JPH0980046A - Analytical element and production thereof - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a good analytical element by adding insoluble particles and Tris(hydroxymethyl)aminomethane into a reagent liquid, thereby suppressing or retarding aggregation or precipitation of insoluble particles and stabilizing dispersion. SOLUTION: In case of preparing a reagent liquid for producing an analytic element analyzing a specified component in a liquid sample, insoluble particles and Tris(hydroxymethyl)aminomethane are added into the reagent liquid. The insoluble particles include titanium dioxide, powder of filter paper, carbon powder, etc., and light reflective insoluble particles, i.e., titanium dioxide, are preferably employed for the purpose of concealing red blood cell and blocking transmission of light. Rutile-type, anatase-type or amorphous titanium dioxide is employed, for example, and rutile-type titanium dioxide is preferably employed because of its high whiteness. The Tris(hydroxymethyl)aminomethane is a chemically stable substance causing no reaction on other reagent and has no effect on the sensitivity and stability of reagent layer.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to an analytical element for analyzing a specific component in a liquid sample, particularly whole blood, and a method for producing the same.

[0002]

2. Description of the Related Art Analytical elements for analyzing a specific component in a liquid sample are already known.

For example, Japanese Patent Laid-Open No. 2-150751 discloses that
A porous film coated with a reagent solution containing light-reflective insoluble particles and dried was placed so as to cover the through-holes of the support and fixed on the support, and then the sample holding layer and the reagent layer were covered. Analytical elements which can be produced by placing them on a support and fixing them on a support are disclosed.

In Japanese Patent Application Laid-Open No. 2-150751, titanium dioxide is mentioned as particles. This is because when whole blood is used as a sample and measured from below the analytical tool,
It is used for the purpose of hiding red blood cells and at the same time not transmitting light.

Titanium dioxide is a white particle that reflects visible light with little absorption and is hardly soluble in any solvent. The size of the particles is widely distributed, and if the diameter is smaller than 0.001 μm, it is 1 m.
There exist up to m or more. Moreover, the smaller the particles, the more easily they aggregate. It is usually mixed with paints, ointments, cosmetics, etc. for the purpose of not transmitting light.

However, titanium dioxide has the property of not dissolving in a solvent, but its particles easily aggregate,
Aggregates are rapidly generated and settled in the liquid, and the concentration distribution of titanium dioxide is not uniform in the upper and lower parts of the liquid.

When a reagent solution for producing an analytical element for analyzing a specific component in a liquid sample in which titanium dioxide is dispersed, is applied, the titanium dioxide in the solution causes aggregation and precipitation, resulting in a uniform distribution. Application becomes impossible. In order to avoid this, it is conceivable to add a polymer or a thickener to the liquid to give viscosity, or to stir.

[0008] However, it is not preferable to give a viscosity that does not cause precipitation of titanium dioxide, which deteriorates fluidity, makes application difficult, and deteriorates accuracy of the analytical element, and stirring the reagent solution is not preferable. It is difficult to mix in. The inclusion of air bubbles causes a problem such that the layer after application is dried while the air bubbles are trapped, or the air bubbles are ruptured and dried in a crater state.

Furthermore, when the reagent solution in which titanium dioxide is dispersed is applied and then dried, agglomeration also occurs in the drying process, and the agglomerates cause the surface of the applied layer to be non-uniform. This phenomenon causes non-uniform reflection of light when titanium dioxide is used as the optically reflective layer, and significantly deteriorates the performance of the reagent layer.

[0010]

DISCLOSURE OF THE INVENTION Problems to be Solved by the Invention As described above, a reagent solution in which insoluble particles such as titanium dioxide are dispersed can be used without stirring without affecting the accuracy of application and without increasing the viscosity of the solution. It has been desired to find a method for uniformly dispersing insoluble particles such as titanium dioxide in a liquid without raising the temperature more than necessary.

[0011]

[Means for Solving the Problems] As a result of earnest research to solve the above problems, when a reagent solution for preparing an analytical element for analyzing a specific component in a liquid sample is prepared, it is insoluble in the reagent solution. It has been discovered that the inclusion of particles and tris (hydroxymethyl) aminomethane can suppress or delay the occurrence of aggregation and precipitation of insoluble particles.

[0012]

BEST MODE FOR CARRYING OUT THE INVENTION The present invention is disclosed in JP-A-2-1507.
It can also be said to be an improved invention for producing the reagent layer shown in No. 51.

As the insoluble particles in the reagent layer of the analytical element, various materials such as filter paper powder and carbon powder are used in addition to titanium dioxide for various purposes, but they do not conceal light while at the same time concealing red blood cells. For the purpose, light-reflecting insoluble particles are preferred.

As the light-reflective insoluble particles, various particles such as titanium dioxide, magnesium oxide and barium sulfate are used. Among them, in terms of high whiteness,
Titanium dioxide is particularly preferred.

The titanium dioxide may have a general grain structure, and for example, rutile type, anatase type, amorphous and the like are used. Among them, the rutile type is particularly preferable in terms of high whiteness.

The particle size of the particles may be such that they can be dispersed by stirring, but the present invention is effective for particles of 100 μm or less. Preferably 0.001 μm to 1
00 μm, more preferably 0.01 μm to 10 μm,
Is good.

Since tris (hydroxymethyl) aminomethane is a chemically stable substance, it does not react with other reagents and has almost no effect on the sensitivity and stability in the reagent layer.

Tris (hydroxymethyl) aminomethane is a substance that is easily soluble in water. The final concentration of tris (hydroxymethyl) aminomethane is preferably 0.01 to 5 mol / l, more preferably 0.1.
It may be ˜1 mol / l.

Further, tris (hydroxymethyl) aminomethane may be added alone or in a buffer solution,
You may use it, mixing with other buffer solutions. For example, a combination of Tris buffer and phosphate buffer may be mentioned. In the case of Tris buffer, acids such as hydrochloric acid and sulfuric acid are usually used.
The pH is adjusted by adding an alkali such as sodium hydroxide or lithium hydroxide.

The method for producing the analytical element after producing the reagent solution is as follows:
A known method may be used. For example, it can be obtained by applying a reagent solution on a film, printing, spraying, etc., and then drying. Examples will be shown below, but the present invention is not limited thereto.

[0021]

Example 1 Aggregation and Precipitation Rate of Titanium Dioxide Five transparent glass bottles having a diameter of 2.4 cm and a height of 15 cm were prepared, and 0.75 g of titanium dioxide (rutile type (manufactured by Wako Pure Chemical Industries)) was placed in each of them. Weigh. Next, 15 ml of the five kinds of liquids shown in Table 1 were put into each transparent glass bottle, and the liquids were mixed well and allowed to stand. The time taken for titanium dioxide to settle to 10 mm and 6 mm from the bottom surface of the transparent glass bottle was measured. The results are shown in Table 2.

[0022]

[Table 1] Types of liquids 1) Distilled water 2) 0.2 mol / l phosphate buffer (pH 7.2) 3) 0.2 mol / l TES buffer (pH 7.2) 4) 0.2 mol / l HEPES buffer (PH 7.2) 5) 0.2 mol / l Tris buffer

[0023]

[Table 2]

[0024]

Example 2 Determination of glucose in blood The following reagents were weighed and sufficiently stirred to prepare a reagent-containing liquid. This reagent solution was applied on a porous film (Celgard (made by Hoechst)) having a thickness of 25 μm with a wet thickness of 100 μm and dried at 40 ° C. for 1 hour. Next, this is 7m
It was cut into m × 7 mm, and was stuck on a polyethylene terephthalate film having a thickness of 250 μm and having a through hole with a diameter of 4 mm so as to cover the through hole to obtain an analytical element. An electron microscopic photograph of the surface of this sample is shown in FIG. In FIG. 1, the left side is 100 times and the right side is 5 times.
It is 00 times. Reagent composition: glucose oxidase (manufactured by Toyobo) 500 ku peroxidase (manufactured by Toyobo) 300 ku 4-aminoantipyrine (manufactured by Kishida Chemical Co., Ltd.) 500 mg N-ethyl-N- (2-hydroxy-3-sulfopropyl) -3,5-dimethylaniline (Manufactured by Dojindo) 1 g 0.2 mol / l Tris buffer (pH 7.5) 100 ml Propiophan (manufactured by BASF Japan) 4 g 50% polyoxyethylene sorbitan monolaurate aqueous solution (manufactured by Nacalai Tesque) 8 ml Titanium dioxide (Wako Pure) Medicinal product) 8g

[0025]

Comparative Example 1 The following reagents were weighed and sufficiently stirred to prepare a reagent-containing liquid. This reagent solution was applied on a porous film (Celgard (made by Hoechst)) having a thickness of 25 μm with a wet thickness of 100 μm and dried at 40 ° C. for 1 hour. Next, this was cut into 7 mm × 7 mm, and a polyethylene terephthalate film having a diameter of 4 mm and having a through hole opened and a thickness of 250 μm was attached so as to cover the through hole to obtain an analytical element. A photograph of the surface of this sample enlarged with an electron microscope is shown in FIG. In FIG. 2, the magnification is 100 on the left side.
Fold, the right side is 500 times. Reagent composition: glucose oxidase (manufactured by Toyobo) 500 ku peroxidase (manufactured by Toyobo) 300 ku 4-aminoantipyrine (manufactured by Kishida Chemical Co., Ltd.) 500 mg N-ethyl-N- (2-hydroxy-3-sulfopropyl) -3,5-dimethylaniline (Manufactured by Dojindo) 1 g 0.2 moI / l phosphate buffer (pH 7.5) 100 ml Propiophan (manufactured by BASF Japan) 4 g 50% polyoxyethylene sorbitan monolaurate aqueous solution (manufactured by Nacalai Tesque) 8 ml Titanium dioxide (sum Kojunyaku) 8g

Comparing FIG. 1 and FIG. 2, in the case where tris (hydroxymethyl) aminomethane was not added, the particles were coarse and the coating surface was non-uniform, whereas tris (hydroxymethyl) aminomethane was added. On the other hand, it can be seen that the finer particles have a uniform coating surface.

20 μl of whole blood having various glucose concentrations was dropped on each analytical element prepared in Example 2 and Comparative Example 1, and 1 minute later, through the through hole of the support, from below (from the porous film side), Color difference meter Nippon Denshoku Industries Σ−
90), and the reflectance at 640 nm was measured.

A calibration curve was prepared from the reflectance, and the measured values were converted into concentrations based on the calibration curve for comparison. Table 1 shows the measurement results.

[0029]

[Table 1]

In the analytical element of the example using the production method according to the present invention, since the coating thickness after drying and the reagent surface were uniform, the reproducibility of measured values was good and good results were obtained. However, the reproducibility of the analysis element of the comparative example was poor.

[0031]

EFFECTS OF THE INVENTION In a reagent solution containing insoluble particles such as titanium dioxide, tris (hydroxymethyl) aminomethane is contained in order to uniformly disperse titanium dioxide, whereby aggregation and precipitation of titanium dioxide are suppressed, The dispersion was stabilized, and during this period, the reagent solution containing titanium dioxide could be applied. Furthermore, the layer obtained by drying the applied material became more uniform, and a good analytical element was obtained.

[Brief description of drawings]

FIG. 1 is an electron micrograph of the surface showing Example 2. The magnification is 100 times on the left side and 500 times on the right side.

2 is an electron micrograph of the surface showing Comparative Example 1. FIG. The magnification is 100 times on the left side and 500 times on the right side.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Takehiro Yamaguchi 57 Kyoto Higashikujo Nishiamita-cho, Minami-ku, Kyoto City Kyoto Prefecture Daiichi Kagaku Co., Ltd.

Claims (10)

[Claims] 1. A method for producing a reagent solution for producing an analytical element for analyzing a specific component in a liquid sample, which comprises insoluble particles and tris (hydroxymethyl) aminomethane. A method for producing a reagent solution, comprising: 2. The method for producing a reagent solution according to claim 1, wherein the insoluble particles are light reflective. 3. The method for producing a reagent solution according to claim 1, wherein the insoluble particles are titanium dioxide. 4. The size of the insoluble particles is 0.001-diameter.
It is 100 micrometers, The manufacturing method of the reagent liquid in any one of Claims 1-3. 5. The method for producing a reagent solution according to claim 1, wherein the final concentration of tris (hydroxymethyl) aminomethane is 0.01 to 5 mol / l. 6. An analytical element for analyzing a specific component in a liquid sample, which comprises an insoluble particle and a reagent containing tris (hydroxymethyl) aminomethane. 7. The analytical element according to claim 6, wherein the insoluble particles are light-reflecting. 8. The analytical element according to claim 6, wherein the insoluble particles are titanium dioxide. 9. The size of the insoluble particles is 0.001-diameter.
The analysis element according to claim 6, which has a size of 100 μm. 10. A final tris (hydroxymethyl) aminomethane concentration of 0.0 when the liquid sample is supplied.
It is 1-5 mol / l, The analysis element in any one of Claims 6-9.