JP2003066040A - Preparation method and composition for liquid reagent raw material - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a preparation method and composition for liquid reagent raw material, particularly, a stable preparation method for a non-prepared liquid reagent used in clinical examination field, and a composition using it. SOLUTION: In this preparation method for liquid reagent raw material, a liquid reagent raw material comprising at least two components is preliminarily prepared, stored for a fixed period, and then dissolved in a prescribed amount of water or a test solution to complete a liquid reagent comprising at least two components mixed therein, which is then used for measurement. The liquid reagent raw material composition is characterized by that the measured sensitivity of a liquid reagent dissolved in a prescribed amount of water or a test solution after storage at 10 deg.C or lower for 6 months keeps 80% or more of the measured sensitivity before storage, and the measured sensitivity after storing the liquid reagent at 35 deg.C for 1 week keeps 80% or more of the measured sensitivity before storage.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method and a composition for producing a liquid reagent raw material. In particular, it relates to a method for producing a stable reagent raw material for an unprepared liquid reagent used in the field of clinical examination, and a composition using the same.

[0002]

2. Description of the Related Art As a diagnostic agent used in the field of clinical examination, there is a biochemical measuring reagent utilizing an enzymatic reaction. In this reagent, since the enzyme is unstable in the solution, the enzyme and the unstable substance other than the enzyme were freeze-dried and dissolved in the solution before use. Therefore, there is a problem in that the manufacturing side requires freeze-drying, which results in cost and manufacturing time, and the user side requires time and effort for dissolution.

On the other hand, in recent years, an unprepared liquid reagent which does not require preparation has been developed to solve the problems associated with freeze-drying. This liquid reagent was completed as a reagent that can be stored in solution for a long period of time by stabilizing the raw material enzymes, coenzymes, etc. by various methods, and is now the mainstream in the domestic biochemical measurement reagent market. There is.

It is considered that the good distribution circumstances in Japan are largely responsible for the reason why this reagent becomes a commercial product. In other words, it is possible to deliver in a refrigerated state within at least two days geographically in Japan. From this,
As for the product design of the liquid reagent, it is necessary and sufficient if it can maintain the quality at low temperature for about one year.

However, the situation is different when the present liquid reagent is exported overseas. In other words, in the case of export, distribution at low temperature is possible, but the number of days required for transportation, including procedures, is required. Furthermore, it is the actual situation that the storage temperature cannot be sufficiently guaranteed at the export destination due to the distribution circumstances of the country. For this reason, the liquid reagent of the present design has a problem that the quality retention period and performance equivalent to those in Japan cannot be obtained at overseas export destinations.

In addition to the quality, since it is a liquid reagent in terms of quality, it is heavier than powder and costs more to transport, and there are problems such as container damage and liquid leakage due to vibration during transportation.

Further, in the manufacturing process of the present liquid reagent, preparation of each raw material reagent requires the longest lead time, and therefore it is necessary to improve the process.

[0008]

The problem to be solved by the present invention is to provide a stable reagent raw material used for an unprepared liquid reagent and a method for producing the same, and further labor saving in the process for producing the liquid reagent. The purpose of the present invention is to provide a stable liquid reagent raw material for a long period of time that contributes to the realization.

[0009]

Means for Solving the Problems As a result of various studies for achieving the above-mentioned object, the present inventors have made it possible to obtain long-term stability by mixing two or more components in a dry state without dissolving them. The present inventors have completed the present invention by producing a simple liquid reagent raw material and finding that a kit using the same is stable for a long period of time. That is, the present invention has the following configurations. (1) In a liquid reagent prepared by mixing at least two kinds of components, a liquid reagent raw material composed of at least two kinds of components is prepared in advance, held for a certain period, and then dissolved with a predetermined amount of water or a test solution. A liquid reagent raw material to be used for measurement after completing the liquid reagent, the liquid reagent raw material after being stored at 10 ° C. or lower for 6 months, the measurement sensitivity of the liquid reagent dissolved in a predetermined amount of water or a reagent solution is A liquid reagent raw material composition, which holds 80% or more of the measurement sensitivity, and further holds 80% or more of the measurement sensitivity after storing the liquid reagent at 35 ° C. for 1 week with respect to the measurement sensitivity before storage. . (2) The liquid reagent raw material composition according to (1), in which two or more components are mixed in a dry state without being dissolved. (3) Liquid reagent raw materials are uniformly mixed (1) to
The liquid reagent raw material composition according to (2). (4) contains at least 10% (w / w) buffer,
The liquid reagent raw material composition according to any one of (1) to (3). (5) Containing 50% (w / w) or more of a buffer, (1)
~ Liquid reagent raw material composition according to (4). (6) The two or more components are substances selected from buffers, enzymes, coenzymes, enzyme substrates, chromogens, preservatives, metal salts, surfactants, and stabilizers (1) to (5) ) Liquid reagent raw material composition. (7) The liquid reagent raw material composition according to (1) to (6), which is used by dissolving it in a predetermined amount of water or a reagent solution. (8) Liquid reagent raw material composition according to (1) to (7), which is used by subdividing a predetermined dose of a liquid reagent raw material into a container to be used in an analyzer, and adding and dissolving a predetermined amount of water or a reagent solution to the container. . (9) The liquid reagent is a creatinine measurement reagent (1) to
The liquid reagent raw material composition according to (7). (10) The liquid reagent is a neutral fat measurement reagent (1) to
The liquid reagent raw material composition according to (7). (11) The liquid reagent raw material composition according to (1) to (7), wherein the liquid reagent is a cholesterol measurement reagent. (12) The liquid reagent is a uric acid measuring reagent (1) to (7)
Liquid reagent raw material composition. (13) In a liquid reagent prepared by mixing at least two kinds of components, a liquid reagent raw material composed of at least two kinds of components is prepared in advance, held for a certain period, and then dissolved with a predetermined amount of water or a test solution. A liquid reagent raw material to be used for measurement after completing the liquid reagent,
After storage for 6 months below, the measurement sensitivity of liquid reagents dissolved in a specified amount of water or reagent solution is 80% of the measurement sensitivity before storage.
In the production of the liquid reagent raw material composition, which retains the above and further retains the liquid reagent at 35 ° C. for 1 week, the measurement sensitivity is 80% or more with respect to the measurement sensitivity before storage, without dissolving each component. A method for producing a liquid reagent raw material, which comprises mixing in a dry state. (14) The method for producing a liquid reagent raw material according to (13), wherein the liquid reagent raw material is uniformly mixed. (15) The method for producing a liquid reagent raw material according to (12) to (13), which contains at least 10% (w / w) of a buffer. (16) Containing 50% (w / w) or more of a buffer, (1
The method for producing a liquid reagent raw material according to 2) to 14).

[0010]

BEST MODE FOR CARRYING OUT THE INVENTION As one embodiment of the present invention, a liquid for clinical examination containing a buffer, an enzyme, a coenzyme, an enzyme substrate, a chromogen, a preservative, a metal salt, a surfactant, a stabilizer and the like. There is a reagent raw material.

The liquid reagent raw material of the present invention comprises a buffer, an enzyme,
It is produced by mixing two or more kinds of components selected from coenzyme, enzyme substrate, chromogen, preservative, metal salt, surfactant and stabilizer in a dry state without being dissolved. Preferably,
It is mixed uniformly based on the performance of the liquid reagent after restoration. As for the performance after reconstitution, for example, a predetermined amount of liquid reagent raw material is randomly weighed at N = 3, and the measurement sensitivity of the liquid reagent dissolved in the predetermined amount of water or reagent solution is within the allowable range. . As the allowable range, the sensitivity range of N = 3 is 40%.
Within, preferably within 20%. As a manufacturing environment, for example, it is preferable to manufacture in a low humidity room or the like in which temperature, humidity and the like are controlled. Specifically, the humidity is 60% or less, preferably 50% or less, more preferably 40% or less, and the temperature is 40 ° C. or less, preferably 30 ° C. or less. In addition, the respective components are mixed by mixing two or more kinds of components selected from a buffer, an enzyme, a coenzyme, an enzyme substrate, a chromogen, an antiseptic, a metal salt, a surfactant, a stabilizer and the like. Is placed in a hermetically sealed container (which is preferably subjected to a discharge treatment for the purpose of suppressing adsorption of each component into the container) and mixed well by using a rotor or the like. At this time, each of the components having a large particle size can be ground in advance with a mortar or the like, or the particles can be mixed more uniformly by sieving so that the particle size is below a certain level. The liquid reagent raw material is not limited to one kind for one constitution of the liquid reagent, but may be completed by mixing two or more kinds of raw materials. Although the method for reconstituting the liquid reagent is not particularly limited, purified water or the like, or a reagent solution containing a component that is not contained in the liquid reagent raw material or a component that cannot maintain a solid state at room temperature is also used. The use of the liquid for the restoration has the advantage of saving labor in the restoration process. In particular, when using purified water, the restoration process is greatly labor-saving.

The buffer used in the present invention is not particularly limited, but examples thereof include Tris buffer, citrate buffer, phosphate buffer, borate borax buffer, GOOD buffer and the like. Among them, the Tris buffer, the citrate buffer, the borax borates buffer, and the phosphate buffer have an advantage that they are inexpensive although the pH easily changes depending on the concentration and temperature. In addition, GOS buffer is MES, Bis-Tr
is, ADA, ACES, BES, PIPES, MOP
S, TES, HEPES, Tricine, Bicin
e, POPSO, TAPS, CHES, CAPS, etc. are exemplified. In addition, these buffers may be prepared in concentrations or salts so that the target pH is obtained after the liquid reagent is reconstituted.
For example, a phosphate buffer may be prepared by mixing a dihydrogen phosphate 2Na salt and a dihydrogen phosphate K salt, a Tris buffer solution with a hydrochloride salt, and a GOOD buffer solution with a mixture ratio with a Na salt.

The concentration of the buffer used in the present invention is not particularly limited, but it is usually at least 10% (w / w) with respect to other coexisting components, preferably (from the viewpoint of uniformly mixing the components). It is desirable that the content is 50% (w / w) or more.

The enzymes used in the present invention are microorganisms, plants,
Examples include those collected from mammals, those produced from genetically modified microorganisms in which these genes are incorporated into other microorganisms, and those containing genetically modified properties are not particularly limited. .

The coenzyme used in the present invention includes nicotinamide adenine dinucleotide, flavin adenine dinuclentide, pyridoxal phosphate, and adenosine 3
Examples include phosphoric acid and adenosine diphosphate.

The enzyme substrate used in the present invention is not particularly limited, but α-ketoglutarate, phosphoenolpyruvate, creatine phosphate, 2-ketoisohexanoic acid, L-aspartic acid, L-α-alanine and the like are used. Can be mentioned.

Examples of the color source used in the present invention include an oxidative coloring reagent, a reducing coloring reagent, a Trinder reagent and a coupler thereof. Oxidative color reagent Reagents include hexa (2-hydroxy-3-sulfopropyl)-
4,4,4-triaminotriphenylmethane, hexa (3-sulfopropyl) -4,4,4-triaminotriphenylmethane, bis (2-hydroxy-3-sulfopropyl) toluidine and the like can be mentioned. Examples of the reducing color reagent include tetrazolium salts.
T, MTT, TB, WST-1, WST-3, WST-
5 and the like, which are used in combination with an electron carrier such as 1-m-PMS. As a Trinder reagent, N-
Ethyl-N-sulfopropyl-m-anisidine, N-ethyl-N-sulfopropylaniline, N-ethyl-N-
Sulfopropyl-3,5-dimethoxyaniline, N-sulfopropyl-3,5-dimethylaniline, N-ethyl-N-sulfopropyl-3,5-dimethylaniline, N
-Ethyl-N-sulfopropyl-m-toluidine, N-
Ethyl-N- (2-hydroxy-3-sulfopropyl)
-M-anisidine, N-ethyl-N- (2-hydroxy-3-sulfopropyl) aniline, N-ethyl-N-
(2-Hydroxy-3-sulfopropyl) -3,5-dimethoxyaniline, N- (2-hydroxy-3-sulfopropyl) -3,5-dimethoxyaniline, N-ethyl-N- (2-hydroxy-3) -Sulfopropyl) -3,
Examples thereof include 5-dimethylaniline, N-ethyl-N- (2-hydroxy-3-sulfopropyl) -m-toluidine, and N-sulfopropylaniline. Examples of the coupler include 4-aminoantipyrine and MBTH.

Examples of the preservatives used in the present invention include azides, chelating agents, antibiotics and antibacterial agents. Antibacterial agents include MIT, HPO, CAA, IZ
U, BND, etc. are mentioned.

The metal salt used in the present invention is not particularly limited, but salts such as sodium, potassium, calcium and the like, such as NaCl, KCl, CaCl 2 and the like can be mentioned.

The surfactant used in the present invention includes:
Non-limiting examples include nonionic surfactants, cationic surfactants, anionic surfactants and zwitterionic surfactants. As the nonionic surfactant, N, N-
Bis (3-D-glucoamidopropyl) colamide, n-
Heptyl-β-D-thioglucoside, n-octyl-β
-D-thiogukoside, MEGA-8, MEGA-9, M
Examples include EGA-10 and sucrose monoca plate. Examples of the cationic surfactant include SDS. Sodium cholate is used as the anionic surfactant, and CHAPS, CH is used as the zwitterionic surfactant.
APSO is mentioned.

The stabilizer used in the present invention is not particularly limited, and examples thereof include known stabilizers such as thiol compounds, flavin-containing compounds, saccharides, albumin and chelating agents.

Another embodiment of the present invention is a liquid reagent raw material which is prepared and held for a certain period of time, and then an additional component is added to complete (reconstitute) the liquid reagent to be used for measurement.
Retention for a period of time includes storing or moving. Specific examples include storage in factories and storage areas and warehouses adjacent to it, transportation by trucks, railroads, aircraft, ships, etc. or by humans, and storage at storage areas such as storage areas and warehouses. To be

Although the holding period is not particularly limited, it is 6 months or longer, and more preferably 1 year or longer at a low temperature of 10 ° C. or lower in consideration of distribution circumstances. The temperature during the holding period is not particularly limited, but is preferably 10 ° C or lower, more preferably 0 ° C or lower, and further -2.
It is 0 ° C or lower. In the actual evaluation, the storage period may be replaced by a short-term storage at high temperature, for example, a severe (accelerated) test at 37 ° C. for 2 weeks. This method has an advantage that it is possible to evaluate not only long-term storage stability but also relatively short-term high temperature resistance considering the possibility of being temporarily exposed to high temperature during transportation. In the present invention, for example, after the liquid reagent raw material is stored at 37 ° C. for 2 weeks, the measurement sensitivity of the liquid reagent dissolved in a predetermined amount of water or a reagent solution is
80% or more of the measurement sensitivity before storage is maintained.

After the liquid reagent raw material of the present invention is held for a certain period of time, the reconstituted liquid reagent exhibits the same quality holding period and performance as the liquid reagent of the same composition prepared by preparing all the components in advance. For example, the measurement sensitivity after storing the liquid reagent at 35 ° C. for 1 week is 80% or more of the measurement sensitivity before storage.

When restoring the liquid reagent raw material of the present invention,
All of them may be restored, or a part of them may be sorted and restored. Since the components of the liquid reagent raw material of the present invention are uniformly mixed, there is an advantage that there is little variation in reagent performance when the process of separating and restoring a part is repeated.

Examples of the use of the liquid reagent raw material according to the present invention include a liquid reagent used for measuring uric acid, creatinine, neutral fat, cholesterol, urea nitrogen and the like in biological components. The configuration of these liquid reagents is not particularly limited, and 1
Used as a test solution or a kit of two or more test solutions.

[0027]

EXAMPLES The present invention will be specifically described below with reference to examples. The present invention is not particularly limited to the examples.

(Example 1) A liquid reagent raw material for measuring uric acid comprising the following two reagents was prepared. <Preparation of Uric Acid Measuring Liquid Reagent Raw Material> First Reagent (1 L as Liquid Reagent) Disodium Hydrogen Phosphate 8.5 g, Sodium Dihydrogen Phosphate 5.5 g, EDTA.3Na 0.1 g, 4-
Aminoantipyrine 0.3 g, peroxidase (TOYOBO PEO-301) 0.02 g, ascorbate oxidase (TOYOBO ASO-311) 0.03 g
Was uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. Second reagent (1 L portion as a liquid reagent) disodium hydrogen phosphate 8.5 g, sodium dihydrogen phosphate 5.5 g, EDTA.3Na 0.1 g, potassium ferrocyanide 0.08 g, N-ethyl-N-
(2-Hydroxy-3-sulfopropyl) -m-toluidine 0.3 g / L, uricase (TOYOBO UAO-2
11) 0.3 g was uniformly mixed in a low humidity chamber to obtain a liquid reagent raw material. The above raw material was immediately dissolved in purified water and stored in a liquid reagent raw material state at -20 ° C for 1 year.
/ DL uric acid aqueous solution was measured.

<Measurement method> A Hitachi 7170 type automatic analyzer was used. 180 μL of the first reagent was added to 4 μL of the sample and 37 ° C.
It was incubated for 5 minutes in the reaction to make it the first reaction. After that, 90 μL of the second reagent was added and incubated for 5 minutes to give a second reaction. The absorbance at 600 nm was measured by the 2-point end method in which the absorbance of the first reaction and the absorbance of the second reaction were liquid-corrected and the difference between the absorbances was taken. The result was determined as the measured absorbance (sensitivity) after subtracting the measured absorbance of purified water.

Results are shown in Table 1. The results show that when the measurement sensitivity of the reagent (Example 1-) in which the liquid reagent raw material was immediately dissolved after preparation was 100, the liquid reagent raw material stored for 1 year at -20 ° C after preparation was restored (Example 1-). ) Shows the relative value of the measurement sensitivity. Good storage stability from the preparation to the restoration was confirmed.

[0031]

[Table 1]

(Example 2) The liquid reagent reconstituted in Example 1 was stored at 10 ° C for 1 year, and 10 mg / mg was stored in the same manner as in Example 1.
A dL aqueous solution of uric acid was measured. The method was the same as in Example 1.

Results are shown in Table 1. In Example 2-, the reagent after dissolution in Example 1- was further added at 10 ° C,
Example 1 of measurement sensitivity when stored for 1 year 100-
Indicates the relative value when. It was possible to confirm good storage stability after restoration.

Example 3 A liquid reagent raw material for creatinine measurement consisting of the following two reagents was prepared. <Preparation of Creatinine Measurement Liquid Reagent Raw Material> First Reagent (1 L as Liquid Reagent) Disodium hydrogen phosphate 11.92 g, potassium dihydrogen phosphate 2.16 g, EDTA.3Na 0.5 g,
NaCl 3 g, N-ethyl-N-sulfopropyl-m
-Toluidine 0.2 g, ascorbate oxidase (TOYOBO ASO-311) 0.03 g, sarcosine oxidase (TOYOBO SAO-341) 0.8 g,
Creatine amidinohydrolase (Toyobo CRH-2
21) 3.5 g was uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. Second reagent (1 L portion as liquid reagent) disodium hydrogen phosphate 11.92 g, potassium dihydrogen phosphate 2.16 g, 4-aminoantipyrine 0.1.
5 g, peroxidase (TOYOBO PEO-301)
0.06 g and 1.8 g of creatinine amide hydrolase (CNH-311 manufactured by Toyobo) were uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. -20 in the state of a liquid reagent and a liquid reagent raw material obtained by immediately dissolving the above raw material with the following dissolution liquid
The solution was stored at 0 ° C for 1 year, and after storage, a 5 mg / dL creatinine aqueous solution was measured using a liquid reagent dissolved in the following solution.

<Dissolution> Dissolution solution for the first reagent PIPES 50 mM (pH 7.5), catalase (CAO-509 manufactured by Toyobo) 100 units / mL, Emulgen 420 0.5% Solution solution PIPES 50 mM for the second reagent (pH 7) .5), Emulgen 42
0 0.5%

<Measurement method> A Hitachi 7170 type automatic analyzer was used. 180 μL of the first reagent was added to 3.3 μL of the sample, and then 3
Incubation at 7 ° C for 5 minutes was used as the first reaction.
After that, 60 μL of the second reagent was added and incubated for 5 minutes to make a second reaction. The absorbance at 546 nm was measured by a 2-point end method in which the absorbance of the first reaction and the absorbance of the second reaction were liquid-corrected and the difference between the absorbances was taken. The result was determined as the measured absorbance (sensitivity) after subtracting the measured absorbance of purified water.

Results are shown in Table 2. The results show that when the measurement sensitivity of the reagent (Example 3-) in which the liquid reagent raw material was immediately dissolved after preparation was set to 100, the reagent obtained by restoring the liquid reagent raw material stored at -20 ° C for 1 year after preparation (Example 3- ) Shows the relative sensitivity of. Good storage stability from the preparation to the restoration was confirmed.

[0038]

[Table 2]

Example 4 The liquid reagent reconstituted in Example 3 was stored at 10 ° C. for 1 year and stored in the same manner as in Example 3 at 5 mg / dL.
The creatinine aqueous solution was measured. The method was the same as in Example 3.

Results are shown in Table 2. In Example 4-, the reagents after dissolution in Example 3-are further added at 10 ° C,
The relative sensitivity when stored for 1 year is shown. It was possible to confirm good storage stability after restoration.

Example 5 A raw material for a liquid reagent for measuring neutral fat consisting of the following two reagents was prepared. <Preparation of Neutral Fat Measuring Liquid Reagent Raw Material> First Reagent (1 L as Liquid Reagent) Trishydroxyaminomethane Hydrochloride (TRIZMA
PRE-SET CRYSTALS pH 7.0) 1
5g, EDTA / 2Na 0.1g, sodium chloride
5.0 g, 4-aminoantipyrine 0.2 g, glycerol kinase (Toyobo GYK-311) 0.03
5 g, glycerophosphate oxidase (Toyobo G3O-
311) 0.4 g, ascorbate oxidase (ASO-311 manufactured by Toyobo) 0.03 g, flavin adenine dinucleotide 0.02 g, adenosine triphosphate
3 g of CHAPS and 1 g of CHAPS were uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. Second reagent (1 L portion as liquid reagent) disodium hydrogen phosphate 11.92 g, potassium dihydrogen phosphate 2.16 g, EDTA.2Na 0.5 g,
Calcium chloride 0.02 g, N-ethyl-N-sulfopropyl-m-anisidine 0.6 g, potassium ferrocyanide 0.03 g, peroxidase (Toyobo P
EO-301) 0.05 g, lipoprotein lipase (Toyobo LPL-314) 1.5 g, adenosine 3
Phosphoric acid (0.1 g) and CHAPS (1 g) were uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. The above raw material was immediately stored in a liquid reagent dissolved in the following dissolution liquid and the state of the liquid reagent raw material at 37 ° C. for 2 weeks, and after storage, a 200 mg / dL triolein aqueous solution was measured using the liquid reagent dissolved in the following dissolution liquid. .

<Dissolution> Dissolution liquid catalase for the first reagent (CAO-509 manufactured by Toyobo) 250 units /
mL, Magnesium chloride 0.2 g / L Solution for second reagent Magnesium chloride 0.6 g / L

<Measurement method> A Hitachi 7170 type automatic analyzer was used. 180 μL of the first reagent was added to 2.1 μL of the sample and 3
Incubation at 7 ° C for 5 minutes was used as the first reaction.
After that, 90 μL of the second reagent was added and incubated for 5 minutes to give a second reaction. The absorbance at 600 nm was measured by the 2-point end method in which the absorbance of the first reaction and the absorbance of the second reaction were liquid-corrected and the difference between the absorbances was taken. The result was determined as the measured absorbance (sensitivity) after subtracting the measured absorbance of purified water.

Results are shown in Table 3. The results show that when the measurement sensitivity of the reagent (Example 5-) in which the liquid reagent raw material was immediately dissolved after preparation was 100, the liquid reagent raw material stored at 37 ° C for 2 weeks after preparation was restored (Example 5-). The relative sensitivity of Good storage stability from the preparation to the restoration was confirmed.

[0045]

[Table 3]

Example 6 The liquid reagent reconstituted in Example 5 was stored at 35 ° C. for 1 week and then stored in the same manner as in Example 5 at 200 mg / mg.
An aqueous dL solution of triolein was measured. The method is Example 5
I went the same way.

Results are shown in Table 3. Example 6- shows the relative value when the reagent of Example 5- was further stored at 35 ° C. for 1 week, and the relative sensitivity when Example 3-was 100. It was possible to confirm good storage stability after restoration.

Example 7 A liquid reagent raw material for measuring total cholesterol consisting of the following two reagents was prepared. <Preparation of Liquid Reagent Raw Material for Measuring Total Cholesterol> First Reagent (1 L as Liquid Reagent) 8.67 g disodium hydrogen phosphate, 5.29 g potassium dihydrogen phosphate, 0.1 g EDTA / 3Na, 10 g sodium chloride, chloride Calcium 0.02g,
N-ethyl-N-sulfopropyl-m-anisidine
0.2 g, cholesterol esterase (TOYOBO CO
E-311) 0.02 g, peroxidase (TOYOBO PEO-301) 0.02 g, ascorbate oxidase (TOYOBO ASO-311) 0.03 g, C
1 g of HAPS was uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. Second reagent (1 L as a liquid reagent) 8.67 g of disodium hydrogen phosphate, 5.29 g of potassium dihydrogen phosphate, 0.2 g of EDTA.3Na, 0.02 g of potassium ferrocyanide, 0.2 g of 4-aminoantipyrine, cholesterol Oxidase (COO-311 manufactured by Toyobo) 0.09 g, CHAPS 1 g
Was uniformly mixed in a low humidity chamber to prepare a liquid reagent raw material. The above raw materials were immediately stored in a liquid reagent dissolved in the following dissolution liquid and a liquid reagent raw material at 37 ° C. for 2 weeks, and after storage, a 200 mg / dL cholesterol aqueous solution was measured using the liquid reagent dissolved in the following dissolution liquid.

<Solution> First reagent solution Magnesium chloride 0.8g / L Second reagent solution purified water

<Measurement method> A Hitachi 7170 type automatic analyzer was used. 180 μL of the first reagent was added to 2.1 μL of the sample and 3
Incubation at 7 ° C for 5 minutes was used as the first reaction.
After that, 90 μL of the second reagent was added and incubated for 5 minutes to give a second reaction. The absorbance at 600 nm was measured by the 2-point end method in which the absorbance of the first reaction and the absorbance of the second reaction were liquid-corrected and the difference between the absorbances was taken. The result was determined as the measured absorbance (sensitivity) after subtracting the measured absorbance of purified water.

Results are shown in Table 4. The results show that when the measurement sensitivity of the reagent (Example 7-) in which the liquid reagent raw material was immediately dissolved after preparation was 100, the liquid reagent raw material stored at 37 ° C for 2 weeks after preparation was restored (Example 7-). The relative sensitivity of Good storage stability from the preparation to the restoration was confirmed.

[0052]

[Table 4]

(Example 8) The liquid reagent reconstituted in Example 7 was stored at 35 ° C for 1 week and stored in the same manner as in Example 7 at 200 mg / mg.
The dL cholesterol aqueous solution was measured. The method is Example 7.
I went the same way.

Results are shown in Table 4. Example 8- shows the relative value when the reagent of Example 7- was further stored at 35 ° C. for 1 week, and the measurement sensitivity of Example 7- was set to 100. It was possible to confirm good storage stability after restoration.

(Example 9) Creatinein measurement liquid reagent raw material consisting of two reagents of Example 3 was used, and first reagent and second reagent were randomly selected as N = 3 from the liquid reagent raw material and the first reagent was 1.10.
6 g (50 mL for liquid reagent), 0.82 second reagent
2g (50mL as liquid reagent) of 70mL Hitachi 7
In the same manner as in Example 3, 5 mg / dL of creatinine aqueous solution was measured using a liquid reagent prepared by weighing in a bottle (made of polyethylene) dedicated to the 170 type automatic analyzer and dissolving the solution of Example 3 in 50 mL each.

Results are shown in Table 5. Example 9-,
Is N = 3 when the measurement sensitivity of Example 9 is 100.
The relative sensitivity of It was confirmed that the components were uniformly mixed.

[0057]

[Table 5]

(Example 10) The liquid reagent prepared in Example 9 was stored at 35 ° C for 1 week, and 5 mg / mg as in Example 3 was stored.
A dL creatinine aqueous solution was measured. The method is Example 3
I went the same way.

Results are shown in Table 5. Example 10-,
, Are the examples of the measurement sensitivity when the liquid reagents of Examples 9-, and were stored at 35 ° C. for 1 week.
Shows the relative value when 100 is set. It was confirmed that the components were uniformly mixed.

[0060]

INDUSTRIAL APPLICABILITY According to the present invention, components such as a buffer, an enzyme, a coenzyme, an enzyme substrate, a chromogen, an antiseptic, a metal salt, a surfactant and a stabilizer are uniformly dissolved in a dry state. It was possible to provide a simple and stable liquid reagent raw material to be used for an unprepared liquid reagent and a composition using the same, by mixing with a conventional lyophilization step.

─────────────────────────────────────────────────── ─── Continuation of front page (51) Int.Cl. ⁷ Identification code FI theme code (reference) G01N 33/92 G01N 33/92 AF term (reference) 2G045 BB01 BB46 BB48 DA01 DA42 DA60 DA69 4B063 QA01 QA19 QQ61 QQ73 QQ76 QQ84 QR02 QR03 QR12 QR50 QR52 QR53 QR65 QR66 QS28 QX01

Claims (16)

[Claims] 1. A liquid reagent prepared by mixing at least two kinds of components, a liquid reagent raw material comprising at least two kinds of components is prepared in advance, held for a certain period, and then dissolved with a predetermined amount of water or a test solution. A liquid reagent raw material to be used for measurement by completing the liquid reagent by
After storage for 6 months at 0 ° C or below, the measurement sensitivity of liquid reagents dissolved in a specified amount of water or reagent solution is 8 times the measurement sensitivity before storage.
The measurement sensitivity after holding the liquid reagent at 0% or more and storing the liquid reagent at 35 ° C for 1 week is 80% of the measurement sensitivity before storage.
A liquid reagent raw material composition, which is held as described above. 2. The liquid reagent raw material composition according to claim 1, wherein two or more components are mixed in a dry state without being dissolved. 3. The liquid reagent raw material composition according to claim 1, wherein the liquid reagent raw material is uniformly mixed. 4. The liquid reagent raw material composition according to claim 1, which contains at least 10% (w / w) of a buffering agent. 5. A buffer containing 50% (w / w) or more,
The liquid reagent raw material composition according to claim 1. 6. A buffer, an enzyme, a coenzyme, comprising two or more components.
The liquid reagent raw material composition according to claim 1, which is a substance selected from an enzyme substrate, a chromogen, an antiseptic, a metal salt, a surfactant, and a stabilizer. 7. The liquid reagent raw material composition according to claim 1, which is used after being dissolved in a predetermined amount of water or a reagent solution. 8. The liquid reagent raw material composition according to claim 1, wherein the liquid reagent raw material is used by subdividing a predetermined dose into a container used in an analyzer, and adding a predetermined amount of water or a test solution to the container for dissolution. object. 9. The liquid reagent raw material composition according to claim 1, wherein the liquid reagent is a creatinine measuring reagent. 10. The liquid reagent raw material composition according to claim 1, wherein the liquid reagent is a neutral fat measuring reagent. 11. The liquid reagent raw material composition according to claim 1, wherein the liquid reagent is a cholesterol measuring reagent. 12. The liquid reagent is a uric acid measuring reagent.
The liquid reagent raw material composition according to any one of 7 to 7. 13. A liquid reagent prepared by mixing at least two kinds of components, a liquid reagent raw material comprising at least two kinds of components is prepared in advance, held for a certain period of time, and then dissolved with a predetermined amount of water or a test solution. A liquid reagent raw material to be used for measurement after completing the liquid reagent by storing the liquid reagent raw material at 10 ° C. or lower for 6 months, and then measuring the sensitivity of the liquid reagent dissolved in a predetermined amount of water or a test solution. 80% or more of the previous measurement sensitivity was maintained, and the measurement sensitivity after storing the liquid reagent at 35 ° C. for 1 week was 80% of the measurement sensitivity before storage.
%, The method for producing a liquid reagent raw material, which comprises mixing the components in a dry state without dissolving them in the production of the liquid reagent raw material composition. 14. The method for producing a liquid reagent raw material according to claim 13, wherein the liquid reagent raw material is uniformly mixed. 15. The method for producing a liquid reagent raw material according to claim 12, which contains at least 10% (w / w) of a buffering agent. 16. The method for producing a liquid reagent raw material according to claim 12, which contains 50% (w / w) or more of a buffer.