Sample analysis method and sample analyzer

Abstract

To provide a sample analysis method and sample analyzer, capable of specifying a component included in a sample regardless of the concentration of the component included in the sample.SOLUTION: A sample analysis method, etc., comprises steps of: separating a plurality of components included in a sample by capillary electrophoresis; obtaining the electropherogram of the sample; acquiring the detection time of each peak and concentration corresponding to each peak from the electropherogram; and specifying a component included in the sample using parameters showing correlation between the detection time and concentration calculated using known components to the acquired detection time and the acquired concentration.SELECTED DRAWING: Figure 1

Description

[Technical field]

[0001] The present disclosure relates to a sample analysis method and a sample analysis device. [Background technology]

[0002] Hemoglobin (hereinafter, also referred to as "Hb") analysis is routinely performed in clinical testing. The Hb species to be analyzed varies depending on the purpose of the test. HbA1c is well known as an analytical target for diagnosing diabetes and understanding the condition. Mutant Hb such as HbS (sickle cell Hb), HbC, HbD, and HbE are used as analytical targets for diagnosing abnormal Hb diseases. In addition, HbA2 and HbF (fetal Hb) are widely used as analytical targets for diagnosing β-thalassemia.

[0003] Hb analysis is carried out by high performance liquid chromatography (HPLC) such as ion exchange chromatography, capillary electrophoresis (CE), or the like. Patent Document 1 describes a method for analyzing HbA2 by anion exchange electrokinetic chromatography using a solution containing a cationic polymer as a pseudostationary phase. Patent Documents 2 and 3 describe analytical methods for identifying components such as HbA1c by ion-exchange electrokinetic chromatography, utilizing the time point at which the components reach the interface. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] JP 2016-136135 A [Patent Document 2] Patent No. 6846324 [Patent Document 3] Patent No. 6871129 Summary of the Invention [Problem to be solved by the invention]

[0005] When the method described in Patent Document 1 is used, it is possible to reduce the size of the device and the measurement time. Furthermore, by combining it with the method described in Patent Document 2 or Patent Document 3, it is possible to identify the Hb species of each peak based on the relative detection time using a simple device. However, with these methods, peaks may overlap when the Hb concentration in the sample fluctuates. When peaks overlap, it is difficult to identify the Hb species.

[0006] An object of one embodiment of the present disclosure is to provide a sample analysis method and sample analysis apparatus that are capable of identifying a component contained in a sample regardless of the concentration of the component contained in the sample. [Means for solving the problem]

[0007] The present disclosure includes the following aspects. <1> Separating a plurality of components contained in the sample by capillary electrophoresis; obtaining an electropherogram of the sample; obtaining the detection time of each peak and the concentration corresponding to each peak from the electropherogram; and identifying a component contained in the sample by using a parameter indicating a correlation between the detection time and the concentration calculated using known components, for the acquired detection time and the acquired concentration. <2> The step of obtaining an electropherogram of the sample comprises: optically measuring the sample to obtain an absorbance; A step of acquiring the absorbance as an original waveform by plotting the absorbance on a two-dimensional plane along a time axis; and acquiring a time-differentiated waveform, which is a waveform obtained by differentiating the original waveform along the time axis; The step of acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram includes: acquiring a detection time of each peak in the time-differentiated waveform, and acquiring an absorbance corresponding to the detection time of each acquired peak, The step of identifying a component contained in the sample includes: and identifying a component contained in the sample by using a parameter indicating a correlation between the detection time and the absorbance calculated using known components, for the acquired detection time and the acquired absorbance. <1> The sample analysis method according to claim 1, <3> The parameter showing the correlation between the detection time and the concentration is A parameter obtained by plotting detection time versus absorbance on a two-dimensional plane using known components. <1> or <2> The sample analysis method according to claim 1, <4> The parameter showing the correlation between the detection time and the concentration is This is data on the slope of the approximate line for each component, obtained by plotting the detection time versus absorbance on a two-dimensional plane using known components. The step of identifying a component contained in the sample includes: correcting the acquired detection time by using the slope data to calculate a corrected detection time; and identifying a component contained in the sample based on the corrected detection time. <3> The sample analysis method according to claim 1, <5> The parameter showing the correlation between the detection time and the concentration is This is data on boundary lines separating each component, which is obtained by plotting the detection time and absorbance on a two-dimensional plane using known components, The step of identifying a component contained in the sample includes: using the data on the boundary line to identify a region to which the acquired detection time and the acquired absorbance belong in each region separated by the boundary line; Identifying a component contained in the sample based on the identified region. <3> The sample analysis method according to claim 1, <6> The component is hemoglobin. <1> ~ <5> 13. A sample analysis method according to any one of the preceding claims. <7> A means for separating a plurality of components contained in a sample by capillary electrophoresis; means for obtaining an electropherogram of said sample; A means for acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram; and means for identifying a component contained in the sample by using a parameter indicating a correlation between the detection time and the concentration calculated using known components, for the acquired detection time and the acquired concentration. Effect of the Invention

[0008] According to one embodiment of the present disclosure, there is provided a sample analysis method and a sample analysis apparatus that are capable of identifying a component contained in a sample regardless of the concentration of the component contained in the sample. [Brief description of the drawings]

[0009] [Figure 1] Fig. 1A is a top view showing one embodiment of a capillary electrophoresis chip, and Fig. 1B is a cross-sectional view of the electrophoresis chip shown in Fig. 1A. [Diagram 2] FIG. 2 is a diagram showing an example of an original waveform. [Diagram 3] FIG. 3 is a diagram showing a time-differentiated waveform of the original waveform of FIG. [Figure 4] FIG. 4 is a graph in which the horizontal axis represents the relative detection time PkTimeRt and the vertical axis represents the absorbance TlAbs. [Diagram 5] FIG. 5 is a diagram showing an example of the relationship between Hb species and corrected detection time. [Figure 6] FIG. 6 is a graph in which the horizontal axis represents the relative detection time PkTimeRt and the vertical axis represents the absorbance TlAbs. [Figure 7] FIG. 7 is a system schematic diagram showing a sample analysis system that can be used to carry out the sample analysis method of this embodiment. [Figure 8] FIG. 8 is a plan view showing an analysis chip used in the sample analysis system of FIG. [Figure 9] FIG. 9 is a cross-sectional view taken along line III-III in FIG. [Figure 10] FIG. 10 is a block diagram showing a hardware configuration of the control unit. [Figure 11] FIG. 11 is a diagram showing the relationship between Hb species and relative detection time. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0010] Hereinafter, an embodiment of the present invention will be described as an example. The description and examples are merely illustrative of the embodiment, and are not intended to limit the scope of the present invention. In the numerical ranges described in the present disclosure in stages, the upper or lower limit value described in one numerical range may be replaced with the upper or lower limit value of another numerical range described in stages. In addition, in the numerical ranges described in the present disclosure, the upper or lower limit value of the numerical range may be replaced with a value shown in the examples.

[0011] When referring to the amount of each component in a composition, if the composition contains multiple substances corresponding to each component, the amount refers to the total amount of those multiple substances present in the composition, unless otherwise specified. In the present disclosure, the term "process" refers not only to an independent process, but also to a process that cannot be clearly distinguished from other processes, as long as the intended purpose of the process is achieved. In the present disclosure, combinations of two or more preferred embodiments are more preferred embodiments. In the present disclosure, when an embodiment is described with reference to the drawings, the configuration of the embodiment is not limited to the configuration shown in the drawings. In addition, the size of the members in each drawing is conceptual, and the relative relationship between the sizes of the members is not limited to this.

[0012] [Sample analysis method] The sample analysis method according to the present disclosure includes a step of separating multiple components contained in a sample by capillary electrophoresis (hereinafter also referred to as a "separation step"), a step of obtaining an electropherogram of the sample (hereinafter also referred to as an "electropherogram obtaining step"), a step of obtaining the detection time of each peak and the concentration corresponding to each peak from the electropherogram (hereinafter also referred to as a "detection time / concentration obtaining step"), and a step of identifying a component contained in the sample using a parameter indicating the correlation between the detection time and concentration calculated using known components, relative to the obtained detection time and concentration (hereinafter also referred to as a "component identification step").

[0013] According to the sample analysis method of the present disclosure, it is possible to identify components contained in a sample regardless of the concentration of the components contained in the sample. For example, the genotype of a variant hemoglobin carrier is either a heterozygote for HbA / HbV (HbV is a variant hemoglobin other than HbA) or a homozygote for HbV / HbV. Compared to the blood of a heterozygote, the blood of a homozygote tends to have a higher Hb (HbV) concentration and an earlier peak. As the peak approaches earlier, multiple peaks may overlap.

[0014] Conventionally, when identifying the Hb species contained in a sample, it can be difficult to identify the Hb species if peaks overlap.

[0015] The present inventors have found that there is a correlation between the detection time of each peak obtained from an electropherogram and the concentration corresponding to each peak, and have also found that by using a parameter that indicates the correlation between the detection time and the concentration calculated using known components, it is possible to accurately identify components contained in a sample regardless of the concentration of each component.

[0016] <Separation process> The sample analysis method according to the present disclosure includes a step of separating a plurality of components contained in a sample by capillary electrophoresis.

[0017] The sample may be in any form, such as liquid, solid, or gas, so long as it contains a component that can be analyzed by capillary electrophoresis and is soluble in such a solvent.

[0018] Among the components contained in the sample, those to be analyzed include hemoglobin (Hb), albumin (Alb), globulin (α1, α2, β, γ globulin), fibrinogen, etc. Examples of the hemoglobin include a plurality of hemoglobin species such as HbA, HbA2, HbF (fetal hemoglobin), and variant hemoglobin (HbC, HbD, HbE, HbG, HbS, etc.). A small amount of HbA2 is contained in whole blood of both healthy individuals and variant hemoglobin carriers. These components are suitable for analysis by capillary electrophoresis because the mutation of the amino acids that are the components is easily electrically reflected in the protein molecules.

[0019] Depending on the components to be analyzed, the sample may be pretreated with an appropriate reagent or may be subjected to a preliminary separation process by a separate method (eg, chromatography).

[0020] Separation of Hb is usually carried out in an electrophoretic solution, which may be either an alkaline solution or an acidic solution, but is preferably an alkaline solution. The sample contains, for example, HbA2, HbE, HbA, HbF, HbC, HbD, HbG, HbS, etc.

[0021] Separation of Hb in a sample by capillary electrophoresis in an alkaline solution can be carried out by using an apparatus equipped with a capillary flow path. Specifically, a sample is introduced into a capillary channel filled with an alkaline solution, and after the sample is introduced, a voltage is applied to the entire or a part of the capillary channel. By applying the voltage, Hb in the sample can be electrophoresed and separated. A voltage can be applied to the capillary flow channel by contacting a negative electrode with the sample introduction side of the capillary flow channel and a positive electrode with the alkaline solution supply side.

[0022] The cross-sectional shape of the capillary channel is not particularly limited, and may be circular, rectangular, or another shape. In the case of a rectangular shape, the capillary channel preferably has a channel height and a channel width of 1 μm to 1000 μm, more preferably 10 μm to 200 μm, and further preferably 25 μm to 100 μm. In the case of a circular shape, the inner diameter of the capillary channel is preferably 10 μm or more or 25 μm or more, and preferably 100 μm or less or 75 μm or less. The length of the capillary channel is preferably 5 mm to 150 mm, more preferably 10 mm to 150 mm, and further preferably 20 mm to 60 mm.

[0023] The material of the capillary channel is not particularly limited, and examples thereof include glass, fused silica, plastic, etc. Examples of plastic include polymethyl methacrylate (PMMA), polycarbonate, polystyrene, polytetrafluoroethylene (PTFE), polyether ether ketone (PEEK), etc.

[0024] In the separation step, a capillary electrophoresis chip in which the capillary flow path is a microchip may be used. The capillary electrophoresis chip shown in Figures 1A and 1B described later and the analytical chip shown in Figures 8 and 9 described later are examples of chips used in capillary electrophoresis.

[0025] The capillary electrophoresis chip can have a sample reservoir, an electrophoretic buffer reservoir, and a capillary flow channel, and the sample reservoir and the electrophoretic buffer reservoir are in communication with each other through the capillary flow channel.

[0026] The size of the capillary electrophoresis chip is not particularly limited and is preferably adjusted appropriately. The size of the capillary electrophoresis chip can be, for example, 10 mm to 200 mm in length, 1 mm to 60 mm in width, and 0.3 mm to 5 mm in thickness.

[0027] The volumes of the sample tank and the electrophoretic buffer tank are determined appropriately according to the inner diameter and length of the capillary flow path. 3 ~1000mm 3 It is preferable that the thickness is 5 mm. 3 ~100mm 3 It is more preferable that: The amount of sample filled in the sample retention tank is not particularly limited, and can be 1 μL to 30 μL. The amount of alkaline solution filled in the electrophoresis buffer holding tank is not particularly limited, and can be 1 μL to 30 μL.

[0028] The voltage applied to both ends of the capillary channel is preferably 500V to 20,000V, more preferably 500V to 10,000V, and even more preferably 500V to 5,000V.

[0029] A liquid flow from the negative electrode side to the positive electrode side may be generated in the capillary flow channel. Examples of the liquid flow include electroosmotic flow.

[0030] The inner wall of the capillary channel is preferably coated with a cationic substance or an anionic substance. By coating the inner wall of the capillary channel with a cationic substance, the inner wall of the capillary channel can be positively charged, which makes it easy to generate an electroosmotic flow from the negative electrode side to the positive electrode side in the capillary channel. When the inner wall of the capillary channel is coated with an anionic substance, the inner wall of the capillary channel is negatively charged, but the cationic polymer contained in the alkaline solution binds to the negatively charged inner wall of the capillary channel, which then becomes positively charged, and an electroosmotic flow from the negative electrode side to the positive electrode side can be easily generated in the capillary channel in the same manner as described above.

[0031] The cationic substance is not particularly limited, and a silane coupling agent having a cationic functional group, etc., can be used. From the viewpoint of improving the separation accuracy, the cationic substance is preferably a polymer having a quaternary ammonium base, and more preferably polydiallyldimethylammonium chloride.

[0032] The anionic substance is not particularly limited, and examples of the anionic substance that can be used include polysaccharides having an anionic group and silane coupling agents having an anionic functional group. Examples of polysaccharides having an anionic group include sulfated polysaccharides, carboxylated polysaccharides, sulfonated polysaccharides, and phosphorylated polysaccharides. Examples of sulfated polysaccharides include chondroitin sulfate, heparin, heparan, fucoidan, salts thereof, etc. Examples of carboxylated polysaccharides include alginic acid, hyaluronic acid, salts thereof, etc.

[0033] 1A and 1B show an embodiment of a capillary electrophoresis chip, in which Fig. 1A is a top view showing the embodiment of the capillary electrophoresis chip, and Fig. 1B is a cross-sectional view of the electrophoresis chip shown in Fig. 1A. 1A and 1B includes a capillary flow channel 1, a sample reservoir 2, and an electrophoretic buffer reservoir 3, and the sample reservoir 2 and the electrophoretic buffer reservoir 3 are in communication with each other through the capillary flow channel 1. A detection unit 4 is formed in the capillary flow channel 1. The sample holding tank 2 and the electrophoretic running solution holding tank 3 may each have an electrode (not shown) for applying a voltage to both ends of the capillary channel 1. Specifically, the sample holding tank 2 (sample introduction side) may have a negative electrode, and the electrophoretic running solution holding tank 3 (alkaline solution supply side) may have a positive electrode.

[0034] The position of the detection unit 4, i.e., the length required for separation (the distance from the sample retention tank 2 to the detection unit 4, x in FIG. 1A) can be appropriately determined depending on the length of the capillary flow channel 1, etc. When the length of the capillary flow channel 1 (x+y in FIG. 1A) is 10 mm to 150 mm, the distance (x) from the sample retention tank 2 to the detection unit 4 is preferably 5 mm to 140 mm, more preferably 10 mm to 100 mm, and even more preferably 15 mm to 50 mm.

[0035] -Alkaline solution- In the present disclosure, "alkaline" means a pH greater than 7.0. The pH of the alkaline solution is preferably higher than the isoelectric point of Hb, and is preferably 7.5 to 12.0, more preferably 8.5 to 11.0, and even more preferably 9.5 to 10.5. In this disclosure, the pH of the alkaline solution is the pH of the alkaline solution at 25° C., and is measured using a pH meter 30 minutes after the electrode is immersed. The pH meter may be an F-72 manufactured by Horiba Ltd. or an equivalent device.

[0036] The alkaline solution contains a cationic polymer. The alkaline solution may contain two or more types of cationic polymers. In this disclosure, "cationic polymer" means a polymer having cationic groups. In the present disclosure, the term "cationic group" includes cationic groups and groups that become cationic groups when ionized. Examples of the cationic group include a primary amino group, a secondary amino group, a tertiary amino group, a quaternary ammonium base, and an imino group.

[0037] The cationic polymer is preferably water-soluble. In the present disclosure, "water-soluble" means that the target substance dissolves in water at 25°C at a concentration of 1% by mass or more.

[0038] Examples of cationic polymers having a primary to tertiary amino group or a cationic group that can be ionized to a primary to tertiary amino group include polyallylamine, polyvinylamine, polylysine, polyarginine, polyhistidine, polyornithine, polydiallylamine, polymethyldiallylamine, polyethyleneimine, diallylamine-acrylamide polymer, dimethylamine-ammonia-epichlorohydrin polymer, allylamine-diallylamine polymer, etc. The above-mentioned cationic polymers may be in the form of a salt, and examples thereof include hydrochloride salts.

[0039] Examples of cationic polymers having an imino group or a cationic group that can be ionized to an imino group include polyethyleneimine.

[0040] Examples of cationic polymers having a quaternary ammonium base or a cationic group that can be ionized to a quaternary ammonium base include polyquaternium, dimethylamine-ammonia-epichlorohydrin polymers, and dimethylamine-epichlorohydrin polymers.

[0041] In the present disclosure, the term "polyquaternium" refers to a cationic polymer containing a constituent unit derived from a monomer having a quaternary ammonium group. Polyquaterniums can be found in the INCI (International Nomenclature for Cosmetic Ingredients) directory. In one or more embodiments, examples of polyquaterniums include polydiallyldimethylammonium salts such as polyquaternium-6 (poly(diallyldimethylammonium chloride), polyquaternium-7 (copolymer of acrylamide and diallyldimethylammonium chloride), polyquaternium-4 (diallyldimethylammonium chloride-hydroxyethylcellulose copolymer), and polyquaternium-22 (copolymer of acrylic acid and diallyldimethylammonium chloride), and polyquaternium-2 (poly[bis(2-chloroethyl)ether-alt-1,3-bis[3-(dimethylamino)propyl]urea]).

[0042] In addition to the above-mentioned ammonium salts, in one or more embodiments, cationic polymers of onium salts such as phosphonium salts, oxonium salts, sulfonium salts, fluoronium salts, and chloronium salts can also be used as the cationic polymer. Examples of cationic polymers having a hydrazide group or a cationic group that can be ionized to a hydrazide group include aminopolyacrylamide.

[0043] From the viewpoint of shortening the separation time, the weight average molecular weight of the cationic polymer is preferably 10,000 to 500,000. In the present disclosure, the weight average molecular weight of the cationic polymer is determined by referring to a catalog value. If there is no catalog value for the weight average molecular weight, the weight average molecular weight is determined as the weight average molecular weight in terms of polyethylene glycol measured by gel permeation chromatography (GPC).

[0044] From the viewpoint of shortening the separation time, the content of the cationic polymer relative to the total mass of the alkaline solution is preferably 0.5% by mass to 10% by mass, more preferably 0.75% by mass to 5.0% by mass, and even more preferably 1.0% by mass to 3.0% by mass.

[0045] The cationic polymer to be used may be one synthesized by a conventionally known method, or a commercially available one.

[0046] The alkaline solution may contain water, such as distilled water, ion-exchanged water, pure water, and ultrapure water. The content of water relative to the total mass of the alkaline solution is not particularly limited, and can be 10% by mass to 99.9% by mass.

[0047] The alkaline solution may contain additives such as non-surfactant type zwitterionic substances, pH buffer substances, and preservatives for suppressing microbial growth, etc. Examples of preservatives include sodium azide, ethylparaben, and proclin.

[0048] -sample- The sum of the Hb content relative to the total mass of the sample is not particularly limited, and can be 0.001% by mass to 100% by mass. The form of the sample is not particularly limited, and may be a preparation of the sample raw material, or the sample raw material itself. Examples of sample raw materials include raw materials containing Hb, biological samples, and the like. Examples of biological samples include blood and blood-derived materials including red blood cell components. Examples of blood include blood collected from a living body, such as blood from mammals other than humans and human blood. Examples of blood-derived substances containing red blood cell components include those separated or prepared from blood and containing red blood cell components, such as blood cell fractions from which plasma has been removed, blood cell concentrates, freeze-dried products of blood or blood cells, hemolyzed samples obtained by hemolyzing whole blood, centrifuged blood, spontaneously precipitated blood, and washed blood cells.

[0049] From the viewpoint of improving separation accuracy, the sample preferably contains an alkaline solution containing a cationic polymer. The sample containing the alkaline solution can be obtained by diluting the sample raw material with the alkaline solution. The dilution ratio is preferably 1.2 to 100 times, more preferably 2 to 60 times, and even more preferably 3 to 50 times, based on mass. The material used for dilution is not particularly limited, and examples thereof include a pH adjuster (e.g., hydrochloric acid, etc.), a surfactant (e.g., Emulgen LS-110 (manufactured by Kao), etc.), a preservative (e.g., sodium azide, etc.), an ionic strength adjuster (e.g., sodium chloride, etc.), and a refractive index adjuster (e.g., sugars such as sucrose). Moreover, the alkaline solution may be the same as or different from the alkaline solution filled in the capillary flow path.

[0050] <Electropherogram acquisition process> The sample analysis method according to the present disclosure includes a step of obtaining an electropherogram of the sample. Specifically, the electropherogram obtaining step preferably includes a step of optically measuring the sample to obtain absorbance, a step of obtaining an original waveform by plotting the absorbance on a two-dimensional plane along a time axis, and a step of obtaining a time-differentiated waveform, which is a waveform obtained by differentiating the original waveform along the time axis. FIG. 2 is a diagram showing an example of an original waveform. FIG. 3 is a diagram showing a time-differentiated waveform of the original waveform of FIG.

[0051] (A step of optically measuring a sample to obtain absorbance) The absorbance is measured, for example, by the following method. The sample is continuously introduced into the capillary flow channel, and multiple components contained in the sample are separated in the capillary flow channel by applying a voltage. In the detection unit 4 provided in the middle of the capillary flow channel, light with a wavelength of 415 to 420 nm is irradiated, and the absorbance is measured by an absorbance measuring device.

[0052] For example, if the component to be analyzed is Hb, its molecular surface is negatively charged in an alkaline solution, so by placing a cathode upstream of the capillary flow path and an anode downstream, the component to be analyzed will migrate toward the anode. At that time, the electrical migration speed varies depending on the charge state of the molecular surface. In other words, the stronger the negative charge on the molecular surface, the higher the migration speed. Therefore, after a sample is introduced into the capillary flow path, components with a high migration speed will reach the detection unit more quickly.

[0053] Then, when the component with the slower migration speed reaches the detection section, the component with the faster migration speed, which is derived from the sample introduced later, also reaches the detection section. That is, in the capillary flow channel, when the component with the faster migration speed reaches the detection section first, the component continues to reach the detection section as long as the sample is continuously introduced into the capillary flow channel. Also, the component with the slower migration speed reaches the detection section later, but once it reaches the detection section, the component continues to reach the detection section as long as the sample is continuously introduced into the capillary flow channel.

[0054] In other words, the components with higher migration speeds arrive at the detection unit first, followed by the components with lower migration speeds, and the absorbance measured at the detection unit increases monotonically and cumulatively over time.

[0055] (A process of acquiring the original waveform by plotting absorbance on a two-dimensional plane along the time axis) The absorbance is plotted on a two-dimensional plane with one axis (e.g., Y-axis) and time on the other axis (e.g., X-axis) to obtain the original waveform. Note that it is sufficient for the absorbance to be obtained as data that can be plotted on a two-dimensional plane, and it is not essential that such data is actually drawn as a graph.

[0056] (A process for obtaining a time-differentiated waveform, which is a waveform obtained by differentiating the original waveform along the time axis) When the original waveform is viewed along the time axis, the portion with a large gradient is due to an increase in absorbance caused by a certain component arriving at the detection unit 4 for the first time. Meanwhile, the portion with a small gradient indicates that the next component has not yet arrived at the detection unit 4. In other words, the portion with a large gradient in the original waveform indicates that the component in the sample solution has arrived at the detection unit 4. Such a portion with a large gradient in the original waveform is represented as a peak waveform that appears in a waveform (called a "time-differentiated waveform") obtained by differentiating the original waveform along the time axis. These peak waveforms are derived from each component contained in the sample. Meanwhile, on the time axis, the component corresponding to the peak waveform further to the left indicates that the electrophoretic speed is higher than the component corresponding to the peak waveform further to the right.

[0057] <Detection time and concentration acquisition process, and component identification process> The sample analysis method according to the present disclosure includes a step of acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram. In addition, the sample analysis method according to the present disclosure includes a step of identifying components contained in the sample using a parameter indicating a correlation between the detection time and the concentration calculated using known components, relative to the acquired detection time and the acquired concentration.

[0058] The detection time vs. concentration acquisition step preferably includes a step of acquiring the detection time of each peak in the time-differentiated waveform, and acquiring the absorbance corresponding to the acquired detection time of each peak. It is preferable that the component identification process includes a process of identifying a component contained in the sample by using a parameter that indicates a correlation between the detection time and the absorbance calculated using known components, relative to the acquired detection time and the acquired absorbance.

[0059] It is preferable that a parameter (hereinafter also referred to as a "specific parameter") indicating the correlation between detection time and absorbance calculated using known components is preset in the sample analyzer prior to analyzing the sample to be analyzed.

[0060] The specific parameter is preferably a parameter obtained by plotting detection time versus absorbance on a two-dimensional plane using a known component.

[0061] A specific method for identifying components contained in a sample using the identification parameters will be described below.

[0062] (Method A) In method A, the specific parameter is data on the slope of an approximation line for each component, which is obtained by plotting the detection time versus absorbance on a two-dimensional plane using known components. In method A, the step of identifying the components contained in the sample includes the steps of correcting the acquired detection time using the slope data and calculating the corrected detection time, and identifying the components contained in the sample based on the corrected detection time.

[0063] The specific parameter in method A is calculated, for example, by the following method.

[0064] Absorbance is obtained by capillary electrophoresis using known Hb species with different concentrations. The absorbance is plotted on a two-dimensional plane along the time axis to obtain original waveform A, and time-differentiated waveform A is obtained by differentiating original waveform A along the time axis. The X-axis represents time and the Y-axis represents the amount of absorbance change.

[0065] The following data is obtained from the time differential waveform A. ·PkTime: The maximum value of the peak is PkTop, and this means the detection time at PkTop. · PkBtTime: The minimum value immediately before PkTop is PkBt, and this means the detection time at PkBt. · PkTlTime: The minimum value immediately after PkTop is PkTl, and this means the detection time at PkTl. ·EOFTime: The earliest detection time among the detection times at PkTop where the change in absorbance exceeds 0.2 mAbs / sec. ·PkArea: This refers to the integrated value of the change in absorbance from PkBtTime to PkTlTime at PkTop. ·Pk%: The percentage (%) of each PkArea in the total PkArea. A2Time: PkTime of HbA2. The PkTime at the peak immediately after EOFTime where the Pk% was 1% or more was defined as A2Time.

[0066] PkTime is converted to a relative detection time PkTimeRt based on the following formula A. PkTimeRt=(PkTime-EOFTime) / (A2Time-EOFTime) …Formula A

[0067] In the above formula A, the relative detection time is calculated using A2Time, but the detection time of HbA in blood or a protein added to the sample from the outside (a protein that absorbs light at a wavelength of 415 nm and behaves in the same way as Hb) may be used instead of A2Time.

[0068] Next, from the original waveform A, the absorbance TlAbs corresponding to PkTlTime is obtained. Instead of the absorbance TlAbs, the absorbance corresponding to PkTime may be acquired.

[0069] FIG. 4 is a graph in which the horizontal axis represents the relative detection time PkTimeRt and the vertical axis represents the absorbance TlAbs.

[0070] As shown in Figure 4, an approximate line is drawn for each Hb species, and the slope of each approximate line is calculated. Then, the average value of the slopes of the approximate lines is calculated.

[0071] The corrected detection time is calculated for each Hb type based on the following formula B. Corrected detection time = PkTimeRt + {TlAbs / (-Average value of the slope of the approximate line)} ... Formula B

[0072] FIG. 5 is a diagram showing an example of the relationship between Hb species and corrected detection time.

[0073] From FIG. 5, the Hb type and the range of the corrected detection time corresponding to the Hb type are determined. HbE: Corrected detection time is 1.010 or greater but less than 1.070. HbC: Corrected detection time is 1.070 or greater but less than 1.110. HbD: Corrected detection time is 1.110 or greater but less than 1.145. HbS: Corrected detection time is 1.145 or greater but less than 1.190. ·HbG: Corrected detection time is 1.190 or greater but less than 1.250. HbA: Corrected detection time is 1.250 or greater but less than 1.450.

[0074] As shown in Figure 5, the corrected detection times do not overlap depending on the Hb species. Therefore, for the sample to be analyzed, the relative detection time PkTimeRt and the absorbance TlAbs are obtained, the corrected detection time is calculated based on formula B, and the Hb species contained in the sample can be identified based on the corrected detection time.

[0075] The method for obtaining the relative detection time PkTimeRt and absorbance TlAbs for a sample to be analyzed (a sample containing an unknown Hb species) is the same as the method for obtaining the relative detection time PkTimeRt and absorbance TlAbs for known Hb species of different concentrations.

[0076] (Method B) In method B, the specific parameter is data on the boundary line separating each component, which is obtained by plotting the detection time versus absorbance on a two-dimensional plane using known components. In method B, the step of identifying the components contained in the sample includes the steps of: using boundary line data to identify, in each region separated by the boundary line, a region to which the acquired detection time and the acquired absorbance belong; and identifying the components contained in the sample based on the identified region.

[0077] The specific parameter in method B is calculated, for example, by the following method.

[0078] As in method A, the relative detection times PkTimeRt and absorbances TlAbs are obtained for known Hb species at different concentrations.

[0079] FIG. 6 is a graph in which the horizontal axis represents the relative detection time PkTimeRt and the vertical axis represents the absorbance TlAbs.

[0080] As shown in Figure 6, a boundary line for separating each Hb type is drawn, and a linear equation for the boundary line is calculated. Eline is the boundary line separating HbE and HbC. Cline is the boundary line separating HbC and HbD. The Dline is the boundary line separating HbD and HbS. The Sline is the boundary line separating HbS and HbG. The Gline is the boundary line that separates HbG and HbA.

[0081] The linear equation for each boundary line is set, for example, as follows: Eline: TlAbs=4583-4250×PkTimeRt Cline: TlAbs=4450-4000×PkTimeRt Dline: TlAbs=4550-3950×PkTimeRt Sline: TlAbs=4794-3967×PkTimeRt Gline: TlAbs=5079-3991×PkTimeRt

[0082] As shown in Figure 6, Hb species are separated by boundaries. Therefore, for the sample to be analyzed, the relative detection time PkTimeRt and absorbance TlAbs are obtained, and in each region separated by the boundary line, the region to which the obtained relative detection time and the obtained absorbance belong is identified. Based on the identified region, the Hb species contained in the sample can be identified.

[0083] When the following formulas E1 and E2 are satisfied, it is identified as HbE. PkTimeRt>1 …Formula E1 TlAbs<4583-4250×PkTimeRt…Formula E2

[0084] If the following formula C1 is satisfied, it is identified as HbC. 4583-4250×PkTimeRt≦TlAbs<4450-4000×PkTimeRt…Formula C1

[0085] If the following formula D1 is satisfied, it is identified as HbD. 4450-4000×PkTimeRt≦TlAbs<4550-3950×PkTimeRt…Formula D1

[0086] If the following formula S1 is satisfied, it is identified as HbS. 4550-3950×PkTimeRt≦TlAbs<4794-3967×PkTimeRt…Formula S1

[0087] If the following formula G1 is satisfied, it is identified as HbG. 4794-3967×PkTimeRt≦TlAbs<5079-3991×PkTimeRt…Formula G1

[0088] If the following formula A1 is satisfied, it is identified as HbA. 5079-3991×PkTimeRt≦TlAbs…Formula A1

[0089] [Sample analysis equipment] The sample analysis apparatus according to the present disclosure includes a means for separating a plurality of components contained in a sample by capillary electrophoresis, a means for obtaining an electropherogram of the sample, a means for acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram, and a means for identifying the components contained in the sample using a parameter indicating the correlation between the detection time and the concentration calculated using known components for the acquired detection time and concentration.

[0090] The means for obtaining an electropherogram of a sample preferably includes a means for optically measuring the sample to obtain absorbance, a means for obtaining an original waveform by plotting the absorbance on a two-dimensional plane along the time axis, and a means for obtaining a time-differentiated waveform, which is a waveform obtained by differentiating the original waveform along the time axis.

[0091] It is preferable that the means for acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram includes a means for acquiring the detection time of each peak in the time differential waveform, and acquiring the absorbance corresponding to the acquired detection time of each peak.

[0092] It is preferable that the means for identifying the components contained in the sample includes a step of identifying the components contained in the sample by using a parameter that indicates a correlation between the detection time and the absorbance calculated using known components, relative to the acquired detection time and the acquired absorbance. Specifically, the parameter indicating the correlation between detection time and concentration is data on the slope of an approximate straight line for each component, obtained by plotting detection time versus absorbance on a two-dimensional plane using known components, and the process of identifying the components contained in the sample preferably includes a means for correcting the obtained detection time using the slope data and calculating a corrected detection time, and a means for identifying the components contained in the sample based on the corrected detection time. Furthermore, the parameter indicating the correlation between detection time and concentration is boundary data separating each component, obtained by plotting the detection time and absorbance on a two-dimensional plane using known components, and it is preferable that the means for identifying the components contained in the sample includes a means for identifying, using the boundary data, an area to which the obtained detection time and the obtained absorbance belong in each area separated by the boundary line, and a means for identifying the components contained in the sample based on the identified area.

[0093] <Sample analysis system> 7 shows a schematic configuration of a sample analysis system A1 that can be used to perform the sample analysis method according to this embodiment. The sample analysis system A1 is configured to include a sample analyzer 100 and an analysis chip 10. The sample analysis system A1 is a system that performs an analysis method based on capillary electrophoresis on a sample Sa.

[0094] The analytical chip 10 holds the sample Sa and provides a place for analysis of the sample Sa when it is loaded into the sample analyzer 100. As shown in Figs. 8 and 9, the analytical chip 10 includes a main body 21, a mixing chamber 22, an introduction chamber 23, a filter 24, a discharge chamber 25, an electrode chamber 26, a flow path 27, and a connection flow path 28. Fig. 8 is a plan view of the analytical chip 10, and Fig. 9 is a cross-sectional view taken along line III-III in Fig. 8. The analytical chip 10 is not limited to a disposable type, and may be one used for multiple analyses. In addition, the sample analysis system of this embodiment is not limited to a configuration including a separate analytical chip 10 loaded into the sample analyzer 100, and may be configured such that a functional portion performing the same function as the analytical chip 10 is incorporated into the sample analyzer 100.

[0095] The main body 21 is the base of the analysis chip 10. The material of the main body 21 is as described above in the material of the capillary channel. In this embodiment, the main body 21 is configured such that the upper part 2A and the lower part 2B in FIG. 9 are formed separately and are joined together. However, this is not limiting, and for example, the main body 21 may be formed integrally.

[0096] The mixing chamber 22 is a place where the sample Sa and the diluent Ld are mixed. The mixing chamber 22 is configured as a recessed portion that opens upward, for example, by a through-hole formed in the upper portion 2A of the main body 21. The introduction chamber 23 is a chamber into which a sample solution obtained by mixing the sample Sa and the diluent Ld in the mixing chamber 22 is introduced. The introduction chamber 23 is configured as a recessed portion that opens upward, for example, by a through-hole formed in the upper portion 2A of the main body 21.

[0097] The filter 24 is provided at an opening of the introduction tank 23, which is an example of an introduction path to the introduction tank 23. The specific configuration of the filter 24 is not limited, and a suitable example is, for example, a cellulose acetate membrane filter (manufactured by ADVANTEC, pore size 0.45 μm).

[0098] The discharge tank 25 is a tank located downstream of the flow path 27. The discharge tank 25 is configured as a recessed portion that opens upward, for example, by a through-hole formed in the upper portion 2A of the main body 21. The electrode tank 26 is a tank into which the electrode 31 is inserted in capillary electrophoresis. The electrode tank 26 is configured as a recessed portion that opens upward, for example, by a through-hole formed in the upper portion 2A of the main body 21. The connection flow path 28 connects the introduction tank 23 and the electrode tank 26, and constitutes a conductive path between the introduction tank 23 and the electrode tank 26.

[0099] The flow channel 27 is formed as a capillary tube connecting the introduction tank 23 and the discharge tank 25, and is a place where electro-osmotic flow (EOF) occurs in electrophoresis. The flow channel 27 is configured as a groove formed in the lower part 2B of the main body 21, for example. In addition, the main body 21 may be appropriately formed with a recess or the like for promoting irradiation of light to the flow channel 27 and emission of light transmitted through the flow channel 27. The cross-sectional shape and size of the flow channel 27 (the flow channel height and flow channel width in the case of a rectangular shape, and the inner diameter and flow channel length in the case of a circular shape) are as described above in the cross-sectional shape and size of the capillary flow channel. The size of the entire analysis chip 10 is appropriately set according to the size of the flow channel 27 and the sizes and arrangements of the mixing tank 22, the introduction tank 23, the discharge tank 25, and the electrode tank 26.

[0100] The analysis chip 10 having the above configuration is merely an example, and any analysis chip capable of component analysis by capillary electrophoresis can be appropriately adopted.

[0101] The sample analyzer 100 performs an analysis process on the sample Sa in a state where an analytical chip 10 on which the sample Sa is deposited is loaded. The sample analyzer 100 includes an electrode 31, an electrode 32, a light source 41, an optical filter 42, a lens 43, a slit 44, a detector 45, a dispenser 6, a pump 61, a diluent tank 71, an electrophoresis liquid tank 72, and a control unit 8. The light source 41, the optical filter 42, the lens 43, and the detector 45 constitute the measurement unit 40 in this embodiment.

[0102] The electrodes 31 and 32 are for applying a predetermined voltage to the flow channel 27 in capillary electrophoresis. The electrode 31 is inserted into the electrode tank 26 of the analytical chip 10, and the electrode 32 is inserted into the discharge tank 25 of the analytical chip 10. The voltage applied to the electrodes 31 and 32 is as described above in the voltage applied to both ends of the capillary flow channel.

[0103] The light source 41 is a part that emits light for measuring absorbance as an optical measurement value in capillary electrophoresis. The light source 41 is equipped with, for example, an LED chip that emits light in a predetermined wavelength range. The optical filter 42 attenuates light of a predetermined wavelength from the light source 41 while transmitting light of the remaining wavelengths. The lens 43 is for collecting the light that has passed through the optical filter 42 to an analysis point in the flow path 27 of the analysis chip 10. The slit 44 is for removing excess light that may cause scattering, etc., from the light collected by the lens 43.

[0104] The detector 45 receives light from the light source 41 that has passed through the flow channel 27 of the analysis chip 10, and is configured with, for example, a photodiode or a photo IC.

[0105] In this way, the path along which the light emitted from the light source 41 reaches the detector 45 is the optical path. Then, at the measurement position 27A where the optical path intersects with the flow path 27, an optical measurement value is measured for the solution (i.e., either the sample solution or the electrophoretic running solution, or a mixture thereof) flowing through the flow path 27. That is, at the measurement position 27A in the flow path 27, the measurement unit 40 measures the optical measurement value of the sample solution. For example, the optical measurement value may be absorbance. The absorbance represents the degree to which the light in the optical path is absorbed by the solution flowing through the flow path 27, and represents the absolute value of the common logarithm of the ratio of the incident light intensity to the transmitted light intensity. In this case, a general-purpose spectrophotometer can be used as the detector 45. It should be noted that, even if the absorbance is not used, any optical measurement value, such as the value of the transmitted light intensity itself, can be used. In the following, an example will be described in which the absorbance is used as the optical measurement value.

[0106] The dispenser 6 dispenses desired amounts of diluent Ld, electrophoretic running liquid Lm, and sample solution, and includes, for example, a nozzle. The dispenser 6 can be freely moved to a plurality of predetermined positions within the sample analyzer 100 by a drive mechanism (not shown). The pump 61 is a suction source and a discharge source for the dispenser 6. The pump 61 may also be used as a suction source and a discharge source for ports (not shown) provided in the sample analyzer 100. These ports are used for filling the electrophoretic running liquid Lm, etc. A dedicated pump separate from the pump 61 may also be provided.

[0107] The diluent tank 71 is a tank for storing the diluent Ld. The diluent tank 71 may be a tank permanently installed in the sample analyzer 100, or a container in which a predetermined amount of diluent Ld is sealed and installed in the sample analyzer 100. The electrophoretic running liquid tank 72 is a tank for storing the electrophoretic running liquid Lm. The electrophoretic running liquid tank 72 may be a tank permanently installed in the sample analyzer 100, or a container in which a predetermined amount of electrophoretic running liquid Lm is sealed and installed in the sample analyzer 100.

[0108] The diluent Ld is mixed with the sample Sa to generate a sample solution. The main component of the diluent Ld is not particularly limited, and examples thereof include water and physiological saline, and preferred examples include a liquid having a similar composition to the electrophoretic liquid Lm described below. In addition to the main component, the diluent Ld may contain additives as necessary.

[0109] The electrophoretic liquid Lm is a medium that fills the discharge tank 25 and the flow path 27 in the analysis step by electrophoresis and generates an electroosmotic flow in the electrophoresis. The preferred embodiments of the electrophoretic liquid Lm are as described above. In addition, additives may be added to the electrophoretic liquid Lm as necessary, as described in the explanation of the diluent Ld.

[0110] The control unit 8 controls each component in the sample analyzer 100. As shown in the hardware configuration of Fig. 10, the control unit 8 has a CPU (Central Processing Unit) 81, a ROM (Read Only Memory) 82, a RAM (Random Access Memory) 83, and a storage 84. Each component is connected via a bus 89 so as to be able to communicate with each other.

[0111] The CPU 81 is a central processing unit that executes various programs and controls each part. That is, the CPU 81 reads a program from the ROM 82 or the storage 84, and executes the program using the RAM 83 as a working area. The CPU 81 controls each of the above components and performs various arithmetic processing according to the program recorded in the ROM 82 or the storage 84.

[0112] The ROM 82 stores various programs and various data. The RAM 83 temporarily stores programs or data as a working area. The storage 84 is configured with an HDD (Hard Disk Drive), SSD (Solid State Drive) or flash memory, and stores various programs including an operating system, and various data. In this embodiment, the ROM 82 or the storage 84 stores programs and various data for executing the sample analysis method according to this embodiment.

[0113] When executing the sample analysis method according to this embodiment, the sample analyzer 100 performs measurement and analysis using the above hardware resources and each of the configurations described above. EXAMPLES

[0114] Examples will be described below, but the present disclosure is not limited to these examples. In the following description, unless otherwise specified, all "parts" and "%" are based on mass.

[0115] <Separation devices and measuring instruments> As the separation device, a resin chip having a capillary flow path 1 with the structure shown in Fig. 1 was used (flow path width: 40 μm, flow path height: 40 μm, flow path length: 30 mm, distance (x) from the sample holding tank 2 to the detection unit 4: 20 mm). The capacities of the sample holding tank 2 and the electrophoresis buffer holding tank 3 were set to 10 μL. The inner wall of the capillary flow path was coated with polydiallyldimethylammonium chloride. The separation device used was the Lab 001A1c HD (manufactured by Arkray, Inc.), and the measuring device used was the Lab 001 (manufactured by Arkray, Inc.).

[0116] <Preparation of Capillary Electrophoresis Solution> The following substances were mixed, and sodium hydroxide and water were added until the pH reached 9.8 to prepare a capillary electrophoresis solution. · Dimethylamine-ammonia-epichlorohydrin polycondensate (cationic polymer, manufactured by Senka, Uniseance KHE 1000L, weight average molecular weight 100,000 - 500,000)…1.5 mass% · Sodium azide…0.02 mass% · Sodium hydroxide · Water

[0117] <Preparation of Dilution Solution> The following substances were mixed, and hydrochloric acid and water were added until the pH reached 8.8 to prepare a capillary dilution solution. · Polyethyleneimine (manufactured by Fujifilm Wako Pure Chemical Corporation, weight average molecular weight 70,000)…1 mass% · Emulgen LS-110 (manufactured by Kao Corporation)…0.4 mass% · Sodium azide…0.02 mass% · Sucrose…7.87 mass% · Sodium chloride…0.26 mass% · Hydrochloric acid · Water

[0118] <Preparation of Hb Specimen> HbA2 is contained in both healthy human whole blood and whole blood of various mutant Hb carriers. · Healthy human whole blood: AA (HbA / HbA) ·Various mutant Hb carrier whole blood: AE (HbA / HbE), AC (HbA / HbC), AD (HbA / HbD), AS (HbA / HbS), AG (HbA / HbG), CC (HbC / HbC), DD (HbD / HbD), SS (HbS / HbS)

[0119] [Example 1] (specific parameter settings) -Separation process- 9 μL of the capillary electrophoresis solution was supplied to the electrophoresis solution holding tank 3 in the capillary electrophoresis chip, and the capillary channel 1 was filled with the capillary electrophoresis solution by capillary action.

[0120] 9 μL of samples prepared by diluting healthy whole blood and whole blood of various mutant Hb carriers with the dilution solution 41-fold or 121-fold were supplied to the sample holding tank 2 . Next, a negative electrode was brought into contact with the sample holding tank 2, and a positive electrode was brought into contact with the electrophoresis buffer holding tank 3, and a voltage was applied under constant current control of 75 μA to start electrophoresis. Electrophoresis was carried out for 90 seconds.

[0121] -Electropherogram acquisition process- During the electrophoresis, the detection unit 4 was irradiated with light of 415 nm, the absorbance was measured, and the original waveform A of the absorbance spectrum was obtained. The original waveform A was differentiated along the time axis to obtain the time-differentiated waveform A. For light irradiation, absorbance measurement, and electropherogram acquisition, Arkray's Xalabo 001 was used. The above measurements were performed on whole blood from healthy subjects and whole blood from carriers of various variants of Hb, and electropherograms were obtained. The capillary electrophoresis chips were disposable after each measurement.

[0122] -Detection time and concentration acquisition process- From the time differential waveform A, PkTime, EOFTime, PkTlTime, and A2Time were obtained. PkTime was converted to relative detection time PkTimeRt based on the following formula A. PkTimeRt=(PkTime-EOFTime) / (A2Time-EOFTime) …Formula A In addition, the absorbance TlAbs corresponding to PkTlTime was obtained from the original waveform A.

[0123] In Figure 4, where the horizontal axis is the relative detection time PkTimeRt and the vertical axis is the absorbance TlAbs, an approximate line was drawn for each Hb species and the slope of the approximate line was calculated. The average slope of the approximate line was -4923. The corrected detection time was calculated based on the following formula B. Corrected detection time = PkTimeRt + {TlAbs / (-Average value of the slope of the approximate line)} ... Formula B

[0124] From FIG. 5, the Hb species and the range of the corrected detection time corresponding to the Hb species were determined. HbE: Corrected detection time is 1.010 or greater but less than 1.070. HbC: Corrected detection time is 1.070 or greater but less than 1.110. HbD: Corrected detection time is 1.110 or greater but less than 1.145. HbS: Corrected detection time is 1.145 or greater but less than 1.190. ·HbG: Corrected detection time is 1.190 or greater but less than 1.250. HbA: Corrected detection time is 1.250 or greater but less than 1.450.

[0125] (component identification process) The Hb species could be identified by determining whether the obtained corrected detection time fell within the above range of corrected detection times.

[0126] [Example 2] Separation was performed in the sample holding tank 2 in the same manner as in the case of using the whole blood of the healthy subject and the whole blood of each of the various mutant Hb carriers, and electropherograms were obtained. The relative detection time PkTimeRt and absorbance TlAbs were obtained from the electropherogram.

[0127] FIG. 6 is a graph in which the horizontal axis represents the relative detection time PkTimeRt and the vertical axis represents the absorbance TlAbs.

[0128] As shown in Figure 6, a boundary line separating each Hb type was drawn, and a linear equation for the boundary line was calculated.

[0129] The linear equations for each boundary line were set as follows: Eline: TlAbs=4583-4250×PkTimeRt Cline: TlAbs=4450-4000×PkTimeRt Dline: TlAbs=4550-3950×PkTimeRt Sline: TlAbs=4794-3967×PkTimeRt Gline: TlAbs=5079-3991×PkTimeRt

[0130] (component identification process) The region to which the obtained relative detection time PkTimeRt and absorbance TlAbs belonged was identified, and the Hb species could be identified based on the identified region.

[0131] [Comparative Example 1] As in Example 1, the relative detection time PkTimeRt was obtained using whole blood from healthy subjects and whole blood from various mutant Hb carriers. FIG. 11 is a diagram showing the relationship between Hb species and relative detection time.

[0132] 11, there are overlapping portions of the relative detection times PkTimeRt for each Hb species. In other words, it is clear that the components contained in a sample cannot be identified based on the relative detection times PkTimeRt alone. [Explanation of symbols]

[0133] 1: Capillary flow path 2: Sample holding tank 3: Running buffer tank 4: Detection section Tx:X peak detection time Ta2: HbA2 peak detection time Ty:Y peak detection time A1: Analysis system Sa: Sample Ld: Diluent Lm: running solution, 100:Component analysis device 10: Analysis chip 2A: Upper part 2B: Lower part 21: Main body 22: Mixing tank 23: Introduction tank 24: Filter 25: Discharge tank 26: Electrode bath 27: Flow path 27A: Measurement position 28: Connection channel 31: Electrode 32: Electrode 40: Measuring part 41:Light source 42: Optical filter 43: Lens 44: Slit 45: Detector 6: Dispenser 61: Pump 71: Dilution tank 72: Electrophoresis tank 8: Control unit 81: CPU 82:ROM 83:RAM 84: Storage 89: Bus

Claims

1. Separating a plurality of components contained in the sample by capillary electrophoresis; obtaining an electropherogram of the sample; obtaining the detection time of each peak and the concentration corresponding to each peak from the electropherogram; and identifying a component contained in the sample using a parameter indicating a correlation between the detection time and the concentration calculated using known components, for the acquired detection time and the acquired concentration.

2. The step of obtaining an electropherogram of the sample comprises: optically measuring the sample to obtain an absorbance; A step of acquiring the absorbance as an original waveform by plotting the absorbance on a two-dimensional plane along a time axis; and acquiring a time-differentiated waveform, which is a waveform obtained by differentiating the original waveform along the time axis; The step of acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram includes: acquiring a detection time of each peak in the time differential waveform, and acquiring an absorbance corresponding to the detection time of each acquired peak, The step of identifying a component contained in the sample includes:

2. The sample analysis method according to claim 1, further comprising a step of identifying a component contained in the sample by using a parameter indicating a correlation between the detection time and the absorbance calculated using known components, for the acquired detection time and the acquired absorbance.

3. The parameter showing the correlation between the detection time and the concentration is 2. The method of claim 1, wherein the parameter is obtained by plotting detection time versus absorbance on a two-dimensional plane using known components.

4. The parameter showing the correlation between the detection time and the concentration is This is data on the slope of an approximate line for each component, which is obtained by plotting the detection time versus absorbance on a two-dimensional plane using known components, The step of identifying a component contained in the sample includes: correcting the acquired detection time by using the slope data to calculate a corrected detection time; The sample analysis method according to claim 3 , further comprising the step of identifying a component contained in the sample based on the corrected detection time.

5. The parameter showing the correlation between the detection time and the concentration is This is data on boundary lines separating each component, which is obtained by plotting the detection time and absorbance on a two-dimensional plane using known components, The step of identifying a component contained in the sample includes: using the data on the boundary line to identify a region to which the acquired detection time and the acquired absorbance belong in each region separated by the boundary line; The sample analysis method according to claim 3 , further comprising the step of identifying components contained in the sample based on the identified region.

6. The sample analysis method according to claim 1 , wherein the component is hemoglobin.

7. A means for separating a plurality of components contained in a sample by capillary electrophoresis; means for obtaining an electropherogram of said sample; A means for acquiring the detection time of each peak and the concentration corresponding to each peak from the electropherogram; and means for identifying a component contained in the sample using a parameter indicating a correlation between the detection time and the concentration calculated using known components, for the acquired detection time and the acquired concentration.

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