JP3487683B2 - Fecal occult blood determination device - Google Patents

Description

Description: BACKGROUND OF THE INVENTION The present invention relates to a method for determining the presence or absence of fecal occult blood.
Fecal occult blood determination device for determining
Fecal occult blood determination system by immunoassay using tomoglobin antibody
About the installation. [0002] Fecal occult blood is caused in the digestive tract due to digestive diseases.
It is expressed when blood is mixed into the stool. Especially the large intestine
Correlation with diseases such as cancer has been confirmed, and fecal occult blood test
An index for early diagnosis of colorectal cancer, which is increasing in Japan
This is a very important and widely used test. [0003] Currently used fecal occult blood test methods include:
There are roughly two methods. The first method is traditional
Chemical occult blood reaction (hereinafter referred to as “chemical method”)
It is called hemoglobin (hereinafter referred to as Hb).
Non-specific reaction to detect peroxidase-like activity
is there. This method uses a chromogen (guaiac oil or ortho)
Trididine) is a peroxidase-like substance
Oxidized in the presence of hydrogen peroxide and hydrogen peroxide
Good. Therefore, in this method, the peroxidase-like activity
Positive by quality drug or food containing Hb
Dosing and food restriction were required to indicate As an inspection device based on the present chemical method, a simple and easy
Measurement systems have been disclosed, some of which are
ing. For example, Japanese Patent Publication No. 2-45826,
9-168370, JP-A-62-266463
In the official gazette and Japanese Utility Model Laid-Open No. Sho 62-153573,
Place a small amount of stool test on a card containing test paper impregnated with yak oil, etc.
After applying the paint, the hydrogen peroxide solution is dropped to check the coloration.
A system is disclosed. In these systems,
Stool sample on a part of guaiac fat attached to the card type body
And apply hydrogen peroxide to the area on the back side of the application area.
By dropping water, guaiac oil at the dropping part develops color
It is determined by whether or not to do. Perform without opening the stool sample application section
It can be handled hygienically and has a simple configuration
It is an advantage, but derived from the measurement principle as described above
The disadvantages have not been solved. [0005] The second method solves the above-mentioned disadvantages of the chemical method.
Was developed in recent years to
(Hereinafter referred to as "immunization method"). This law is anti-hi
This method is based on an antigen-antibody reaction using an Hb antibody.
High specificity, requiring dietary restrictions such as chemical methods
do not do. Judgment: latex agglutination, enzyme-labeled immunoassay, etc.
Of the general immunoserologic test
The most commonly used method is the latex agglomeration method.
You. In the latex agglutination method, a stool sample is placed on a judgment plate.
The solution and latex reagent are added dropwise and mixed to form an aggregated image.
Is determined based on the presence or absence of. Principle compared to chemical method
Is highly specific, but skill is required for the determination of the agglutination method. Ma
Inspection system based on the immunization method
Kits containing reagents, stir bars,
However, the composition is complicated and the above chemical method is used.
A simple system such as has not been developed. [0007] In the immunization method, there is a unique immunization method which is not available in the chemical method
Process, i.e., dissolving the stool sample in a buffer solution
Step of obtaining and undissolved solids in the stool sample solution by filtration
It is necessary to carry out a removing step. These operations are simplified.
Various attempts have been made for convenience. For example,
A device for quantitative collection of stool was developed.
No., Japanese Utility Model Laid-Open No. 1-158964, etc.
The device for obtaining the removed stool sample solution is disclosed in Japanese Utility Model Laid-Open No. 2-21.
557, Japanese Utility Model Application Laid-Open No. 2-140468, Japanese Utility Model
It is disclosed in JP-A-2-13977. As described above, the steps required to obtain a stool sample solution
Various ideas have been disclosed. But
Mix the fecal sample solution and the determination reagent to detect fecal occult blood.
Inspection device that enables a series of operations to be performed easily
Has not been realized to date. That is, a fecal occult blood measurement system using an immunoassay.
The simplification of the system is achieved by integrating the above-mentioned steps with the antigen-antibody reaction.
It is likely that this technology will be fulfilled,
Not realized. [0010] In recent years, self-care is called
With the spread of the concept, a simple test system based on the immune response
Is now available at pharmacies as a general test drug.
Was. For example, in pregnancy diagnostics, immunochromatography
A simple inspection system that utilizes the system has been established. The immunity
The most common use of chromatographic methods for fecal occult blood testing
A major problem is that the stool sample dissolution and filtration steps described above
It is difficult to incorporate into the system. An object of the present invention is to provide an immunochromatography
This is a fecal occult blood determination device using the
Preparation of stool sample solution from which insoluble solids were removed, and stool
Determination of the presence or absence of occult blood can be easily performed even by unskilled persons.
It is an object of the present invention to provide a fecal occult blood determination device that can perform the following. [0012] The fecal occult blood determination apparatus of the present invention.
Is a roughly bar-shaped tip with a semi-quantitative
Feces collection device and feces collection device inserted through opening at one end and built-in
The stool is internally moved in its length direction.
And a cylindrical container main body that is configured to be able to be used. Up
The opening on the opposite side of the container body
The mouth is formed so that the tip of the stool can enter and exit
Have been. In addition, a buffer storage
, A filter, and a development layer. The buffer solution storage section dissolves stool and
Where the buffer solution for preparing the feed solution is stored.
is there. Further, the filter is used for filtering a stool sample solution.
It is provided for. The spreading layer is filtered by a filter.
The stool sample solution is supplied.
In the spreading layer, sensitization with anti-human hemoglobin antibody (first antibody)
A carrier is provided, and the first antibody is recognized.
Anti-human hemoglobin antibodies with different pitopes (second antibody)
Is immobilized, and the anti-human hemoglobin antibody
Different from the sensitizing carrier installation position and the second antibody immobilization position.
Reacts with the anti-human hemoglobin antibody-sensitized carrier
Substance is immobilized. In addition, the container body
Enables external determination of fecal occult blood presence
Therefore, a judgment window was formed, and it was confirmed that the judgment was completed.
An end window for confirmation is formed. Further, in the fecal occult blood determination apparatus of the present invention,
When the toilet device is moved in its length direction within the container body,
To prevent the stool sample solution from leaking outside the container body
A liquid-tight seal ring near the other end opening of the container body.
Is attached. Also, from the buffer storage to the filter
Flow path to enable the supply of stool sample solution
Tool and / or flow path forming means provided in the container body
Is formed. In the fecal occult blood determination device of the present invention,
Cover the other end opening of the container body and the tip of the stool
In addition, a lid is detachably attached to the container body. (Function) The fecal occult blood determination apparatus of the present invention comprises:
It is used as follows. In other words, there is no
Remove the lid from the container body
By piercing and pulling out the stool sample,
A semi-quantitative sample of the stool is collected. Next, in the container body
Move the stool to pull it back,
Move the distal end of the stool collection instrument from the opening at the other end of the container body to the container.
Move to the buffer storage section inside the main body and shake the entire container body.
By giving or giving appropriate vibrations,
Dissolve sample in buffer. Made as above
Stool sample solution into a filter using the above channel forming means.
Lead. In the filter, the stool sample solution is filtered and filtered.
Stool sample solution is supplied to the developing layer,
The presence or absence of fecal occult blood is determined by immunochromatography
You. This judgment is made by the judgment window provided on the container
This can be done by observing through a window. As described above, with the fecal occult blood determination device of the present invention,
Is a fecal occult blood test consisting of the above-mentioned fecal collection device, container body and lid
Stool samples were collected using only a measuring device.
Perform all operations up to removal of digested solids and determination of fecal occult blood.
It is possible to That is, the present invention relates to immunochromatography
This is a simple fecal occult blood determination device using the
And stool sample solution to immunochromatography
All processes up to the judgment method using the method of the present invention
It is characterized in that it can be performed by using. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, the present invention will be described with reference to the drawings.
By describing the fecal occult blood determination device according to one embodiment,
The present invention will be elucidated. FIG. 1 shows a fecal occult blood test according to one embodiment of the present invention.
It is a longitudinal cross-sectional view which shows a fixing device. The fecal occult blood determination device 1
A toilet device 2, a container body 3 and a lid 4 are provided. Stool collection device 2
Has a substantially rod-like shape as a whole, and is formed on the outer surface of the tip 2a.
Irregularities are provided. Unevenness on the outer surface of this tip 2a
Is provided for semi-quantitatively collecting stool samples.
You. However, this stool sample must be collected semi-quantitatively.
The shape of the tip is not limited to the unevenness shown in FIG.
As shown in (a) to (e), a groove 21 or a cut is formed in the tip 2a.
It may be one in which an appropriate concave portion such as a notch 22 is formed.
No. A tip 2a for collecting a stool sample
And the above-mentioned feces collection device 2 to constitute the flow path forming means.
Is the diameter of both between the distal end portion 2a and the stool collection device main body 2b.
It has a connecting portion 2c having a smaller diameter. Stool collection device
At the upper end of the main body 2b, a handle 2d having a relatively large diameter is provided.
It is configured integrally. The handle 2d is further
Large diameter portion 2e and a lower end of the large diameter portion 2e.
And a small diameter portion 2f. The diameter of the small diameter portion 2f is
3, and the diameter of the large diameter portion 2e is
The outer diameter of the container body 3 is set larger. Therefore,
In the state shown in FIG. 1, the toilet device 2 has a large diameter portion 2e.
Because the lower end of the container is in contact with the upper end of the container body 3,
Is configured so that it cannot move further downward from the state of
Have been. The stool collection device 2 semi-quantitatively collects a stool sample.
And do not adversely affect the determination of fecal occult blood.
It can be made of a rigid material such as a synthetic resin.
The container body 3 has an opening 3a at the lower end and an opening 3b at the upper end.
Having. The inner diameter of the lower opening 3a is the tip of the
It is configured to have a diameter substantially equal to the maximum diameter portion of the portion 2a
Have been. That is, it is clear from the usage method described below.
As described above, the collected stool sample was transferred into the buffer solution.
The tip 2a is moved into the container body 3 from the opening 3a.
It is. In this case, remove the excess stool sample and semi-quantitatively
In order to dissolve the stool sample in the buffer, the diameter of the opening 3a
Has a size almost equal to the maximum diameter portion of the tip 2a.
Or slightly larger than the maximum diameter of the tip 2a.
It is configured to have a large diameter. In the vicinity of the opening 3a, a container
O-rings 5a and 5b as liquid-tight seal rings on main body 3
Is attached. O-ring mounted double
5a and 5b are for transferring the buffer solution in the container body 3 to the outside of the container body.
Installed to prevent leakage. The diameter of the opening 3b at the upper end of the container body 3 is
The outer diameter of the small diameter portion 2f of the device 2 is substantially equal to the outer diameter. Ma
A male screw is formed on the outer peripheral surface of the small diameter portion 2f.
And formed on the inner peripheral surface near the opening 3b of the container body 3.
It is configured to mesh with a female screw. That is,
The toilet device 2 includes the female screw and the male screw in the container body 3.
It is configured to be screwable by use. However, the small diameter portion 2f is located inside the container body 3.
It may be press-fitted, in which case the container body
1 shows a small diameter portion 2f in a state where the inside of the device 3 is liquid-tightly sealed.
Even configured to be able to move upwards
Just fine. The lower part of the container body 3 is opened.
Tapered so that the diameter decreases toward the mouth 3a
Have been. The container body 3 is made of polyethylene or polypropylene.
Film-like resin such as
Supports determination of fecal occult blood in addition to synthetic rubber such as butadiene rubber.
It is made of appropriate material unless it causes any obstacle
be able to. In the container body 3, a buffer solution storage section 3
c is provided. A stool sample is stored in the buffer storage section 3c.
Buffer solution 6 for dissolving is stored. Buffer storage
The volume of the retaining part 3c is required for mixing the stool sample and the buffer solution 6.
It is not particularly limited as long as it has a sufficient size.
No. The buffer 6 may be a saline solution or various buffer solutions.
Can be used, but this may affect the determination of fecal occult blood.
Any buffer can be used, unless otherwise noted. Also,
The amount of buffer 6 was determined by dissolving the collected stool sample.
To be able to prepare a stool sample solution with an appropriate concentration for occult blood determination
It is sufficient if it is a necessary and sufficient amount, usually about 1 to 10 ml.
Buffer solution 6 is used. Above the buffer storage section 3c, a partition wall 3 is provided.
d is formed. The partition wall 3d is aligned with the container body 3
The body is configured with an opening in the center. Of this opening
The diameter is almost the same as the outer diameter of the body 2b of the stool device 2
It has been. Also, O-rings are provided above and below the partition wall 3d.
7a and 7b are attached. O-rings 7a, 7b
Means that the buffer solution 6 or the stool sample solution
To prevent leakage above the partition wall 3d
It is provided in. That is, the opening of the partition wall 3d
Is provided for liquid-tight sealing. Above the partition wall 3d, a filter 8 and
The spreading layer 9 is attached in that order. This fill
Tab 8 is fixed on the partition wall 3d, and the development layer 9
It is fixed to the upper surface of the filter 8. Filter 8
And the developing layer 9 has a cylindrical shape and has the above-described stool collection inside.
The feces collection device main body 2b of the device 2 is inserted. The filter 8 is used to filter insoluble solids in the stool sample solution.
Is provided to eliminate the Filter 8
The position to be provided is a stool sample prepared in the container body 3.
As long as the solution can pass through the filter 8 and be supplied to the spreading layer 9
And it is not particularly limited. The filter 8 is made of, for example, filter paper or sponge.
It is composed of a known porous material for removing solids.
Can be In addition, remove undigested solids
And absorb substances that are of interest in determining fecal occult blood.
Use appropriate material for filtration unless it wears
Can be. The spreading layer 9 spreads the stool sample solution,
Provided for performing chromatography. Exhibition
The open layer 9 contains a test substance, ie, an antibody against human Hb.
(Hereinafter, the first antibody) sensitized carrier, ie, the first antibody
A sensitizing carrier 10 is provided. In addition, primary antibody sensitization
Above the position where the body 10 is installed, the first antibody and
Human Hb antibody 11 recognizing different epitopes (hereinafter referred to as
The second antibody is immobilized, and the second antibody 11 is immobilized.
3f, the judgment window 3f is formed in the container body 3.
Has been established. In addition, the judgment window 3f is used for observation from outside.
The container body is provided for ease of use.
3, an opening is formed in the wall surface, and a transparent resin material
The opening is closed. The second antibody 11 is further immobilized on the developing layer 9.
The first antibody-sensitized carrier 10 above the
A substance 12 which reacts with
In the part where is fixed, the end window is
3 g are formed. In addition, 3 g of the end window is
Determines whether the liquid has been correctly supplied onto the spreading layer 9
It is easy to observe from the outside whether or not
The wall of the container body 3
An opening is formed on the surface and the opening is made of a transparent resin material
Is closed. In the developing layer 9, a stool sample solution containing human Hb
Is developed, human Hb is converted to the second antibody of the first antibody-sensitized carrier 10
Causes an immune reaction with one antibody. As a result, human Hb-
The first antibody-sensitized carrier complex moves on the spreading layer 9 and
The antibody 11 reaches the portion where the antibody 11 is immobilized. Second antibody
The immune complex reaches the part where 11 is immobilized
Then, human Hb and the second antibody 11 cause an antigen-antibody reaction.
The human Hb-first antibody-sensitized carrier complex is then
11 is trapped in the portion where it is immobilized. This and
The carrier provides a clear contrast to the spreading layer 9
If indicated, the presence of the carrier, i.e. the carrier and the carrier adsorb
The presence of human Hb that has been
You. In addition, the first antibody-sensitized carrier is usually a human stool sample.
Since it is used in sufficient excess with respect to Hb, human Hb
The first antibody-sensitized carrier that did not react with
The second antibody 11 is immobilized along with the sample solution.
Moved further upward than the site, and the first antibody-sensitized carrier 10
Reaching the site where the reacting substance 12 is immobilized,
Reacts with the substance 12 and is trapped. As a result,
If the carrier shows a clear contrast to the spreading layer 9
In this case, the presence of the carrier can be visually confirmed. You
That is, it can be confirmed that this measurement was performed correctly.
When no human Hb was present in the stool sample,
All the first antibody-sensitized carriers are associated with the stool sample solution,
Since it moves to the position 12, the measurement end is confirmed in the same way.
It can be recognized. As a material for forming the spreading layer 9,
Suitable for the developing layer in epidemiological chromatography
Materials such as cellulosic materials and synthetic polymer materials
Materials that are widely used as known membrane filters
Can be used. The spreading layer 9 has a clear color tone of the carrier.
It is necessary to have a high contrast. like this
Most preferred as carriers are latex particles.
However, since latex particles are usually white,
When used for a white spreading layer, apply latex particles.
Coloring obtained by a method of coloring by contact with a coloring agent
It is desirable to use latex particles. The first antibody used in the present invention is a rabbit
Immunization of human Hb on goats, sheep, goats, horses, mice, etc.
Antiserum obtained by affinity chromatography
It can be obtained by purifying with a gel.
Known cell fusion using immunized mouse spleen cells
It is also possible to produce monoclonal antibodies by the method
is there. In any case, the first antibody used is sufficient
It is preferable that it is purified. The method of sensitizing the first antibody to the carrier is known in the art.
Operation method, for example, physical adsorption method or chemical bonding method.
I can. For example, anti-human Hb is added to latex particles.
Add buffer solution of antibody and at 37 ° C for 30-120 minutes
The first antibody is mixed with the latex particles
Physical adsorption can be performed on the surface. Also, carboxyl
When latex particles having a group are used, water-soluble carbodi
Latex the first antibody by covalent bonding via imide etc.
It can be bound to particles. The first antibody sensitizing agent obtained as described above
To provide the body on the spreading layer 9, for example, the first antibody
0.05-1 ml of a 0.01-1% by weight dispersion of the carrier
Drop it in place and let it dry at room temperature for at least 12 hours.
And can be performed by: The second antibody 11 may be the first antibody
Uses anti-human Hb antibodies with different epitopes from the body
You can. As a method for immobilizing the second antibody on the spreading layer 9
Contains 0.1 to 10 μl of 1 to 5 mg / ml of the second antibody.
A method of dropping at a fixed position and drying at room temperature for 12 hours or more
Etc. can be adopted. A substance which reacts with the first antibody-sensitized carrier 10
As the quality 12, for example, human Hb, producing the first antibody
(Eg, the first antibody is a goat)
If it is an antibody of origin, anti-goat IgG antibody) and the like.
It is. For example, a method of immobilizing human Hb on the spreading layer 9
Is 1 to 10 μg / ml human Hb 0.1 to 1 m
1 in a predetermined position, and dried at room temperature for 12 hours or more.
Can be adopted. In the fecal occult blood determination apparatus 1 of this embodiment, the buffer
Supply the stool sample solution prepared in the storage section 3c to the filter 8
In order to achieve this, the connecting portion 2c is used as a flow path forming means.
Tool 2. That is, from the usage described below
As can be seen, the stool sample solution was introduced into the filter 8.
The connecting portion 2c is located at the position where the partition wall 3d is provided.
To the position. As a result, from the opening of the partition wall 3d
Also, since the diameter of the connecting portion 2c is small, the stool sample solution
It will be possible to move to the filter 8 through the 3d opening.
As described above, the connecting portion 2 c allows the stool sample solution to pass through the filter 8.
The flow path forming means for supplying
The means for forming is not limited to the connecting portion 2c, but is always the buffer storage portion 3.
c can be separated from the filter 8 and, if necessary,
The stool sample solution is supplied from the buffer storage section 3c to the filter 8.
It can be configured by an appropriate structure to obtain
It is not always necessary to provide the flow path forming means on the feces collection instrument 2 side.
Instead, it may be provided on the container body 3 side. The lid 4 is provided with an opening 3 at the lower end of the container body 3.
a and the state shown in FIG.
It is provided for. The lid 4 is used for determining fecal occult blood.
It is provided to do it rawly,
The material forming the lid 4 is not particularly limited.
No. However, the lid 4 is made of the same material as the container body 3.
Is preferable because of easy production. FIG.
Although it is not clear, the inner peripheral surface near the upper end opening of the lid 4
Is formed with a female screw, while the lid 4 of the container body 3 is
At the part that comes into contact with the inner peripheral surface,
Screws are formed. Therefore, the lid body 4 is
Can be removably fixed to the device. However, the lid 4
The structure for detachably fixing to the container body 3 is as described above.
It is not limited to the one that uses screws.
The body 3 is pressed into the lid 4 to fix them together.
It may be made. Next, the method of using the fecal occult blood determination device 1 will be described.
This will be described with reference to FIGS. First, the stool shown in FIG.
The lid 4 of the occult blood determination device 1 is removed from the container body 3. Next
Then, as shown in FIG.
The tip 2a of the instrument 2 is pierced into the fecal sample 13 and pulled out.
Good. In this way, a stool sample is collected at the tip 2a.
Next, in the container body 3, take the stool by holding the handle 2d.
The instrument 2 is moved upward, and the tip 2a is moved to the buffer storage section 3.
Move into c. At this time, it adheres around the tip 2a.
The excess stool sample that has been removed is removed when passing through the opening 3a.
And the amount of stool sample corresponding to the concave portion of the tip 2a is buffered.
It is guided into the storage part 3c (see FIG. 3). Next, cover 4
Is reattached to the feces collection instrument main body 2. And the whole
The stool sample can be mixed by overturning or shaking.
Is dissolved in buffer solution 6 to prepare a stool sample solution. Then figure
As shown in FIG. 4, the stool collection device 2 is further pulled up,
A partition wall 3d is provided to divide the connecting portion 2c of the stool device 2.
Move to the position As a result, the buffer storage section 3c and
A flow path A is formed between the filter and the filter 8. Therefore, FIG.
To tilt the entire fecal occult blood determination device 1 from the state shown in FIG.
Thus, the stool sample solution is guided to the filter 8. The undigested solid in the stool sample solution by the filter 8
The filtered stool sample solution is supplied to the developing layer 9.
Is done. The fecal sample solution spreads in the spreading layer 9 and
As described above, the stool sample solution is accompanied by the first antibody-sensitized carrier 10.
The end of the development layer 9, that is, the filter 8 is provided.
Move towards the end opposite the side. At this time,
When human Hb is contained in the feed solution,
Due to the immune reaction, the human Hb-first antibody sensitized carrier complex is
The complex is formed and transferred to the immobilization site of the second antibody 11.
To the second antibody 11 at the second antibody immobilization site.
Be wrapped. Accordingly, the color tone of the carrier is determined from the judgment window 3f.
Can be observed, and can be determined to be positive. On the other hand, human
If Hb was not included in the stool sample solution,
No immune complex was formed and only the first antibody-sensitized carrier was
Move to the antibody 11 side, and do not get trapped by the second antibody 11.
No. Therefore, the first antibody-sensitized carrier is entirely on the end side of the spreading layer 9.
To observe the color of the carrier from the judgment window 3f.
Can not. Therefore, it can be determined to be negative. Also,
Thus, if human Hb is contained in the stool sample solution, the first
A part of the antibody-sensitized carrier 10 contains human Hb in the stool sample solution.
If not, all of the first antibody-sensitized carrier 10 is
Reacts with the first antibody-sensitized carrier with each stool sample solution
Substance 12 that reaches the site where the
The color tone of the carrier is observed from the end window 3g.
Can be In other words, to observe this color tone
Therefore, it can be confirmed that this equipment is operating normally.
You. In addition, the stool sample solution reached the position of the end window 3g.
That is, the reaction was performed at the determination site of the determination window 3f.
Observe the change in color tone at the end window 3g.
By doing so, the display in the determination window 3f can be determined. The judgment window 3f and the end window 3g are
As long as the judgment can be made clearly,
The shape and size are not particularly limited.
No. As described above, the fecal occult blood determination device 1 is relatively simple.
Exempt from the collection of stool samples
Up to determination of fecal occult blood by epidemiological chromatography
Can be reliably performed by the relatively simple operation described above.
Wear. Next, a specific example using the fecal occult blood determination device 1 will be described.
An experimental example will be described. (1) Preparation of first antibody-sensitized carrier As a first antibody, an anti-human Hb monoclonal antibody (mouse
MU-110: manufactured by Japan Biotest Laboratory)
Acid buffer solution (pH 7.2: antibody concentration is 200 μg / m
l) 10 ml was prepared. In addition, the latex described above
Polystyrene obtained by methods such as contacting particles with a colorant
Len blue latex (average particle size 0.12 μm: solid content
5% by weight) and stirred at 37 ° C for 60 minutes.
did. The resulting dispersion is treated at 50 ° C. at 18000 rpm at 4 ° C.
Centrifuged for minutes. After removing the supernatant, 1 wt% bovine
Serum albumin (Sigma) phosphate buffer (p
H7.2) to bring the latex concentration to 1% by weight.
It was adjusted so that. (2) Installation of first antibody-sensitized carrier, second antibody
Preparation of Immobilized and Human Hb-Immobilized Spreading Layer Anti-human Hb monoclonal antibody (mouse
MU-107: 1 mg of Japan Biotest Laboratory)
/ Ml phosphate buffer (pH 7.2) with 5 μl
Trocellulose membrane: Biorat, size
1 × 5 cm) planned position of immobilization of the second antibody in FIG. 1
And dried at room temperature for 12 hours. Next, one weight
% Bovine serum albumin in phosphate buffer (pH 7.2)
Soak at 37 ° C. for 30 minutes, dry again at room temperature for 12 hours
Was. In addition, the first antibody sensitivity obtained in the above (1)
0.5 ml of the developed carrier in the same manner as the second antibody
The sample was adsorbed to the position of the first antibody-sensitized carrier in 1. In addition, human Hb (manufactured by Sigma)
0.5 μl of 0 μg / ml phosphate buffer (pH 7.2)
Substance Reacting with the First Antibody-Sensitized Carrier in FIG.
Is dropped at the position to be fixed and dried at room temperature for 12 hours.
Was. (3) Preparation of Fecal Occult Blood Determination Device The fecal occult blood determination device 1 shown in FIG. 1 was prepared. Stool collection equipment
And made of polyethylene shaped as shown in Fig. 1.
And the container body is made of polyether
Use a product made of Tylene (with an internal volume of 10 ml) and add phosphoric acid
3 ml of buffer was injected. Also, a filter 8 for filtration
For liquid chromatography with an average pore size of 2 μm
A filter (manufactured by Sekisui Chemical Co., Ltd.) was used. Test Example 1 In this example, a sensitivity test was performed using a human standard solution. Human
Hb (manufactured by Sigma) in phosphate buffer (pH 7.2)
To a concentration of 0-1000 μg / ml.
3 ml prepared as described above was prepared in the above example.
Used in place of 3 ml of phosphate buffer solution for fecal occult blood test
As a result, a fecal occult blood determination device was prepared. [0053] The container body of the fecal occult blood determination device is shaken lightly.
And then pull the handle of the stool to the end,
The solution was spread on a developing layer. After standing for 5 minutes, the container
The second antibody immobilization site from the body judgment window and the human from the end window
Observe the Hb immobilization site and observe the blue color of the colored latex
Positive or negative judgment was made depending on whether it was performed. No.
Table 1 shows the results of observation of the two antibody-immobilized sites. Table 1
In addition, + indicates positive and-indicates negative. From the end window
The results of observing the human Hb immobilization site were all blue.
Was. [Table 1] From the results shown in Table 1, the determination device of the present invention
The measurement sensitivity was found to be 0.25 μg / ml. Test Example 2 In this example, a fixed amount of human Hb standard was added to a human stool sample
Then, the sample was applied to the same determination device as in Test Example 1. As in Test Example 1, 0-100 Hb of human Hb
3 ml of phosphate buffer at a concentration of 0 μg / ml was
3 ml of the phosphate buffer of the fecal occult blood determination device prepared in Example
Instead, a fecal occult blood determination device was prepared. Above feces
Using a blood determination device, the stool of a healthy person (ie,
Human Hb is not included)
A sample was collected and dissolved in a buffer in the container body. The above trial
The same operation as in Experimental Example 1 was carried out, and the observation of the second antibody-immobilized site was performed.
The results are shown in Table 1 above. From the end window, human Hb
The observation of the immobilized site was all blue. From Table 1
As is clear, almost the same results as in Test Example 1 were obtained.
Was. Therefore, the determination device of the present invention can be used for testing occult blood in a stool sample.
It can be seen that it can actually be applied. As described above, according to the present invention, the toilet bowl
Using the container body where the utensil is set,
Pierce the stool sample, and then place the stool sample in the buffer reservoir.
By guiding and shaking the whole or giving vibration,
A stool sample solution can be easily prepared. In addition,
Continue to operate the stool device in its length direction,
Guide the sample solution to the filter and remove insoluble solids with the filter.
After removal, follow the immunochromatography method in the developing layer.
Thus, the presence or absence of fecal occult blood can be determined. That is, according to the present invention, collection of a stool sample
Of stool sample solution from milk and immunochromatography
All the work up to the determination of the presence or absence of fecal occult blood by the method of the present invention
Performed as a series of steps using a simple fecal occult blood determination device
be able to. Therefore, even unskilled people can easily
Hygienically and reliably determine the presence or absence of fecal occult blood
You.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a longitudinal sectional view of a fecal occult blood determination device according to one embodiment of the present invention. FIG. 2 is a schematic perspective view for explaining a process of collecting a stool sample using the fecal occult blood determination device of the present embodiment. FIG. 3 is a cross-sectional view for explaining a step of preparing a stool sample solution. FIG. 4 is a cross-sectional view for explaining a step of guiding a stool sample solution to a filter. FIGS. 5 (a) to 5 (e) are partial cutaway side views showing a modification of the shape of the distal end portion. [Description of Signs] 1 Fecal occult blood determination device 2 Fecal sampling device 2a Tip 2c Connection part 3 constituting flow path forming means Container main body 3a Opening 3b Opening 3c Buffer storage part 3f Determination window 3g End window 4 Cover 5a, 5b O-ring as a liquid-tight seal ring 6 Buffer 8 Filter 9 Spreading layer 10 First antibody-sensitized carrier 11 Second antibody 12 Substance reacting with anti-human hemoglobin antibody-sensitized carrier

Continued on the front page (56) References JP-A-8-320320 (JP, A) (58) Fields investigated (Int. Cl. ⁷ , DB name) G01N 33/50 G01N 31/22 G01N 33/48 G01N 33 / 53 G01N 33/72

Claims (1)

(57) [Claim 1] An apparatus for judging fecal occult blood, comprising a substantially rod-shaped stool having a distal end capable of semi-quantitatively collecting stool; A cylindrical container body in which the device is inserted and built in through an opening at one end, and the inside of which is configured so that the stool can be moved in the longitudinal direction thereof; and the other end of the container body. The opening is formed so that the distal end of the stool collection instrument can enter and exit, and a buffer for dissolving stool and storing a buffer solution for preparing a stool sample solution inside the container body. The liquid reservoir, and a filter for filtering the stool sample solution, arranged so that the stool sample solution filtered by the filter is supplied, and provided with an anti-human hemoglobin antibody sensitized carrier, Differences between anti-human hemoglobin antibody and recognition epitope An anti-human hemoglobin antibody is immobilized, and a substance that reacts with the anti-human hemoglobin antibody-sensitized carrier is fixed to a position different from the anti-human hemoglobin antibody-sensitized carrier installation position and the anti-human hemoglobin antibody-immobilized position. And a determination window for enabling determination of fecal occult blood from outside, and an end window for confirming that the determination has been completed. In order to prevent the stool sample solution from leaking to the outside of the container main body when the stool collection instrument is moved in the length direction within the container main body, the other end opening of the container main body is formed. A liquid-tight seal ring attached to the vicinity, and provided on the stool collection device and / or the container body to form a flow path for supplying a stool sample solution from the buffer storage section to the filter. And a lid body detachably attached to the container body so as to cover the other end opening of the container body and the distal end portion of the stool. .