JP3292211B2 - Sampling rod and test equipment for detecting hemoglobin or transferrin - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention provides a feces collection rod for collecting feces and a test device used for performing occult blood tests.

[0002]

2. Description of the Related Art Conventionally, fecal occult blood testing has been carried out in such a procedure that a person in charge of testing uses a stool collected by a subject to detect hemoglobin and transferrin in the stool. Instruments used for such a stool collection include a stool collection rod for stool collection, a container for holding a buffer solution for extracting hemoglobin and the like in the stool, and a device attached to the back side of the lid of the container. A stool collection container made of a stool collection rod is used. After the stool is collected using such a container, the inspector removes the buffer from the container and detects hemoglobin or the like by an antigen-antibody reaction using a microtiter plate or the like.

[0003] Since the above-described method has problems such as the need for human analysis and difficulty in mass processing, a stick having a spherical tip portion coated uniformly with an anti-human hemoglobin antibody and an anti-human transferrin antibody. And a case containing a solution containing a lytic enzyme (Japanese Utility Model Application Laid-Open No. 2-59466). This test tool is adapted to be applied to an automatic analyzer of the bead method by cutting the spherical portion after the reaction between the hemoglobin in the sample and the antibody.

[0004] Conventionally, pregnancy test drugs are only for physicians, but in recent years, various pregnancy test kits capable of self-test have been marketed. These have a urine sampling rod that absorbs urine, and a filter paper that performs chromatography of urine absorbed from the urine sampling rod, and detect HCG antigen in urine by a monoclonal antibody applied to the filter paper. is there.

[0005]

As described above, various instruments for occult blood tests have been proposed, and efforts have been made to improve the analysis efficiency.
At present, the subject cannot detect hemoglobin or the like, and cannot perform an occult blood test without requesting a doctor or a laboratory.

[0006] If a certain period of time elapses after the subject collects the stool and performs the test, hemoglobin and the like are liable to degenerate particularly in the summertime when the temperature is high, so that an accurate test cannot be expected. Therefore, it is preferable to perform an inspection immediately after collecting the stool, but it is difficult to respond quickly with the conventional method.

[0007] The present invention has been made in view of the above, and it is necessary to take a stool and to promptly determine the hemoglobin and the like so that the subject can easily perform an occult blood test or the like without skill. An object of the present invention is to provide an inspection tool capable of performing detection and to provide a stool collection rod used for such an inspection tool.

[0008]

Means for Solving the Problems In order to solve the above-mentioned problems, the present inventor has configured an inspection tool as follows. That is, the test device of the present invention is a test device for detecting hemoglobin or transferrin in feces by immunochromatography using an anti-hemoglobin antibody or an anti-transferrin antibody, wherein the test device includes a stool collection rod and a stool collection rod. Chromatography piece in contact with one end of the
And a lid provided on the stool collection rod side of the test tool , wherein the stool collection rod is made of a material having a hardness capable of collecting feces and absorbing an aqueous solution, and further, a chromatographic piece. The opposite end of the end in contact with is formed of a material that can move hemoglobin together with the aqueous solution to the chromatographic piece when immersed in an aqueous solution containing hemoglobin, and the chromatographic piece can detect hemoglobin or transferrin by immunochromatography. , The lid is placed inside the lid at the time of inspection,
It is possible to accommodate the buffer while covering the characterized Rukoto.

In other words, the stool collection rod of the present invention is a stool collection rod for collecting stool for detecting hemoglobin or transferrin in stool, wherein the stool collection rod has a hardness capable of collecting stool. And a material that absorbs the aqueous solution and is formed of a material that can move hemoglobin or transferrin with the aqueous solution at the other end when the one end of the stool collection rod is immersed in an aqueous solution containing hemoglobin or transferrin. It is characterized by.

[0010] Further, the test device of the present invention is a test device for detecting hemoglobin or transferrin in feces by immunochromatography using an anti-hemoglobin antibody or an anti-transferrin antibody. Having a chromatographic piece in contact with one end of the stool collection rod, wherein the stool collection rod is made of a material having a hardness capable of collecting feces and absorbing an aqueous solution, and further having an end in contact with the chromatography piece. When the opposite end of the part is immersed in an aqueous solution containing hemoglobin, the chromatographic piece is formed of a material capable of moving hemoglobin together with the aqueous solution, and the chromatographic piece is capable of detecting hemoglobin or transferrin by immunochromatography. I do.

Hereinafter, the present invention will be described.

<1> Stool collection rod The stool collection rod of the present invention has a hardness capable of collecting feces and is made of a material that absorbs an aqueous solution. Further, one end of the stool collection rod is made of hemoglobin or transferrin. (Hereinafter referred to as “hemoglobin”) is formed of a material capable of moving these substances together with the aqueous solution at the other end when immersed in an aqueous solution containing the same. this is,
If you do not have enough hardness, it is difficult to collect stool,
It is also for guiding the aqueous solution containing hemoglobin and the like in feces to a chromatography piece described later.

The material is not particularly limited as long as it satisfies the above-mentioned requirements, but natural fibers such as wool and paper; semi-synthetic fibers such as nitrocellulose and cellulose acetate; and synthetic materials such as nylon and polyester. In addition to fibers, inorganic fibers such as glass fibers and the like can be mentioned, and the form is preferably a fiber which easily causes a capillary phenomenon such as a hard felt, a fiber bundle, a woven fabric, and a nonwoven fabric.

The shape of the stool collection bar of the present invention may be a bar shape, and the cross section may be any shape such as a rectangle, a circle and an ellipse, but a thin rectangular parallelepiped is preferred. In addition, it is preferable that one end of the stool collection rod is formed in a shape that allows stool to be collected. For example, the tip is spoon-shaped, screw-shaped,
Alternatively, a tip having a zigzag vertical section and a groove having a groove at the tip can be exemplified.

Usually, as a method of using a stool collection rod, a method of piercing feces several times is used. However, hemoglobin and the like are said to be mainly localized on the surface portion of the defecation feces, so that a method of scraping the surface is preferable to a method of piercing. Therefore, it is desirable that the tip of the stool collection rod has a shape that facilitates scraping of feces. From this viewpoint, among the above shapes, those having a spoon-like shape, a zigzag shape in a vertical section, and a groove-like notch in the tip end are particularly preferable.

<2> Test Tool of the Present Invention The test tool of the present invention has the above-mentioned feces sampling rod and a chromatography piece in contact with one end of the feces sampling rod.

The chromatographic piece indicates the presence or absence of hemoglobin or the like by an immunological method. For this, the same configuration as that already on the market for a pregnancy test kit can be used. However, a monoclonal antibody against HCG antigen is used in the pregnancy test kit, whereas a monoclonal antibody against hemoglobin or the like is used in the present invention. Hereinafter, a test device for detecting hemoglobin will be described. However, if hemoglobin is replaced with transferrin, a test device for detecting transferrin can be obtained.

For example, a labeled first anti-hemoglobin monoclonal antibody (hereinafter, referred to as "first antibody") is applied to one side of the flaky chromatography base, and the monoclonal antibody is applied to the other side. A second anti-hemoglobin monoclonal antibody or polyclonal antibody having a different antigenic determinant from the antibody (hereinafter, referred to as "second antibody").
A region where the anti-immunoglobulin antibody against the immunoglobulin of the animal used to produce the antibody is fixed on the side opposite to the side in contact with the stool collection bar across the region where the fixed second antibody is located. Is provided. The region where the anti-immunoglobulin antibody is immobilized is not necessarily required, but is preferably provided for accurate testing.

Examples of the chromatographic base include membrane filter materials such as nitrocellulose and cellulose acetate, nylon membranes, filter paper, sintered silica gel, glass fibers, etc., and hemoglobin, hemoglobin and antibodies together with an aqueous solution. Any material can be used as long as it can move the combined material. Further, using the same material as the above-mentioned stool collection rod, the stool collection rod and the chromatography piece may be integrally formed.

The principle of the chromatography piece is as follows. When an aqueous solution containing hemoglobin comes into contact with the end of the chromatography piece on the side to which the first antibody is applied, the aqueous solution is absorbed and developed by the chromatography piece, and permeates toward the opposite end. When reaching the area coated with the first antibody, the hemoglobin binds to the first antibody and travels further on the chromatographic strip. When the conjugate of the hemoglobin and the first antibody reaches the region where the second antibody is immobilized, the hemoglobin binds to the second antibody and is trapped. As a result, the first antibody stops in this region with hemoglobin and does not move any further.

On the other hand, when hemoglobin is not contained, the first antibody passes through the region where the second antibody is immobilized,
Continue moving. At this time, if a region to which the anti-immunoglobulin antibody is fixed is provided, the second antibody is trapped in this region. In addition, protein A derived from Staphylococcus may be used instead of the anti-immunoglobulin antibody.

Which of the above regions is trapped depends on
For example, it can be visually determined by labeling the first antibody with a metal colloid such as gold colloid, colored latex, or an enzyme such as β-galactosidase or horseradish peroxidase. However, when an enzyme is used, it is necessary to develop a color by an enzyme immunoassay using a dye that develops the color of the enzyme.

The method of preparing the antibody, the method of labeling the above-mentioned labeling substance with the first antibody, and the method of immobilizing the antibody on the chromatographic piece are not particularly limited, and may be performed by a conventional method. For example, a method for labeling an antibody with colloidal gold is disclosed in
No. 100 and Japanese Patent Application Laid-Open No. 1-503174. Examples of a method for immobilizing the antibody on the chromatography piece include a method described in Japanese Patent Publication No. 1-503174 or the like.

The monoclonal antibody can be obtained by culturing fusion cells (hybridomas) of spleen cells of mice immunized with hemoglobin or the like and myeloma cells.
Various antibodies are commercially available and may be used.

A labeling method using a metal colloid is disclosed in
No. 41665. In order to immobilize the second antibody and the anti-immunoglobulin antibody on the chromatographic piece, when using a nitrocellulose membrane or a nylon membrane, the material of the chromatographic piece is activated in advance with cyanogen bromide and the immobilization is attempted. An antibody solution is dropped on the site to covalently bind the antibody. Thereafter, in order to prevent the nonspecific adsorption of the first antibody and hemoglobin to the membrane, it is preferable to mask the sample by immersing it in a solution such as casein or bovine serum albumin.

As a form of the inspection tool of the present invention, for example,
The chromatographic fee piece is covered with a container, a stool collection rod is provided on one side of the chromatography piece, and the stool collection rod is projected from the container (FIG. 3).

[0027]

The operation of the inspection tool and the stool collection bar of the present invention will be described together with the usage. Stool is scraped off with the tip of the stool collection tool, and the stool collection tool is immersed in a buffer solution or the like. For example, if a lid is provided on the side of the test tool container from which the stool sampling rod protrudes, a buffer is stored in the lid during the test, and the stool collection rod is immersed therein, the inspection can be performed easily. . Alternatively, the lid may be pre-filled with a gel containing a buffer solution.

When the stool bar is immersed in a buffer solution or gel and stirred, hemoglobin and the like are absorbed by the stool bar together with the buffer solution and guided to the chromatography piece. Thereafter, the presence or absence of hemoglobin or the like is determined according to the above-described principle of the chromatography piece.

When the test device of the present invention is used, an occult blood test can be performed quickly and easily.

[0030]

Embodiments of the present invention will be described below with reference to the drawings.

[0031]

Embodiment 1 First, an embodiment of the stool collection bar of the present invention will be described. FIG. 1 shows an example of a stool collection rod. In this method, the tip of a hard felt molded into a thin rectangular shape is processed so that the cross section becomes zigzag.

FIG. 2 shows another example of the stool collection rod. This is obtained by processing the end of a polyethylene fiber bundle bundled in a cylindrical shape into a spoon shape. In use, since the stool collection rod is immersed in the buffer solution, the processed portion at the tip does not necessarily have to have water absorbency. Therefore, strength may be imparted by coating the tip with plastic or the like.

FIG. 3 shows another example of the shape of the stool collection bar. This has a groove-like cut at the tip.

[0034]

Embodiment 2 Next, an embodiment of the inspection tool of the present invention will be described (FIGS. 4 and 5). This is for hemoglobin detection,
If the hemoglobin antibody is replaced with a transferrin antibody, it can be used for transferrin detection.

The chromatography piece 20 is accommodated in the plastic container body 31, and the feces collection rod 10 is set on the container body 31 so as to be in contact with the end of the chromatography piece 20. The lid part 32 of the container body 31 is fitted to the side where the stool collection rod is installed.

The container body 31 covers the chromatography piece 20 and a part of the stool collection rod 10, and a region 22 to which an anti-immunoglobulin polyclonal antibody is fixed and a region 23 to which an anti-immunoglobulin antibody is fixed, which will be described later, are externally provided. A window 33 on which a transparent plastic is attached is provided so as to be visible.

The chromatographic strip 20 is a thin section of nitrocellulose, a region 21 coated with an anti-human hemoglobin monoclonal antibody (mouse) labeled with colloidal gold on the side in contact with the stool collection rod, and an anti-human hemoglobin polyclonal antibody on the other side. (Rabbit) is immobilized, and a region 23 on which an anti-mouse immunoglobulin antibody (rabbit) is immobilized is further provided outside.

Anti-human hemoglobin monoclonal antibody (mouse), anti-human hemoglobin polyclonal antibody (rabbit), and anti-mouse immunoglobulin antibody (rabbit) are commercially available (SU-104, manufactured by Nippon Biotest, A118, respectively). , Z456: manufactured by Dako Japan Co., Ltd.).

Labeling of an anti-human hemoglobin monoclonal antibody with colloidal gold was carried out according to the method described in JP-A-1-503174. The colloidal gold was adjusted to pH 7 with 0.1 M K ₂ CO ₃ through a 0.22 μm filter and 20 ml was placed in a clean beaker. In addition, 0.
An anti-human hemoglobin monoclonal antibody solution (1 mg / ml, pH 9) passed through a 22 μm filter was added and stirred for 2 minutes. The pH of the mixture of antibody and gold sol was adjusted to 9 using 0.1 M K ₂ CO ₃ and 2 ml of 10%
(V / V) BSA was added. This at 4 ° C for 30 minutes,
Purification was performed by repeating centrifugation at 12000 × g and resuspending the precipitate three times. Further, a 20 mM Tris buffer (150 mM NaCl, pH 8.2, 1% BS
Suspended in A), a colloidal gold-labeled antibody was obtained.

Anti-human hemoglobin polyclonal antibody (rabbit), anti-mouse immunoglobulin antibody (rabbit)
Was fixed using an immunodyne manufactured by Pall Corporation. In addition, a 1 (W / W)% BSA solution (50 mM Tris, pH
Using 7.5), portions other than those on which the antibodies were immobilized were masked.

The stool collection rod 10 is a glass fiber formed into a cylindrical shape and in which water moves in one direction by capillary action, and has a groove-shaped notch at the tip. On the inside of the lid portion 32, a scale is provided as a guide when the buffer solution contained in a separate container is injected. When performing the inspection, the faeces surface is scraped off with the tip of the stool collection rod 10, a buffer solution is injected up to the scale inside the lid 32, and the stool collection rod is immersed in this and allowed to stand. Another lid 3 is attached to the opposite end of the lid 32 of the container body 31.
If a buffer solution or a buffer-containing gel 36 sealed with a plastic film 35 or the like is accommodated therein, the inspection can be performed simply by replacing the lid when used. That is, if the lid 34 is put on the stool collection rod side of the container main body with the container main body standing, the stool collection rod 10 pierces the film 35, and immunochromatography can be performed only by gently shaking the entire container.

If a desiccant is contained in the container, for example, on the opposite side of the chromatography piece from the stool collection rod, inactivation of the antibody can be prevented.

[0043]

[Embodiment 3] An example of use of the inspection tool of the present invention will be described. Hemoglobin was detected using the inspection tool of Example 2 (FIG. 4) (the tip of the stool collection bar has a groove-shaped notch).

Human hemoglobin was mixed with feces of a Beagle dog at various concentrations, and the feces were scraped off using the test device of the present invention, and detection was carried out by the above procedure. The concentrations of human hemoglobin were 0, 0.25, and 0.75 mg / (g feces). In addition, iatrohem check (manufactured by Yatron Corporation) was used as a control. Table 1 shows the determination results based on the following criteria.

++: Detected very well +: Detected ±: Difficult to judge-: Not detected

[0046]

[Table 1]

As is clear from the results, the test device of the present invention can measure human hemoglobin with high sensitivity and easily.

[0048]

The test device of the present invention enables a subject to take his stool and detect hemoglobin or transferrin thereafter. As a result, a fresh blood test can be performed quickly and easily. The stool collection rod of the present invention enables stool collection and immunochromatography.

[Brief description of the drawings]

FIG. 1 is a perspective view showing an example of a stool collection rod of the present invention.

FIG. 2 is a side view showing another example of the stool collection rod of the present invention.

FIG. 3 is a side view showing another example of the stool collection rod of the present invention.

FIG. 4 is a partially broken front view showing an example of the inspection tool of the present invention.

FIG. 5 is a partially cutaway front view showing another example of the inspection tool of the present invention.

[Explanation of symbols]

 DESCRIPTION OF SYMBOLS 10 ... Stool collection rod 11 ... Tip part 20 ... Chromatography piece 21 ... Area | region which applied the 1st anti-hemoglobin antibody 22 ... Area | region which fixed the 2nd anti-hemoglobin antibody 23. ..Area where anti-IgG antibody is fixed 30 ・ ・ ・ Container 31 ・ ・ ・ Container body 32 ・ ・ ・ Cover 33 ・ ・ ・ Window 34 ・ ・ ・ Cover 35 ・ ・ ・ Plastic film 36 ・ ・ ・ Buffer Or buffer-containing gel

──────────────────────────────────────────────────続 き Continued on front page (58) Field surveyed (Int.Cl. ⁷ , DB name) G01N 33/48 G01N 33/50 G01N 33/543 G01N 33/72

Claims (3)

(57) [Claims] 1. A test tool for detecting hemoglobin or transferrin in feces by immunochromatography using an anti-hemoglobin antibody or an anti-transferrin antibody, wherein the test tool is one of a stool collection rod and a stool collection rod. It is provided on the chromatography piece in contact with the end and on the stool collection rod side of the test tool.
The stool collection rod has a hardness capable of collecting feces and is made of a material that absorbs an aqueous solution.Furthermore, the opposite end of the end in contact with the chromatography piece is made of hemoglobin. The chromatographic piece is formed of a material that can move hemoglobin together with the aqueous solution when immersed in the aqueous solution containing the chromatographic piece, and the chromatographic piece can detect hemoglobin or transferrin by immunochromatography , and the lid is placed in the lid during the test. , With the stool bar covered
Inspection instrument according to claim Rukoto can be accommodated buffer. 2. The test device according to claim 1, wherein the chromatography piece has a region coated with a first anti-hemoglobin monoclonal antibody or an anti-transferrin monoclonal antibody on an end portion in contact with the stool collection portion. Has an area in which a second anti-hemoglobin antibody or anti-transferrin antibody having a different antigenic determinant from the monoclonal antibody is immobilized. An inspection device having a region to which an immunoglobulin antibody is immobilized. 3. The test device according to claim 2, wherein the first monoclonal antibody is labeled with one kind selected from the group consisting of a metal colloid, a colored latex, and an enzyme.