JP4344743B2 - Immunoassay test equipment - Google Patents

Description

  The present invention relates to an immunoassay test instrument used for an assay using an immunochromatography method such as an enzyme-labeled immunochromatography method. More specifically, the present invention relates to an immunoassay test instrument that is low in production cost, can stably hold a liquid reagent until the start of reaction, and can start an immunoassay by an easy operation.

  In recent years, an immunochromatographic method that can easily detect various substances in a sample using an immunological reaction such as an antibody-antigen reaction has attracted attention in a wide range of fields including clinical diagnosis.

  For example, in enzyme-labeled immunochromatography, a test substance in a sample, for example, a first immunological substance that reacts specifically with an antigen, for example, an antibody is immobilized on a stationary phase comprising a water-absorbing substrate. In addition, a sample containing the test substance is dripped upstream from the fixing part to form a complex with an enzyme-labeled second immunological substance that can bind to the test substance, for example, a labeled antibody. And by allowing a developing solution containing a chromogenic substrate for the labeling enzyme to flow from the upstream side, the complex is developed and moved to the position of the immobilized first immunity substance, and is bound to the first immunity substance. An immunoassay is performed.

  By the way, a disposable test instrument that can store a drug, a reagent, and the like necessary for a test such as the immunochromatography method in a compact casing and can start a test simply and quickly is used for clinical diagnosis and the like. It is useful in the field and various types have been developed.

  For example, Patent Document 1 discloses a stationary phase made of a water-absorbing base material stretched in the longitudinal direction, a case that accommodates the stationary phase, and a case having a specimen dropping window and a determination window on the upper surface thereof. A liquid container covered with a covering member housed in the case, a portion of the case located at the top of the liquid container having a protrusion on the inner surface for breaking the covering member, and A test instrument is disclosed that is deformable so that the projection can be pressed against the covering member.

  If a test instrument as shown in Patent Document 1 is used, it is stored in the test instrument in advance without requiring an operation such as adding a drug, a reagent, or the like required for a separately prepared test to the reaction system. The inspection can be started by easily breaking the liquid storage container containing the medicine, reagent, and the like, from the outside of the test instrument.

However, in the test instrument shown in Patent Document 1, in order to form the deformable portion as described above, a part of the wall surface of the case must be molded as a thin wall, and technically in terms of the molding. In general, a test container having such a deformable portion will be covered with some protective material before use, but these protective materials are used during transportation and storage. If any external pressure exceeding the capacity of the tester is applied to the test container, the liquid container will be broken and the medicine, reagent, etc. stored inside will leak into the container, and the test instrument will no longer be usable. There was a high possibility that the problem would occur.
JP-A-10-104236

  Therefore, an object of the present invention is to provide an immunoassay test instrument that solves the above-described problems in the prior art. Another object of the present invention is to provide an immunoassay test instrument that is low in production cost, can stably hold a liquid reagent until the start of the reaction, and can start an immunoassay with an easy operation.

That is, the present invention for solving the above problems is an immunoassay test instrument used for an assay using an immunochromatographic method,
A stationary phase consisting of a water-absorbing substrate stretched in the longitudinal direction;
A liquid storage container that is disposed near one end in the longitudinal direction of the stationary phase, stores liquid therein, and is formed of a film material that can be broken at least partially;
A casing containing the stationary phase and the liquid storage container, the casing main body having a sample dropping window and a determination window on the upper surface thereof, and having an opening at a position engageable with the liquid storage container And a casing that is slidably attached to the opening and includes a slider having a breaking means for the liquid container.

  In the present invention, the stationary phase has a detection line formed by immobilizing a first immunity substance that specifically reacts with a test substance in a sample under the determination window, and under the sample dropping window. And a conjugate pad containing an enzyme-labeled second immunity substance that can bind to a test substance in a dry state, and the liquid container contains the second immunity substance. 1 shows an immunoassay test instrument characterized by containing a developing solution containing a chromogenic substrate for a labeled enzyme labeled.

  According to the present invention, the stationary phase further comprises another immunity substance capable of reacting with a second immunity substance for confirming whether a series of immune reactions are normally performed under the determination window. 1 shows an immunoassay test instrument characterized by having a confirmation line (hereinafter referred to as a control line) fixed.

  According to the present invention, a liquid container containing a reagent such as a developing solution housed inside an immunoassay test instrument can be easily moved from the outside by moving a slider slidably attached to a casing. It is possible to start an immunoassay reaction by developing the reagent inside the casing and developing the reagent inside the casing, and the slider and the casing body are difficult in terms of molding technology, such as forming a particularly thin wall portion or the like. The manufacturing cost is low because it can be manufactured without difficulty, and the slider functions by sliding. For example, even if an external force is applied from the outside of the package containing the product, it is less likely to operate erroneously. The liquid reagent can be held stably until the reaction starts. As described above, the immunoassay test instrument of the present invention is low in production cost, can stably hold a liquid reagent until the reaction starts, and can start an immunoassay with an easy operation.

  Hereinafter, the present invention will be described in detail based on specific embodiments.

  FIG. 1 is a plan view showing an assembly process of an embodiment of a test instrument for immunoassay according to the present invention.

  An immunoassay test instrument 10 according to the present invention is an immunoassay test instrument used for an assay using an immunochromatographic method, and as shown in FIG. A liquid container 30 which is arranged in the vicinity of one end portion in the longitudinal direction of the stationary phase 20 and the stationary phase and which contains a liquid therein and at least part of which can be broken; and the stationary phase And a casing 40 containing the liquid container.

  In this embodiment, the casing 40 includes a casing body 41 including a pair of matable upper casing 41a and lower casing 41b, and a slider 42 slidably attached to the upper surface of the casing 41a. is there.

  On the upper surface of the upper casing 41a, a sample dropping window 43 is opened at a substantially central portion in the longitudinal direction, and a determination window 44 is provided in the vicinity of one end portion in the longitudinal direction, and further in the vicinity of the other end portion. An opening 45 for attaching the slider 42 is provided. The slider 42 has, for example, grooves that can be slidably fitted to both side walls in the longitudinal direction of the opening 45, and these members are made of a flexible material such as a resin material. Thus, it can be attached to the opening 45.

  Further, when the slider 42 slides and comes into contact with the liquid storage container 30, it can break the breakable portion of the liquid storage container 30, for example, a relatively sharp pressing portion (not shown) such as a nail. Is provided on the inside of the container.

  The water-absorbing base material constituting the stationary phase 20 housed in such a casing is not particularly limited as long as it is a porous material or a fiber material exhibiting capillary action. For example, nitrocellulose, cellulose, glass fiber, etc. Filter paper containing can be used.

  Further, the stationary phase 20 does not need to be constituted by an integral water-absorbing substrate. For example, in the embodiment shown in FIG. 1, the absorbent filter paper 21 and the conjugate disposed from the liquid storage container side (upstream side) are used. The gate pad 22 and the nitrocellulose film 23 are attached to the backing sheet 24, and the developed solution is completely applied to the end surface portion on the opposite side (downstream side) from the arrangement position of the liquid container. An absorption pad 25 for absorbing is placed.

  The conjugate pad 22 is located immediately below the sample dropping window 43 provided on the upper surface of the upper casing 41a, and is labeled with an enzyme that can bind to a test substance contained in the dropped sample. It contains a second immunity substance in a dry state.

  In addition, immediately below the determination window 44 provided on the upper surface of the upper casing 41a, a first immunity substance that specifically reacts with the test substance in the sample is immobilized on the stationary phase 20 (nitrocellulose film 23). And a control line (not shown) in which another immunity substance capable of reacting with the enzyme-labeled second immunity substance is immobilized downstream of the detection line (not shown). Z).

  Further, if necessary, on the stationary phase 20, for example, an activator for promoting the reaction between the enzyme labeled with the second immunity substance and the chromogenic substrate, or a substrate supplied from upstream. It is possible to provide a site formed by applying an enzyme or the like that can react to produce the activator.

  Next, the measurement principle using the immunoassay test instrument according to the present invention will be described by taking as an example the case of detecting Ddimer which is a degradation product of fibrin.

  First, as a basic arrangement in the test apparatus, as shown in FIG. 2A, the substrate solution, conjugate pad 22, glucose oxidase 26 accommodated in the liquid container 30 from the upstream side on the stationary phase 20, as shown in FIG. A detection line 27 on which an anti-D dimer monoclonal antibody is immobilized, a control line 28 on which an anti-mouse antibody is immobilized, and an absorption pad 25 are arranged. Here, the substrate solution is composed of an aqueous solution containing glucose as a substrate for glucose oxidase and a benzidine derivative such as tetramethylbenzidine as a chromogenic substrate for peroxidase, and the conjugate pad has a peroxidase-labeled anti-D dimer monoclonal. The antibody is placed in a dry state.

  In the assay, first, after dropping a sample (plasma) from a sample dropping window 43 provided on the upper surface of the casing 40 as shown in FIG. The container 30 is broken, and the substrate solution stored in the container 30 is developed. As shown in FIG. 2 (b), the target substance D dimer contained in the sample dropped from the sample dropping window 43 is peroxidase-labeled anti-D dimer monoclonal in the conjugate pad 22 located immediately below the sample dropping window. The reaction begins with the antibody and begins to penetrate onto the stationary phase 20 (nitrocellulose film 23). The mixture of the sample on the conjugate pad 22 and the peroxidase-labeled anti-D dimer monoclonal antibody is washed away toward the downstream side (right side in the figure) by the substrate solution penetrating from the upstream side (left side in the figure). .

  Next, as shown in FIG. 2C, the glucose flowing on the stationary phase 20 becomes a substrate of the glucose oxidase 26 and generates active oxygen (hydrogen peroxide). The target substance D dimer that has reacted with the target substance D dimer and the peroxidase-labeled anti-D dimer monoclonal antibody is captured by the anti-D dimer monoclonal antibody immobilized on the detection line 27. Further, the unreacted peroxidase-labeled anti-D dimer monoclonal antibody is captured by the anti-mouse antibody immobilized on the control line 28 located further downstream. Further, excess components are absorbed by the absorbent pad 25 located on the more downstream side.

  As shown in FIG. 2 (d), the peroxidase-labeled anti-D dimer monoclonal antibody captured on the detection line and the control line is generated by the benzidine derivative contained in the substrate solution and glucose oxidase by the action of peroxidase. Hydrogen peroxide is used to produce a blue insoluble colored material. The test result is visually observed by observing this color development state from the outside of the casing 40 as shown in FIG. Note that blue color development on the detection line indicates that the target substance is present in the sample, and blue color development on the control line indicates that the reaction system itself has proceeded appropriately.

  That is, when both the detection line and the control line are colored, it is determined that the target substance is positive and when only the control line is colored, it is negative. When only the detection line is colored, when neither line is colored, it indicates that some trouble has occurred.

  The measurement principle using the immunoassay test instrument according to the present invention has been described above with an example. However, the immunoassay test instrument according to the present invention is not limited to the immune system as in the above example. It can be applied to various immunoassays. For example, as an enzyme immunoassay method, either a sandwich method or a competitive method can be used. Furthermore, when the labeled test substance (for example, antigen) is bound to the immobilized immune substance (for example, antibody), the test substance is not directly bound, but the test substance. In advance, an immune substance (for example, a primary antibody) is reacted in advance, and an immune substance (for example, a secondary antibody) that specifically binds to the immune substance is immobilized, and a test substance is indirectly bound to the test substance. It is also possible to use a double or multiple sandwich method or a double or multiple competition method for bonding. The test substance is not particularly limited as long as it can be detected by the sandwich method or the competitive method as described above. For example, fibrin degradation products in detecting thrombosis, acute leukemia, α2-plasmin inhibitor / plasmin complex (PIC), vitamin K deficient protein-II (PIVKAII) in detecting severe liver disease, etc. It is also a marker for diseases associated with vitamin K deficiency and liver cancer); pathogenic bacteria such as Escherichia coli O-157, methicillin-resistant Staphylococcus aureus, Salmonella, Vibrio parahaemolyticus, Helicobacter pylori and Chlamydia trichomatis, and components thereof Toxins produced by bacteria such as verotoxin and streptolysin O; for example, proteins such as human transferrin, human albumin, human immunoglobulin, microglobulin and C-reactive protein; HBs, HBc, HBe antigens and antibodies of hepatitis B virus; , Hepatitis C virus antigen And a wide variety of types such as human immunodeficiency virus antigens and antibodies, rotavirus antigens and antibodies, and viral antigens and antibodies such as adenovirus antigens and antibodies; and other yeasts, fungi and other microorganisms or antibodies thereto Can be targeted.

It is a top view which shows the assembly process of one Embodiment of the test device for immunoassay which concerns on this invention. It is process drawing which shows typically the measurement principle using the test device for immunoassay which concerns on this invention.

Explanation of symbols

DESCRIPTION OF SYMBOLS 10 Immunoassay test instrument 20 Stationary phase 30 Liquid container 40 Casing 41 Casing main body 42 Slider.
43 Sample dropping window 44 Judgment window

Claims (3)

An immunoassay test instrument used for an assay using an immunochromatographic method,
A stationary phase consisting of a water-absorbing substrate stretched in the longitudinal direction;
A liquid storage container that is disposed near one end in the longitudinal direction of the stationary phase, stores liquid therein, and is formed of a film material that can be broken at least partially;
A casing containing the stationary phase and the liquid storage container, the casing main body having a sample dropping window and a determination window on the upper surface thereof, and having an opening at a position engageable with the liquid storage container If, slidably mounted along the upper surface of the casing body to the opening, the test instrument for immunoassays, characterized in that it comprises a casing comprising a slide elements having a breaking means for said liquid container . The stationary phase has a detection line formed by immobilizing a first immunological substance that specifically reacts with a test substance in a sample under the determination window, and a test substance under the sample dropping window. A conjugate pad containing the enzyme-labeled second immunity substance in a dry state, which can bind to the liquid- containing container, and the label containing the second immunity substance labeled The test apparatus for immunoassay according to claim 1, wherein a developing solution containing a chromogenic substrate for the enzyme is contained. The stationary phase, the lower determination window, for further immunoassay according to claim 2, characterized in that it comprises a separate confirmation line the immunogenic substance comprising immobilized capable of reacting with a second immunogenic substance Test equipment.