Method for suppressing blank changes

Abstract

In biological sample measurement reagents, pH changes can occur due to opening the container or contamination with other reagents, which can result in changes to the blank. This invention provides a method for suppressing such changes in the blank caused by pH changes, and a biological sample measurement reagent utilizing this method. [Solution] A surfactant is included in the biological sample measurement reagent. This biological sample measurement reagent can suppress changes in the blank caused by pH changes.

Description

Technical Field

[0001] The present invention relates to a biological sample measurement reagent characterized by containing a surfactant in order to suppress changes in blanks derived from pH changes. The present invention also relates to a method for suppressing changes in blanks derived from pH changes, characterized by containing a surfactant in a biological sample measurement reagent. The present invention is particularly useful in fields such as chemistry, life science, analytical science, and clinical examinations.

Background Art

[0002] Various metals exist in the living body, each exerting diverse functions. Therefore, measuring the metal concentration in a biological sample and observing its fluctuations are essential for disease diagnosis, treatment, early detection, and prevention, and are widely carried out in hospitals, testing facilities, etc. For example, it is known that symptoms such as growth disorders and taste disorders occur when zinc deficiency develops. Therefore, when these symptoms are observed, zinc concentration measurement in serum etc. may be performed.

[0003] Known methods for measuring zinc include atomic absorption spectrometry, ICP emission spectrometry, and colorimetric measurement methods. Among these, the colorimetric measurement method using a chelating chromogenic agent measures the zinc concentration by utilizing the change in absorbance when a chelating chromogenic agent such as 2-(5-bromo-2-pyridylazo)-5(N-propyl-N-sulfopropylamino)phenol sodium (hereinafter sometimes abbreviated as 5-Br-PAPS) or 2-(5-nitro-2-pyridylazo)-5(N-propyl-N-sulfopropylamino)phenol sodium (hereinafter sometimes abbreviated as nitro-PAPS) forms a chelate with zinc. Using this measurement method, it is possible to measure the zinc concentration in a biological sample with a general-purpose automatic analyzer etc. even without an atomic absorption photometer, and it is used in hospital laboratories etc.

[0004] Incidentally, biological sample measurement reagents are not always used immediately after purchase. They may be stored for long periods in general-purpose automated analyzers without stoppers before use, and in such cases, the pH of the biological sample measurement reagent may change over time from its initial value, i.e., from the pH intended by the developer. Furthermore, while general-purpose automated analyzers can often accommodate multiple biological sample measurement reagents simultaneously, components used to transfer the biological sample measurement reagents to the reaction vessel, such as the reaction vessel and agitator, may be reused. As a result, the removal of biological sample measurement reagents and detergents adhering to these components may not be completely achieved, leading to mixing of biological sample measurement reagents with other biological sample measurement reagents and detergents, resulting in a pH value different from that intended by the developer. Thus, due to various factors, the pH of biological sample measurement reagents can differ from the value intended by the developer. If the pH of a biological sample measurement reagent differs from the value assumed by the developer, that is, if a pH change occurs, phenomena such as changes in the blank, decreased stability making long-term storage impossible, and changes in reactivity with the substance being measured may occur, potentially leading to problems such as incorrect measurement results. To address the above issues, methods such as constructing a reagent group consisting of multiple reagents with high-buffering agent concentration reagents and low-buffering agent concentration reagents are known (see Patent Document 1). [Prior art documents] [Patent Documents]

[0005] [Patent Document 1] Japanese Patent Application Publication No. 09-294597 [Overview of the Initiative] [Problems that the invention aims to solve]

[0006] However, since the methods described above cannot be used for all biological sample measurement reagents, it is difficult to completely prevent pH changes with some biological sample measurement reagents, and therefore it is also difficult to completely prevent changes in the blank resulting from pH changes. Such biological sample measurement reagents have problems such as the possibility of not obtaining accurate measurements or even being unable to perform measurements at all.

[0007] The inability to obtain accurate measurements, or even the inability to perform measurements at all, is a serious problem that can lead to misdiagnosis or delays in disease diagnosis. Therefore, there is a need for biological sample measurement reagents that can provide accurate measurements. Consequently, there is a demand for biological sample measurement reagents that are less susceptible to blank changes. [Means for solving the problem]

[0008] As a result of diligent research into the above-mentioned problems and their solutions, the inventors discovered that by including a certain type of surfactant in the biological sample measurement reagent, it is possible to suppress changes in the blank resulting from pH changes, thus completing the present invention.

[0009] In other words, the present invention is as follows: (1) A biological sample measurement reagent characterized by containing a surfactant to suppress changes in the blank caused by pH changes. (2) A biological sample measurement reagent as described in (1) above, wherein the surfactant is at least one of the following: a nonionic surfactant, an amphoteric surfactant, or an anionic surfactant. (3) A biological sample measurement reagent as described in (1) above, wherein the surfactant is at least one of the following: polyoxyethylene lauryl ether, polyoxyethylene alkyl ether, polyether-modified silicone oil, betaine-based surfactant, amine oxide-based surfactant, taurine-based surfactant, higher alcohol-based surfactant, or amino acid-based surfactant. (4) A biological sample measurement reagent as described in any one of items (1) to (3) above, wherein the biological sample measurement reagent is a biochemical test reagent. (5) A biological sample measurement reagent as described in any one of items (1) to (3) above, wherein the biological sample measurement reagent is a zinc measurement reagent. (6) A method for suppressing changes in the blank caused by pH changes, characterized by containing a surfactant in the biological sample measurement reagent. (7) The inhibitory method described in (6) above, wherein the surfactant is at least one of the following: a nonionic surfactant, an amphoteric surfactant, or an anionic surfactant. (8) The inhibitory method described in (6) above, wherein the surfactant is at least one of the following: polyoxyethylene lauryl ether, polyoxyethylene alkyl ether, polyether-modified silicone oil, betaine-based surfactant, amine oxide-based surfactant, taurine-based surfactant, higher alcohol-based surfactant, and amino acid-based surfactant. (9) The suppression method described in any one of items (6) to (8) above, wherein the biological sample measurement reagent is a biochemical test reagent. (10) The suppression method according to any one of items (6) to (8) above, wherein the biological sample measurement reagent is a zinc measurement reagent. [Effects of the Invention]

[0010] This invention makes it possible to suppress changes in the blank caused by pH changes. Therefore, accurate measurement values ​​can be obtained even in situations where pH changes occur. [Modes for carrying out the invention]

[0011] The present invention will now be described in detail. The following embodiments are illustrative for illustrating the present invention and are not intended to limit the present invention to these embodiments only. The present invention can be implemented in various forms without departing from its spirit.

[0012] 1. Overview Biological sample measurement reagents are not always used immediately after purchase. They may be stored for extended periods in general-purpose automated analyzers without stoppers before use, and in such cases, the pH of the biological sample measurement reagent may change over time from its initial value, i.e., from the pH intended by the developer. Furthermore, while general-purpose automated analyzers can often accommodate multiple biological sample measurement reagents simultaneously, components used to transfer the reagents to reaction vessels, such as reaction vessels and agitators, may be reused. As a result, the removal of biological sample measurement reagents and detergents adhering to these components may not be completely achieved, leading to mixing of biological sample measurement reagents with other reagents and detergents, resulting in a pH value different from the developer's intended value. Thus, due to various factors, the pH of biological sample measurement reagents can differ from the value intended by the developer. If the pH of a biological sample measurement reagent differs from the value intended by the developer, that is, if a pH change occurs, phenomena such as changes in the blank, decreased stability making long-term storage impossible, and changes in reactivity with the substance being measured may occur. As a result, problems such as obtaining incorrect measurement results may arise. With some biological sample measurement reagents, it is difficult to completely prevent pH changes, and therefore, it is also difficult to completely prevent changes in the blank resulting from pH changes. Such biological sample measurement reagents have problems such as the possibility of not obtaining accurate measurements or even being unable to perform measurements at all. The inability to obtain accurate measurements, or even the inability to perform measurements at all, is a serious problem that can lead to misdiagnosis or delays in disease diagnosis. Therefore, there is a need for biological sample measurement reagents that can provide accurate measurements. Consequently, there is a demand for biological sample measurement reagents that are less susceptible to blank changes. As a result of diligent research into the above-mentioned problems and their solutions, the inventors have discovered that by including a certain type of surfactant in the biological sample measurement reagent, it is possible to suppress changes in the blank caused by pH changes. This invention was made based on these findings.

[0013] 2. Changes in the blank In this invention, "blank" refers to the absorbance obtained when purified water is measured as a sample in the biological sample measurement reagent, and "blank change" refers to a change in the absorbance obtained when purified water is measured as a sample in the biological sample measurement reagent. Blank changes include both cases where the blank value increases and cases where it decreases. Therefore, suppressing blank changes means reducing or eliminating such fluctuations in the value.

[0014] Various factors can cause changes in the blank, including, for example, changes in the pH or components of the biological sample measurement reagent, and concentration. Deterioration of the reaction vessel or light source of the measurement device can also be a contributing factor.

[0015] 3. Surfactants The surfactant to be included in the biological sample measurement reagent of the present invention is not particularly limited as long as it has the effect of suppressing changes in the blank due to pH changes, but examples include nonionic surfactants, amphoteric surfactants, and anionic surfactants. In particular, nonionic surfactants include polyoxyethylene lauryl ether, polyoxyethylene alkyl ether, and polyether-modified silicone oil, amphoteric surfactants include betaine-based surfactants and amine oxide-based surfactants, and anionic surfactants include taurine-based surfactants, higher alcohol-based surfactants and amino acid-based surfactants. Furthermore, acylmethyl taurine salts are preferred for taurine-based surfactants, alkyl sulfates are preferred for higher alcohol-based surfactants, and acyl sarcosine salts are preferred for amino acid-based surfactants. Examples of polyoxyethylene lauryl ethers include Brij (registered trademark, same hereinafter)-35 (Kishida Chemical), examples of polyoxyethylene alkyl ethers include BT-9 (Nikko Chemicals) and Em1108 (Kao), examples of polyether-modified silicone oils include KF-351A (Shin-Etsu Silicone) and KF-355A (Shin-Etsu Silicone), and examples of betaine-based surfactants include 3-[(3-colamidopropyl)dimethylammonio]-1-pro Examples of surfactants include pansulfonates (sometimes abbreviated as CHAPS) (Fujifilm Wako Pure Chemical Industries), AM301 (Nikko Chemicals), etc., as amine oxide surfactants, examples include Anhitol (registered trademark, same applies hereinafter) 20N (Kao), etc., as acylmethyl taurate salts, examples include LMT (Nikko Chemicals), etc., as alkyl sulfates, examples include DSNa (Nikko Chemicals), etc., and as acyl sarcosine salts, examples include sarcosinate LN-30 (Nikko Chemicals). Surfactants may be included for purposes other than obtaining the effects described above.

[0016] The reagent for measuring biological samples only needs to contain at least one type of surfactant, and may contain two or more types.

[0017] The concentration of the surfactant to be included in the biological sample measurement reagent is not particularly limited as long as it exhibits an effect of suppressing changes in the blank due to pH changes. The concentration of the surfactant may be, for example, in a concentration range of 0.05 - 0.1%, 0.05 - 0.5%, 0.05 - 1%, 0.05 - 5%, 0.05 - 10%, 0.1 - 0.5%, 0.1 - 1%, 0.1 - 5%, 0.1 - 10%, 0.5 - 1%, 0.5 - 5%, 0.5 - 10%, 1 - 5%, 1 - 10%, or 5 - 10%.

[0018] When the biological sample measurement reagent is composed of multiple reagents, the surfactant may be included in at least one reagent, and may also be included in other reagents.

[0019] 4. Biological Sample Measurement Reagent In the present invention, the biological sample measurement reagent is one that can measure the enzyme activity, substance concentration, etc. in a biological sample, and is not particularly limited as long as it is such a reagent. Examples of biological sample measurement reagents include biochemical biological sample measurement reagents using biochemical reactions, immunological biological sample measurement reagents using antigen - antibody reactions, genetic biological sample measurement reagents using gene analysis techniques, etc. In the present invention, biochemical biological sample measurement reagents are preferred. Examples of biochemical biological sample measurement reagents include zinc measurement reagents.

[0020] The biological sample measurement reagent of the present invention may perform measurement by the endpoint method or by the reaction rate method, and may be appropriately selected.

[0021] The biological sample measurement reagent of the present invention may be a one - reagent method in which a biological sample and one reagent are mixed for measurement, a two - reagent method in which a biological sample and two reagents are mixed simultaneously or in an appropriate order for measurement, or a multi - reagent method in which a biological sample and three or more reagents are mixed simultaneously or in an appropriate order for measurement, and may be appropriately selected.

[0022] In the biological sample measurement reagent of the present invention, the measurement may be performed by a manual method, or it may be performed using a device such as a general-purpose automated analyzer.

[0023] The biological sample measurement reagent of the present invention may contain all or part of its constituent reagents as liquid reagents.

[0024] The biological sample measurement reagent of the present invention can be sold on its own or used for measuring target substances in biological samples.

[0025] The biological sample measurement reagent of the present invention can also be sold or used for measuring target substances in biological samples in combination with other reagents other than the biological sample measurement reagent described above. Examples of other reagents other than the biological sample measurement reagent described above include buffer solutions, sample diluents, reagent diluents, reagents containing substances for calibration, or reagents containing substances for quality control.

[0026] The biological sample measurement reagent of the present invention may be a biological sample measurement reagent kit consisting of a first reagent, a second reagent, or a plurality of other reagents.

[0027] The pH of the biological sample measurement reagent of the present invention can be appropriately selected considering the stability of the contained components during storage, the reaction rate during biological sample measurement, etc. Furthermore, if the biological sample measurement reagent is composed of multiple reagents, the pH of each reagent constituting the biological sample measurement reagent can be appropriately selected considering the stability of the contained components during storage, the reaction rate during sample measurement, etc.

[0028] For example, in the case of zinc measurement reagents, the lower limit of the pH when measuring the zinc concentration in the sample is preferably 7.0, and particularly preferably 8.0. Furthermore, the upper limit is preferably 11.0, and particularly preferably 10.0. Regarding the pH of this zinc measurement reagent, for example, if the lower limit is 7.0, then a range of 7.0 to 10.0 or 7.0 to 11.0 can be given, and if the lower limit is 8.0, then a range of 8.0 to 10.0 or 8.0 to 11.0 can be given.

[0029] The biological sample measurement reagent of the present invention may contain, in addition to surfactants, known surfactants, buffers, enzymes, coenzymes, reaction substrates, pH adjusters, preservatives, etc., as needed. The concentration of each of these components can be appropriately selected considering the stability of the contained components during storage, the reaction rate during biological sample measurement, etc. Furthermore, when the biological sample measurement reagent is composed of multiple reagents, the concentration of each of these components in each reagent constituting the biological sample measurement reagent can be appropriately selected considering the stability of the contained components during storage, the reaction rate during biological sample measurement, etc.The buffering agent is not particularly limited, but examples include acetic acid, citric acid, tartaric acid, carbonic acid, boric acid, phosphoric acid, trishydroxymethylaminomethane, imidazole, glycylglycine, bistris, tricine, bicine, bistrispropane (hereinafter sometimes abbreviated as BTP), N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid (hereinafter sometimes abbreviated as TES), 2-morpholinoethanesulfonic acid (hereinafter sometimes abbreviated as MES), N-(2-acetamide)iminodiacetic acid (hereinafter sometimes abbreviated as ADA), N-(2-acetamide)-2- Aminoethanesulfonic acid (hereinafter sometimes abbreviated as ACES), piperazine-1,4-bis(2-ethanesulfonic acid) (hereinafter sometimes abbreviated as PIPES), 2-hydroxy-3-morpholinopropanesulfonic acid (hereinafter sometimes abbreviated as MOPSO), 3-morpholinopropane-1-sulfonic acid (hereinafter sometimes abbreviated as MOPS), N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (hereinafter sometimes abbreviated as BES), 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid (hereinafter sometimes abbreviated as HEPES) (Also available), 3-[N,N-bis(2-hydroxyethyl)amino]-2-hydroxypropanesulfonic acid (hereinafter sometimes abbreviated as DIPSO), 3-[N-tris(hydroxymethyl)methylamino]-2-hydroxypropanesulfonic acid (hereinafter sometimes abbreviated as TAPSO), piperazine-1,4-bis(2-hydroxypropanesulfonic acid) (hereinafter sometimes abbreviated as POPSO), N-(2-hydroxyethyl)piperazine-N'-(3-propanesulfonic acid) (hereinafter sometimes abbreviated as HEPPS), 4-(2-hydroxyethyl)piperazine Examples include -1-(2-hydroxypropane-3-sulfonic acid) (sometimes abbreviated as HEPPSO), N-tris(hydroxymethyl)methyl-3-aminopropanesulfonic acid (sometimes abbreviated as TAPS), 2-cyclohexylaminoethanesulfonic acid (sometimes abbreviated as CHES), 3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid (sometimes abbreviated as CAPSO), or N-cyclohexyl-3-aminopropanesulfonic acid (sometimes abbreviated as CAPS), or salts thereof.

[0030] 5. Biological samples In the present invention, a sample refers to an object that is the subject of measurement, such as enzyme activity or substance concentration, and a biological sample refers to an object contained in a living body that is the subject of measurement, such as enzyme activity or substance concentration. The biological sample is not particularly limited as long as it is one of these types of objects. Examples of such biological samples include human or animal blood, serum, plasma, urine, breast milk, feces, semen, cerebrospinal fluid, saliva, sweat, tears, ascites, amniotic fluid, organs such as the brain, and tissues and cells such as hair, skin, nails, muscles, or nerves.

[0031] The present invention will be described in detail below with reference to examples, but the present invention is not limited to these examples. [Examples]

[0032] [Example 1] (Confirmation of the effect of surfactants on suppressing blank changes) The effect of surfactants in suppressing blank changes was confirmed by varying the pH of the zinc measurement reagent, measuring the blank, and examining the magnitude of the change in the blank.

[0033] 1. Preparation of reagents

[0034] (1) Zinc measurement reagent: Preparation of first reagent, control X-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare control X-1 of the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) (2) Zinc measurement reagent First reagent Preparation of control X-2 A solution was prepared by changing only the pH of control X-1 to 9.5 (20°C), and this was used as control X-2 for the first reagent of the zinc measurement reagent. (3) Zinc measurement reagent First reagent Preparation of control X-3 A solution was prepared by changing only the pH of control X-1 to 9.7 (20°C), and this was used as control X-3 for the first reagent of the zinc measurement reagent. (4) Zinc measurement reagent First reagent Preparation of reagent A1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent A1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Brij-35 0.1% (Kishida Chemical) (5) Zinc measurement reagent First reagent Preparation of reagent A1-2 A solution was prepared by changing only the pH of reagent A1-1 to 9.5 (20°C), and this was designated as reagent A1-2, the first reagent for zinc measurement. (6) Zinc measurement reagent First reagent Preparation of reagent A1-3 A solution was prepared by changing only the pH of reagent A1-1 to 9.7 (20°C), and this was designated as reagent A1-3, the first reagent in the zinc measurement reagent. (7) Zinc measurement reagent First reagent Preparation of reagent A2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent A2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Brij-35 0.5% (Kishida Chemical) (8) Zinc measurement reagent First reagent Preparation of reagent A2-2 A solution was prepared by changing only the pH of reagent A2-1 to 9.5 (20°C), and this was designated as reagent A2-2, the first reagent in the zinc measurement reagent. (9) Zinc measurement reagent First reagent Preparation of reagent A2-3 A solution was prepared by changing only the pH of reagent A2-1 to 9.7 (20°C), and this was designated as reagent A2-3, the first reagent in the zinc measurement reagent. (10) Zinc measurement reagent First reagent Preparation of reagent A3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent A3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Brij-35 1% (Kishida Chemical) (11) Zinc measurement reagent First reagent Preparation of reagent A3-2 A solution was prepared by changing only the pH of reagent A3-1 to 9.5 (20°C), and this was designated as reagent A3-2, the first reagent for zinc measurement. (12) Zinc measurement reagent First reagent Preparation of reagent A3-3 A solution was prepared by changing only the pH of reagent A3-1 to 9.7 (20°C), and this was designated as reagent A3-2, the first reagent in the zinc measurement reagent. (13) Zinc measurement reagent First reagent Preparation of reagent A4-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent A4-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Brij-35 5% (Kishida Chemical) (14) Zinc measurement reagent First reagent Preparation of reagent A4-2 A solution was prepared by changing only the pH of reagent A4-1 to 9.5 (20°C), and this was designated as reagent A4-2, the first reagent for zinc measurement. (15) Zinc measurement reagent First reagent Preparation of reagent A4-3 A solution was prepared by changing only the pH of reagent A4-1 to 9.7 (20°C), and this was designated as reagent A4-3, the first reagent in the zinc measurement reagent. (16) Zinc measurement reagent First reagent Preparation of reagent B1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent B1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) BT-9 0.1% (Nikko Chemicals) (17) Zinc measurement reagent First reagent Preparation of reagent B1-2 A solution was prepared by changing only the pH of reagent B1-1 to 9.5 (20°C), and this was designated as reagent B1-2, the first reagent in the zinc measurement reagent. (18) Zinc measurement reagent First reagent Preparation of reagent B1-3 A solution was prepared by changing only the pH of reagent B1-1 to 9.7 (20°C), and this was designated as reagent B1-3, the first reagent in the zinc measurement reagent. (19) Zinc measurement reagent First reagent Preparation of reagent B2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent B2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) BT-9 0.5% (Nikko Chemicals) (20) Zinc measurement reagent First reagent Preparation of reagent B2-2 A solution was prepared by changing only the pH of reagent B2-1 to 9.5 (20°C), and this was designated as reagent B2-2, the first reagent in the zinc measurement reagent. (21) Zinc measurement reagent First reagent Preparation of reagent B2-3 A solution was prepared by changing only the pH of reagent B2-1 to 9.7 (20°C), and this was designated as reagent B2-3, the first reagent in the zinc measurement reagent. (22) Zinc measurement reagent First reagent Preparation of reagent B3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent B3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) BT-9 1% (Nikko Chemicals) (23) Zinc measurement reagent First reagent Preparation of reagent B3-2 A solution was prepared by changing only the pH of reagent B3-1 to 9.5 (20°C), and this was designated as reagent B3-2, the first reagent in the zinc measurement reagent. (24) Zinc measurement reagent First reagent Preparation of reagent B3-3 A solution was prepared by changing only the pH of reagent B3-1 to 9.7 (20°C), and this was designated as reagent B3-3, the first reagent in the zinc measurement reagent. (25) Zinc measurement reagent First reagent Preparation of reagent B4-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent B4-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) BT-9 5% (Nikko Chemicals) (26) Zinc measurement reagent First reagent Preparation of reagent B4-2 A solution was prepared by changing only the pH of reagent B4-1 to 9.5 (20°C), and this was designated as reagent B4-2, the first reagent in the zinc measurement reagent. (27) Zinc measurement reagent First reagent Preparation of reagent B4-3 A solution was prepared by changing only the pH of reagent B4-1 to 9.7 (20°C), and this was designated as reagent B4-3, the first reagent in the zinc measurement reagent. (28) Zinc measurement reagent First reagent Preparation of reagent C1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent C1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Em1108 0.1% (Kao) (29) Zinc measurement reagent First reagent Preparation of reagent C1-2 A solution was prepared by changing only the pH of reagent C1-1 to 9.5 (20°C), and this was designated as reagent C1-2, the first reagent in the zinc measurement reagent. (30) Zinc measurement reagent First reagent Preparation of reagent C1-3 A solution was prepared by changing only the pH of reagent C1-1 to 9.7 (20°C), and this was designated as reagent C1-3, the first reagent in the zinc measurement reagent. (31) Zinc measurement reagent First reagent Preparation of reagent C2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent C2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Em1108 0.5% (Kao) (32) Zinc measurement reagent First reagent Preparation of reagent C2-2 A solution was prepared by changing only the pH of reagent C2-1 to 9.5 (20°C), and this was designated as reagent C2-2, the first reagent in the zinc measurement reagent. (33) Zinc measurement reagent First reagent Preparation of reagent C2-3 A solution was prepared by changing only the pH of reagent C2-1 to 9.7 (20°C), and this was designated as reagent C2-3, the first reagent in the zinc measurement reagent. (34) Zinc measurement reagent First reagent Preparation of reagent C3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent C3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Em1108 1% (Kao) (35) Zinc measurement reagent First reagent Preparation of reagent C3-2 A solution was prepared by changing only the pH of reagent C3-1 to 9.5 (20°C), and this was designated as reagent C3-2, the first reagent in the zinc measurement reagent. (36) Zinc measurement reagent First reagent Preparation of reagent C3-3 A solution was prepared by changing only the pH of reagent C3-1 to 9.7 (20°C), and this was designated as reagent C3-3, the first reagent in the zinc measurement reagent. (37) Zinc measurement reagent First reagent Preparation of reagent C4-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent C4-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Em1108 5% (Kao) (38) Zinc measurement reagent First reagent Preparation of reagent C4-2 A solution was prepared by changing only the pH of reagent C4-1 to 9.5 (20°C), and this was designated as reagent C4-2, the first reagent in the zinc measurement reagent. (39) Zinc measurement reagent First reagent Preparation of reagent C4-3 A solution was prepared by changing only the pH of reagent C4-1 to 9.7 (20°C), and this was designated as reagent C4-3, the first reagent in the zinc measurement reagent. (40) Zinc measurement reagent First reagent Preparation of reagent D1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent D1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) KF-351A 0.1% (Shin-Etsu Silicone) (41) Zinc measurement reagent First reagent Preparation of reagent D1-2 A solution was prepared by changing only the pH of reagent D1-1 to 9.5 (20°C), and this was designated as reagent D1-2, the first reagent in the zinc measurement reagent. (42) Zinc measurement reagent First reagent Preparation of reagents D1-3 A solution was prepared by changing only the pH of reagent D1-1 to 9.7 (20°C), and this was designated as reagent D1-3, the first reagent in the zinc measurement reagent. (43) Zinc measurement reagent First reagent Preparation of reagent D2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent D2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) KF-351A 0.5% (Shin-Etsu Silicone) (44) Zinc measurement reagent First reagent Preparation of reagent D2-2 A solution was prepared by changing only the pH of reagent D2-1 to 9.5 (20°C), and this was designated as reagent D2-2, the first reagent in the zinc measurement reagent. (45) Zinc measurement reagent First reagent Preparation of reagent D2-3 A solution was prepared by changing only the pH of reagent D2-1 to 9.7 (20°C), and this was designated as reagent D2-3, the first reagent for zinc measurement. (46) Zinc measurement reagent First reagent Preparation of reagent D3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent D3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) KF-351A 5% (Shin-Etsu Silicone) (47) Zinc measurement reagent First reagent Preparation of reagent D3-2 A solution was prepared by changing only the pH of reagent D3-1 to 9.5 (20°C), and this was designated as reagent D3-2, the first reagent in the zinc measurement reagent. (48) Zinc measurement reagent First reagent Preparation of reagent D3-3 A solution was prepared by changing only the pH of reagent D3-1 to 9.7 (20°C), and this was designated as reagent D3-3, the first reagent in the zinc measurement reagent. (49) Zinc measurement reagent First reagent Preparation of reagent E1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent E1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) KF-355A 0.1% (Shin-Etsu Silicone) (50) Zinc measurement reagent First reagent Preparation of reagent E1-2 A solution was prepared by changing only the pH of reagent E1-1 to 9.5 (20°C), and this was designated as reagent E1-2, the first reagent for zinc measurement. (51) Zinc measurement reagent First reagent Preparation of reagent E1-3 A solution was prepared by changing only the pH of reagent E1-1 to 9.7 (20°C), and this was designated as reagent E1-3, the first reagent for zinc measurement. (52) Zinc measurement reagent First reagent Preparation of reagent E2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent E2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) KF-355A 0.5% (Shin-Etsu Silicone) (53) Zinc measurement reagent First reagent Preparation of reagent E2-2 A solution was prepared by changing only the pH of reagent E2-1 to 9.5 (20°C), and this was designated as reagent E2-2, the first reagent in the zinc measurement reagent. (54) Zinc measurement reagent First reagent Preparation of reagent E2-3 A solution was prepared by changing only the pH of reagent E2-1 to 9.7 (20°C), and this was designated as reagent E2-3, the first reagent for zinc measurement. (55) Zinc measurement reagent First reagent Preparation of reagent E3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent E3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) KF-355A 5% (Shin-Etsu Silicone) (56) Zinc measurement reagent First reagent Preparation of reagent E3-2 A solution was prepared by changing only the pH of reagent E3-1 to 9.5 (20°C), and this was designated as reagent E3-2, the first reagent for zinc measurement. (57) Zinc measurement reagent First reagent Preparation of reagent E3-3 A solution was prepared by changing only the pH of reagent E3-1 to 9.7 (20°C), and this was designated as reagent E3-3, the first reagent in the zinc measurement reagent. (58) Zinc measurement reagent First reagent Preparation of reagent F1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent F1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) CHAPS 0.1% (Fujifilm Wako Pure Chemical Industries) (59) Zinc measurement reagent First reagent Preparation of reagent F1-2 A solution was prepared by changing only the pH of reagent F1-1 to 9.5 (20°C), and this was designated as reagent F1-2, the first reagent for zinc measurement. (60) Zinc measurement reagent First reagent Preparation of reagent F1-3 A solution was prepared by changing only the pH of reagent F1-1 to 9.7 (20°C), and this was designated as reagent F1-3, the first reagent in the zinc measurement reagent. (61) Zinc measurement reagent First reagent Preparation of reagent F2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent F2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) CHAPS 0.5% (Fujifilm Wako Pure Chemical Industries) (62) Zinc measurement reagent First reagent Preparation of reagent F2-2 A solution was prepared by changing only the pH of reagent F2-1 to 9.5 (20°C), and this was designated as reagent F2-2, the first reagent in the zinc measurement reagent. (63) Zinc measurement reagent First reagent Preparation of reagent F2-3 A solution was prepared by changing only the pH of reagent F2-1 to 9.7 (20°C), and this was designated as reagent F2-3, the first reagent in the zinc measurement reagent. (64) Zinc measurement reagent First reagent Preparation of reagent F3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent F3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) CHAPS 1% (Fujifilm Wako Pure Chemical Industries) (65) Zinc measurement reagent First reagent Preparation of reagent F3-2 A solution was prepared by changing only the pH of reagent F3-1 to 9.5 (20°C), and this was designated as reagent F3-2, the first reagent in the zinc measurement reagent. (66) Zinc measurement reagent First reagent Preparation of reagent F3-3 A solution was prepared by changing only the pH of reagent F3-1 to 9.7 (20°C), and this was designated as reagent F3-3, the first reagent in the zinc measurement reagent. (67) Zinc measurement reagent First reagent Preparation of reagent F4-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent F4-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) CHAPS 5% (Fujifilm Wako Pure Chemical Industries) (68) Zinc measurement reagent First reagent Preparation of reagent F4-2 A solution was prepared by changing only the pH of reagent F4-1 to 9.5 (20°C), and this was designated as reagent F4-2, the first reagent in the zinc measurement reagent. (69) Zinc measurement reagent First reagent Preparation of reagent F4-3 A solution was prepared by changing only the pH of reagent F4-1 to 9.7 (20°C), and this was designated as reagent F4-3, the first reagent in the zinc measurement reagent. (70) Zinc measurement reagent First reagent Preparation of reagent G1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent G1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) AM301 0.1% (Nikko Chemicals) (71) Zinc measurement reagent First reagent Preparation of reagent G1-2 A solution was prepared by changing only the pH of reagent G1-1 to 9.5 (20°C), and this was designated as reagent G1-2, the first reagent for zinc measurement. (72) Zinc measurement reagent First reagent Preparation of reagent G1-3 A solution was prepared by changing only the pH of reagent G1-1 to 9.7 (20°C), and this was designated as reagent G1-3, the first reagent for zinc measurement. (73) Zinc measurement reagent First reagent Preparation of reagent G2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent G2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) AM301 0.5% (Nikko Chemicals) (74) Zinc measurement reagent First reagent Preparation of reagent G2-2 A solution was prepared by changing only the pH of reagent G2-1 to 9.5 (20°C), and this was designated as reagent G2-2, the first reagent in the zinc measurement reagent. (75) Zinc measurement reagent First reagent Preparation of reagent G2-3 A solution was prepared by changing only the pH of reagent G2-1 to 9.7 (20°C), and this was designated as reagent G2-3, the first reagent in the zinc measurement reagent. (76) Zinc measurement reagent First reagent Preparation of reagent G3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent G3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) AM301 1% (Nikko Chemicals) (77) Zinc measurement reagent First reagent Preparation of reagent G3-2 A solution was prepared by changing only the pH of reagent G3-1 to 9.5 (20°C), and this was designated as reagent G3-2, the first reagent for zinc measurement. (78) Zinc measurement reagent First reagent Preparation of reagent G3-3 A solution was prepared by changing only the pH of reagent G3-1 to 9.7 (20°C), and this was designated as reagent G3-3, the first reagent in the zinc measurement reagent. (79) Zinc measurement reagent First reagent Preparation of reagent G4-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent G4-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) AM301 5% (Nikko Chemicals) (80) Zinc measurement reagent First reagent Preparation of reagent G4-2 A solution was prepared by changing only the pH of reagent G4-1 to 9.5 (20°C), and this was designated as reagent G4-2, the first reagent for zinc measurement. (81) Zinc measurement reagent, first reagent, preparation of reagent G4-3 A solution was prepared by changing only the pH of reagent G4-1 to 9.7 (20°C), and this was designated as reagent G4-3, the first reagent in the zinc measurement reagent. (82) Zinc measurement reagent First reagent Preparation of reagent H1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent H1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Anchitol 20N 0.5% (Kao) (83) Zinc measurement reagent First reagent Preparation of reagent H1-2 A solution was prepared by changing only the pH of reagent H1-1 to 9.5 (20°C), and this was designated as reagent H1-2, the first reagent in the zinc measurement reagent. (84) Zinc measurement reagent First reagent Preparation of reagent H1-3 A solution was prepared by changing only the pH of reagent H1-1 to 9.7 (20°C), and this was designated as reagent H1-3, the first reagent in the zinc measurement reagent. (85) Zinc measurement reagent First reagent Preparation of reagent H2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent H2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Anchitol 20N 1% (Kao) (86) Zinc measurement reagent First reagent Preparation of reagent H2-2 A solution was prepared by changing only the pH of reagent H2-1 to 9.5 (20°C), and this was designated as reagent H2-2, the first reagent in the zinc measurement reagent. (87) Zinc measurement reagent First reagent Preparation of reagent H2-3 A solution was prepared by changing only the pH of reagent H2-1 to 9.7 (20°C), and this was designated as reagent H2-3, the first reagent in the zinc measurement reagent. (88) Zinc measurement reagent First reagent Preparation of reagent H3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent H3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Anchitol 20N 5% (Kao) (89) Zinc measurement reagent, First reagent, Preparation of reagent H3-2 A solution was prepared by changing only the pH of reagent H3-1 to 9.5 (20°C), and this was designated as reagent H3-2, the first reagent in the zinc measurement reagent. (90) Zinc measurement reagent First reagent Preparation of reagent H3-3 A solution was prepared by changing only the pH of reagent H3-1 to 9.7 (20°C), and this was designated as reagent H3-3, the first reagent in the zinc measurement reagent. (91) Zinc measurement reagent First reagent Preparation of reagent I1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent I1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) LMT 0.1% (Nikko Chemicals) (92) Zinc measurement reagent First reagent Preparation of reagents I1-2 A solution was prepared by changing only the pH of reagent I1-1 to 9.5 (20°C), and this was designated as reagent I1-2, the first reagent for zinc measurement. (93) Zinc measurement reagent First reagent Preparation of reagents I1-3 A solution was prepared by changing only the pH of reagent I1-1 to 9.7 (20°C), and this was designated as reagent I1-3, the first reagent in the zinc measurement reagent. (94) Zinc measurement reagent First reagent Preparation of reagent I2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent I2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) LMT 0.5% (Nikko Chemicals) (95) Zinc measurement reagent First reagent Preparation of reagent I2-2 A solution was prepared by changing only the pH of reagent I2-1 to 9.5 (20°C), and this was designated as reagent I2-2, the first reagent in the zinc measurement reagent. (96) Zinc measurement reagent First reagent Preparation of reagents I2-3 A solution was prepared by changing only the pH of reagent I2-1 to 9.7 (20°C), and this was designated as reagent I2-3, the first reagent in the zinc measurement reagent. (97) Zinc measurement reagent First reagent Preparation of reagent I3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent I3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) LMT 1% (Nikko Chemicals) (98) Zinc measurement reagent First reagent Preparation of reagent I3-2 A solution was prepared by changing only the pH of reagent I3-1 to 9.5 (20°C), and this was designated as reagent I3-2, the first reagent in the zinc measurement reagent. (99) Zinc measurement reagent First reagent Preparation of reagent I3-3 A solution was prepared by changing only the pH of reagent I3-1 to 9.7 (20°C), and this was designated as reagent I3-3, the first reagent in the zinc measurement reagent. (100) Zinc measurement reagent First reagent Preparation of reagent I4-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent I4-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) LMT 5% (Nikko Chemicals) (101) Zinc measurement reagent First reagent Preparation of reagent I4-2 A solution was prepared by changing only the pH of reagent I4-1 to 9.5 (20°C), and this was designated as reagent I4-2, the first reagent in the zinc measurement reagent. (102) Zinc measurement reagent First reagent Preparation of reagent I4-3 A solution was prepared by changing only the pH of reagent I4-1 to 9.7 (20°C), and this was designated as reagent I4-3, the first reagent in the zinc measurement reagent. (103) Zinc measurement reagent First reagent Preparation of reagent J1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent J1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) DSNa 0.5% (Nikko Chemicals) (104) Zinc measurement reagent First reagent Preparation of reagent J1-2 A solution was prepared by changing only the pH of reagent J1-1 to 9.5 (20°C), and this was designated as reagent J1-2, the first reagent for zinc measurement. (105) Zinc measurement reagent, First reagent, Preparation of reagent J1-3 A solution was prepared by changing only the pH of reagent J1-1 to 9.7 (20°C), and this was designated as reagent J1-3, the first reagent in the zinc measurement reagent. (106) Zinc measurement reagent First reagent Preparation of reagent J2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (at 20°C) to prepare reagent J2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) DSNa 1% (Nikko Chemicals) (107) Zinc measurement reagent, First reagent, Preparation of reagent J2-2 A solution was prepared by changing only the pH of reagent J2-1 to 9.5 (20°C), and this was designated as reagent J2-2, the first reagent in the zinc measurement reagent. (108) Zinc measurement reagent, First reagent, Preparation of reagent J2-3 A solution was prepared by changing only the pH of reagent J2-1 to 9.7 (20°C), and this was designated as reagent J2-3, the first reagent in the zinc measurement reagent. (109) Zinc measurement reagent, first reagent, preparation of reagent J3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent J3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) DSNa 5% (Nikko Chemicals) (110) Zinc measurement reagent, first reagent, preparation of reagent J3-2 A solution was prepared by changing only the pH of reagent J3-1 to 9.5 (20°C), and this was designated as reagent J3-2, the first reagent in the zinc measurement reagent. (111) Zinc measurement reagent, first reagent, preparation of reagent J3-3 A solution was prepared by changing only the pH of reagent J3-1 to 9.7 (20°C), and this was designated as reagent J3-3, the first reagent in the zinc measurement reagent. (112) Zinc measurement reagent, First reagent, Preparation of reagent K1-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent K1-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Sarcosinate LN-30 0.5% (Nikko Chemicals) (113) Zinc measurement reagent, first reagent, preparation of reagent K1-2 A solution was prepared by changing only the pH of reagent K1-1 to 9.5 (20°C), and this was designated as reagent K1-2, the first reagent in the zinc measurement reagent. (114) Zinc measurement reagent, First reagent, Preparation of reagent K1-3 A solution was prepared by changing only the pH of reagent K1-1 to 9.7 (20°C), and this was designated as reagent K1-3, the first reagent in the zinc measurement reagent. (115) Preparation of Zinc Measurement Reagent, First Reagent, Reagent K2-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent K2-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Sarcosinate LN-30 1% (Nikko Chemicals) (116) Zinc measurement reagent, First reagent, Preparation of reagent K2-2 A solution was prepared by changing only the pH of reagent K2-1 to 9.5 (20°C), and this was designated as reagent K2-2, the first reagent in the zinc measurement reagent. (117) Zinc measurement reagent, First reagent, Preparation of reagent K2-3 A solution was prepared by changing only the pH of reagent K2-1 to 9.7 (20°C), and this was designated as reagent K2-3, the first reagent in the zinc measurement reagent. (118) Preparation of Zinc Measurement Reagent, First Reagent, Reagent K3-1 The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent K3-1, the first reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) Bistrispropane 8.47g / L (MP Bio) Sarcosinate LN-30 5% (Nikko Chemicals) (119) Zinc measurement reagent, First reagent, Preparation of reagent K3-2 A solution was prepared by changing only the pH of reagent K3-1 to 9.5 (20°C), and this was designated as reagent K3-2, the first reagent in the zinc measurement reagent. (120) Zinc measurement reagent, first reagent, preparation of reagent K3-3 A solution was prepared by changing only the pH of reagent K3-1 to 9.7 (20°C), and this was designated as reagent K3-3, the first reagent in the zinc measurement reagent.

[0035] (121) Zinc measurement reagent, Second reagent, Preparation of Reagent A The following components were dissolved in pure water to the concentrations indicated, and the pH was adjusted to 9.1 (20°C) to prepare reagent A, the second reagent for zinc measurement. Sodium carbonate 21.20 g / L (Kanto Chemical) 5-Br-PAPS 0.0696g / L (Actec) (122) Zinc measurement reagent, Second reagent, Preparation of reagent B A solution was prepared by changing only the pH of reagent A to 9.5 (20°C), and this was designated as reagent B, the second reagent for zinc measurement. (123) Zinc measurement reagent Second reagent Preparation of reagent C A solution was prepared by changing only the pH of reagent A to 9.7 (at 20°C), and this was used as reagent C, the second reagent for zinc measurement.

[0036] 2. Measurement of the blank

[0037] Using reagent 1, purified water was used as the measurement sample and measurements were performed as follows to identify the blanks. These blanks are shown in Table 1. (Measurement method) A Hitachi High-Tech 7180 automated analyzer was used for the measurements. 10 μL of the sample was mixed with 150 μL of the first zinc measurement reagent (control X-1), and reacted at 37°C for 5 minutes. Then, 50 μL of the second zinc measurement reagent (reagent A) was added and reacted at 37°C for another 5 minutes. The difference between the absorbance at the primary wavelength of 546 nm and secondary wavelength of 700 nm at 4 minutes and 30 seconds (16th point) after the addition of the first reagent and the absorbance at the primary wavelength of 546 nm and secondary wavelength of 700 nm at 5 minutes and 8 seconds (34th point) after the addition of the second reagent was calculated, and this value multiplied by 10,000 was used as the blank. Similarly, for the first reagent of the zinc measurement reagent, reagents A1-1, A2-1, A3-1, A4-1, B1-1, B2-1, B3-1, B4-1, C1-1, C2-1, C3-1, C4-1, D1-1, D2-1, D3-1, E1-1, E2-1, E3-1, F1-1, F2-1, F3-1, F4-1, G1-1, G2-1, G3-1, G4-1, H1-1, H2-1, H3-1, I1-1, I2-1, I3-1, I4 -1, J1-1, J2-1, J3-1, K1-1, K2-1 or K3-1 combined with reagent A of the second reagent of the zinc measurement reagent, control X-2 of the first reagent of the zinc measurement reagent, reagents A1-2, A2-2, A3-2, A4-2, B1-2, B2-2, B3-2, B4-2, C1-2, C2-2, C3-2, C4-2, D1-2, D2-2, D3-2, E1-2, E2-2, E3-2, F1-2, F2-2, F3-2 F4-2, G1-2, G2-2, G3-2, G4-2, H1-2, H2-2, H3-2, I1-2, I2-2, I3-2, I4-2, J1-2, J2-2, J3-2, K1-2, K2-2 or K3-2 combined with reagent B of the second reagent of the zinc measurement reagent, control X-3 of the first reagent of the zinc measurement reagent, reagents A1-3, A2-3, A3-3, A4-3, B1-3, B2-3, B3-3, B4-3, C1-3, C Measurements were also performed using combinations of 2-3, C3-3, C4-3, D1-3, D2-3, D3-3, E1-3, E2-3, E3-3, F1-3, F2-3, F3-3, F4-3, G1-3, G2-3, G3-3, G4-3, H1-3, H2-3, H3-3, I1-3, I2-3, I3-3, I4-3, J1-3, J2-3, J3-3, K1-3, K2-3, or K3-3 with reagent H of the second reagent of the zinc measurement reagent.

[0038] [Table 1]

[0039] 3. Checking for blank changes

[0040] The blank change when the pH of control X-2 decreased was calculated by subtracting the blank measurement using control X-2 from the blank measurement using control X-1, and the blank change when the pH of control X-2 increased was calculated by subtracting the blank measurement using control X-2 from the blank measurement using control X-3. Similar calculations were performed for the combinations of reagents A1-1, A1-2, A1-3, A2-1, A2-2, A2-3, A3-1, A3-2, A3-3, A4-1, A4-2, A4-3, B1-1, B1-2, B1-3, B2-1, B2-2, B2-3, B3-1, B3-2, B3-3, B4-1, B4-2, B4-3, C1-1, C1-2, C1-3, and reagents. Combinations of reagents C2-1, C2-2, C2-3, combinations of reagents C3-1, C3-2, C3-3, combinations of reagents C4-1, C4-2, C4-3, combinations of reagents D1-1, D1-2, D1-3, combinations of reagents D2-1, D2-2, D2-3, combinations of reagents D3-1, D3-2, D3-3, combinations of reagents E1-1, E1-2, E1-3, combinations of reagents E2-1, E2-2, E2-3, combinations of reagents E3-1, E3-2, E3-3, combinations of reagents F1-1, F1-2 , combination of F1-3, combination of reagents F2-1, F2-2, F2-3, combination of reagents F3-1, F3-2, F3-3, combination of reagents F4-1, F4-2, F4-3, combination of reagents G1-1, G1-2, G1-3, combination of reagents G2-1, G2-2, G2-3, combination of reagents G3-1, G3-2, G3-3, combination of reagents G4-1, G4-2, G4-3, combination of reagents H1-1, H1-2, H1-3, combination of reagents H2-1, H2-2, H2-3 Combinations of reagents H3-1, H3-2, H3-3, I1-1, I1-2, I1-3, I2-1, I2-2, I2-3, I3-1, I3-2, I3-3, I4-1, I4-2, I4-3, J1-1, J1-2, J1-3, J2-1, J2-2, J2-3, J3-1, J3-2, J3-3, K1-1, K1-2, K1-3, K2-1,The tests were also performed using the combinations of reagents K2-2 and K2-3, and reagents K3-1, K3-2, and K3-3, respectively, resulting in reagents A1-2, A2-2, A3-2, A4-2, B1-2, B2-2, B3-2, B4-2, C1-2, C2-2, C3-2, C4-2, D1-2, D2-2, D3-2, E1-2, E2-2, E3-2, and F The blank change when the pH decreased and the blank change when the pH increased were defined for 1-2, F2-2, F3-2, F4-2, G1-2, G2-2, G3-2, G4-2, H1-2, H2-2, H3-2, I1-2, I2-2, I3-2, I4-2, J1-2, J2-2, J3-2, K1-2, K2-2, and K3-2. These values ​​are shown in Table 2.

[0041] [Table 2]

[0042] 4. Summary

[0043] Regarding the change in the blank, when the pH of control X-2 decreased, the absolute value was 363, but when the pH of reagents A1-2, A2-2, A3-2, A4-2, B1-2, B2-2, B3-2, B4-2, C1-2, C2-2, C3-2, C4-2, D1-2, D2-2, D3-2, E1-2, E2-2, E3-2, F1-2, F2-2, F3-2, F4-2, G1-2, G2-2, G3-2, G4-2, H1-2, H2-2, H3-2, I1-2, I2-2, I3-2, I4-2, J1-2, J2-2, J3-2, K1-2, K2-2, and K3-2 decreased, the absolute value was less than 363. Furthermore, when the pH of control X-2 increased, its absolute value was 288, but when the pH of reagents A1-2, A2-2, A3-2, A4-2, B1-2, B2-2, B3-2, B4-2, C1-2, C2-2, C3-2, C4-2, D1-2, D2-2, D3-2, E1-2, E2-2, E3-2, F1-2, F2-2, F3-2, F4-2, G1-2, G2-2, G3-2, G4-2, H1-2, H2-2, H3-2, I1-2, I2-2, I3-2, I4-2, J1-2, J2-2, J3-2, K1-2, K2-2, and K3-2 increased, their absolute values ​​were less than 288.

[0044] Regarding the blank change, reagents A1-2, A2-2, A3-2, A4-2, B1-2, B2-2, B3-2, B4-2, C1-2, C2-2, C3-2, C4-2, D1-2, D2-2, D3-2, E1-2, E2-2, E3-2, F1-2, F2-2, F3-2, F4-2, G1-2, G2-2, G3-2, G4-2, H1-2, H2-2, H3-2, I1-2, I2-2, I3-2, I4-2, J1-2, J2-2, J3-2, K1-2, K2-2, and K3-2, which contain surfactants, show that the blank change is suppressed by the inclusion of surfactants.

[0045] This confirms that surfactants have the effect of suppressing changes in the blank caused by pH changes.

[0046] [Example 2] (Measurement of sample in surfactant-containing reagent) A sample was measured using the zinc measurement reagent of the present invention.

[0047] 1. Preparation of reagents

[0048] (124) Zinc measurement reagent Preparation of the first reagent Similar to [Example 1], reagents A1-1, A1-2, A1-3, A2-1, A2-2, A2-3, A3-1, A3-2, A3-3, A4-1, A4-2, and A4-3 of the first reagent for zinc measurement were prepared.

[0049] (125) Zinc measurement reagent: Preparation of the second reagent Reagents A, B, and C of the second reagent for zinc measurement were prepared in the same manner as in [Example 1].

[0050] (126) Preparation of zinc measurement reagents We prepared the first and second reagents for Accuras Auto Zn (Sinotest).

[0051] 2. Measurement of the sample

[0052] Using reagent 1, a zinc control (Sinotest) with a zinc concentration of 100 μg / dL was used as the measurement sample, and the following measurements were performed to confirm the measured zinc concentration. These values ​​are shown in Table 3. (Measurement method) A Hitachi High-Tech 7180 automated analyzer was used for the measurement. 10 μL of the sample was mixed with 150 μL of reagent A1-1, the first reagent of the zinc measurement reagent, and reacted at 37°C for 5 minutes. Then, 50 μL of reagent I, the second reagent of the zinc measurement reagent, was mixed in and reacted at 37°C for another 5 minutes. The zinc concentration (μg / dL) of the sample was determined by proportionally calculating the difference between the absorbance at the primary wavelength of 546 nm and secondary wavelength of 700 nm at 4 minutes and 30 seconds (16th point) after the addition of the first reagent and the absorbance at the primary wavelength of 546 nm and secondary wavelength of 700 nm at 5 minutes and 8 seconds (34th point) after the addition of the second reagent, and the difference in absorbance when using a Zn standard solution (Sinotest) with a zinc concentration of 200 μg / dL instead of the sample. Similarly, measurements were also performed using the following combinations of reagents: reagent A2-1, A3-1, or A4-1 from the first reagent of the zinc measurement reagent and reagent I from the second reagent of the zinc measurement reagent; reagent A1-2, A2-2, A3-2, or A4-2 from the first reagent of the zinc measurement reagent and reagent II from the second reagent of the zinc measurement reagent; reagent A1-3, A2-3, A3-3, or A4-3 from the first reagent of the zinc measurement reagent and reagent III from the second reagent of the zinc measurement reagent; and Accuras Auto Zn.

[0053] [Table 3]

[0054] 3. Summary

[0055] Regarding the measured values, reagents A1-1, A1-2, A1-3, A2-1, A2-2, A2-3, A3-1, A3-2, A3-3, A4-1, A4-2, A4-3, and Accuras Auto Zn were all within the range of 98 μg / dL to 101 μg / dL.

[0056] Since the measured values ​​of reagents A1-1, A1-2, A1-3, A2-1, A2-2, A2-3, A3-1, A3-2, A3-3, A4-1, A4-2, and A4-3, which utilize the present invention, were almost the same as the measured values ​​of Accuras Auto Zn, it can be seen that the zinc measurement reagents utilizing the present invention accurately measure zinc concentration.

[0057] This confirms that surfactants have the effect of suppressing changes in the blank caused by pH changes, and that accurate measurements can be obtained even when surfactants are present. In zinc measurement reagents, the inclusion of surfactants has the effect of obtaining accurate measurements even in situations where the pH changes.

Claims

1. A reagent for measuring biological samples, characterized by containing a surfactant to suppress changes in the blank caused by pH changes.

2. The biological sample measurement reagent according to claim 1, wherein the surfactant is at least one of a nonionic surfactant, an amphoteric surfactant, or an anionic surfactant.

3. The biological sample measurement reagent according to claim 1, wherein the surfactant is at least one of the following: polyoxyethylene lauryl ether, polyoxyethylene alkyl ether, polyether-modified silicone oil, betaine-based surfactant, amine oxide-based surfactant, taurine-based surfactant, higher alcohol-based surfactant, and amino acid-based surfactant.

4. A biological sample measurement reagent according to any one of claims 1 to 3, wherein the biological sample measurement reagent is a biochemical test reagent.

5. A biological sample measurement reagent according to any one of claims 1 to 3, wherein the biological sample measurement reagent is a zinc measurement reagent.

6. A method for suppressing changes in the blank caused by pH changes, characterized by including a surfactant in the biological sample measurement reagent.

7. The suppression method according to claim 6, wherein the surfactant is at least one of the following: a nonionic surfactant, an amphoteric surfactant, or an anionic surfactant.

8. The suppression method according to claim 6, wherein the surfactant is at least one of the following: polyoxyethylene lauryl ether, polyoxyethylene alkyl ether, polyether-modified silicone oil, betaine-based surfactant, amine oxide-based surfactant, taurine-based surfactant, higher alcohol-based surfactant, and amino acid-based surfactant.

9. The suppression method according to any one of claims 6 to 8, wherein the biological sample measurement reagent is a biochemical test reagent.

10. The suppression method according to any one of claims 6 to 8, wherein the biological sample measurement reagent is a zinc measurement reagent.

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