it may have a horizontal portion and a vertical portion with respect to the structure 2. Alternatively, although not shown, it may be sloped from the fluid inlet toward the outlet or liquid reservoir.

The planar shape of the passage may be completely arbitrary, for example, as shown in FIGS. 1a and 1b, it may be linear,
It may also be curved, as shown in FIGS. 2a, 4, or 3b and 3c. Furthermore, the sixth
The channels may have corners and/or branches, as shown in Figures b, 7b, 8b, 9b and 11.

Note that the passage generally has a flat side surface (wall surface) as shown in FIG. 13a, but is not limited to this. The side (
It may have a protrusion 63 on the wall surface.

The cross-sectional shape of the passage 50 may be U-shaped as shown in FIG. 14a, square as shown in FIG. 14b, convex as shown in FIG. 14c, or
In addition to the ■-shape shown in Figure 14d and the W-shape shown in Figure 14e, any shape may be used, such as a circle or an ellipse.
In the example of Fig. c, the convex portion is the capillary portion 51.52, and in the examples of Figs. 14d and 14e, the sharp tip is the capillary portion 51.52.
Thanks to capillary action, the added liquid moves smoothly. Alternatively, as shown in FIG. 14f, the passage may be formed as a space with a width sufficient to support the liquid flowing therethrough.

In addition, in the present invention, the "capillary portion of the passage" refers to the first
4c, 14d and 14e,
It does not necessarily refer to a portion of the cross-sectional shape of the passage, but may also refer to a portion of the planar shape of the passage. In any case, any part of the passage whose shape is such that capillary action occurs is called a capillary part.

The cross-sectional area of the passage may be of the same size over its entire length, as shown in FIGS. 1a and 1b, i.e. all of the passage 50 may be a capillary section 51, or the cross-sectional area of the passage 50 may be of the same size throughout its length, as shown in FIGS.
As shown in Figures 3b and 3c, capillary portion 51.
In addition to 52, 53, and 54, it may have a narrow portion 60.61, a liquid retention portion 70.71, etc. Furthermore, as shown in FIG. 5, FIG. 6b, FIG. 7b, and FIG. It may have regions S and T, a reagent fixing region X having a reagent fixing portion, and an M body storage portion retaining portion 90. Furthermore, as shown in FIGS. 7b and 8b, a portion of the passage may be replaced by a hydrophilic strip 59.

Incidentally, in the present invention, the liquid retention section is provided near the fluid inlet, and serves to retain a liquid such as a sample and facilitate its addition.

The size of such a liquid retention part is determined by the amount of liquid added and the volume of the entire passage, but in particular, the passage has a liquid retention part, which will be described later, so that the added liquid is retained within the structure. In a reaction vessel, if the volume of the liquid reservoir is defined as the amount of liquid added at one time, the size is such that the following relationship holds true: (Volume of the liquid reservoir x Number of liquid additions <Volume of the liquid reservoir).

In addition, if you want to increase the volume of the liquid retention part,
As shown in Figure C, it is sufficient to take measures such as making the liquid retaining portion 70 protrude beyond the surface of the split mold 4 having the passage.

Further, the narrow portion plays the role of adjusting the inflow speed of the liquid flowing inside the passage and the role of preventing backflow of the liquid.

Therefore, FIG. 2a, FIG. 4, or FIG. 3a, 3b
As shown in the example showing the exploded plan view of each component in FIG. 3C and FIG.
．． 71 and a narrow portion 60, 61 in the vicinity thereof, it is easy to adjust the inflow speed of the liquid to be added.

In addition, in the example showing exploded plan views of each component in FIGS. 3a, 3b, and 3c, the narrow portion 60 is
A liquid retention section 70 in which liquid flowing from the most upstream fluid inlet 11 communicates with the fluid inlet 10 located downstream of the most upstream side.
It also plays a role in preventing inflows into the country.

The liquid storage part is a part that stores the specimen, reaction solution, washing liquid, etc. after the reaction is completed. Therefore, the liquid reservoir is
It is formed on the downstream side of the reagent fixing part, which will be described later.

In the example shown in FIG. 4, the liquid storage section 90 is located downstream of the reagent fixing section 30 in the capillary section 52.

In the example shown in FIG. 5, the liquid storage part 90 is located downstream of the reagent fixing part 30, but in this example, in order to store a large amount of liquid, the cross-sectional area of a part of the liquid storage part 90 is reduced. It is enlarged to have a large volume, and that part is used as a water-absorbing material storage area 80, in which a water-absorbing material 81 is stored. Also, Figures 6b, 7b, 8b and 9
The example shown in FIG.
It is 0.

The water-absorbing material refers to filter paper, a polymeric material called a water-absorbing polymer, natural fibers such as cotton, etc., and is used in the present invention by being stored in a part or the whole of the liquid storage section.

Note that examples of the water-absorbing polymer include a copolymer of polyvinyl alcohol and sodium acrylate, cellulose, and the like. Also, it is preferable that the volume thereof is not too large.

The water-absorbing material may be simply stored without being fixed, but if it is fixed, it may be done by a known method such as adhesion or encapsulation.

Further, the amount of water-absorbing material to be used is determined according to the amount of liquid to be added, and is determined to be an amount that can absorb all of the liquid.

Note that water-absorbing materials generally have air permeability, but as the amount of liquid they absorb and retain increases, the air permeability may decrease. , and the upper surface of the passage is sealed with a lid 3, as shown in FIGS. 6b and 7b, an outlet 20 is provided.
It is preferable to provide the liquid on the upstream side of the water-absorbing material storage area 80 so that the liquid can be exhausted even after the water-absorbing material has absorbed and retained the liquid.

When a liquid storage part is provided, even in the case of a configuration with an outlet, the outlet is used only for evacuation, and the entire reaction can be completed before the liquid such as the sample added sequentially flows out of the reaction vessel. This makes it easier to dispose of the sample, especially when there is a risk that the sample may contain infectious or contaminant substances. When stored, these water-absorbing materials absorb and retain the liquid such as the sample, ensuring that the added liquid such as the sample does not leak out of the container.
It is further preferred that the water-absorbing material absorbs the liquid and facilitates the movement of the liquid added from the fluid inlet within the passage.

By the way, the passage of the reaction container of the present invention includes a capillary part or a passage part which is not a capillary but has a relatively small cross-sectional area, and a part which has a large cross-sectional area and a large volume (hereinafter referred to as "passage part"). It is called the large volume section.The large volume section is
It functions as reagent adhesion regions S, T, reagent fixation region X, etc. ), it is preferable that the transition portion from the capillary portion etc. to the large volume portion widens at a predetermined acute angle, as shown in FIGS. 6b and 7b.
This is an important factor because the liquid that has flowed through the capillary portion continues to wet the large volume portion. If the transition part widens at an obtuse angle, as in the example shown in Figure 8b, the large volume part should be sloped (higher on the side transitioning from the capillary part to the larger volume part, lower on the other side, It is preferable to configure the structure so that the liquid flowing through the capillary portion or the like continues to wet the large volume portion and flows along with the action of gravity.

In addition, in the example (FIGS. 7b and 8b) in which a part of the passageway is replaced with the hydrophilic strip 59, the hydrophilic strip 59
plays the role of arbitrarily adjusting and controlling the flow rate of the liquid flowing in the passage.

That is, in a reaction container, the flow rate of the liquid flowing inside the container is closely related to the precision of the reaction. To describe this in the case of performing an immune reaction (EIA method), the flow rate is: (1) a speed suitable for completely performing the immune reaction, (2) a speed suitable for completely performing B/F separation, (2) The substrate becomes a coloring body, and the accuracy of the reaction will be increased by controlling the speed, speed, and slowest speed suitable for firmly depositing the coloring body in a predetermined position.

The flow velocity is controlled by selecting the material of the structure and adjusting the cross-sectional area of the passage, but adjusting the cross-sectional area of the passage is
There may be processing or technical difficulties, which may lead to increased costs. In such a case,
By replacing a portion of the passage with a hydrophilic strip, it is possible to easily and precisely control the flow rate simply by selecting the thickness of the hydrophilic strip. In particular, if the site replaced by the hydrophilic strip is located immediately before the liquid reservoir, the rate of all reaction steps can be controlled.

The part replacing the hydrophilic strip may or may not form a passage in the structure itself, but even if it does,
Since the passage formed in the structure itself can be rough-processed, there is no problem in terms of processing cost and technology.

In addition, as in the example shown in FIGS. 8b and 8e, when the hydrophilic strip 59 is extended into a passage formed in the structure, at least a portion of the hydrophilic strip 59 is in the hollow chamber formed in the passage. 58. To explain FIG. 8e, in the area other than the hollow chamber 58, the hydrophilic strips 5
9, the cross-sectional area of the passage will be the cross-sectional area of the capillary part 55, but in the hollow chamber 58 part, only the hydrophilic strip 59, which has a smaller cross-sectional area than the capillary part 55, becomes a passage. The flow rate can be completely controlled and the necessary and sufficient time for the reaction can be secured.

Examples of the material of the hydrophilic stripes include thread, paper, cloth, etc.
Further, the cross-sectional area is appropriately selected depending on the required reaction time, but for example, in the case of an immune reaction, when the cross-sectional shape of the hydrophilic strip is circular, the diameter is preferably about 0.2 to 1 mm.

The passage of the reaction vessel of the present invention has been described above, and next, the method for forming the passage will be described.

As in the example shown in FIG. 1a, if there is a passage 50 inside the structure 2, it may be formed by cutting the structure 2 or the like. Also, for example, as in the example shown in FIG. 2a,
When the structure 2 is composed of split molds 4.5, it is manufactured by making a mold corresponding to each of the split molds 4.5 having passages, filling the mold with a resin composition, curing it, and removing it from the mold. be able to. Alternatively, as in the example shown in FIG. 7C, when the structure 2 is composed of the lid 3 and the split mold 4, the lid 3 and the split mold 4 having the passage can be similarly molded.

Of course, even in the case of a split mold or a lid and a split mold, the passage may be formed by cutting or the like.

Then, the split molds or the lid body and the split mold are adhered with a suitable adhesive to form a structure.

By the way, when the structure 2 is composed of the split mold 4.5, or when it is composed of the lid body 3 and the split mold 4.5, the lid body 3,
The mold halves 4.5 do not have to be in contact with each other in all planes other than the passage 50.

FIG. 15 is a cross-sectional view of one embodiment of the invention having a passageway defined by a septum. As shown in Figure 15, the split mold 4
Only the portion of the partition wall 67 that defines the passage 50 formed in the
It may be bonded to the lid body 3 with an adhesive 65.

Note that in order to apply the adhesive 65 sufficiently and uniformly, it is preferable that the width W of the partition wall 67 for forming the passage 50 is small.

In addition, the examples shown in FIGS. 9a, 9b, and 9c are as follows:
This is an embodiment of the invention in which the upper surface has a passageway that is not directly closed.

As shown in FIG. 9C, the lid 3 may be bonded only to the four circumferential parts of the split mold 4.

Furthermore, as another method, there is a method of forming a passage between flat plate-like split molds by using the adhesive itself as a partition wall.

FIGS. 16a and 16b are diagrams for explaining a method of forming a passage between the flat lid 3 and the split mold 4 by using the adhesive 65 itself as a partition wall. still,
FIG. 16a is an exploded perspective view, and FIG. 16b is a partial sectional view.

In this method, an adhesive 65 is applied to the split mold 4 in a manner corresponding to the shape of the passage 50 so as to serve as a partition wall for forming the passage 50,
Next, a spacer (not shown) having the same thickness as the height of the passage 50 is sandwiched between the lid 3 and the split mold 4.
are pressed together, and the adhesive 65 is cured. As a result, the cured adhesive 65 itself serves as a partition wall, creating a reaction vessel in which the passage 50 is defined.

The adhesive used for adhering the lid and the split molds or the split molds to each other is preferably a room temperature curing adhesive that has an appropriate viscosity and does not shrink during curing. The viscosity is preferably low when the bonding area is large, and slightly high viscosity when the bonding area is small. When forming, it is preferable to use an adhesive that can be bonded in a raised manner. Examples of adhesives include epoxy adhesives, vinyl acetate adhesives, synthetic rubber adhesives, and cyanoacrylate adhesives.

Incidentally, it is preferable that adhesion of the split molds to each other or formation of the passage with an adhesive is the final step in the manufacturing process of the reaction container of the present invention.

By the way, it is good if the material of the structure is hydrophilic, but if it is not, it is better to make at least a part of the passage hydrophilic.This allows the added liquid to wet the inside of the passage and move smoothly inside the passage. will begin to flow into the country.

Methods for making the passages hydrophilic include, but are not particularly limited to, methods using a material with hydrophilic groups introduced into the surface of the horizontal body material for the passage portion, rough surface treatment such as blasting, plasma treatment, laser treatment, frost treatment, etc. Examples include the use of chemical treatment, or the application of an antistatic agent such as a cationic surfactant or a hydrophilic substance such as protein. For hydrophilization, if the horizontal body material is (meth)acrylic resin, a copolymer of methyl (meth)acrylate and sulfuric acid (meth)acrylate should be used, and if the horizontal body material is a styrene resin, the same Styrenic copolymers are preferably used.

In addition, if the structure is composed of three or more parts (split molds, or a lid body and a split mold), and a passage is formed between the split molds, for example, FIGS. 3a, 3b, and 3c. , 3rd d
As in the example shown in Figures 3e and 3f, the split mold 4
The capillary portion 52 of the passage formed in the split mold 5 is configured to communicate with the passage communication portion 57 of the capillary portion 54 of the passage formed in the split mold 5 at a passage communication portion 56 . As a result, even if the area of the structure composed of the lid 3 and the split mold 4.5 is small, a reaction vessel having a long passage can be obtained.

Furthermore, as in the examples shown in Figures 7a, 7b, 7C, and 7d, and Figures 8a, 8b, 8C, 8d, and 8e, a portion of the passage may Hydrophilic striae 59
In this case, the ends or bending points of the hydrophilic strips 59 may be fixed using an adhesive or the like in a conventional manner.

One reaction unit of the reaction container of the present invention has at least one reagent fixing portion in the passage configured as described above.

The reagent fixing part is a part located in the passage where a substance (reagent) that specifically binds to the substance to be measured is fixed, and the final reaction in the reaction container of the present invention is carried out here. Observe this part (qualitative reaction)
By this or by measuring (quantitative reaction), it is possible to detect and measure the presence or absence of the analyte in the sample or the amount of the analyte.

The reagent immobilized on the reagent immobilization part is an antibody, an antigen, or a habten, or a derivative thereof when the measurement is based on an immune reaction, and is DNA or RNA when the measurement is based on a nucleic acid piperidization reaction. In addition, any substance that specifically binds to the substance to be measured, such as lectins, receptors, and ligands, can be used as the fixation reagent.

The reagent immobilization portion is within the passageway and downstream of all fluid inlets. However, the length of the passage downstream of the reagent fixing portion is not particularly limited. If the added liquid is discharged, the reagent fixing part is provided on the most downstream side, and the added liquid is not discharged from the sample (the downstream side of the reagent fixing part of the passage is the liquid storage part). configuration), it is preferable to provide the reagent fixing part on the upstream side.

The shape of the reagent fixing portion is not particularly limited, and may be any suitable shape such as a square, circle, ellipse, or hexagon.

By the way, as mentioned above, the reagent fixing part is a place for detecting and measuring the presence or absence of the analyte in the sample or the amount of the analyte, so the reagent fixing part is a place for detecting and measuring the presence or absence of the analyte in the sample or the amount of the analyte. If the passage is configured so that the passage does not overlap with the reagent fixing portion 30 as shown in FIG.

In addition, the portion 6.7 corresponding to the reagent fixing portion 30 of the split mold 4 that does not have the lid body 3 and the reagent fixing portion 30 is the same (as in the examples shown in FIGS. 3a, 3b, and 3c). If it is made of a colorless and transparent material, the measurement accuracy will be high, especially when making judgments or measurements based on color.

Furthermore, if the split mold 5 having the reagent fixing portion 30 is also made of a colorless and transparent material, the measurement accuracy will be high, especially when the measurement is performed using transmitted light.

By the way, the reagent fixing part can be formed by fixing the immobilizing reagent in the passage, but a part of the passage is made into a large volume part, the large volume part is made into a reagent fixing area, and one reagent is fixed in the reagent fixing area. Alternatively, a plurality of reagent fixing parts may be provided. Furthermore, the number of reagent immobilization areas is not limited to one, but may be multiple.

When there are multiple reagent-fixed parts, by devising the arrangement pattern of the reagent-fixed parts, the test results can be determined more easily and with higher precision. or,
Simultaneous measurement of multiple items is possible.

Here, an example of the arrangement pattern of the reagent fixing parts in the case of having a large number of reagent fixing parts will be explained based on the drawings.

FIG. 12 shows an example in which one reagent fixing region X has two reagent fixing portions 30.31, and FIG. 7b shows an example in which one reagent fixing region 8b and 10, the capillary portion of the passage has two
An example with three or three reagent fixing sections 30, 31, 32, and FIG. 11 shows branched capillary sections 52, 53, .
54, one reagent fixing part 30.31.32
This is an example with

In the examples shown in FIGS. 7b, 8b, 10, and 12, the reagent fixing portion is provided with two reagents that do not interfere with each other.
A species or three species are fixed. That is, for example, in the case of a reaction vessel used for measuring a substance to be measured in a specimen by immunological reaction, a plurality of antibodies, antigens, or hubtains that do not have cross-reactivity with each other are immobilized.

Furthermore, in the example shown in FIG. 11, five immobilized reagents may interfere with each other.

In this way, when multiple reagents are immobilized, one type may be used as a detection reagent and the other as a control reagent, or different types of detection reagents may be immobilized to perform multi-item simultaneous measurements. good. Of course, one type of reagent may be immobilized at multiple locations.

Furthermore, in FIGS. 17a, 17b, and 17c,
An example of the arrangement pattern of a plurality of reagent fixing parts within the reagent fixing area X is shown.

In the example shown in Fig. 17a, the reagent fixing part is provided in a fan shape. For example, the detection reagent is placed at the key position (30 in the figure) and at the arc position (31 in the figure). Fix the standard product (preferably in a dilution series). When the analyte in the sample and the standard dilution series are simultaneously measured using a container with such a layout pattern of the reagent fixing part in the reagent fixing area can be compared with standard products, making it possible to perform more accurate semi-quantification.

In the example shown in FIG. 17b, the reagent fixing portions are arranged in a "+" shape. In this example, for example, 3
One reagent fixing part (31 in the figure) contains a reagent that binds or reacts with a substance that is always present in the sample and does not cross-react with the analyte. (30 in the figure) is immobilized with a reagent that selectively binds to or reacts with the substance to be measured. With this arrangement, if the substance to be measured is present in the sample, all (5)
If a binding or reaction occurs in the reagent-fixed part of the sample and the binding or reaction is detected by color development, etc., it will read as a "+" sign, and if there is no analyte in the sample, it will be detected as 17b.
In the figure, binding or reaction occurs only at the three reagent-immobilized portions indicated by 31, which can be read as an rJ shape. Therefore, it becomes easy to judge the results.

In addition, in the example shown in FIG. 8b described above, the reagent fixing part 31 lying horizontally in the same figure has a substance that binds to a substance that is always present in the sample and does not cross-react with the analyte. Alternatively, if a reagent that selectively binds to or reacts with the analyte is immobilized on the vertical reagent fixing portion 30 in the same figure, the analyte can be fixed as in the example shown in FIG. 17b. If the substance to be measured is present, it can be read as a "+" sign, and if not, it can be read as a "-" sign.

In the example shown in FIG. 17c, in addition to the same five reagent fixing parts (30 and 31 in the figure) as shown in FIG. 17b,
It has four reagent fixing parts (32 in the figure). In Figure 17c, if binding or reaction occurs at the reagent immobilization part indicated by 32, and color development based on this is observed, this indicates an operational error such as insufficient washing operation. An appropriate reagent may be immobilized on the reagent immobilization portion indicated by 32.

In this way, with a configuration having a plurality of reagent fixing parts, simultaneous measurement of multiple items and simultaneous measurement of the analyte and the control substance in the sample are possible.

In addition, fixing the reagent to the capillary part of the passage or the reagent fixing area,
Any known method may be used, and the fixation may be by chemical bonding or physical adsorption as long as it can be fixed so that the reagent does not come off during normal handling. -For example, there is a method of adsorption by heating.

In addition, the size of the reagent fixing area when providing the reagent fixing area is related to the size of other parts of the passage. preferable.

In addition to the above configuration, if there is a reagent adhering part in the passage of the reaction container of the present invention upstream of the reagent fixing part, it is possible to reduce the number of times a reaction solution, etc. is dispensed when one reaction container is used. This makes the operation even easier.

Here, the reagent adhering part is the part to which the reagent involved in the reaction is attached, and the adhesion force must be such that the adhering reagent can be removed by flowing the liquid through the reagent adhering part. Therefore, 4. For example, it is preferable to apply an aqueous solution of the reagent to an appropriate location within the passageway and freeze-dry the solution to make the reagent adhere to the reagent-attached portion.

Note that the reagent-attached portion only needs to be within the passageway and upstream of the reagent-fixed portion, so for example, as shown in FIGS.
3c, 3d, 3e, and 3f, the reagent adhering portion 40 is in the middle of the capillary portion 52, but in this example, the reagent adhering portion is inside the liquid retention portion 71. There may be.

Further, as in the example shown in FIG. 9b, a reagent adhering portion 40 may be provided in the passage upstream of the fluid inlet 10.

With such a configuration, the reagent attached to the reagent attachment portion 40 will cause the analyte in the specimen to be absorbed by the reagent fixation portion 30.
After the reagent has sufficiently reacted, it reaches the reagent fixing part 3o.

Furthermore, as in the examples shown in FIGS. 6b, 7b, and 8b, a large volume part is provided in the middle of the passage, and the large volume part is used as the reagent adhesion area S, T. Reagent adhesion parts 4o and 41 may be provided inside. The reagent is not necessarily attached to only one place in one reagent attachment area, and there may be two reagent attachment parts 41 in the reagent attachment area T, as shown in FIG. 8b. Such a reaction container can, for example, analyze antigens in a sample by EIA using the Sand-Deutsch method.
If the container is used for measurement using a method, if an enzyme-labeled antibody and an enzyme substrate are attached to the container, all reactions can be carried out simply by adding the sample.

The reagents attached to the reagent attachment portion include labeled antigens, labeled antibodies, labeled habuten,
Examples include labeled DNA, etc., or enzyme substrates when the labeling agent is an enzyme.

Further, it is preferable that a reagent adhering region is provided and its cross-sectional area is larger than the cross-sectional area of the capillary portion of the passage, since the reaction can be carried out sufficiently.

By the way, the reagent fixing part or the reagent adhering part is formed by simply fixing or adhering a predetermined reagent to the capillary part of the passage, the reagent fixing area, or the reagent adhesion slope. and/or forming recesses and/or small protrusion aggregates in the reagent attachment area;
By fixing or adhering a reagent to the recess or small protrusion to form a reagent fixing portion or reagent adhering portion, contact and reaction between the analyte in the sample and the reagent can be more reliably performed.

FIG. 18a is a sectional view of a recess-forming portion of the specimen 2 in which a recess 33a is provided in the passageway 50. Also, Section 18b
The figure is a cross-sectional view of the small protrusion forming portion of the structure 2 in which the small protrusions 35a, 35b, and 35c are provided in the passage 50. Furthermore, FIG. 18c shows that a recess 33a is provided in the passage 50, and small protrusions 35a, 35b, 35c are provided in the recess 33a.
, 35d, and 35e are sectional views of the concave portion and the small protrusion forming portion of the structure 2. Such recesses and collections of small protrusions can be made by known methods, and they may be made at the same time as the passageway is formed or afterward.

Note that when a recess is provided at a predetermined position in the passage and a reagent is fixed in the recess, a reagent fixing region X is formed as shown in FIGS. 17a, 17b, and 17c described above,
In the reagent fixing region X, recesses 33a, 33b, 33c,
33d, 33e, 33f, 33g, 33h, 33i,
It is preferable to form it in a fan shape, an r+J shape, or the like.

In addition, when the reagent fixing part is formed by a collection of small protrusions,
It is preferable that the small protrusion aggregates be arranged in a fan shape, a +j shape, or the like.

Individual small protrusions 35a constituting the small protrusion assembly 35,
35b, 35c, 35d, 35e, 35
As shown in FIGS. 19a, 19b, and 19c, the shape of f is suitably a cylinder or a square cylinder, or even a cylinder with a convex tip.

The diameter or side size of the cross section of the small protrusion is 0.3 μm
”-□ is preferably about 1.0 mm.

The height of the small protrusion is related to the cross-sectional area of the passage, but is between 0.5 and
It is preferably about 2.0 mm.

The spacing between the small protrusions is such that liquid can be retained between the small protrusions.

This kind of liquid retention is thought to be caused by surface tension and capillary action, so it is preferable that the size is such that surface tension and capillary action work.However, if it is too small, liquids such as specimens and reaction solutions may This may cause problems such as the intrusion between the small protrusions not being carried out smoothly and the cleaning during B/F separation being insufficient, so it is necessary to have a size that does not cause such problems. be. For such a size, the distance between the small protrusions is 0.
．． It is preferably about 5 to 1.5 mm.

Providing a collection of recesses or small protrusions increases the surface area, which increases the amount of reagent that can be immobilized or attached, and since the liquid is retained between the recesses or small protrusions, it is especially difficult to fix the reagent in the area. When determining or measuring by color, the color appears dark due to its depth (height).
Measurement accuracy increases.

In addition to this, the present invention also includes a reaction vessel in which a plurality of reaction units having various configurations as described above are arranged in parallel. For example, it is a reaction vessel whose partial plan view is shown in FIGS. 20 and 21.

In a reaction vessel in which a plurality of reaction units of the present invention are arranged in parallel, a large number of specimens, or a specimen and a control solution or standard solution are reacted simultaneously under the same conditions.

Also, if you change the reagent fixed for each passage, it is possible to
It is possible to simultaneously measure more items than using the container shown in FIG. 11 or 12.

The structure of the reaction vessel of the present invention has been described above, and next, the movement of liquid when the reaction vessel of the present invention is used will be explained.

When using the reaction vessel of the present invention, the movement of the added liquid can be roughly divided into five types.

The first is the case where, for example, a container shown in Fig. 1a or Fig. 2a is used, in which liquids such as specimens are added one after another.
When the passage is filled, liquid is sequentially discharged.

The second is that, as in the example shown in FIG. 4, the reagent fixing part 30 is located relatively upstream of the passage 50, and the part downstream of the reagent fixing part 30 of the passage 50 is a liquid storage part 90. is the movement of liquid in a container.

The substance to be measured in the specimen is an antigen, and a monoclonal antibody against the substance to be measured is immobilized on the reagent fixing portion 30.
FIG. 4 will be explained using an example in which an enzyme-labeled monoclonal antibody is attached to the reagent attachment portion 40.

■ The sample is added to the position indicated by ■ in the figure.

■ The washing solution is added, and the tip of the specimen is at the position indicated by ■ in the figure.

(2) The enzyme substrate solution is added, and the tip of the specimen is at the position indicated by m in the figure.

(2) A washing solution is added, and the tip of the specimen is at the position indicated by +V in the figure.

■ The chromogen solution is added, and the tip of the specimen is at the position indicated by ■ in the figure.

In this way, all of the sequentially added liquids are retained within the reaction vessel and do not flow out.

In addition, as in the example shown in FIG. 5, when at least a part of the liquid storage section 90 is a water-absorbent material storage area 80 in which the water-absorbent material 8 is stored, the added liquid is sequentially
Everything is absorbed and retained by the absorbent material 81, but the movement of the liquid is similar to the second case.

The third problem is the movement of liquid in a container with a single liquid inlet and branched passages, such as the containers shown in FIGS. 6b and 8b.

The analyte in the specimen is an antigen, a monoclonal antibody against the analyte is immobilized on the reagent fixing portion 30 in the reagent fixing region X, and the reagent attaching portion 4 in the reagent attaching region S
FIG. 6b will be explained using an example in which an enzyme-labeled monoclonal antibody is attached to 0 and an enzyme substrate is attached to the reagent attachment portion 41 in the reagent attachment region T.

■ A sample is added from the fluid inlet 10, and the sample is added to the liquid retention section 70.
satisfy.

(2) The specimen passes through the capillary sections 51 and 52 to reach the reagent attachment region S, and simultaneously passes through the capillary sections 51 and 55 to reach the reagent attachment region T.

After the capillary portion 55 and the reagent adhesion region T are filled with the specimen, the specimen is transferred from the liquid retention portion 70 to the capillary portion 51 to the capillary portion 52 to the reagent adhesion region S to the capillary portion 53 to the reagent fixing region X to the capillary portion. 54-Movement like water absorbent material storage area 80 (.

■ When the liquid retention part 70 and the capillary part 51 are emptied, the sample filled in the capillary part 55 and the reagent adhesion area T is continuously transferred from the capillary part 52 to the reagent attachment area S to the capillary part 53 to the reagent. Fixed region

In this way, by branching the passage and attaching the reagents to appropriate positions within the passage so that the reagents are supplied according to the order of the reaction, it is possible to complete the entire reaction by adding only the sample. .

The fourth case is a modification of the third case, for example, the ninth case.
As in the example shown in Figure b, the reagent attachment portion 40 is connected to the fluid inlet 1.
This is the movement of liquid in a container upstream of zero.

As described above, in the examples shown in FIGS. 9a, 9b, and 9c, the upper surface of the passage is not directly sealed, so no outlet is provided.

The substance to be measured in the specimen is an antigen, and a monoclonal antibody against the substance to be measured is immobilized on the reagent fixing portion 30.
FIG. 9b will be explained by taking as an example a case where a fluorescently labeled monoclonal antibody is attached to the reagent attachment portion 40.

■ A sample is added from the fluid inlet 10, and the sample is added to the liquid retention section 70.
satisfy.

(2) The sample flows into the left and right passages 50a and 50b of the liquid boiling section 70 in the figure, toward the liquid storage section 90 and the reagent adhesion region T.

(2) The specimen flowing into the passage 50a reaches the reagent immobilization region X, the antigen in the specimen is immobilized there, and the specimen further moves to the passage 50c-absorbent material storage area 80.

- When the liquid storage part 70 is emptied, the sample containing the fluorescently labeled monoclonal antibody dissolved in the passage 50b is transferred seamlessly from the passage 50a to the reagent immobilization area X to the passage 50.
c→absorbent material storage area 80.

In this way, if the reagent adhesion region T is provided upstream of the fluid inlet lO, the difference in time between the analyte and the label in the sample to reach the reagent immobilization region X becomes large (the analyte becomes is sufficiently immobilized in the reagent immobilization region X.

Note that if necessary, a cleaning liquid may be added from the fluid inlet 10,
It is advisable to sufficiently remove unreacted fluorescently labeled monoclonal antibodies from the reagent immobilization area.

The fifth example is an example showing a plan view of each disassembled component in Figures 3a, 3b, and 3C, and Figure 7a.
The movement of liquid in a container with multiple fluid inlets is illustrated in the example shown in FIG.

In the example shown in FIGS. 3a, 3b, and 3c, which are exploded plan views of each component, the sample added from the fluid inlet 10 is distributed between the liquid retention section 70, the capillary section 53, and the narrow section 6.
〇-capillary portion 54-moves like outlet 20, while buffer solution or reaction solution etc. added from fluid inlet 11,
It moves as follows: liquid retention part 71 - capillary part 51 - narrow part 61 - capillary part 52 - passage communication part 56 - passage communication part 57 - capillary part 54 - outlet 20. Therefore, fluid inlet lO
After the sample added from the fluid inlet 11 passes through the reagent fixing part 30, the liquid added from the fluid inlet 11 passes through the reagent fixing part 30.
pass through.

In the example shown in FIGS. 7a and 7b, which show plan views of the disassembled components, the sample added from the fluid inlet IO is
First, it passes through the reagent fixation area X, and then the fluid inlet 1.
The sample, buffer solution, reaction solution, etc. added from step 1 pass through the reagent immobilization region X.

Further, FIG. 11 also shows a container having multiple fluid inlets, and such a container is used to mix and add reagents and specimens that react with the reagents fixed to each of the reagent fixing parts 30, 31, and 32. This is advantageous when it is not possible to do so.

In the container shown in FIG. 11, after the sample added from the fluid inlet 10 passes through the reagent fixing part 30, 31, 32, the reaction solution etc. added from the fluid inlet 11, 12, 13 passes through the reagent fixing part. 30.31.32 or vice versa.
After adding different reagents and specimens from 13 and passing through the reagent fixing part 30.31.32, the reaction solution added from the fluid inlet 10 passes through the reagent fixing part 30.31.3.
It is also possible to have a configuration that passes through 2.

Next, two methods of using the reaction container of the present invention will be explained using an example in which the substance to be measured is an antigen.

The operating procedure for measuring an antigen in a specimen by the Sand-Deutsch method using a reaction vessel as shown in FIG. 1a, on which the antibody of the present invention is immobilized, is as follows.

(2) A sample predicted to contain an antigen, which is a substance to be measured, is introduced through the fluid inlet 10, and the antibody immobilized on the reagent fixing portion 30 in the passageway 50 is bonded to the antigen in the sample.

(2) A labeled antibody solution is introduced through the fluid inlet 10, and the antigen bound to the antibody immobilized on the reagent fixing portion 30 is bonded to the labeled antibody.

■ After washing if necessary, measure the amount of antigen or determine the presence or absence of antigen based on the signal shown by the label.

Further, the operating procedure when measuring by competitive method is as follows.

(2) A specimen is introduced through the fluid inlet 10, and the antibody immobilized on the reagent fixing portion 30 in the capillary passage 50 is bonded to the antigen in the specimen.

■ Pour the labeled antigen solution through the fluid inlet 10 and pass through the passage 5.
The reagent is bound to the antibody immobilized on the reagent immobilization portion 30 in 0.

■ After washing if necessary, measure the amount of antigen or determine the presence or absence of antigen based on the signal shown by the label.

In addition, the specimen and the labeled antibody in the Sand-Deutsch method, and the specimen and the labeled antigen in the competitive method, may be simultaneously introduced from a single fluid inlet.

Here, the label refers to a so-called labeling agent such as a dye, a radioactive isotope, an enzyme, a fluorescent substance, or a luminescent substance, and these labeling agents can be bound to antibodies, antigens, habuten, etc. by known methods. .

In addition, to measure the signal indicated by the label, if the labeling agent is an enzyme, add a substrate and measure the enzymatic activity, if the labeling agent is a radioactive isotope, measure the radioactivity,
If the labeling agent is a dye, fluorescence, or luminescent substance, each of them may be measured.

The operating procedure when measuring an antigen by the Sand-Deutsch method using a reaction vessel having a reagent attachment portion 40 as shown in FIG. 2a is as follows.

■ Inject the sample through the fluid inlet 10 and pass through the capillary section 5 of the passage.
The labeled antibody attached to the reagent attachment portion 40 in 2 is bound to the antigen in the specimen, and then to the antibody immobilized to the reagent fixation portion 30.

■ After washing if necessary, measure the amount of antigen or determine the presence or absence of antigen based on the signal shown by the label.

In the case of the competitive method, a reaction vessel is used in which a labeled antigen is attached to the reagent attachment portion 40 and an antibody is immobilized to the reagent fixation portion 30, and the sample is allowed to flow into the passage as in the case of the sandwich method. After the antigen in the specimen and the labeled antigen compete with each other to bind to the antibody immobilized on the reagent immobilization portion 30, the signal indicated by the label may be measured.

Further, a reagent adhering portion 40 as shown in FIGS. 3a, 3b, 3c, 3d, 3e and 3f.
When measuring an antigen by the Sand-Deutsche method using a reaction container to which a fluorescently labeled antibody is attached, by flowing the sample through the fluid inlet 10 and the buffer through the fluid inlet 11, the reagent fixing portion 30 is filled with a sample. First, the antigen in the specimen arrives and binds with the antibody immobilized there, and then the fluorescently labeled antibody arrives and reacts with the antigen-antibody complex immobilized here, so this is necessary. If so, the fluorescence intensity may be measured after washing.

In the example shown in FIGS. 6a, 6b, and 6c,
It is only necessary to introduce the sample from the fluid inlet 10. In other words, in this example, if the container is for the Sand-Deutsch method using an enzyme-labeled antibody, the monoclonal antibody is immobilized on the reagent immobilization portion 30 of the reagent immobilization area X, and the reagent adhesion area is An enzyme-labeled antibody is attached to the reagent attachment portion 40 of S, and an enzyme substrate is attached to the reagent attachment portion 41 of the reagent attachment region T. Therefore, when a specimen is allowed to flow in from the fluid inlet IO, the antigen in the specimen and the enzyme-labeled antibody located at the reagent attachment portion 40 of the reagent attachment region S are first combined, and this is bonded to the enzyme-labeled antibody located at the reagent attachment portion 30 of the reagent attachment region X. The substrate bound to and immobilized by the monoclonal antibody and attached to the reagent attaching portion 41 of the reagent attaching region T reaches here and shows a signal such as color development, so that signal can be measured.

When performing multi-item simultaneous measurements, a reaction vessel as shown in FIG. 10 is used, for example, and the following operations are performed.

■ A sample is introduced through the fluid inlet 10, and the three types of antigens in the sample are combined with the antibodies corresponding to each antigen that do not have cross-reactivity with each other and are immobilized on the reagent fixing portions 30, 31, and 32. let

■ Pour a mixture of three labeled antibodies from the fluid inlet IO,
Each antigen bound to each antibody immobilized on the reagent immobilization portion 30, 31, and 32 is combined with the corresponding labeled antibody.

■ After washing if necessary, measure the signal indicated by the label.

The types of labeling agents for the three types of labeled antibodies may be the same or different. Also, reagent immobilization portion 30.3
1.32, if the antibody corresponding to the analyte in the specimen and the antibody corresponding to the control substance are immobilized, the analyte and the control substance can be measured simultaneously.

Also, for example, FIG. 7a, FIG. 7b, FIG. 7C, and FIG.
When three types of substances to be measured are to be measured simultaneously using a container as shown in Figure d, the procedure is as follows.

(2) Inject a sample through the fluid inlet 10 and stir as necessary to obtain common recognition between the three types of antigens in the sample and the three types of antigens attached to the reagent attachment portion 40 of the reagent attachment area S, for example. An enzyme-labeled antibody formed against the site is bound to the site. Subsequently, the three types of antigen-enzyme-labeled antibody conjugates are combined with antibodies formed against non-common recognition sites of the three types of antigens immobilized on the reagent immobilization portions 30, 31, and 32 of the reagent immobilization region X. and fix it.

(2) A sample or a buffer solution is introduced through the fluid inlet 11 to dissolve the enzyme substrate attached to the reagent attachment portion 41 of the reagent attachment region T. The enzyme substrate is such that the antigen-enzyme-labeled antibody conjugate is in the reagent-immobilized portion 30.31 of the reagent-immobilized region X.
．． After being fixed at 32, the reagent fixing region X is reached.

■ After cleaning if necessary, the condition is shown in Figures 7c to 7.
When it changes to figure d, measure the signal shown by the substrate.

Such a reaction container is effective when the sample is a small amount, such as when the sample is infant serum.

In addition, in the reaction container, a mixture of three types of labeled antibodies formed against non-common recognition sites of three types of antigens may be attached to the reagent attachment area S. Furthermore, in the reaction container, one or two types of reagents are immobilized in the reagent fixation area X and the reagent adhesion area S, respectively, in order to measure one or two types of analyte. Good too.

When measuring the specimen and the standard solution at the same time, for example,
Using a reaction vessel as shown in the figure, operate as follows.

■ Sample from fluid inlet 11, fluid inlet 12 and 13
Then, put standard solutions with different concentrations into the reagent fixing part 30.
．． The same antibody immobilized on 31.32 is bound to the antigen in each sample (specimen and standard solution).

■ Pour a solution of a single labeled antibody from the fluid inlet 10,
The antigen bound to the antibody immobilized on the reagent immobilization portion 30, 31, and 32 is made to bind.

■ After washing if necessary, measure the signal indicated by the label.

The reaction vessel of the present invention has been described above, and now the features of the illustrated preferred embodiment will be briefly described.

The example shown in Figures 1a and 1b is the simplest version of the reaction vessel of the invention.

In the example shown in FIGS. 2a, 2b and 2c, the liquid reservoir 70. Since it has the narrow part 60 and the reagent adhesion part 40, the flow rate is controlled, sufficient reaction time is ensured, and the number of additions of the reaction reagent can be reduced.

The examples shown in Figures 3a, 3b, 3c, 3d, 3e and 3f have two fluid inlets, allowing multiple liquids to be added simultaneously.

Fig. 4, Fig. 5, Fig. 6a, Fig. 6b and Fig. 6C,
7a, 7b, 7c and 7d, as well as 8a, 8b, 8c, 8d and 8.
The example shown in Figure e, Figures 9a, 9b and 9c
In the example shown in the figure, since there is a liquid reservoir, the added liquid does not flow out of the container. Therefore, it is preferable because no contamination or infection occurs.

In addition, Fig. 6a, Fig. 6b, Fig. 6C, Fig. 7a,
Figures 7b, 7c and 7d, 8a and 8b
8c, 8d and 8e, the upstream passage portion including the liquid retention portion of the passage and the downstream passage portion including the liquid retention portion are located on opposite sides of the center of gravity of the structure. A reaction vessel in which the liquid that flows into the liquid reservoir from the fluid inlet flows through the passage in the bridge body toward the liquid reservoir while performing a predetermined reaction, and the predetermined reaction is substantially completed. Occasionally, a swinging means is provided so that as a result of the movement of the inflowing liquid, the side of the structure where the liquid reservoir is present is lowered. The swinging means is a leg (60th
98 in the figure), a table with a curved surface (see Figure 7C, however,
In FIG. 7C, the split mold and the base are integrally molded), or the base has a flat surface (9b in FIG. 8D).

Furthermore, the examples shown in Figures 6a, 6b, and 6C have the feature that the reaction of the entire process can be completed with a single addition of only the sample, and such containers can be molded. During molding, since the outlet 20 is located at the position shown in FIG. 6b, it is easy to release the mold.

The examples shown in Figures 7a, 7b, 7C, and 7d also have the advantage that simultaneous measurement of multiple items can be performed even with a small amount of sample, and that part of the passageway is provided with hydrophilic conditions. Since it is replaced by the body 59, the flow rate is precisely controlled and the reaction accuracy is high. Moreover, the mold releasability in the case of molding is the same as the example shown in FIGS. 6a, 6b, and 6c.

Figures 8a, 8b, 8c, 8d and 8e
The example shown in the figure is characterized in that the reaction of all steps can be completed by adding only the specimen once, and that two types of analytes can be measured simultaneously. Furthermore, this example also
Similar to the examples shown in Figures 7a, 7b, 7c and 7d, part of the passageway is replaced by a hydrophilic strip 59, so that the flow rate can be precisely controlled.

The examples shown in FIGS. 9a, 9b, and 9c are characterized in that a sufficient reaction time between the substance to be measured and the reagent immobilized in the reagent immobilization area X can be ensured.

The examples shown in FIG. 1O and FIG. 12 have a simple configuration, but are characterized in that simultaneous measurement of multiple items can be performed.

If the example shown in Fig. 11 is used for simultaneous multi-item measurement,
It has the feature of being able to simultaneously measure substances to be measured that interfere with each other, and also has the feature of being able to be used for simultaneous measurement of multiple types of analytes as another application.

The examples shown in FIGS. 20 and 21 are containers that are useful for group medical examinations and the like, and are capable of simultaneously measuring multiple items and multiple samples.

As described above, the reaction container of the present invention has many variations in its configuration and can be adapted to various measurement methods.

The reaction container of the present invention can be used not only for manual measurement and determination, but also for measurement and determination using automated equipment. For example, in the chemical reaction apparatus described in JP-A No. 63-69539, if the reaction vessel of the present invention is held in the conveying means instead of the capillary tube, automated measurement is possible.

Specifically, antibodies are immobilized in a passageway and enzyme-labeled antibodies are attached to a device that has a conveyance means such as a belt conveyor, a supply section for specimens, reagents, washing liquid, etc., and, for example, optical measurement means. After placing the reaction container of the present invention on the container and automatically injecting a sample predicted to contain an antigen, a washing liquid, and an enzyme substrate solution in sequence, the color shown by the substrate may be measured as absorbance. Note that the injected liquid may be removed by suction from the outlet. Measured values will be analyzed by computer if necessary and will be part of the diagnosis.

<Example> The present invention will be specifically described below based on Examples.

(Example 1) A pregnancy reaction using a reaction vessel having the shape shown in FIG. 2a, FIG. 2b, and FIG. 2C will be described.

■ Manufacture of reaction container A monoclonal anti-human chorionic gonadotropin (hCG) antibody is bound to the reagent fixing part 30 of the capillary part 52 of the channel of the split mold 5 made of white plastic (made of polyacrylic resin), and the antibody is bound to the insolubilized carrier. Immobilization is performed by a known method.

Next, a solution (50 μg/m° C.) of a monoclonal anti-hCG antibody labeled with alkaline phosphatase (hereinafter referred to as labeled antibody A) was added to the reagent attachment portion 40 of the capillary portion 52 of the passageway of the split mold 5 at 0.05 mβ. Drip.

After freeze-drying this, a split mold 4 made of colorless and transparent plastic (made of polyacrylic resin) is adhered to the top.

The capillary portions 51 and 52 of the passage each have a width of 3 mm.
, the depth is 0.2 mm.

■ Measurement Collect a small amount of urine from a pregnant woman into a pipette and insert it into fluid inlet 1.
The liquid is dripped from zero to fill the liquid retention section 70. Urine is
The entrance speed is controlled by the narrow portion 60, and it gradually enters the capillary portion 52 of the passage. Urine is attached to the reagent part 40
When reaching this point, the labeled antibody A attached thereto is dissolved, and the labeled antibody A is combined with hCG in the urine, while further entering the capillary portion 52 of the passage.

When the urine reaches the reagent fixing part 30, hCG and labeled antibody A
The conjugate is bound to and immobilized with the monoclonal anti-hCG antibody immobilized here. Thereafter, the urine further enters the capillary portion 52 of the passage, and the urine in which a large amount of unimmobilized excess labeled antibody A is dissolved exits the outlet 2.
reaches 0. Note that when the liquid retention section 70 becomes empty, the entry of urine stops.

Next, chromogen B (jP (5-
bromo-4-chloro-3-indolyl
When the liquid retaining portion 70 is filled with a solution of phosphate (hereinafter referred to as substrate solution A), the substrate solution A enters the capillary portion 52 of the passage, and at the same time, urine enters again, and finally all of the urine is drained. Ru.

When the substrate solution A reaches the reagent immobilized portion 30, the BCIP changes color to blue due to the action of the enzyme immobilized here.

This indicates the presence of hCG in the urine and can be determined to be pregnancy. When urine from a non-pregnant person is used, since hCG is not present in the urine, the labeled antibody A is not immobilized on the reagent fixing portion 30 and all flows out from the outlet 20. Therefore, the reagent fixing portion 30 is not colored.

(Example 2) A pregnancy reaction using reaction vessels having the shapes shown in FIGS. 3a, 3b, 3c, 3d, 3e and 3f will be described. However, in addition to the illustrated reagent attachment portion 40, a container was used that had a reagent attachment portion in which a reagent was also adhered within the liquid retention portion 70.

■Manufacture of reaction container A monoclonal anti-hCG antibody is placed in the reagent fixing portion 30 of the capillary portion 54 of the channel of the split mold 5 made of white plastic (made of polystyrene resin) using a known method for binding the antibody to an insolubilized carrier. to be fixed.

Next, the labeled antibody A is placed in the liquid retention portion 70 of the split mold 5.
(Already mentioned) solution (100 μg/mA) was added to 0.05 m42
It is dripped and freeze-dried, being careful not to enter beyond the narrow part 60. Separately, substrate solution A (already mentioned, 10 μg/m Q ) was added to the reagent adhering portion 40 of the capillary portion 52 of the channel of the split mold 4, which is made of transparent plastic in the part indicated by 7 in Figure 1 and the rest is white. Drop and freeze-dry.

The split mold 4 is glued onto the split mold 5 which has undergone the above treatment, and the lid body 3 made of transparent plastic (made of polystyrene resin) where the part indicated by 6 in the figure is transparent and the rest is white is placed on top of it. Glue.

The capillary portions 51, 52, 53, and 54 of the passage each have a width of 2 mm and a depth of 0.2 mm.

■ Measurement Collect a small amount of urine from a pregnant woman into a pipette and insert it into fluid inlet 1.
At the same time, the liquid is dripped from zero to fill the liquid retention section 70.
Labeled antibody A attached here is dissolved. Then, the same urine is immediately dripped from the fluid inlet 11 to fill the liquid retention section 71.

The urine filled in the liquid retention part 70 passes through the narrow part 60,
It gradually enters the capillary portion 54 of the passage. At this time, hCG in the urine and labeled antibody A enter the capillary portion 54 of the passageway while binding to each other. When the urine reaches the reagent immobilization area 30, the combination of hCG and labeled antibody A combines with the monoclonal anti-hCG antibody immobilized here, and is immobilized. Thereafter, the urine further enters the capillary portion 54 of the passage, and the urine, in which a large amount of unimmobilized excess labeled antibody A is dissolved, reaches the outlet 20.

On the other hand, the urine filled in the liquid retention part 71 passes through the narrow part 61 and advances in a meandering manner within the capillary part 52 of the passage, reaches the reagent attachment part 40, and the BCIP attached thereto.
(substrate, already mentioned) is dissolved. Urine is passed through the communication section 56 of the passage.
．． 57 into the capillary portion 54 of the passageway. Note that the total length of the capillary portion 52 of the passage is longer than the total length of the capillary portion 54 of the passage. After that, the reagent fixing part 30 is reached.

When the urine in which the substrate has been dissolved reaches the reagent fixing part 30, the substrate turns blue over time due to the action of the enzyme fixed there, indicating the presence of hCG in the urine, that is, the used urine. It is confirmed through the 6 parts of the lid body 3 and the 7 parts of the split mold 4 that the urine is pregnant urine. If hCG is not present in the urine, the labeled antibody A will not be immobilized on the reagent immobilization portion 30 and will all flow out from the outlet 20. Therefore, the reagent fixing portion 30 does not turn blue.

(Example 3) Simultaneous automated measurement of three cancer markers using a reaction container having the shape shown in FIG. 20 will be described.

■ Manufacture of reaction container Reagent fixing part 30.31.3 in one passage 50 of a split mold made of white plastic (made of polystyrene resin)
2, hCG, CEA, a-fetoprotein (hereinafter 3)
A monoclonal antibody directed against a cancer marker (species cancer marker) is each immobilized by a known method for binding antibodies to an insolubilizing carrier. Similarly, in other passages 50,
Immobilize three types of cancer markers.

Next, place a colorless and transparent plastic (
Glue the lid made of polystyrene resin.

The reaction vessel 1 thus obtained has ten passages 50 each having a width of 3 mm and a depth of 0.3 mm arranged in parallel.

■ Automated measuring device The automated measuring device used here consists of a transport means that can be moved vertically and horizontally at a constant pitch, a supply section for specimens, reagents, washing liquid, etc., and a suction means that sucks the fluid in the capillary passage from the outlet. , and optical measuring means.

The automated measuring device used here, excluding the measuring means,
The structure is such that it can process 10 samples at the same time.

■ Measurement The reaction container is placed on the transport means of the automated device.

When the reaction container is transported and reaches a predetermined position, human serum NIILI to Nll0 is supplied to each passage 50 from the 10 sample supply nozzles in the sample supply section through each fluid inlet 10.
The substance to be measured in the serum (3 types of cancer markers)
binds to each antibody immobilized on the reagent immobilization portion 30, 31, 32 and is immobilized.

After 5 minutes have passed (during which time the reaction container is transported to the washing liquid supply section), human serum NLL1 to NILIO are suctioned out from each outlet 20 of each passage 50. Subsequently, a cleaning liquid is supplied to each passage 50, and this is also removed by suction.
Note that the washing and suction operations are performed five times.

When the reaction container is transported to the reagent supply section, a buffer solution containing an enzyme-labeled antibody (in this case, a mixture of antibodies against hCG, CEA, and α-fetoprotein labeled with alkaline phosphatase) is supplied.

The enzyme-labeled antibody is immobilized by binding to the antibody immobilized on the reagent immobilization portion 30, 31, and 32 by binding to the corresponding cancer marker. It does not bind to immobilized antibodies that do not have cancer markers immobilized on them.

After 5 minutes (during which time the reaction container is transported to the washing liquid supply section), the enzyme-labeled antibody solution is removed by suction. Subsequently, a cleaning liquid is supplied to each passage 50, and this is also removed by suction. The cleaning, washing, and suction operations are performed five times.

When this operation is completed and the reaction container is transported to the reagent supply section, substrate solution A (already introduced) is supplied to each passage 50.

In the case of this example, the reagent-immobilized portion on which the enzyme-labeled antibody is immobilized turns blue due to the presence of the cancer marker.
This blue coloration is proportional to the concentration of cancer marker contained in the sample serum.

Next, the reaction container is transported to the position of the measuring means. Here, the color development of the reagent-fixed portions 30, 31, 32 is continuously measured photometrically, and each cancer marker in each serum is quantified.

The reaction containers are automatically fed one after another, and a large number of serums are processed one after another.

(Example 4) Simultaneous automated measurement of three cancer markers using a reaction container having the shape shown in FIG. 21 will be described.

③ Manufacture of Reaction Vessel Capillary portions 52, 53 of passages of first unit 100 in split mold made of white plastic (made of polystyrene resin).
h to each reagent fixing part 30, 31, and 32 in 54, respectively.
Monoclonal antibodies against CG, CEA, and α-fetoprotein (hereinafter referred to as three types of cancer markers) are immobilized by a known method for binding antibodies to an insolubilized carrier. Similarly, three types of cancer markers are immobilized at the same position in the passages of the second unit 200 and subsequent units.

Next, place a colorless and transparent plastic (
Glue the lid made of polystyrene resin.

Note that the capillary portions 51, 52, 53, and 54 of the passage of the reaction vessel I obtained in this manner each have a width of 5 mm and a depth of 0.
．． It is 5mm.

■ Automated measuring device The automated measuring device used here has a transport means that can be moved at a constant pitch, a supply section for specimens, reagents, washing liquid, etc., and a suction means that sucks the fluid in the capillary passage from the outlet. .

■ Measurement The reaction container is placed on the transport means of the automated device.

Once the reaction vessel has been transported and reaches the predetermined position, human serum seed 1 is transferred from the specimen supply through the fluid inlet 10 of the first unit 100 to the capillary section 5 of the passageway of the first unit 100.
1.

Serum flows from the capillary portion of the passageway 51 to the capillary portion of the passageway 52.
53.54 and reaches the reagent fixing part 30.31.32, the substance to be measured (three types of cancer markers) in the serum is detected.
binds to each antibody immobilized on the reagent immobilization portion 30, 31, 32 and is immobilized.

After 2.5 minutes have passed (during this time, the reaction vessel 1 is transported so that the fluid inlet 1o of the first unit 100 is located in the cleaning liquid supply section A, and the fluid inlet 10 of the second unit 200 is located in the sample supply section). , human serum seed 2 is extracted from the specimen supply section from the second human serum seed 2.
The second unit 20 via the fluid inlet lO of the unit 200
0 to the capillary section 51 of the passage.

At the same time, the suction removal operation of human serum No. 1 from the outlet 20.21.22 of the first unit 100, followed by the supply of washing liquid from the fluid inlet 10 of the first unit 100 and the operation from the outlet 20.21.22. Suction removal (5 times)
will be carried out.

Furthermore, after 2.5 minutes have elapsed after the suction removal operation of human serum Nα,1 (during this period, the reaction vessel 1 is connected to the reagent supply section A and the fluid inlet 11, 12, 13 of the first unit 100 is connected to the sample supply section). ), the fluid inlet 1 of the first unit 100 is conveyed so that the fluid inlet of the third unit (not shown) is located
1, a buffer solution in which an antibody against hCG labeled with alkaline forsterase is introduced from the fluid inlet 12,
A buffer in which an antibody against CEA labeled with alkaline forsterase is dissolved is supplied from the fluid inlet 13, and a buffer in which an antibody against α-fetoprotein is labeled with alkaline forsterase is supplied from the fluid inlet 13.

These enzyme-labeled antibodies are immobilized by binding to the antibodies immobilized on the reagent immobilization portions 30, 31, and 32 by binding to the corresponding cancer markers. It does not bind to immobilized antibodies that do not have cancer markers immobilized on them.

After 2.5 minutes have passed after supplying the enzyme-labeled antibody (during this time,
The reaction container 1 is transported so that the fluid inlet 10 of the first unit 100 is located in the cleaning liquid supply section B, and the fluid inlet of the fourth unit (not shown) is located in the sample supply section), and the outlet 20, 21 of the first unit 100 is located in the sample supply section. The buffer solution in which the enzyme-labeled antibody was dissolved is removed by suction from .22, followed by the supply of washing liquid from the fluid inlet IO of the first unit 100 and the suction removal from the outlet 20.21.22 (5 times). be exposed.

Furthermore, after 2.5 minutes have elapsed after the suction removal operation of the buffer solution in which the enzyme-labeled antibody has been dissolved (during this period, the fluid inlet 10 of the first unit lOO of the reaction vessel 1 is connected to the reagent supply section B, and the fluid inlet 10 of the first unit lOO is connected to the sample supply section). (transferred so that the fluid inlet of the fifth unit (not shown) is located), the fluid inlet 1 of the first unit 100
From 0, substrate solution A (already described) is supplied.

In the case of this example, since the cancer marker is present, the substrate is immobilized on the reagent immobilization region where the enzyme-labeled antibody is immobilized, and the region is colored blue.

This coloration remains even after substrate solution A is removed, so
After 2.5 minutes have passed after the supply of substrate solution A (during this period, the fluid inlet 10 of the first unit 100 is located in the washing liquid supply section C of the reaction vessel 1, and the fluid inlet of the sixth unit (not shown) is located in the sample supply section). ), the first unit 1
Suction removal operation of substrate solution A from outlet 20.21.22 of 00 followed by fluid inlet 1 of first unit 100
After the automated device performs the supply of washing liquid from 0 and suction removal from the outlet 20.21.22 (5 times), the reagent fixed part 30.31.32 is observed with the naked eye to check for the presence or absence of cancer markers. judge.

Incidentally, the serum supplied to the passages after the second unit 200 is also sequentially processed in the same manner with a one-step delay.

(Example 5) Diagnosis of hepatitis B using reaction vessels having the shapes shown in FIGS. 2a, 2b, and 2c will be described.

■ Manufacture of reaction container A heat-denatured solution of DNA fragments corresponding to hepatitis B virus DNA (
5 μg/ml2) was added dropwise in an amount of 201 L, left at 25° C. for 24 hours, removed by suction, and further irradiated with ultraviolet rays to fix the DNA piece.

Next, biotin-labeled hepatitis B virus DNA is added to the reagent attachment portion 40 of the capillary portion 52 of the passageway of the split mold 5.
l of probe solution (0,02 u g/mE)
Oμnf4 down.

After freeze-drying this, a lid 3 made of colorless and transparent glass (made of polystyrene resin) is adhered to the upper part.

The capillary portions 51 and 52 of the passage each have a width of 3 mm.
, the depth is 0.2 mm.

■ Measurement DNA was extracted from the serum of a hepatitis patient and used as a specimen.

The sample is dropped from the fluid inlet 10 to fill the liquid retention section 70. The specimen gradually enters the capillary portion 52 of the passageway, with its entry speed being controlled by the narrow portion 60.
When the specimen reaches the reagent attachment portion 40, it dissolves the biotin-labeled DNA probe attached thereto, and further enters the capillary portion 52 of the passage while binding the biotin-labeled DNA probe and hepatitis B virus. When the specimen reaches the reagent immobilization part 30, it binds to the DNA piece corresponding to the hepatitis B virus immobilized there and is immobilized. Thereafter, the sample further enters the capillary portion 52 of the passageway, and the sample that has dissolved a large amount of unimmobilized excess biotin-labeled probe reaches the outlet 20. Note that when the liquid retention section 70 becomes empty, the specimen stops entering.

Next, a previously prepared avidin-biotin labeled peroxidase conjugate solution (lOOug/ml2
) is dripped from the fluid inlet 10 to fill the liquid reservoir 70. As the avidin-biotin labeled peroxidase conjugate solution enters the capillary portion 51 of the channel, the analyte re-enters and eventually all of the analyte is expelled.

When the avidin-biotin-labeled peroxidase conjugate reaches the reagent immobilization portion 30, it binds to the biotin immobilized there and is immobilized. Thereafter, the avidin-biotin labeled peroxidase conjugate solution further enters the capillary portion 52 of the passage and reaches the outlet 2o. still,
When the liquid reservoir 70 is emptied, the avidin-biotin labeled peroxidase conjugate solution stops entering.

After this, 0.076M phosphate buffered saline (pH 7,
0) (PBS) is dripped from the fluid inlet 10 to fill the liquid retention section 7o.

The PBS gradually enters the capillary portion 51 of the passageway through the narrowing 60 and into the capillary portion 52 of the passageway, forcing out the avidin-biotin labeled peroxidase conjugate solution. P.B.
When S reaches the reagent fixing portion 30, cleaning is started.

Thereafter, the PBS further enters the capillary section 52 of the passage and reaches the outlet 2o. Note that when the liquid retention section 70 becomes empty, the PBS stops entering.

When the liquid retention part 7o is empty, the liquid retention part 7o is filled with a mixed solution of water peroxide (substrate) and orthophenylenediamine (chromogen). As soon as the mixed solution enters the capillary section 51 of the channel, the PBS is expelled.

When the mixed solution reaches the reagent immobilization part 30, the chromogen changes color to yellow due to the action of the enzyme immobilized here. This indicates the presence of hepatitis B virus in the serum, and can be determined to be highly infectious.
If hepatitis B virus is not present in the serum, the enzyme will not be immobilized on the reagent immobilization portion 30 and will all flow out from the outlet 20. Therefore, the reagent fixing portion 30 is not colored.

(Example 6) Figures 8a, 8b, 8c, 8d and 8e
Measurement of LH using a reaction vessel having the shape shown in the figure will be explained.

(2) Manufacture of reaction container A lid 3 and split molds 4 and S made of colorless and transparent plastic (made of epoxy resin) are molded using a mold. The capillary portions 51, 52, 53, 54 and 55 of the passage each have a width of 0.7 mm and a depth of 0.7 mm. Next, the capillary portion 55 containing the hollow chamber 58 of the passageway of the split mold 4 is filled with a diameter of 0.
．． Cotton yarn with a circular cross section of 5 mm (hydrophilic strips 5
9) are fixed with adhesive.

Anti-LH-
After the β antibody is immobilized by a known method for binding antibodies to an insolubilizing carrier, the split mold 5 is adhered. Further, an anti-mouse IgG antibody is immobilized on the reagent immobilization portion 31 (in this portion, a passage is formed above the split mold 4) by a known method for binding the antibody to an insolubilized carrier.

Next, a monoclonal anti-LH-〇 antibody labeled with alkaline phosphatase (hereinafter referred to as labeled antibody B) solution (lolig/mg
) is added in 2 drops of 0.5 mj.

Furthermore, the substrate solution A (
Two drops of 0.2 mj of 2 mg/m 12 ) were added and freeze-dried.

Thereafter, 30 mg of nonwoven fabric is similarly stored in the water-absorbent material storage area 80 of the split mold 4, the lid 3 is adhered, and the stand 9b is further adhered.

■Measurement Collect 400μρ of urine into a pipette, drop it from the fluid inlet 10, and fill the liquid retention part 70. Urine enters the capillary portion 51 of the passageway, passes through the capillary portion 52 of the passageway, reaches the reagent adhesion region T, and absorbs the BCrP (
Dissolve the substrate (already mentioned). When the reagent adhesion area T is filled with urine, the urine in the liquid retention part 70 reaches the reagent adhesion area S and dissolves the labeled antibody B attached there, but at the same time, the LH in the urine dissolves into the labeled antibody B. Join. The urine is further collected in the capillary portion 53 of the passage, the reagent fixing portion 30, the capillary portion 54 of the passage, the reagent fixing portion 31 . It passes through the cotton yarn (hydrophilic strip 59) and reaches the water absorbent material storage area 80, where it is absorbed and retained by the nonwoven fabric (water absorbent material 8). In this process, a conjugate of LH and labeled antibody B (L H-labeled anti-LH-
〇Antibody) is anti-LH immobilized on the reagent immobilization part 30
- It binds to the β antibody and at the same time binds to the anti-mouse IgG antibody immobilized on the reagent immobilization portion 31, thereby immobilizing each of them. When the liquid retention section 70 is emptied, the urine with dissolved substrate flows out from the reagent adhesion area T, and the passage capillary portion 52, the passage capillary portion 51, the reagent adhesion area S, and the passage capillary portion 5.
3. It passes through the reagent fixing part 30, the capillary part 54 of the passage, the reagent fixing part 31, and the cotton thread (hydrophilic strip 59) to reach the water absorbent material storage area 80, and is absorbed and retained by the nonwoven fabric (water absorbent material 8i). be done. During this process, the substrate changes color to blue due to the action of the enzyme immobilized on the reagent immobilization parts 30 and 31, which can be read as "+". This indicates the presence of LH in the urine (sample) and can be determined to be positive.
On the other hand, when urine from a person lacking LH is used, labeled antibody B
is not immobilized on the reagent fixing part 30 and is attached to the reagent fixing part 31.
It is fixed only in

Therefore, the substrate changes color to blue due to the action of the enzyme immobilized on the reagent immobilized portion 31, but the reagent immobilized portion 30 is not colored and can be read as "-".

The total reaction time was approximately 5 minutes, and LH in urine was 50 m
If the concentration is IU/mρ or more, it can be determined as "+".

<Effects of the Invention> The present invention provides a reaction vessel in which highly sensitive measurements and operations, particularly B/F separation operations, can be performed accurately and easily.

The reaction container of the present invention is widely applicable to various reactions, such as detection methods using mechanisms such as ETA and nucleic acid hybridization, and is also applicable to multi-item measurements with simple operations, and furthermore, it can be applied to detection methods using the mechanisms of ETA and nucleic acid hybridization. It is very useful because it can be applied to both measurements using automatic measurement equipment and measurements using automatic measurement equipment.

The reaction container of the present invention is applicable not only to qualitative determination but also to quantitative measurement, and is particularly useful in that it has achieved the simplification of quantitative measurement, which has been difficult in the past.

[Brief explanation of drawings]

FIG. 1a is a perspective view of an embodiment of the present invention, and FIG. 1b is a perspective view of an embodiment of the present invention.
It is a sectional view of the passage part. FIG. 2a is a perspective view of one embodiment of the present invention, and FIGS. 2b and 2c are views taken along the line A-A and line B-B. Figures 3a, 3b and 3c are plan views of exploded components of an embodiment of the present invention, Figure 3d is a side view thereof, and Figures 3e and 3f are C-- C line and D-D I! FIG. FIG. 4 is a perspective view of one embodiment of the present invention. FIG. 5 is a plan view of one embodiment of the present invention. FIGS. 6a and 6b are plan views of exploded components of an embodiment of the present invention, and FIG. 6c is a side view thereof. 7a and 7b are plan views of exploded components of an embodiment of the present invention, and FIGS. 7c and 7d are side views thereof. Figures 8a, 8b, and 8C are plan views of exploded components of an embodiment of the present invention, Figure 8d is a side view thereof, and Figure 8e is a portion A of Figure 8b. It is an enlarged sectional view. FIGS. 9a and 9b are plan views of exploded components of an embodiment of the present invention, and FIG. 9c is a cross-sectional view taken along the line X--X. FIG. 10 is a plan view of one embodiment of the present invention. FIG. 11 is a plan view of one embodiment of the present invention. FIG. 12 is a plan view of one embodiment of the present invention. FIGS. 13a, 13b, and 13c are plan views showing examples of the shape of passages in the reaction container of the present invention. FIGS. 14a, 14b, 14c, 14d, 14e, and 14f are cross-sectional views showing an example of the shape of the passageway of the reaction vessel of the present invention. FIG. 15 is an exploded sectional view for explaining one method of forming a reaction container of the present invention. Fig. 1.6a is an exploded perspective view for explaining one method of forming a reaction vessel of the present invention, and Fig. 16b is a sectional view showing a passage portion of the reaction vessel. FIG. 1.7a, FIG. 17b, and FIG. 17c are schematic diagrams for explaining the arrangement pattern of a plurality of reagent fixing parts. FIG. 18a shows the recess provided in the passage, FIG. 18b shows the small projection assembly provided in the passage, and FIG.
The figure is a schematic cross-sectional view for explaining a recess and a small projection assembly provided in a passage. FIG. 19a, FIG. 19b, and FIG. 19c are schematic diagrams showing an example of a small protrusion aggregate. FIG. 20 shows the following. FIG. 21 is as follows. Partial plan view of one embodiment of the present invention Partial plan view of one embodiment of the present invention Description of symbols 1... Reaction vessel, 2... Structure, 3... Lid (of the structure), 4.5 ...split mold (of the structure), 6.7... part corresponding to the reagent fixing part. 8... Seat, 9a... Leg, 9b... Stand, io, 11.12.13... Fluid inlet, 20.21.
22... Outlet, x, y, z... Reagent fixing area, 30.31.32... Reagent fixing part, 33a, 33b
, 33c, 33d, 33e, 33f, 33g, 33h,
33i... recess, 35... small protrusion aggregate, 35a, 35b, 35c, 35d, 35e, 35f...
・Small protrusion, S, T...Reagent adhesion area, 40.41.42...Reagent adhesion part, 50.50a,
50b, 50c-Aisle, 51.52.53.54.5
5... Capillary part, 56.57... Passage communication part, 58... Hollow chamber, 59... Hydrophilic stripe, 60.61... Narrow part, 63... Protruding part 65...Adhesive, 67...Partition wall, 70.71...Liquid retention section, 80...Water absorbent material storage area, 81...Water absorbent material, 90...Liquid storage section, 100 ... 1st unit, 200... 2nd unit FIG, 1a 〆FIG, 1b 50 (51) 2 0 76053 FIG, 3d 1 0] Mouth 0 1 7° 7 1) 4 FIG, 4 ＼FIG, 5 0 FIG, 6a 6b 0 FIG, 6c FIG, 8d FIG, 8e 8 FIG, 9a 0 FIG, 9b FIG, 10 FIG, 12 2 FIG, 13a FIG , 13b FIG, 13c F I 0.14a F I 0.14b lG44c 1 F I G, 14d FIG, 14e FIG, 14f F I G, 16b 50 33b (31) 3a 5d 5a ”F 35a 35b 35c \ \ 5a 5b lG20/

Claims (1)

Scope of Claims: (1) The structure has a passage having at least one fluid inlet, and at least one reagent fixing part is provided in the middle of the passage and downstream of all the fluid inlets. have, and
A reaction vessel comprising at least one reaction unit having an exhaust mechanism communicating with a passage. (2) The reaction vessel according to claim 1, wherein the evacuation mechanism is at least one evacuable outlet provided in the passageway. (3) At least one
The reaction container according to claim 1 or 2, having a reagent attachment portion. (4) The reaction container according to claim 3, wherein at least one of the reagent-attached portions is located upstream of the fluid inlet. (5) Claims 1 to 3, wherein the reagent fixing portion and/or the reagent adhering portion are a recess and/or a small protrusion aggregate.
4. The reaction container according to any one of 4. (6) The reaction vessel according to any one of claims 1 to 5, further comprising at least one liquid retention portion near the fluid inlet of the passage. (7) The reaction container according to any one of claims 1 to 6, further comprising a liquid storage portion downstream of the reagent fixing portion of the passage. (8) The reaction vessel according to claim 7, wherein a water-absorbing material is housed in the liquid storage section. (9) The reaction vessel according to claim 8, wherein the water-absorbing material is absorbent cotton. (10) The reaction vessel according to claim 8 or 9, wherein the outlet that can be evacuated is provided near the water-absorbing material storage section. (11) Claims 8 to 1 in which at least a part of the passage between the reagent fixing part and the water-absorbing material is made of a hydrophilic strip.
0. The reaction container according to any one of 0. (12) The reaction container according to claim 11, wherein at least a portion of the hydrophilic strip is expanded into a hollow chamber formed in the passage. (13) The reaction vessel according to any one of claims 1 to 12, wherein the passage is horizontal. (14) The reaction vessel according to any one of claims 1 to 13, wherein the passage has a capillary portion. (15) Claims 1 to 1, wherein a part of the passage has a narrow part.
4. The reaction container according to any one of 4. (16) In the passage,
A reagent fixing area having a reagent fixing part and/or a reagent adhering area having a reagent adhering part are formed, and the cross-sectional area of the reagent fixing area and/or the reagent adhering area is larger than that of the passage liquid retaining part and the liquid retaining part. The reaction vessel according to any one of claims 1 to 15, wherein the reaction vessel has a cross-sectional area larger than that of the portion. (17) The reaction vessel according to any one of claims 1 to 16, wherein the structure is composed of a plurality of split molds each having at least one of the passages. (18) The reaction vessel according to claim 17, wherein the structure includes a split mold having the passage and a lid having no passage. (19) The reaction container according to claim 18, wherein the lid body is in close contact with the split mold having the passage. (20) The reaction container according to claim 18, wherein a space is provided between the lid and the split mold having the passage. (21) Claims 1 to 20 wherein the structure is composed of three or more split molds or a lid and a split mold, and a passage formed between adjacent split molds communicates with another adjacent passage. The reaction container according to any one of. (22) Of the at least one fluid inlet, the liquid flowing in from the fluid inlet downstream of the most upstream fluid inlet is configured to travel in substantially the same direction as the liquid flowing in from the most upstream fluid inlet. The reaction vessel according to any one of claims 1 to 21. (23) The reaction vessel according to any one of claims 1 to 22, wherein at least a portion of the structure is made of a hydrophilic material. (24) an upstream passage portion including a liquid retention portion of the passage;
The downstream passage portion including the liquid storage portion is a reaction vessel located on opposite sides of the center of gravity of the structure, and the liquid flowing into the liquid storage portion from the fluid inlet undergoes a predetermined reaction through the passage within the structure. 24. A structure according to claim 7, wherein the liquid flows toward the liquid storage part, and when a predetermined reaction is substantially completed, the side of the structure where the liquid storage part is present descends as a result of the movement of the inflowing liquid. The reaction vessel described in Crab. (25) The structure has a swinging means at a position where the side where the downstream passage portion including the liquid storage portion exists is lowered as a result of the liquid movement according to claim 24.
The reaction vessel described in . (26) The swinging means is a leg provided at the bottom of the structure, or a table having a flat or curved surface.
The reaction vessel described in . (27) A reaction vessel comprising a plurality of reaction units according to any one of claims 1 to 26 arranged in parallel.