Bilirubin standard solution for urine qualitative test, kit for urine qualitative test, method for urine qualitative test, and method for controlling accuracy of urine qualitative test paper

Abstract

Provided are: a bilirubin standard solution for a urine qualitative test, the bilirubin standard solution containing at least one substance selected from the group consisting of indirect bilirubin and ditaurobilirubin, a yellow pigment, and a solvent; a kit for a urine qualitative test, the kit containing the standard solution; and a method for a urine qualitative test and a method for controlling the accuracy of urine qualitative test paper, each method comprising using the standard solution.

Description

Bilirubin Standard Solution for Urine Qualitative Test, Kit for Urine Qualitative Test, Urine Qualitative Test Method, and Accuracy Management Method for Urine Qualitative Test Strip

[0001] The present disclosure relates to a bilirubin standard solution for urine qualitative test, a kit for urine qualitative test, a urine qualitative test method, and an accuracy management method for urine qualitative test strips.

[0002] As a method for detecting bilirubin in a biological sample, various methods are used, such as a method using the absorbance of a specimen and a method using a color reaction. Japanese Patent Laid-Open No. 02-238897 describes a method for measuring total bilirubin in which conjugated bilirubin in a specimen is deconjugated by an enzyme and the amount of unconjugated bilirubin is determined based on the change in absorbance.

[0003] Among them, as a method for examining bilirubin in urine, a method of detecting bilirubin by bringing a urine specimen into contact with a urine qualitative test strip that colors with bilirubin is known. As a method for determining the amount of bilirubin using a urine qualitative test strip, a method of visually comparing the color development with a color tone table and a method of measuring the reflectance by irradiating light on the test strip with an analyzer are known.

[0004] In an examination using a urine qualitative test strip, generally, a standard solution is used to evaluate the sensitivity and accuracy of the test strip. Bilirubin that appears in urine is mainly glucuronide-conjugated bilirubin (also called "direct bilirubin"), but glucuronide-conjugated bilirubin is not commercially available due to problems such as stability. Therefore, in a general standard solution, ditaurobilirubin, which is taurocholate-conjugated bilirubin and has excellent stability, water solubility, and easy handling, is widely used as a substitute for glucuronide-conjugated bilirubin.

[0005] In Analytical Chemistry Vol. 49, No.11, pp.901-905 (2000), the chemical properties of ditaurobilirubin used as a commercially available bilirubin standard substance are analyzed.

[0006] Since ditaurobilirubin is not a biological component, it may not necessarily reflect the reactivity of biological bilirubin. For example, in a urine qualitative test strip, ditaurobilirubin does not show the same color development as glucuronide-conjugated bilirubin.

[0007] When measuring reflectance using an analytical instrument, measurement is possible regardless of differences in color development. However, when judging the color development of test strips visually or through imaging, ditaurobilirubin is difficult to use as a standard solution. On the other hand, there is a need for the development of a standard solution that can be used not only for judgment methods using reflectance measurement with an analytical instrument, but also for judgment methods using color analysis.

[0008] In view of the above circumstances, this disclosure relates to a bilirubin standard solution for urine qualitative testing, a urine qualitative testing kit containing the standard solution, and a urine qualitative testing method using the standard solution, as well as a method for quality control of urine qualitative test strips, all applicable to both reflectance measurement and color analysis.

[0009] Means for solving the above problems include the following embodiments: <1> A bilirubin standard solution for urine qualitative testing, comprising at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin, a yellow pigment, and a solvent. <2> The bilirubin standard solution for urine qualitative testing according to <1>, wherein the yellow pigment is a yellow pigment that does not affect the reaction between bilirubin and a compound selected from (1) to (5) below: (1) 2-methyl-5-nitroaniline and sodium nitrite (2) 2,4-dichlorobenzenediazonium tetrafluoroborate (3) p-diazobenzenesulfonic acid (4) 2,6-dichlorobenzenediazonium tetrafluoroborate (5) 2,4-dichloroaniline and sodium nitrite <3> The bilirubin standard solution for urine qualitative testing according to <1> or <2>, wherein the yellow pigment is a yellow pigment having an absorption peak in the range of 400 to 480 nm. <4> A bilirubin standard solution for urine qualitative testing according to any one of <1> to <3>, wherein the reflectance of light at a wavelength of 420 nm is 60% or less of an aqueous solution of 20 mg / 100 mL of the yellow pigment and the reflectance of light at a wavelength of 560 nm is 80% or more of the yellow pigment. <5> A bilirubin standard solution for urine qualitative testing according to any one of <1> to <4>, wherein the yellow pigment comprises at least one selected from the group consisting of Acid Yellow 23, Acid Yellow 11, and Titanium Yellow. <6> A bilirubin standard solution for urine qualitative testing according to any one of <1> to <5>, wherein the amount of the yellow pigment is 50 to 350% by mass relative to the amount of the indirect bilirubin and the ditaurobilirubin. <7> A bilirubin standard solution for urine qualitative testing according to any one of <1> to <6>, wherein the concentrations of the indirect bilirubin and the ditaurobilirubin are 0.5 to 20 mg / dL. <8> A bilirubin standard solution for urine qualitative testing according to any one of <1> to <7>, used to bring into contact with a pad of urine qualitative test paper containing a compound that reacts with bilirubin. <9> A urine qualitative testing kit comprising a bilirubin standard solution for urine qualitative testing according to any one of <1> to <8> stored in a container, and urine qualitative test paper. <10> A urine qualitative testing kit according to <9>, wherein the urine qualitative test paper is a urine qualitative test paper having a pad containing a compound that reacts with bilirubin.<11> The urine qualitative test kit according to <10>, wherein the yellow dye contained in the bilirubin standard solution for urine qualitative testing does not inhibit the reaction between the compound that reacts with bilirubin and bilirubin. <12> A urine qualitative test method comprising: contacting the bilirubin standard solution for urine qualitative testing according to any one of <1> to <8> with the pad of a first urine qualitative test paper containing a compound that reacts with bilirubin; and contacting a urine sample with the pad of a second urine qualitative test paper containing a compound that reacts with bilirubin. <13> A method for quality control of a urine qualitative test paper, comprising contacting the bilirubin standard solution for urine qualitative testing according to any one of <1> to <8>, which contains at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin in a known concentration, with the pad containing a compound that reacts with bilirubin.

[0010] This disclosure provides a bilirubin standard solution for urine qualitative testing, a urine qualitative testing kit containing the standard solution, and a urine qualitative testing method and a method for quality control of urine qualitative test strips, all applicable to either reflectance measurement or color analysis.

[0011] This shows the reflectance spectrum of a urine test strip pad soaked in an indirect bilirubin solution, or a solution containing indirect bilirubin and Acid Yellow 23. This shows the reflectance spectrum of a urine test strip pad soaked in a ditaurobilirubin solution, or a solution containing ditaurobilirubin and Acid Yellow 23. This shows the spectrum of a real urine sample that tested positive for bilirubin. This shows the spectrum of a real urine sample that tested positive for bilirubin. This shows the spectrum of a real urine sample that tested positive for bilirubin.

[0012] The embodiments for carrying out the embodiments of this disclosure will be described in detail below. However, the embodiments of this disclosure are not limited to the embodiments described below. In the embodiments described below, the components (including elemental steps, etc.) are not essential unless otherwise specified. The same applies to numerical values ​​and their ranges, and do not limit the embodiments of this disclosure.

[0013] In this disclosure, numerical ranges indicated using "~" include the numbers before and after "~" as the minimum and maximum values, respectively. In numerical ranges described in stages in this disclosure, the upper or lower limit of one numerical range may be replaced by the upper or lower limit of another numerical range described in stages. Also, in numerical ranges described in this disclosure, the upper or lower limit of that range may be replaced by the values ​​shown in the examples. Even when an element is expressed in the singular form in this disclosure, unless otherwise explicitly stated, it does not preclude the existence of multiple elements unless it causes a technical inconsistency. In this disclosure, each component may contain multiple types of the corresponding substance. If multiple types of the substance corresponding to each component exist in a composition, the content or amount of each component means the total content or amount of the multiple types of substances present in the composition unless otherwise specified. In this disclosure, a combination of two or more preferred embodiments is a more preferred embodiment. In this disclosure, when simply referred to as "bilirubin," it includes various types of bilirubin, such as direct bilirubin, indirect bilirubin, and ditaurobilirubin, unless otherwise specified.

[0014] <Bilirubin Standard Solution for Qualitative Urine Testing> The bilirubin standard solution for qualitative urine testing according to this disclosure comprises at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin, a yellow dye, and a solvent. Bilirubin in urine is mainly direct bilirubin. In qualitative urine testing, when measuring bilirubin using the reflectance of a test strip, even if indirect bilirubin or ditaurobilirubin is used as the standard solution, there is no problem in measuring bilirubin in the urine sample as long as the reflectance at the measurement wavelength is roughly the same for the standard solution and the urine sample of the same concentration. On the other hand, in the case of color analysis of qualitative urine test strips, the color development differs between direct bilirubin and indirect bilirubin or ditaurobilirubin, making it difficult to use a standard solution containing indirect bilirubin or ditaurobilirubin as an indicator of bilirubin concentration in a urine sample. The inventors investigated the composition of standard solutions that can also be used for color analysis and found that a standard solution containing indirect bilirubin or ditaurobilirubin and a yellow dye exhibits a color reaction similar to that of bilirubin in urine samples and can be suitably used for color analysis. This is thought to be because the addition of the yellow dye reduces the reflectance in the low-wavelength range, especially around 420 nm, when the standard solution is brought into contact with a white pad, while the reflectance of light in the mid-to-long wavelength range remains unchanged, resulting in a color similar to that of direct bilirubin. Furthermore, since the yellow dye does not affect the reflectance of light in the mid-to-long wavelength range, the bilirubin concentration of the standard solution can also be used as an indicator of the bilirubin concentration in the urine sample by measuring the reflectance in the mid-to-long wavelength range (e.g., wavelength 560 nm). Therefore, the bilirubin standard solution for urine qualitative testing of this disclosure is applicable to both reflectance measurement and color analysis. Hereinafter, the bilirubin standard solution for urine qualitative testing described herein will also be referred to simply as "the standard solution described herein" or "the standard solution."

[0015] A urine qualitative test is a test that determines the presence or absence of a target substance in a urine sample. The presence or absence of the target substance is determined, for example, as positive or negative. In a urine qualitative test, the presence or absence of the target substance in the urine sample is typically confirmed by contacting the urine sample with a urine qualitative test strip equipped with a pad containing a compound that reacts with bilirubin, and observing the color development of the pad or measuring its reflectance. In the urine qualitative test described herein, in addition to determining the presence or absence of bilirubin in the urine sample, the bilirubin concentration may also be determined. The bilirubin concentration may be a reference value of bilirubin concentration, i.e., a semi-quantitative result. The results of the qualitative test may be expressed as intermittent indicators such as 1+, 2+, 3+, etc. The semi-quantitative result may be expressed, for example, as the bilirubin concentration (mg / dL) in the urine sample.

[0016] The result may be determined by visually observing or imaging the color of the pad, or by measuring the reflectance using an analytical instrument. When visually observing the color, the color of the urine sample and the color of the standard solution may be visually observed, and the presence or absence of bilirubin, and / or its concentration, may be determined by the similarity between the two. Alternatively, the color of the urine sample and the color chart may be visually observed, and the presence or absence of bilirubin, and / or its concentration, may be determined by the similarity between the two. When visually observing the color, the color of the urine sample and the color of the standard solution may be photographed, and the presence or absence of bilirubin, and / or its concentration, may be determined by the similarity between the two on the image. Alternatively, the color of the urine sample and the color chart may be photographed, and the presence or absence of bilirubin, and / or its concentration, may be determined by the similarity between the two on the image. Similarity may be determined, for example, by the similarity of hue. Hue can be calculated by the method described in the examples. In the case of determination by reflectance measurement, a urine qualitative analyzer (e.g., AUTION MAX AX-4060) may be used to irradiate the pad with light of a specific wavelength and measure its reflectance. From the viewpoint of minimizing the influence of yellow pigment, the wavelength of the irradiated light is preferably 500 to 650 nm, and more preferably 550 to 600 nm.

[0017] A standard solution is a solution containing a known concentration of the substance to be measured or a substitute thereof. The standard solutions of this disclosure contain, at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin, in a known concentration, as a substitute for bilirubin in urine. The standard solution may be used to determine the sensitivity and / or accuracy of a urine test strip. Alternatively, the standard solution may be used to determine the presence or absence and / or concentration of bilirubin in a urine sample by comparing the color or reflectance of a urine test strip with the standard solution with the color or reflectance of a urine test strip with a urine sample. Calibration curves may be created using standard solutions of various concentrations. The standard solution may be used for calibration of an analytical instrument.

[0018] (Indirect bilirubin and ditaurobilirubin) The standard solution contains at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin. That is, the standard solution may contain only indirect bilirubin or ditaurobilirubin, or both. From the viewpoint of ease of preparation, it is preferable that the standard solution contains only indirect bilirubin or ditaurobilirubin. Indirect bilirubin is unconjugated bilirubin and exists in the blood as bilirubin bound to albumin, but is not usually excreted in the urine. Indirect bilirubin can be synthesized and is also commercially available. Ditaurobilirubin is a synthesized taurine-conjugated bilirubin that is widely used as a substitute for bilirubin in the body. Ditaurobilirubin can be synthesized and is also commercially available.

[0019] The concentrations of indirect bilirubin and ditaurobilirubin in the standard solution (the concentration of one if one is present, or the total concentration if both are present) are preferably above the upper limit of the target detection concentration. For example, if the upper limit of the target detection concentration is 5 mg / dL, the concentrations of indirect bilirubin and ditaurobilirubin in the standard solution are preferably 5 mg / dL or higher. The concentrations of indirect bilirubin and ditaurobilirubin in the standard solution may be 0.5 mg / dL or higher, 1 mg / dL or higher, or 5 mg / dL or higher. The concentrations of indirect bilirubin and ditaurobilirubin in the standard solution may be 20 mg / dL or lower, 15 mg / dL or lower, or 10 mg / dL or lower. From this viewpoint, the concentrations of indirect bilirubin and ditaurobilirubin in the standard solution may be 0.5 to 20 mg / dL, 1 to 15 mg / dL, or 5 to 10 mg / dL. When using the standard solution, one concentration may be used, or a combination of multiple concentrations may be used. One or more concentrations of the standard solution may be diluted with a solvent at the time of use to adjust to the desired concentration.

[0020] (Yellow Pigment) The standard solution contains a yellow pigment. The yellow pigment is preferably one having an absorption peak at 400 to 480 nm, and more preferably one having an absorption peak at 400 to 450 nm. Note that the yellow pigment in this disclosure does not include bilirubin (including various types of bilirubin such as direct bilirubin, indirect bilirubin, and ditaurobilirubin).

[0021] Generally, urine test strips for bilirubin measurement consist of a pad containing a compound that reacts with bilirubin (typically a compound that azo-couples with bilirubin), and bilirubin is measured by observing the color reaction on this pad. Examples of such compounds include: (1) 2-methyl-5-nitroaniline and sodium nitrite (2) 2,4-dichlorobenzenediazonium tetrafluoroborate (3) p-diazobenzenesulfonic acid (4) 2,6-dichlorobenzenediazonium tetrafluoroborate (5) 2,4-dichloroaniline and sodium nitrite

[0022] The yellow pigment is preferably a yellow pigment that does not affect the color reaction on the urine qualitative test strip, for example, a yellow pigment that does not affect the reaction between the above compound and bilirubin, for example, a yellow pigment that does not react with the above compound. The fact that the yellow pigment does not affect the color reaction, for example, does not affect the reaction between the above compound and bilirubin, for example, does not react with the above compound, can be confirmed by the fact that the urine qualitative test strip does not show a positive result with the yellow pigment alone (for example, an aqueous solution of 20 mg / 100 mL of yellow pigment).

[0023] For example, (1) In a urine test strip containing 2-methyl-5-nitroaniline and sodium nitrite, a diazonium salt is produced by the reaction of 2-methyl-5-nitroaniline with sodium nitrite, and the resulting diazonium salt reacts with bilirubin to produce a reddish-brown azo dye through a diazo coupling reaction. When using such a urine test strip, it is preferable that the yellow dye is a yellow dye that does not affect the reaction between 2-methyl-5-nitroaniline and sodium nitrite, and the reaction between the diazonium salt and bilirubin. Similarly, (2) When using a urine test strip containing 2,4-dichlorobenzenediazonium tetrafluoroborate, it is preferable that the yellow dye is a yellow dye that does not affect the coupling reaction between 2,4-dichlorobenzenediazonium tetrafluoroborate and bilirubin. (3) When using a urine test strip containing p-diazobenzenesulfonic acid, it is preferable that the yellow dye is a yellow dye that does not affect the coupling reaction between p-diazobenzenesulfonic acid and bilirubin. (4) When using urine test strips containing 2,6-dichlorobenzenediazonium tetrafluoroborate, it is preferable that the yellow pigment is a yellow pigment that does not affect the reaction between 2,6-dichlorobenzenediazonium tetrafluoroborate and bilirubin. (5) When using urine test strips containing 2,4-dichloroaniline and sodium nitrite, it is preferable that the yellow pigment is a yellow pigment that does not affect the reaction between 2,4-dichloroaniline and sodium nitrite, and the reaction between the diazonium salt and bilirubin.

[0024] The reflectance of a 20 mg / 100 mL aqueous solution of yellow dye at a wavelength of 420 nm is preferably 60% or less, and more preferably 50% or less, from the viewpoint of suitably reducing the reflectance of the standard solution in the low-wavelength range. The reflectance may be 10% or more, or 20% or more. Therefore, the reflectance may be 10 to 60%, or 20 to 50%. In this disclosure, reflectance represents the reflectance at the measurement wavelength, with the reflectance at a wavelength of 760 nm being 100%. Here, the reflectance measurement is performed using a white pad so that the color of the pad does not affect the measurement result. As a white pad, a color correction pad from Arkray, Inc. (registered trademark) or a similar pad may be used.

[0025] The reflectance of a 20 mg / 100 mL aqueous solution of yellow dye at a wavelength of 560 nm is preferably 80% or more, and more preferably 90% or more. The reflectance may be 105% or less, or 100% or less. Therefore, the reflectance may be between 80% and 105%, or between 90% and 100%.

[0026] Examples of yellow pigments include water-soluble yellow pigments, with yellow dyes being preferred. The yellow pigment may be used alone or in combination of two or more. Examples of yellow dyes include C.I. Acid Yellow 2,3,4,5,6,7,8,9,9:1,10,11,11:1,12,13,14,15,16,17,17:1,18,20,21,22,23,25,26,27,29,30,31,33,34,36,38,39,40,40:1,41,42,42:1,43,44,46,48,51,53,55,56,60,63,65,66,67,68,69,72,7 Examples include 6, 82, 83, 84, 86, 87, 90, 94, 105, 115, 117, 122, 127, 131, 132, 136, 141, 142, 143, 144, 145, 146, 149, 153, 159, 166, 168, 169, 172, 174, 175, 178, 180, 183, 187, 188, 189, 190, 191, 192, 199, and Titanium Yellow. Among these, at least one selected from the group consisting of Acid Yellow 23, Acid Yellow 11, Riboflavin, and Titanium Yellow is preferred.

[0027] The amount of yellow pigment in the standard solution is preferably 50 to 350% by mass, and more preferably 100 to 300% by mass, relative to the amount of indirect bilirubin and ditaurobilirubin (the amount of one if one is included, or the total amount if both are included). If the standard solution contains indirect bilirubin, the amount of yellow pigment in the standard solution is preferably 50 to 200% by mass, more preferably 60 to 150% by mass, and even more preferably 70 to 130% by mass, relative to the amount of indirect bilirubin. If the standard solution contains ditaurobilirubin, the amount of yellow pigment in the standard solution is preferably 50 to 350% by mass, and more preferably 100 to 300% by mass, relative to the amount of ditaurobilirubin. When the amount of yellow pigment is within the above range relative to the amounts of indirect bilirubin and ditaurobilirubin, the hue of the urine qualitative test strip tends to be particularly similar to that when direct bilirubin is used.

[0028] (Solvent) Examples of solvents include aqueous solvents such as water, salt solutions, buffer solutions, and artificial urine solvents.

[0029] Examples of salt solutions include physiological saline, sodium carbonate aqueous solution, potassium chloride aqueous solution, potassium acetate aqueous solution, potassium sulfate aqueous solution, ammonium sulfate aqueous solution, ammonium chloride aqueous solution, and ammonium acetate aqueous solution. The salt concentration of the salt solution is preferably 5 to 500 mM at the final concentration.

[0030] Examples of buffer solutions include phosphate buffer, tris-HCl buffer, succinate buffer, glycylglycine buffer, MES (2-morpholinoethanesulfonic acid) buffer, HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) buffer, TES (N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid) buffer, MOPS (3-(N-monophorino)propanesulfonic acid) buffer, and PIPES (piperazine-1,4-bis(2-ethanesulfonic acid)) buffer. The concentration of the buffering agent in the buffer solution is preferably 5 to 500 mM at the final concentration.

[0031] Artificial urine solvents are solvents having a composition that mimics urine of biological origin, and may contain various components such as water, sodium chloride, urea, creatinine, uric acid, phosphates (sodium dihydrogen phosphate, potassium dihydrogen phosphate, etc.), sulfates (sodium sulfate, etc.), and ammonium salts (ammonium chloride, etc.). However, the artificial urine solvent used in the standard solution of this disclosure does not contain bilirubin.

[0032] The amount of solvent is preferably adjusted so that the indirect bilirubin and / or ditaurobilirubin concentrations reach the desired levels.

[0033] From the viewpoint of stably detecting bilirubin by color reaction, the pH of the solvent is preferably between 5 and 9.

[0034] (Other components) The standard solution may or may not contain components other than indirect bilirubin or ditaurobilirubin, yellow dye, and solvent. Other components include preservatives, chelating agents, buffering agents, etc. Examples of preservatives include sodium azide, sodium benzoate, and ethylparaben. Examples of chelating agents include EDTA (ethylenediaminetetraacetic acid).

[0035] From the viewpoint of reliably detecting bilirubin by color reaction, the pH of the standard solution is preferably between 5 and 9.

[0036] From the viewpoint of ensuring that the specific gravity of the standard solution is in a range similar to that of a urine sample derived from a living organism, it is preferably 1.00 to 1.10, and more preferably 1.00 to 1.05.

[0037] The standard solution of this disclosure is used for a urine qualitative test. Details of the urine qualitative test are as described above. In one embodiment, the standard solution is preferably used to contact a urine qualitative test strip that is used to contact a pad of urine qualitative test strips containing a compound that reacts with bilirubin.

[0038] <Urine Qualitative Test Kit> The urine qualitative test kit of this disclosure comprises the aforementioned standard solution of this disclosure, stored in a container, and a urine qualitative test strip.

[0039] There are no particular restrictions on the containers used to store the standard solution; examples include glass, polyethylene, polypropylene, fluororesin, and polyvinyl chloride containers.

[0040] A urine qualitative test strip typically comprises a pad containing a compound that reacts with bilirubin. Compounds that react with bilirubin include compounds that can undergo an azo coupling reaction with bilirubin, such as diazonium salts or compounds that are converted to diazonium salts by reaction. More specifically, the following compounds are included: (1) 2-methyl-5-nitroaniline and sodium nitrite (2) 2,4-dichlorobenzenediazonium tetrafluoroborate (3) p-diazobenzenesulfonic acid (4) 2,6-dichlorobenzenediazonium tetrafluoroborate (5) 2,4-dichloroaniline and sodium nitrite. Among these, (1) and (2) are preferred, and (1) is more preferred.

[0041] It is preferable that the yellow dye contained in the standard solution does not affect (e.g., does not inhibit) the reaction between the compound that reacts with bilirubin and bilirubin.

[0042] The bilirubin concentration measurable by a urine qualitative test strip may be, for example, in the range of 0.5 to 15 mg / dL, or in the range of 1 to 10 mg / dL.

[0043] A urine qualitative test strip may measure only bilirubin, or it may measure bilirubin and one or more other substances. Other substances that can be measured in urine include glucose, protein, urobilinogen, creatinine, occult blood, ketones, nitrite, and leukocytes. In addition to bilirubin and, if necessary, one or more other substances, a urine qualitative test strip may also measure the pH, specific gravity, and other properties of the urine. When measuring other substances and / or properties, a single urine qualitative test strip may have a pad for measuring bilirubin and pads for measuring the other substances and / or properties.

[0044] A urine qualitative test kit may include, in addition to standard solutions and urine qualitative test strips, instructions, a color chart, etc. The instructions may include instructions to perform the urine qualitative test according to any method described herein (in particular, the urine qualitative test method described herein).

[0045] <Urine qualitative test method> The urine qualitative test method of the present disclosure includes contacting the above-mentioned standard solution of the present disclosure with a pad of a first urine qualitative test paper containing a compound that reacts with bilirubin, and contacting a urine sample with a pad of a second urine qualitative test paper containing a compound that reacts with bilirubin. Details of the standard solution and the compound that reacts with bilirubin are as described above. The compound that reacts with bilirubin used in the first urine qualitative test paper and the second urine qualitative test paper is the same.

[0046] The first urine qualitative test paper is a test paper for observing the color development due to the reaction between indirect bilirubin and / or ditauro bilirubin in the standard solution and a compound that reacts with bilirubin. The second urine qualitative test paper is a test paper for observing the color development due to the reaction between bilirubin in the urine sample and a compound that reacts with bilirubin. By contacting the standard solution with the pad of the first urine qualitative test paper containing a compound that reacts with bilirubin, the performance of the urine qualitative test paper may be confirmed. Also, the measurement result may be determined by comparing the color development in the second urine qualitative test paper with the color development in the first urine qualitative test paper. Details of the urine qualitative test method are as described in the section "<Urine qualitative test bilirubin standard solution>".

[0047] <Accuracy management method for urine qualitative test paper> The accuracy management method for the urine qualitative test paper of the present disclosure includes contacting a standard solution of the present disclosure containing at least one selected from the group consisting of indirect bilirubin and ditauro bilirubin at a known concentration with a pad containing a compound that reacts with bilirubin. Details of the standard solution are as described above, and the standard solution contains at least one selected from the group consisting of indirect bilirubin and ditauro bilirubin at a known concentration. Details of the compound that reacts with bilirubin are as described above.

[0048] In the accuracy control method of the urine qualitative test strip of the present disclosure, for example, when a standard solution containing at least one selected from the group consisting of indirect bilirubin and ditauro bilirubin at a known concentration is brought into contact with the pad, accuracy control is performed by checking whether there is any abnormality in the color development. For example, when a bilirubin standard solution for urine qualitative test at 2 mg / dL is brought into contact with the pad, it is possible to determine whether there is an abnormality in the urine qualitative test strip based on whether the color development of a prescribed degree (for example, 2+) to be determined is exhibited. The determination of whether there is an abnormality in the urine qualitative test strip may be performed visually. Also, when using an analytical instrument, it is possible to determine whether there is an abnormality in the analytical instrument.

[0049] The concentration of the standard solution can be selected, for example, from the concentrations described above. Accuracy control may be performed using a plurality of types of standard solutions containing at least one selected from the group consisting of indirect bilirubin and ditauro bilirubin at different concentrations.

[0050] Next, embodiments of the present disclosure will be specifically described with reference to examples, but the embodiments of the present disclosure are not limited to these examples.

[0051] <Test Example 1> Using the following components, Standard Solutions 1 to 8 having the compositions shown in Table 1 were prepared. In Table 1, the numerical values indicate the final concentrations of the respective components, and "-" indicates that the component is not blended.

[0052] (Indirect bilirubin or ditauro bilirubin) - Indirect bilirubin (FUJIFILM Wako Pure Chemical Corporation) - Ditauro bilirubin (Funakoshi Corporation)

[0053] (Yellow pigment) - Acid Yellow 23 (Absorption peak wavelength: 420 nm; The reflectance of light with a wavelength of 420 nm when in contact with the pad for color tone correction of Ocean Sticks (registered trademark) is 36%, and the reflectance of light with a wavelength of 560 nm is 102%.)

[0054] (Solvent) - 100 mM aqueous sodium carbonate solution - Artificial urine solvent (pH 7.0)

[0055]

[0056] [Evaluation using urine qualitative test strips] A bilirubin measurement pad from a urine qualitative test strip (Autionsticks®, Arkray, Inc.) was immersed in each standard solution for 1 to 2 seconds. After removing the excess liquid, the color change was checked after approximately 1 minute and compared with the color chart. The color chart was created based on the bilirubin concentration of actual urine.

[0057] Visual inspection revealed that standard solutions 1-3, which do not contain yellow pigment, produced light pink, pink, and dark pink pads, respectively, which were clearly different from the color chart (light brown to dark brown). On the other hand, standard solutions 4-6, which contain yellow pigment, produced yellow, brown, and brown pads, respectively, which were similar to the color chart.

[0058] Furthermore, with standard solution 7, which does not contain yellow pigment, the pad turned purple, showing a color distinctly different from that of the color chart. On the other hand, with standard solution 8, the pad turned dark brown, confirming a color similar to 3+ to 4+ on the color chart.

[0059] From this, it can be seen that standard solutions 4-6 and 8, to which yellow dye was added to indirect bilirubin or ditaurobilirubin, showed a color similar to the color chart compared to standard solutions 1-3 and 7, to which no yellow dye was added. Therefore, it was found that adding yellow dye to indirect bilirubin or ditaurobilirubin makes it easier to accurately reflect the bilirubin concentration of urine of biological origin.

[0060] The reflectance spectra were measured using a reflectance photometric system with pads immersed in either a 20 mg / dL indirect bilirubin solution or a solution of 20 mg / dL indirect bilirubin solution with 20 mg / dL Acid Yellow 23 added (solvent: the above-mentioned sodium carbonate aqueous solution). The results are shown in Figure 1. Similarly, the reflectance spectra were measured using a reflectance photometric system with pads immersed in either a 20 mg / dL ditaurobilirubin solution or a solution of 20 mg / dL ditaurobilirubin solution with 20 mg / dL Acid Yellow 23 added (solvent: the above-mentioned artificial urine solvent). The results are shown in Figure 2. From the measurement results, it was found that the yellow dye compensates for absorption around 400-500 nm, and does not affect the reflectance around 560 nm.

[0061] On the other hand, Figures 3A-C show the spectra of three urine samples that tested positive for bilirubin. It can be seen that the reflectance of each solution to which the yellow dye was added is similar to the spectrum of the urine that tested positive for bilirubin. From this, it is thought that by adding the yellow dye to indirect bilirubin or ditaurobilirubin in the standard solution, a spectrum similar to that of bilirubin-positive urine is obtained, and this results in a color similar to that of bilirubin-positive urine being displayed on the urine qualitative test strip.

[0062] <Test Example 2> As shown in Tables 2 and 3, standard solutions 11 to 22 were prepared by changing the amount of yellow dye added to indirect bilirubin or ditaurobilirubin, and were brought into contact with the urine qualitative test paper pad in the same manner as in Test Example 1. The pad was photographed, and the RGB values ​​were calculated from the captured images to calculate the hue H. Generally, the RGB values ​​are data values ​​of 0 to 255 levels depending on the amount of light of the three primary colors. The hue H is calculated from the RGB values ​​as follows.

[0063]

[0064] Similarly, the hue of the color chart was calculated. The hues of standard solutions 11-13 and 17-19, which have an indirect bilirubin or ditaurobilirubin concentration of 5 mg / dL, were compared with the 2+ hue (31) in the color chart, and the hues of standard solutions 14-16 and 20-22, which have an indirect bilirubin or ditaurobilirubin concentration of 10 mg / dL, were compared with the 3+ hue (20) in the color chart, and the difference in hue of each standard solution was calculated.

[0065]

[0066]

[0067] The test results show that the hues of the other standard solutions containing yellow pigment are closer to the hues on the color chart compared to the hues of standard solutions 11, 14, 17, and 20, which do not contain yellow pigment. Furthermore, in the case of indirect bilirubin solutions, the hues are particularly close when 100% by mass of yellow pigment is added relative to the amount of indirect bilirubin. In the case of ditaurobilirubin solutions, the hues are particularly close when 100-300% by mass of yellow pigment is added relative to the amount of ditaurobilirubin.

[0068] The disclosure of Japanese Patent Application No. 2024-190062, filed on 29 October 2024, is incorporated herein by reference in its entirety. All documents, patent applications, and technical standards described herein are incorporated herein by reference to the same extent as if each individual document, patent application, and technical standard were specifically and individually noted to be incorporated by reference.

Claims

1. A bilirubin standard solution for urine qualitative testing, comprising at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin, a yellow dye, and a solvent.

2. The bilirubin standard solution for urine qualitative testing according to claim 1, wherein the yellow pigment is a yellow pigment that does not affect the reaction between bilirubin and a compound selected from (1) to (5) below: (1) 2-methyl-5-nitroaniline and sodium nitrite (2) 2,4-dichlorobenzenediazonium tetrafluoroborate (3) p-diazobenzenesulfonic acid (4) 2,6-dichlorobenzenediazonium tetrafluoroborate (5) 2,4-dichloroaniline and sodium nitrite 3. The bilirubin standard solution for urine qualitative testing according to claim 1, wherein the yellow pigment is a yellow pigment having an absorption peak in the range of 400 to 480 nm.

4. The bilirubin standard solution for urine qualitative testing according to claim 1, wherein the reflectance of light at a wavelength of 420 nm is 60% or less of the aqueous solution of the yellow dye at 20 mg / 100 mL, and the reflectance of light at a wavelength of 560 nm is 80% or more.

5. The bilirubin standard solution for urine qualitative testing according to claim 1, wherein the yellow dye comprises at least one selected from the group consisting of Acid Yellow 23, Acid Yellow 11, and Titanium Yellow.

6. The bilirubin standard solution for urine qualitative testing according to claim 1, wherein the amount of the yellow pigment is 50 to 350% by mass relative to the amounts of the indirect bilirubin and the ditaurobilirubin.

7. The bilirubin standard solution for urine qualitative testing according to claim 1, wherein the concentrations of the indirect bilirubin and the ditaurobilirubin are 0.5 to 20 mg / dL.

8. The bilirubin standard solution for urine qualitative testing according to claim 1, used to bring into contact with a pad of urine qualitative test paper containing a compound that reacts with bilirubin.

9. A urine qualitative test kit comprising a bilirubin standard solution for urine qualitative testing according to any one of claims 1 to 8, stored in a container, and a urine qualitative test strip.

10. The urine qualitative test kit according to claim 9, wherein the urine qualitative test paper comprises a pad containing a compound that reacts with bilirubin.

11. The urine qualitative test kit according to claim 10, wherein the yellow dye contained in the bilirubin standard solution for urine qualitative testing does not affect the reaction between the compound that reacts with bilirubin and bilirubin.

12. A method for performing a urine qualitative test, comprising: contacting a bilirubin standard solution for urine qualitative testing according to any one of claims 1 to 8 with a pad of a first urine qualitative test paper containing a compound that reacts with bilirubin; and contacting a urine sample with a pad of a second urine qualitative test paper containing the compound that reacts with bilirubin.

13. A method for quality control of a urine qualitative test strip, comprising contacting a pad containing a compound that reacts with bilirubin with a bilirubin standard solution for urine qualitative testing according to any one of claims 1 to 8, which contains at least one selected from the group consisting of indirect bilirubin and ditaurobilirubin in a known concentration.

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