Biological sample analysis device, biological sample analysis system, and state verification method for biological sample analysis device

Abstract

The purpose of the present disclosure is to provide a new technology for verifying the state of a device. The present disclosure provides a biological sample analysis device equipped with an information processing unit that performs information processing using signal intensity data of light generated by irradiating light onto a flow channel through which particles flow. In a verification process for verifying a device state, an information processing unit performs a data replacement step, and a data replacement step for replacing a data set of signal intensity data of light generated by irradiating light to a sample containing a plurality of types of particle swarms having gradually different fluorescence intensity levels with signal intensity data obtained by irradiating light to a flow channel through which particles do not flow, and signal intensity data of light generated by irradiating the particle swarm of any type with light.

Description

Technical Field

[0001] The present disclosure relates to a biological sample analysis device, a biological sample analysis system, and a method for verifying the state of a biological sample analysis device. More specifically, the present disclosure relates to a biological sample analysis device and a biological sample analysis system that perform analysis based on light generated by irradiating particles flowing through a flow channel with light, and a method for verifying the state of the biological sample analysis device. Background Art

[0002] For example, a group of particles such as cells, microorganisms, and liposomes are labeled with a fluorescent dye, and the intensity and / or pattern of fluorescence generated by the fluorescent dye excited by irradiating each particle of the group of particles with a laser is measured, thereby measuring the characteristics of the particles. Examples of particle analyzers that perform the measurement include a flow cytometer and a cell sorter.

[0003] A flow cytometer and a cell sorter can be configured to irradiate particles flowing in a line through a flow channel with a laser (excitation light) having a specific wavelength and detect fluorescence and / or scattered light emitted from each particle to analyze a plurality of particles one by one. Such a device can convert the light detected by a photodetector into an electrical signal, quantify the electrical signal, and perform statistical analysis to determine characteristics such as the type, size, structure, etc. of each particle.

[0004] From the viewpoint of maintaining data quality, quality control (QC) is important for such particle analyzers. For example, regarding quality control, Patent Document 1 below discloses an information processing device including an information processing unit that obtains a plurality of fluorescence intensities based on fluorescence signals from a sample containing a plurality of particles labeled with fluorescent dyes having different fluorescence intensities, identifies the intensity range of each of the detected plurality of fluorescence intensities based on the fluorescence intensity ratio of the sample, and calculates information regarding the sensitivity of the fluorescence detection unit.

[0005] Citation List

[0006] Patent Document

[0007] Patent Document 1: WO 2016 / 185755 A Summary of the Invention

[0008] Problems to be Solved by the Invention

[0009] In order to verify the state of a biological sample analysis device such as a flow cytometer or a cell sorting system, a device state verification sample such as eight-peak beads flows into the flow channel, and a light detection process is performed. For example, evaluation values such as linearity, molecule of equivalent soluble fluorescent dye (MESF), Q value, and B value are calculated based on the data obtained as a result of the light detection process.

[0010] The eight-peak beads include seven-level fluorescent beads and one type of non-fluorescent beads. Generally, MESF is an evaluation value based on the mean fluorescence intensity (MFI) of Dim1, and the components constituting these data mainly include the fluorescent components of non-fluorescent beads and the background of the device.

[0011] The background of the device is directly related to its performance, so it is considered necessary to verify this value. Here, non-fluorescent beads produce fluorescence, although the fluorescence produced is significantly weaker than that produced by fluorescent beads. In the case where the fluorescence level of non-fluorescent beads is high, it may be difficult to accurately evaluate the performance of the background of the device. In addition, when processing (such as sterilizing) a verification sample (such as eight-peak beads), the fluorescence level of non-fluorescent beads can change, which may affect the performance evaluation of the background of the device.

[0012] Therefore, an object of the present disclosure is to provide a new technique for verifying the state of a device. In particular, the object is to provide a technique for appropriately verifying the state of a device not only using a standard verification sample but also using a verification sample whose fluorescence level has changed as described above.

[0013] Solution to the problem

[0014] The present disclosure provides

[0015] A biological sample analysis device, comprising:

[0016] An information processing unit configured to perform information processing on light generated by irradiating a flow channel through which particles flow with light using signal intensity data, where

[0017] The information processing unit performs a data replacement step, in which, in a verification process for verifying the state of the device, the signal intensity data regarding light generated by irradiating any one type of particle group in a dataset of signal intensity data with light is replaced with signal intensity data obtained by irradiating a flow channel through which no particles flow with light. The signal intensity data is regarding light generated by irradiating a sample containing multiple types of particle groups, and the multiple types of particle groups have varying fluorescence intensity levels.

[0018] The information processing unit can use the dataset obtained after the data replacement process to calculate one or more evaluation values for verifying the state of the device.

[0019] One or more evaluation values may include at least any one of an accuracy evaluation value of the device, a linearity evaluation value of the device, or a sensitivity evaluation value of the device.

[0020] In the data replacement step, the signal intensity data of the particle group with the lowest signal intensity among the multiple types of particle groups can be replaced.

[0021] A sample containing multiple types of particle groups may contain one type of non-fluorescent particle group and one or more types of fluorescent particle groups, and

[0022] In the data replacement step, the signal intensity data of one type of non-fluorescent particle group can be replaced.

[0023] The biological sample analysis device can be configured to perform a trigger plate setting step of setting a trigger plate using a data set.

[0024] The biological sample analysis device can be configured to acquire signal intensity data regarding light generated by irradiating a flow channel through which no particles flow with light using a trigger plate.

[0025] The biological sample analysis device can be configured to perform a background data acquisition step on the light generated by irradiating a flow channel through which no particles flow with light: acquire signal intensity data.

[0026] In the background data acquisition step, the acquisition of signal intensity data can be performed in a state where the sample does not flow into the flow channel but only the sheath fluid flows into the flow channel.

[0027] The acquisition of signal intensity data in the background data acquisition step can be performed at a predetermined time interval.

[0028] The number of signal intensity data acquired in the background data acquisition step can be the same as the number set based on the number of event data in the data set and the number of peaks formed by the verification sample.

[0029] The number of signal intensity data acquired in the background data acquisition step can be the same as the number set based on the number of signal intensity data replaced in the data replacement step.

[0030] The biological sample analysis device can be configured to further perform a data separation step that separates the data set into data corresponding to each type of multiple types of particle groups.

[0031] The biological sample analysis device can be configured to perform data separation through clustering processing in the data separation step.

[0032] The biological sample analysis device can be configured to, in the data replacement step, replace the signal intensity data of the particle group with the lowest fluorescence level among multiple types of particle groups with the signal intensity data obtained by irradiating a flow channel through which no particles flow with light.

[0033] A sample containing multiple types of particle groups may contain one type of non-fluorescent particle group and one or more types of fluorescent particle groups, and

[0034] The biological sample analysis device may be configured to replace the signal intensity data of one type of non-fluorescent particle population with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light in the data replacement step.

[0035] The biological sample analysis device can be used as a flow cytometer.

[0036] This disclosure further provides

[0037] A biological sample analysis system, comprising:

[0038] An information processing unit configured to perform information processing on the light generated by irradiating the flow channel through which particles flow with light using the signal intensity data, wherein

[0039] The information processing unit performs a data replacement step, in which in the verification process of verifying the state of the device, the signal intensity data of the light generated by irradiating any one type of particle population in the data set of the signal intensity data of the light generated by irradiating the flow channel through which particles flow with light is replaced with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light. The signal intensity data is for the light generated by irradiating a sample containing multiple types of particle populations with different fluorescence intensity levels.

[0040] This disclosure also provides a method for verifying the state of a biological sample analysis device, the method comprising:

[0041] A data replacement step, in which a sample containing multiple types of particle populations with different fluorescence intensity levels is made to flow into the flow channel, and the signal intensity data of the light generated by irradiating any one type of particle population in the data set of the signal intensity data of the light generated by irradiating the particles flowing through the flow channel with light is replaced with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light. Description of the Drawings

[0042] Figure 1 Is an example of curve data for describing fluorescence level fluctuations.

[0043] Figure 2 Is a diagram showing an example of the configuration of the biological sample analysis device of the present disclosure.

[0044] Figure 3 Is an example of a flowchart of the verification process.

[0045] Figure 4 Is a diagram for describing the trigger piece.

[0046] Figure 5 Is a diagram for describing the difference in the data separation results using different fluorescence dye channels.

[0047] Figure 6 It is a diagram for describing clamping.

[0048] Figure 7 It is a table showing MFI and MEF.

[0049] Figure 8 It is an example of curve data for describing the fluctuation of fluorescence level.

[0050] Figure 9 It is a table showing the difference value and the difference rate of the evaluation value.

[0051] Figure 10 It is a diagram showing a configuration example of the biological sample analysis device of the present disclosure.

[0052] Figure 11 It is a diagram showing an example of a flowchart of a biological particle sorting process.

[0053] Figure 12 It is a schematic enlarged view of the particle sorting unit.

[0054] Figure 13 It is a diagram showing a schematic configuration example of the control unit. Detailed Description of the Invention

[0055] Hereinafter, preferred modes for implementing the present disclosure will be described. It should be noted that the embodiments described below show representative embodiments of the present disclosure, and the scope of the present disclosure is not limited to these embodiments only. It should be noted that the present disclosure will be described in the following order.

[0056] 1. First Embodiment (Biological Sample Analysis Device)

[0057] (1) Basic Concept

[0058] (2) Configuration Example

[0059] (3) Verification Process

[0060] (4) Examples

[0061] (5) Configuration Example of Biological Particle Sorting Device

[0062] 2. Second Embodiment (Biological Sample Analysis System)

[0063] 3. Third Embodiment (Verification Method of Biological Sample Analysis Device and Program for Executing Verification Method)

[0064] 1. First Embodiment (Biological Sample Analysis Device)

[0065] (1) Basic Concept

[0066] Examples of samples (hereinafter, also referred to as "device status verification samples" or "verification samples") for verifying the device status of a biological sample analysis device include samples containing multiple types of particle populations with varying fluorescence intensity levels. The eight-peak beads, as an example of a sample, contain seven types of fluorescent beads with different fluorescence levels and one type of non-fluorescent bead as described above.

[0067] As described above, when a treatment such as sterilization is performed on a verification sample (such as eight-peak beads), the fluorescence level of the beads (such as non-fluorescent beads) can change. Examples of how the fluorescence level changes will be described with reference to Figure 1 Figure A in the figure shows the plotted data of the signal intensity obtained by flow cytometry of non-sterilized eight-peak beads. Figure B of this figure shows the plotted data of the signal intensity obtained by flow cytometry of sterilized eight-peak beads. The horizontal axis of this plotted data represents the signal intensity, and the vertical axis represents the number of events.

[0068] Figure A in the drawings shows that the eight peaks are separated from each other. On the other hand, Figure B of the drawings shows fluctuations in the fluorescence level of the particle population with a lower fluorescence intensity level; in particular, the fluorescence intensity level of the particle population Dim1 with the lowest fluorescence intensity fluctuates significantly and is increasing. Such fluctuations in the fluorescence intensity level adversely affect evaluation values (such as MESF) used to evaluate the device status, for example. The particle population Dim1 in the drawings includes non-fluorescent beads, and even non-fluorescent beads may experience fluctuations in the fluorescence intensity level due to sterilization.

[0069] The biological sample analysis device according to the present disclosure performs a verification process for verifying its device status using a verification sample. In the verification process, signal intensity data regarding one type of particle population among multiple types of particle populations (signal intensity data belonging to the data set obtained as a result of the light detection process performed on the sample) is replaced with the signal intensity data on the background. For example, in the verification process using eight-peak beads, for example, the signal intensity data regarding non-fluorescent beads in the data set obtained as a result of the light detection process performed on the eight-peak beads is replaced with the signal intensity data on the background (the signal intensity data obtained by irradiating the flow channel through which no particles flow with light).

[0070] Using the data set obtained as a result of the replacement, the evaluation value of the device status becomes a value reflecting the background. Therefore, for example, even when the beads are sterilized, the device status can be appropriately evaluated without being affected by changes in the bead state.

[0071] In addition, this replacement can also be applied to the verification process using non-sterilized beads. That is, according to the present disclosure, the device status can be appropriately evaluated in both the case of using non-sterilized beads and the case of using sterilized beads.

[0072] In the present disclosure, for example, in order to use beads having a fluorescence level that has fluctuated due to sterilization or the like, data based only on the background can be applied to Dim1. Thus, an evaluation value of a device state such as MESF reflects the background to represent the performance of the device and is not affected by the state of the beads. Further, this technique can be applied to eight-peak beads having a non-fluctuating fluorescence level.

[0073] When the fluorescence level of non-fluorescent beads is high, it may not be possible to appropriately evaluate the device state. In such a case, MESF cannot accurately represent the device state. In particular, an increase in the level of non-fluorescent beads due to sterilization or the like tends to make the evaluation difficult. This is one of the reasons why the evaluation of the device state using sterilized beads has not been performed on an analyzer (such as a flow cytometer).

[0074] In one embodiment, a biological sample analysis device according to the present disclosure may include: an information processing unit configured to perform information processing on light generated by irradiating light on a flow channel through which particles flow using signal intensity data; wherein, the information processing unit may be configured to perform a data replacement step of replacing signal intensity data of light generated by irradiating light on any type of particle population in a data set of signal intensity data of light generated by irradiating light on a sample containing a plurality of types of particle populations, the plurality of types of particle populations having different fluorescence intensity levels, the sample having signal intensity data obtained by irradiation, in a verification process of verifying the state of the device, using light, a flow channel through which no particles flow. The data set subjected to replacement obtained as a result of the data replacement step is useful for appropriately verifying the device state.

[0075] For example, a biological sample analysis device may include: a light irradiation unit that irradiates light on a flow channel through which particles flow; a detection unit that detects light generated by the light irradiation; and an information processing unit that controls the light irradiation unit and the detection unit.

[0076] Hereinafter, first, a configuration example of a biological sample analysis device according to the present disclosure will be described, and then, a verification process performed by the device will be described.

[0077] (2) Configuration example

[0078] Figure 2 A configuration example of a biological sample analysis device of the present disclosure is shown. Figure 2The biological sample analysis device 6100 shown in the figure includes: a light irradiation unit 6101 that irradiates a biological sample S flowing through a flow channel C with light; a detection unit 6102 that detects the light generated by irradiating the biological sample S with light; and an information processing unit 6103 that processes information regarding the light detected by the detection unit. For example, the biological sample analysis device 6100 is a flow cytometer or an imaging cytometer. The biological sample analysis device 6100 may include a sorting unit 6104 that sorts specific biological particles P in the biological sample. The biological sample analysis device 6100 including the sorting unit is, for example, a cell sorter.

[0079] (Biological sample)

[0080] The biological sample S may be a liquid sample containing biological particles. The biological particles are, for example, cells or non - cell biological particles. The cells may be living cells, and more specific examples thereof include blood cells (such as red blood cells and white blood cells) and germ cells (such as semen and fertilized eggs). In addition, the cells may be those directly collected from a sample such as whole blood, or may be cultured cells obtained after culturing. For example, the non - cell biological particles are extracellular vesicles, or particularly, exosomes and microvesicles. The biological particles can be labeled with one or more labeling substances such as dyes (particularly, fluorescent dyes) and antibodies labeled with fluorescent dyes. It should be noted that particles other than biological particles can be analyzed by the biological sample analysis device of the present disclosure, and beads and the like can be analyzed for calibration and the like.

[0081] (Flow channel)

[0082] The flow channel C is designed such that a flow of the biological sample S is formed. Specifically, the flow channel C can be designed such that a flow in which the biological particles contained in the biological sample are aligned substantially in a line is formed. The flow channel structure including the flow channel C can be designed to form a laminar flow. Specifically, the flow channel structure is designed to form a laminar flow in which the flow of the biological sample (sample flow) is surrounded by the flow of the sheath fluid. The design of the flow channel structure can be appropriately selected by those skilled in the art, or a known design can be adopted. The flow channel C can be formed into a flow channel structure such as a microchip (a chip having a flow channel with a micron - scale) or a flow cell. The width of the flow channel C is 1 mm or less, or particularly, it can be not less than 10 μm and not more than 1 mm. The flow channel C and the flow channel structure including the flow channel C can be formed of a material such as plastic or glass.

[0083] The biological sample analysis device of the present disclosure is designed such that a biological sample flowing in the flow channel C, or specifically, biological particles in the biological sample, is irradiated with light from the light irradiation unit 6101. The biological sample analysis device of the present disclosure can be designed such that the irradiation point (probe point) of light on the biological sample is located in the flow channel structure in which the flow channel C is formed, or can be designed such that the irradiation point is located outside the flow channel structure. An example of the former case can be a configuration in which light is emitted onto the flow channel C in a microchip or a flow cell. An example of the latter case can be a configuration in which biological particles after leaving the flow channel structure (specifically, its nozzle portion) are irradiated with light, such as an air-jet flow cytometer.

[0084] (Light irradiation unit)

[0085] The light irradiation unit 6101 includes a light source unit that emits light and a light guiding optical system that guides the light to the irradiation point. The light source unit includes one or more light sources. The type of the light source is, for example, a laser light source or an LED. The wavelength of the light emitted from each light source can be any wavelength of ultraviolet light, visible light, and infrared light. For example, the light guiding optical system includes optical components such as a beam splitter, a mirror, or an optical fiber. The light guiding optical system may also include a lens group for condensing light and includes, for example, an objective lens. There may be one or more irradiation points where the biological sample and the light intersect. The light irradiation unit 6101 can be designed to collect light emitted from one light source or different light sources onto one irradiation point.

[0086] (Detection unit)

[0087] The detection unit 6102 includes at least one photodetector that detects light generated by emitting light onto the biological particles. For example, the light to be detected can be fluorescence or scattered light (such as one or more of the following: forward scattered light, backward scattered light, and lateral scattered light). For example, each photodetector includes one or more light receiving elements and has an array of light receiving elements. Each photodetector may include one or more photomultiplier tubes (PMTs) and / or photodiodes such as APDs and MPPCs as light receiving elements. The photodetector includes, for example, a PMT array in which a plurality of PMTs are arranged in a one-dimensional direction. The detection unit 6102 may also include an image sensor such as a CCD or a CMOS. Through the image sensor, the detection unit 6102 can acquire an image of the biological particles (for example, a bright field image, a dark field image, or a fluorescence image).

[0088] The detection unit 6102 includes a detection optical system that causes light of a predetermined detection wavelength to reach a corresponding photodetector. The detection optical system includes a spectroscopic unit such as a prism or a diffraction grating, or a wavelength separation unit such as a dichroic mirror or a filter. For example, the detection optical system is designed to disperse the light generated by light irradiation to biological particles, and detect the dispersed light using more photodetectors than the number of fluorescent dyes that label the biological particles. A flow cytometer including such a detection optical system is called a spectral flow cytometer. Further, for example, the detection optical system is designed to separate the light corresponding to the fluorescence wavelength band of a specific fluorescent dye from the light generated by light irradiation to biological particles, and cause the corresponding photodetector to detect the separated light.

[0089] The detection unit 6102 may further include a signal processing unit that converts the electrical signal obtained by the photodetector into a digital signal. The signal processing unit may include an A / D converter as a device that performs the conversion. The digital signal obtained by the conversion performed by the signal processing unit may be sent to the information processing unit 6103. The digital signal processing unit 6103 may process the digital signal as data related to light (hereinafter, also referred to as "light data"). For example, the light data may be light data including fluorescence data. More specifically, the light data may be data of light intensity, and the light intensity may be light intensity data of light including fluorescence (the light intensity data may include characteristic quantities such as area, height, and width).

[0090] (Information processing unit)

[0091] For example, the information processing unit 6103 includes a processing unit that performs processing of various types of data (e.g., light data) and a storage unit that stores various types of data. In the case where the processing unit acquires light data corresponding to a fluorescent dye from the detection unit 6102, the processing unit may perform fluorescence leakage correction (compensation processing) on the light intensity data. In the case of a spectral flow cytometer, the processing unit also performs fluorescence separation processing on the light data and acquires light intensity data corresponding to the fluorescent dye. For example, the fluorescence separation process may be performed by the unmixing method disclosed in JP 2011-232259 A. In the case where the detection unit 6102 includes an image sensor, the processing unit may acquire morphological information about the biological particles based on the image acquired by the image sensor. The storage unit may be designed to be able to store the acquired light data. The storage unit may be designed to be able to further store spectral reference data to be used in the unmixing process.

[0092] In the case where the biological sample analysis device 6100 includes the sorting unit 6104 described later, the information processing unit 6103 may determine whether to sort biological particles based on optical data and / or morphological information. Then, the information processing unit 6103 controls the sorting unit 6104 based on the determination result, and can sort biological particles by the sorting unit 6104.

[0093] The information processing unit 6103 can be designed to be capable of outputting various types of data (e.g., such as optical data and images). For example, the information processing unit 6103 can output various data generated based on optical data (e.g., such as two-dimensional curves or spectral curves). For example, the information processing unit 6103 can also be designed to be capable of accepting the input of various types of data, and accepting the gating process of the user for plotting. The information processing unit 6103 may include an output unit (e.g., such as a display) or an input unit (e.g., such as a keyboard) for performing output or input.

[0094] The information processing unit 6103 can be designed as a general-purpose computer, and can be designed as an information processing device including, for example, a CPU, a RAM, and a ROM. The information processing unit 6103 can be included in a housing including the light irradiation unit 6101 and the detection unit 6102, or can be located outside the housing. In addition, various processes or functions to be executed by the information processing unit 6103 can be implemented by a server computer or cloud connected via a network.

[0095] (Sorting unit)

[0096] The sorting unit 6104 sorts biological particles according to the determination result made by the information processing unit 6103. The sorting method can be a method of generating droplets containing biological particles by vibration, applying an electric charge to the droplets to be sorted, and controlling the moving direction of the droplets by an electrode. The sorting method can be a method for sorting by controlling the traveling direction of biological particles in a flow channel structure. For example, the flow channel structure has a control mechanism based on pressure (injection or suction) or charge. An example of the flow channel structure can be a chip having a flow channel structure in which the flow channel C branches into a recovery flow channel and a waste liquid flow channel on the downstream side, and specific biological particles are collected in the recovery flow channel (e.g., the chip disclosed in JP2020-76736A).

[0097] (3) Verification process

[0098] (3-1) Overview of the verification process

[0099] The biological sample analysis device according to the present disclosure can be configured to perform an analysis process on a biological sample based on light generated by irradiating particles flowing through a flow channel with light. For example, the biological sample analysis device can perform a verification process for verifying the device state before or during the analysis process. It should be noted that the biological sample analysis device can perform a verification process after the analysis process.

[0100] In the verification process, for example, a device state verification sample flows into the flow channel. Then, each particle contained in the verification sample is irradiated with light, and an evaluation value for evaluating the device state is calculated based on the light generated by the light irradiation. The verification sample can be a sample containing a plurality of types of particle populations having varying fluorescence intensity levels, and can be, for example, a sample that allows multiple peaks to be observed through a fluorescence separation process. The number of peaks is, for example, 2 to 10, particularly 3 to 10. Examples of the verification sample can include a bead population having multiple peaks, and more specifically, can include three-peak beads, four-peak beads, five-peak beads, six-peak beads, seven-peak beads, eight-peak beads, nine-peak beads, and ten-peak beads.

[0101] Any one of the plurality of types of particle populations contained in the verification sample can contain non-fluorescent particles. That is, the plurality of types of particle populations can also contain one type of non-fluorescent particle population and more than one type of fluorescent particle population. For example, the eight-peak beads contain one type of non-fluorescent beads and seven types of fluorescent beads. The present disclosure is particularly useful for performing a verification process using a verification sample containing a non-fluorescent particle population.

[0102] In the verification process, a data set of signal intensity data regarding light generated by irradiating a sample containing a plurality of types of particle populations having varying fluorescence intensity levels with light is acquired.

[0103] Then, the signal intensity data of the light generated by irradiating any one type of particle population with light in the data set is replaced with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light.

[0104] The data set subjected to the replacement is useful for evaluating the state of the biological sample analysis device. For example, the data set subjected to the replacement can be used to calculate an evaluation value of at least any one of the accuracy of the device, the linearity of the device, or the sensitivity of the device. The evaluation value calculated using the data set subjected to the replacement is useful for appropriately evaluating the device state, and is particularly useful from the perspective of quality control (QC).

[0105] Note that in this document, the data set not subjected to the replacement is also referred to as the "data set before replacement".

[0106] The data set subjected to the replacement is also referred to as the "data set after replacement".

[0107] In addition, the signal intensity data obtained by irradiating the flow channel through which no particles flow with light and used for replacement is also referred to as "background data".

[0108] That is, in the present disclosure, a pre-replacement data set and background data are obtained, and a post-replacement data set is generated from these data. Then, the post-replacement data set is used to calculate an evaluation value for evaluating the device state. The evaluation value calculated as described above is useful for appropriately evaluating the device state even when the verification sample is sterilized. Needless to say, this verification process is also applicable to the case where the verification sample is not sterilized.

[0109] In the case where the verification sample contains one type of non-fluorescent particle group and one or more types of fluorescent particle groups, the pre-replacement data set contains the signal intensity data of one type of non-fluorescent particle group and the signal intensity data of each of one or more types of fluorescent particle groups.

[0110] In this case, preferably, the signal intensity data of the non-fluorescent particle group in the pre-replacement data set is replaced with the background data. That is, by replacement, a post-replacement data set including the background data and the signal intensity data of one or more types of fluorescent particle groups is generated. The post-replacement data set thus generated is suitable for verifying the device state. In particular, since the fluorescence level of the non-fluorescent particle group is easily affected by variations caused by sterilization and the like, for appropriate verification processing, it is particularly preferable to replace the signal intensity data of the non-fluorescent particle group with the background data.

[0111] As described above, in the verification process, a pre-replacement data set is obtained. The pre-replacement data set can be a data set of the signal intensity of light (specifically, fluorescence) generated by flowing the verification sample through the flow channel and irradiating each particle contained in the verification sample with light. Each particle contained in the verification sample can be contained in the sample flow in the above-mentioned laminar flow. Then, as described above, the sample flow can be surrounded by the sheath liquid flow.

[0112] And, in the verification process, background data is obtained. The background data can be a data set of the signal intensity of light generated by irradiating the flow channel through which the test sample does not flow with light. In a state where only the sheath liquid flows through the flow channel without a sample (i.e., without sample liquid), the background data can be obtained as a result of the periodic light detection process.

[0113] As described above, the pre-replacement data set and the background data can be obtained.

[0114] (3-2) Examples of Verification Processing

[0115] Examples of the verification process will be described below with reference to Figure 3 The accompanying drawings are examples of the flowchart of the verification process.

[0116] As shown in the figure, the verification process may include a data set acquisition step S11, a trigger piece acquisition step S12, a background data acquisition step S13, a data separation step S14, a data replacement step S15, and an evaluation value calculation step S16. Each step will be described below.

[0117] (3-2-1) Data set acquisition step S11

[0118] In the data set acquisition step S11, the biological sample analysis device 6100 performs the following process: causing a verification sample to flow through a flow channel and acquiring signal intensity data of light generated by irradiating each particle flowing through the flow channel with light. That is, in the data set acquisition step S11, flow cytometry is performed on the verification sample.

[0119] For example, the light irradiation unit performs light irradiation, and the detection unit detects the light generated due to the light irradiation. The signal intensity data of the detected light is transmitted from the detection unit to the information processing unit.

[0120] For example, in the case where the verification sample is a population of beads having multiple peaks, signal intensity data of fluorescence generated by irradiating each bead included in the bead population with light is acquired. Examples of the bead population are as described above.

[0121] For example, the size and fluorescence intensity of the predetermined fluorescent beads are preferably uniform. The predetermined fluorescent beads may be beads capable of displaying fluorescence in a wavelength range of, for example, 400 nm to 800 nm.

[0122] The verification sample may be, for example, a sterilized verification sample. Sterilization may cause a change in the fluorescence intensity of the particles included in the verification sample. Even in the case of using a verification sample that has undergone a change in fluorescence intensity, an accurate evaluation value can be obtained by performing the verification process according to the present disclosure.

[0123] The number of events of the signal acquired in the data set acquisition step may be, for example, greater than or equal to 10,000 events, greater than or equal to 20,000 events, greater than or equal to 30,000 events, greater than or equal to 40,000 events, or greater than or equal to 50,000 events. The above number of events enables a more accurate evaluation of the device state.

[0124] In addition, from the viewpoint of improving the efficiency of the verification process, the number of events may be, for example, less than or equal to 1,000,000 events, less than or equal to 500,000 events, less than or equal to 300,000 events, or less than or equal to 100,000 events.

[0125] The number of events can be the number of all events obtained as a result of the optical detection process, or can be the number of singlet data among all events. Specifically, the number of events can be the number of segments of singlet data. That is, the data set can be a data set of singlet data.

[0126] In the data set acquisition step, the process of acquiring singlet data can be further performed. Techniques known in the art can be used to obtain singlet data, and for example, data processing can be performed by software (such as AutoGate).

[0127] In the data set acquisition step, the verification sample can flow into the flow channel as a sample liquid. The sample liquid of the verification sample can be surrounded by the sheath liquid, and specifically, a laminar flow in which the sample liquid is surrounded by the sheath liquid flows into the flow channel. The biological sample analysis device acquires signal intensity data by irradiating each particle contained in the flowing verification sample described above with light.

[0128] (3-2-2) Trigger slice setting step S12

[0129] In the trigger slice setting step S12, the biological sample analysis device uses the data set acquired in the data set acquisition step to set a trigger slice for background data to be described later. For example, the trigger slice setting can be performed by the information processing unit.

[0130] For example, as described above, the biological sample analysis device obtains singlet data from all events acquired in the data set acquisition step. The biological sample analysis device calculates the median of the trigger slice (TriggerSlice) from the singlet data. The median can be set as the trigger slice for the background data to be described later.

[0131] Reference will be made to Figure 4 the trigger slice. The drawings schematically show the change in the signal intensity of each event on a graph, where the signal intensity of the fluorescence detected by the photodetector is on the vertical axis and time is on the horizontal axis.

[0132] In particle analysis (such as flow cytometry), a photodetector can also detect signals that are not required for analysis. To exclude such signals, a threshold is set to determine whether a signal originates from a target particle. In the case where the signal intensity of the light generated from a specific particle is greater than or equal to a predetermined threshold, the particle is determined to be a target particle, and in the case where the signal intensity of the light generated from a specific particle is less than the predetermined threshold or greater, the particle is determined not to be a target particle. The signal intensity data regarding the particles determined to be target particles is used for analysis, and the signal intensity data regarding the particles determined not to be target particles is not used for analysis. The threshold can be preset in the light detector. Then, an event that makes the signal greater than or equal to the threshold (i.e., the signal intensity data regarding the target particle) is generated and acquired as the data to be analyzed.

[0133] As shown in the figure, the trigger slice is the time when the signal is greater than or equal to the threshold, that is, the time when the signal crosses the threshold.

[0134] In the trigger slice setting step, the trigger slice is obtained from each event included in the singlet data. Then, based on the trigger slices of each obtained event, the median of the trigger slices is calculated. The median calculated in this way is used in the background data acquisition step described later.

[0135] As described above, by setting the time called the trigger slice and acquiring the background data in the set trigger slices, the verification process can be performed more appropriately.

[0136] (3-2-3) Background data acquisition step S13

[0137] In the background data acquisition step S13, the biological sample analysis device acquires signal intensity data (i.e., background data) regarding the light generated by irradiating the flow channel through which the test sample does not flow with light.

[0138] To acquire background data, for example, light irradiation and detection of the light generated by the light irradiation are performed in a state where the verification sample does not flow into the flow channel and only the sheath fluid flows into the flow channel.

[0139] For example, the light irradiation unit performs light irradiation, and the detection unit detects the light generated by the light irradiation. The signal intensity data regarding the detected light is transmitted from the detection unit to the information processing unit.

[0140] The light irradiation for acquiring background data can be performed under the same conditions as the light irradiation performed in the dataset acquisition step S11. For example, the laser power of the light irradiation for acquiring background data can be the same as the light irradiation performed in the dataset acquisition step S11.

[0141] In order to detect the light generated by the light irradiation for obtaining background data, the trigger piece set in the trigger piece setting step S12 can be used. That is, the signal intensity detected within the time of the trigger segment can be used as one piece of event data.

[0142] The detection of the light generated by the light irradiation for obtaining background data can be performed using an automatic trigger. That is, the light detection can be performed at a predetermined time interval.

[0143] The number of pieces of signal intensity data constituting the obtained background data can preferably be greater than or equal to [(the number of pieces of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)], preferably greater than [(the number of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)], more preferably greater than or equal to [(the number of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)×1.1], still more preferably greater than or equal to [(the number of copies of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)×1.2], and still more preferably greater than or equal to [(the number of copies of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)×1.3].

[0144] The number of pieces of signal intensity data constituting the obtained background data can, for example, be less than or equal to [(the number of pieces of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)×2], less than or equal to [(the number of pieces of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)×1.8], or less than or equal to [(the number of pieces of event data obtained in the data set acquisition step) / (the number of peaks formed by the verification samples)×1.5].

[0145] The number of pieces of signal intensity data constituting the obtained background data can particularly correspond to the number of times of detection using an automatic trigger instrument.

[0146] For example, the number of particles forming each peak included in the verification sample (such as the eight-peak bead) is almost the same. Therefore, when the number of pieces of signal intensity data constituting the background data is within the above range, the background data becomes data corresponding to the data to be replaced in the data replacement step described later, which helps to appropriately perform the verification process.

[0147] As described above, the number of pieces of signal intensity data obtained in the background acquisition step can be set based on the number of pieces of event data obtained in the data set acquisition step and the number of peaks formed by the verification samples.

[0148] For example, assume that the number of events of data obtained using eight-peak beads in the dataset acquisition step S11 is 80,000. In this case, these eight-peak beads contain almost the same number of beads for each peak, so the number of events for each peak is 80,000 / 8 = 10,000. Considering that there are slight differences in the number of beads for each peak, for example, 13,333 signal intensity data with an increase of 1 / 3 of 10,000 can be obtained as background data.

[0149] Note that in the case where the number of background data is insufficient, the same data can be cycled and applied.

[0150] In addition, in the case where the number of signal intensity data to be replaced is specified in the dataset acquisition step S11 (or before performing the background data acquisition step S13), the number of signal intensity data on the background equal to (or greater than) the number of signal intensity data to be replaced can be obtained in the background data acquisition step.

[0151] In this case, a data separation step for each peak (a step similar to the data separation step S14 described later) can be performed in the dataset acquisition step S11, and the number of signal intensity data to be replaced can be specified in the data separation step.

[0152] For example, in the trigger chip setting step S12, in addition to acquiring singlet data, classification can also be performed into each particle group described in the data separation step S14 described later. Then, for example, the number of events of the particle group classified as having the lowest fluorescence intensity (e.g., non-fluorescent beads) can be obtained. Background data can be obtained in the same quantity as the number of events.

[0153] As described above, the number of signal intensity data obtained in the background acquisition step can be set based on the number of signal intensity data to be replaced by background data in the data replacement step described later.

[0154] In addition, the signal intensity data to be replaced can be obtained in the dataset acquisition step in the same amount as the signal intensity data obtained in the background data acquisition step.

[0155] For example, the number of signal intensity data obtained in the background data acquisition step can be preset. The dataset acquisition step and / or the data separation step can be performed such that the number of events classified into the particle group to be replaced is equal to the set number.

[0156] For example, in the case where the number of signal intensity data obtained in the background data acquisition step is set to 13,333 as described above, the dataset acquisition step and / or the data separation step can be performed such that the number of events classified into the particle group to be replaced by background data is up to 13,333.

[0157] In the background data acquisition step, the biological sample analysis device allows only the sheath fluid to flow through the flow channel and does not allow the sample fluid (verification sample) to flow through the flow channel. That is, in a state where the verification sample does not flow through the flow channel, light is used to irradiate the flow channel and the light generated by the light irradiation is detected.

[0158] To allow only the sheath fluid to flow through the flow channel as described above, for example, the biological sample analysis device can stop the pump for supplying the verification sample to the flow channel before (and after) performing the background data acquisition step, or can close the valve provided in the flow channel for supplying the verification sample to the flow channel. The state where only the sheath fluid flows through the flow channel can be achieved by stopping the pump or the valve.

[0159] (3-2-4) Data separation step S14

[0160] In the data separation step S14, the biological sample analysis device performs data separation processing on the data set of the signal intensity data acquired in the data set acquisition step S11. In the data separation step S14, data separation processing can be performed on the singlet data in the data set of the signal intensity data acquired in the data set acquisition step S11. For example, the data separation step can be performed by the information processing unit.

[0161] In the data separation step S14, the signal intensity data regarding the verification sample is separated into data for each of a plurality of types of particle groups having varying fluorescence intensity levels. For example, in the case where the verification sample has eight peaks, the data set of the high signal intensity data acquired for the verification sample is separated into data for eight particle groups corresponding to each peak. This separation is useful for identifying the signal intensity data to be replaced by the background data.

[0162] As described above, the biological sample analysis device can perform the data separation step to identify the signal intensity data to be replaced by the background data in the data replacement step described later. By replacing the identified signal intensity data with the background data, a signal intensity data set useful for obtaining an accurate evaluation value can be obtained.

[0163] In the data separation step, the signal intensity data acquired in the fluorescence dye channel can be clustered for each event. Clustering allows each event to be classified into any one of a plurality of types of particle groups. For example, in the case where the verification sample has eight peaks, each event data included in the data set of the high signal intensity data acquired for the verification sample is clustered into one of eight groups. As described above, the biological sample analysis device can be configured to perform data separation through the clustering process in the data separation step.

[0164] As a fluorescent dye channel for aggregation, for example, one or more fluorescent dye channels that can satisfactorily separate multiple types of particle groups can be selected.

[0165] When sterilizing the validation sample, fluctuations in the level of the acquired fluorescence signal intensity may occur in all fluorescent dye channels. In some fluorescent dye channels, two peaks in the low signal intensity region may merge, making it difficult to separate the data between the two peaks. Therefore, in the data separation step, each event is clustered using a fluorescent dye channel with good separation performance to classify the events into multiple types of particle groups. For example, by performing clustering in the data separation step, the eight-peak beads are classified into eight particle groups Dim1 to Dim8.

[0166] Refer to Figure 5 Describe data separation. A to C of this figure are two-dimensional plotted data, where each event is plotted based on the signal intensities of the eight-peak beads in two fluorescent dye channels. In A of the accompanying drawings, the X-axis represents the signal intensity in the FITC channel, and the Y-axis represents the signal intensity in the PE channel. In B of the accompanying drawings, the X-axis represents the signal intensity in the VioBlue channel, and the Y-axis represents the signal intensity in the PE channel. In C of the accompanying drawings, data separation itself is performed using the VioBlue / PE channel with good separation performance, and the plotted data is shown based on FITC / PE.

[0167] In A of the accompanying drawings, some events to be classified as Dim1 are classified as Dim2, and some events to be classified as Dim2 are classified as Dim1. That is, the separation between Dim1 and Dim2 is not appropriately performed. On the other hand, regarding B of the figure, the separation is appropriately performed, and C corresponds to a two-dimensional plot created using FITC / PE based on the clustering information in the separation of B, and regarding C, the separation is also appropriately performed, as in B, which is better than A.

[0168] Based on such plotted data, the VioBlue channel and the PE channel can be selected as the fluorescent dye channels to be used in the data separation step.

[0169] By performing clustering using a fluorescent dye channel that can perform appropriate separation as described above, an accurate evaluation value can be obtained.

[0170] In one embodiment, the fluorescent dye channel can be selected based on, for example, two-dimensional plotted data, where the signal intensity in one fluorescent dye channel on the X-axis and the signal intensity in another fluorescent dye channel on the Y-axis.

[0171] For selection, the biological sample analysis device can generate the above two-dimensional graph data using, for example, the signal intensity data obtained from any two of the multiple fluorescent dye channels included in the detection unit of the biological sample analysis device. For example, two-dimensional plotting data can be created for each pair among the multiple fluorescent dye channels.

[0172] Therefore, the user of the biological sample analysis device can select the fluorescent dye channels to be used in the data separation step.

[0173] In addition, based on the generated two-dimensional plotting data, the biological sample analysis device can select the fluorescent dye channels to be used in the data separation step.

[0174] In another embodiment, the fluorescent dye channels can be selected based on the separation index (SI) of each fluorescent dye channel. For example, the biological sample analysis device can calculate the SI of each fluorescent dye channel and select two fluorescent dye channels with the highest SI from the calculated SIs. The signal intensity data obtained from the two selected fluorescent dye channels can be used for clustering.

[0175] For example, the SI can be calculated using the signal intensities of two types of particle populations that produce two adjacent peaks in the signal intensity data obtained from any two fluorescent dye channels. For example, the SI can be calculated using the median and standard deviation of the signal intensities of each of the two types of particle populations.

[0176] For example, assume that two types of particle populations among multiple types of particle populations include the first particle population Dim1 and the second particle population Dim2, and the signal intensity of the second particle population Dim2 is higher than that of the first particle population Dim1. In this case, the SI is represented by the following formula:

[0177] SI = (Dim2_median - Dim1_median) / (rSD_Dim2 + rSD_Dim1).

[0178] In this formula, Dim2_median represents the median of the signal intensity of the second particle population Dim2, and Dim1_median represents the median of the signal intensity of the second particle population Dim1. In addition, rSD_Dim2 represents the standard deviation of the signal intensity of the second particle population Dim2, and rSD_Dim1 represents the standard deviation of the signal intensity of the first particle population Dim1.

[0179] The clustering process can be performed using a known method (e.g., the k-means method). For example, an example of a more specific method of the clustering process is disclosed in WO 2016 / 185755A.

[0180] For example, in the clustering process, the first central intensity of the signal intensity data of each particle group can be calculated based on the signal intensity data obtained as the result of the light detection, and clustering can be repeated based on the first central intensity. This allows each event to be appropriately classified into the corresponding particle group.

[0181] (3-2-5) Data replacement step S15

[0182] In the data replacement step S15, the biological sample analysis device replaces the signal intensity data of the light generated by irradiating any type of particle group in the data set obtained in the data set acquisition step S11 with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light. That is, as described above, the signal intensity data of any type of particle group in the data set before replacement is replaced with the background data. As a result, a replaced data set including the background data is generated. For example, the data replacement step can be executed by the information processing unit.

[0183] The signal intensity data to be replaced in the data replacement step can be any one of the signal intensity data of the multiple types of particle groups separated in the data separation step. Specifically, the signal intensity data to be replaced in the data replacement step can be the signal intensity data of the particle group with the lowest signal intensity. That is, the biological sample analysis device can be configured to replace the signal intensity data of the particle group with the lowest fluorescence level among the multiple types of particle groups with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light.

[0184] Since the particle group with a lower fluorescence level is particularly sensitive to fluctuations in the fluorescence level, the effect of data replacement according to the present disclosure becomes particularly obvious by replacing the signal intensity data of the particle group.

[0185] In particular, preferably, the data set before replacement includes the signal intensity data of the non-fluorescent particle group as described above, and in the data replacement step, the signal intensity data of the non-fluorescent particle group is replaced with the background data. That is, in the case where a sample containing multiple types of particle groups contains one type of non-fluorescent particle group and one or more types of fluorescent particle groups, the biological sample analysis device can be configured to, in the data replacement step, replace the signal intensity data of one type of non-fluorescent particle group with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light.

[0186] Because the non-fluorescent particle group is particularly vulnerable to fluctuations in the fluorescence level, the effect of data replacement according to the present disclosure becomes particularly obvious by replacing the signal intensity data of the particle group.

[0187] (3-2-6) Evaluation value calculation step S16

[0188] In the evaluation value calculation step S16, the biological sample analysis device calculates one or more evaluation values for evaluating the device state using the replaced dataset obtained in the data replacement step. The one or more evaluation values may include, for example, at least any one of the accuracy evaluation value of the device, the linearity evaluation value of the device, and the sensitivity evaluation value of the device. For example, the evaluation value calculation step may be executed by the information processing unit.

[0189] Preferably, the accuracy evaluation value is calculated in the evaluation value calculation step. The accuracy evaluation value may be, for example, MESF. Preferably, the accuracy evaluation value (specifically MESF) is calculated using the background data in the replaced dataset, specifically only the positive signal intensity data in the background data.

[0190] The output of each fluorescent dye channel is clamped to ensure that the long-term average value of the signal intensity data on the background becomes zero when no sample is flowing. The background increases when the optical noise or electrical noise becomes larger. In addition, as Figure 6 shown, the background shows a swing similar to alternating current (AC). When the level of the AC-like swing increases, the statistics based only on positive values (such as the average value) increase and can be used as an evaluation value for the device state.

[0191] In the evaluation value calculation step, in addition to the accuracy evaluation value, the linearity evaluation value and / or the sensitivity evaluation value of the device may also be calculated. For example, the linearity evaluation value may be linearity. The sensitivity evaluation value may be, for example, the Q value and / or the B value. The Q value represents the detection efficiency. The B value represents the background.

[0192] The details and calculation methods of these evaluation values will be described below based on the case of using octaplex beads. For other verification samples, those skilled in the art can appropriately calculate each evaluation value.

[0193] (a) Definition

[0194] Particle group Dim1: The particle group whose fluorescence intensity belongs to the lowest peak

[0195] Particle group Dim2: The particle group whose fluorescence intensity belongs to the second lowest peak

[0196] Particle group Dim3: The particle group whose fluorescence intensity belongs to the third lowest peak

[0197] Particle group Dim4: The particle group whose fluorescence intensity belongs to the fourth lowest peak

[0198] Particle group Dim5: The particle group whose fluorescence intensity belongs to the fifth lowest peak

[0199] Particle group Dim6: The particle group whose fluorescence intensity belongs to the sixth lowest peak

[0200] Particle swarm Dim7: The particle swarm with the fluorescence intensity belonging to the seventh lowest peak value

[0201] Particle swarm Dim8: The particle swarm with the fluorescence intensity belonging to the eighth lowest peak value

[0202] (Dim1 to Dim8 are also respectively called Peak1 to Peak8.)

[0203] MFI: Mean fluorescence intensity (the value obtained by converting the maximum and minimum average fluorescence signal intensities to 1 and 0 respectively)

[0204] MEF2 to MEF8: The reference MESF values of Dim2 to Dim8

[0205] MESF: Molecules of equivalent soluble fluorescent dye (the number of fluorescent molecules per particle)

[0206] Average values of Dim1 to Dim8: MFI1 to MF18

[0207] Logarithmic values of MFI1 to MFI 8: LogMFI1 to LogMFI8

[0208] Reference values of Dim1 to Dim8: MEF1 to MEF8

[0209] Logarithmic values of MEF2 to MEF8: LogMEF2 to LogMEF8

[0210] MESF value calculated for Dim1: MESF

[0211] MESF of Dim2 to Dim4: MESF2 to MESF4

[0212] Coefficient of variation of Dim2 to Dim8: CV2 to CV8

[0213] Standard deviation of Dim2 to Dim8: SD2 to SD8

[0214] (b) Linearity

[0215] Determine R using LogMFI2 to LogMFI8 and LogMEF2 to LogMEF8 2 (Coefficient of determination), and define the R obtained by taking the square root of R 2 as linearity.

[0216] The closer the linearity is to 100%, the better.

[0217] (c) Precision (molecules of equivalent soluble fluorescent dye (MESF),

[0218] also called fluorescence detection sensitivity)

[0219] [1] Determine the regression line using LogMFI2 to LogMFI8 and LogMEF2 to LogMEF8.

[0220] [2] Determine the fluorescence detection sensitivity (MESF) using MFI1 from the regression line.

[0221] The closer the fluorescence detection sensitivity (MESF) is to the reference value of Dim1 (usually 0), the better.

[0222] (d) Sensitivity (Q value and B value)

[0223] Determine MESF2 to MESF4 for Dim2 to Dim4.

[0224] Determine (SD2) 2 = ((CV2) 2 - (CV8) 2 ) × (MESF2) 2 , and also determine (SD3) 2 and (SD4) 2 .

[0225] Plot MESF2 to MESF4 on the horizontal axis and plot (SD2) 2 to (SD4) 2 on the vertical axis to determine the regression line, and represent the slope and intercept of the regression line as a and b, respectively.

[0226] The Q value is the reciprocal of the slope of the regression line (1 / a).

[0227] The B value is the ratio of the slope to the intercept of the regression line (b / a).

[0228] To calculate one or more evaluation values, the molecules of the equivalent fluorescent dye (MEF) of the validation sample can be used. The MEF can be pre-held by the biological sample analysis device and can be stored, for example, in the information processing unit (specifically, the storage unit) of the biological sample analysis device.

[0229] In one embodiment, the validation sample for the present disclosure can be a sample having different fluorescence levels for each peak as described above, and can be, for example, a sterilized validation sample. In this case, the MEF used to calculate one or more evaluation values is the MEF of the validation sample having different fluorescence levels, particularly the MEF of the sterilized validation sample.

[0230] In this embodiment, due to the MEF used to calculate one or more evaluation values, the MEF of the validation sample with unchanged fluorescence level is not suitable for use, that is, the MEF of the unsterilized validation sample is not suitable for use.

[0231] Therefore, the biological particle analyzer of the present invention can pre-store the MEF of the verification sample, or can obtain the MEF of the verification sample as follows.

[0232] First, as described in the dataset acquisition step S11, acquire the signal intensity dataset of the reference sample. As described in the data separation step S14, separate the signal intensity dataset of the reference sample for each type of particle population, and acquire the MFI of each of the multiple types of particle populations (specifically, each fluorescent particle population).

[0233] Similarly, for the verification sample as well, acquire the signal intensity dataset, and then acquire the MFI of each of the multiple types of particle populations (specifically, each fluorescent particle population).

[0234] For each particle population, the MFI of each particle population of the verification sample is calculated from the ratio between the MFI of the reference sample and the MFI of the verification sample. Specifically, the MFI is calculated using the following formula:

[0235] (MEF of a certain particle population of the verification sample) = (MEF of a certain particle population of the reference sample) × (MFI of the certain particle population of the verification sample) / (MFI of the certain particle population of the reference sample).

[0236] Figure 7 Shows examples of the (measured) MFI and (known) MEF of the reference sample and examples of the (measured) MFI and (calculated) MEF of the verification sample. For example, regarding Dim2, to calculate the MEF of the verification sample, use the MFI and MEF of Dim2 of the reference sample and the MFI of the verification sample to calculate the MEF of 205×86283 / 123916 = 143.

[0237] Preferably apply the MEF of the verification sample to the verification samples of the same batch. The biological sample analysis device can acquire the MEF of the verification sample via a network from, for example, a two-dimensional code such as a QR code (registered trademark).

[0238] (3-3) Other steps

[0239] The biological sample analysis device can perform an output step of outputting the calculated evaluation value to a display device after the evaluation value calculation step. By displaying the evaluation value in the output step, the user of the device can be prompted to determine whether to perform biological sample analysis and whether to calibrate the biological sample analysis device. The biological sample analysis device can cause the display device to display a screen for prompting such determination.

[0240] After the evaluation value calculation step, the biological sample analysis device can determine whether to perform biological sample analysis based on the calculated evaluation value. That is, it can automatically determine whether the device performs biological sample analysis after the verification process. For the determination, the biological sample analysis device can pre-have a predetermined threshold for determining whether to allow the execution of biological sample analysis, and can perform biological sample analysis based on whether the value is greater than or equal to or less than or equal to the threshold.

[0241] (4) Embodiment

[0242] In the case of no sterilization, a commercially available eight-peak bead is used to perform a verification process for verifying the device state of the cell sorter. Similarly, the eight-peak bead is sterilized, and the verification process for verifying the device state of the cell sorter is performed in a similar manner. The eight-peak bead contains one type of non-fluorescent bead and seven types of fluorescent beads, and forms eight peaks as a result of flow cytometry. As described in (2) above, the cell sorter includes a light irradiation unit, a detection unit, an information processing unit, and a sorting unit.

[0243] First, the fluorescence separation results of the non-sterilized eight-peak bead and the sterilized eight-peak bead are shown in Figure 8 . The fluorescence separation result of the non-sterilized eight-peak bead is shown as A on the upper side of the figure, and the fluorescence separation result of the sterilized eight-peak bead is shown as B on the lower side of the figure. As these results show, the non-sterilized eight-peak bead is well separated in terms of fluorescence. On the other hand, for the sterilized eight-peak bead, for example, the peaks of the non-fluorescent bead and the adjacent peaks are close to each other, preventing these peaks from being separated.

[0244] In addition, for the non-sterilized eight-peak bead and the sterilized eight-peak bead, the obtained signal intensity data sets are used to calculate their respective evaluation values for verifying the device state. The calculated evaluation values include a linear evaluation value (linearity), a precision evaluation value (MESF), and a sensitivity evaluation value (Q value and B value).

[0245] The difference (ΔX: X represents each evaluation value) and the difference rate (X difference rate: X represents each evaluation value) between the evaluation value obtained using the non-sterilized eight-peak bead and the evaluation value obtained using the sterilized eight-peak bead are shown in Figure 9 in the upper table A above. The difference and the difference rate are calculated using the following formulas:

[0246] ΔX = (evaluation value of the sterilized eight-peak bead) - (evaluation value of the non-sterilized eight-peak bead); and

[0247] X difference rate (%) = ((evaluation value of the sterilized eight-peak bead) - (evaluation value of the non-sterilized eight-peak bead)) / ((evaluation value obtained using the sterilized eight-peak bead) + (evaluation value obtained using the non-sterilized eight-peak bead) / 2) × 100.

[0248] As shown in the figure, the linear evaluation values of the sterilized eight-peak beads and the non-sterilized eight-peak beads are different and the linear evaluation values decrease. The accuracy evaluation values of the sterilized eight-peak beads and the non-sterilized eight-peak beads are different and the accuracy evaluation values increase. The sensitivity evaluation values of the sterilized eight-peak beads and the non-sterilized eight-peak beads are different and decrease or increase.

[0249] As described above, due to sterilization, the eight-peak beads experience fluctuations in fluorescence levels, and the evaluation values used to evaluate the device state change accordingly.

[0250] Next, a verification process according to the present disclosure is performed using the signal intensity data sets obtained for both non-sterilized eight-peak beads and sterilized eight-peak beads. That is, evaluation values are calculated using the replacement data set obtained by replacing the signal intensity data of non-fluorescent beads in the signal intensity data set with the background data of the cell sorter. In a similar manner as described above, the calculated evaluation values include a linear evaluation value (linearity), an accuracy evaluation value (MESF), and a sensitivity evaluation value (Q value and B value).

[0251] The differences and difference rates between the evaluation values obtained using non-sterilized eight-peak beads and the evaluation values obtained using sterilized eight-peak beads are shown in Figure 9 Table B below. As shown in the figure, the linearity evaluation values of the sterilized eight-peak beads and the non-sterilized eight-peak beads are the same and are significantly improved compared to the linearity evaluation values shown in Chart A. As shown in the figure, the accuracy evaluation value of the sterilized eight-peak beads is roughly the same as that of the non-sterilized eight-peak beads and is significantly improved compared to the accuracy evaluation value shown in Table A in the figure. As shown in the figure, the sensitivity evaluation value of the sterilized eight-peak beads is roughly the same as that of the non-sterilized eight-peak beads and is significantly improved compared to the sensitivity evaluation value shown in Table A in the figure.

[0252] As shown above, by performing the verification process according to the present disclosure, the device state can be appropriately evaluated even when using a sterilized verification sample.

[0253] In addition, it is considered that in addition to applying background data, using the MEF calculated as described above helps to improve linearity and sensitivity.

[0254] In addition, it is considered that in addition to applying background data, using the MEF calculated as described above and using a fluorescence dye channel suitable for separation helps to improve accuracy.

[0255] (5) Configuration example of a biological particle sorting device

[0256] In one embodiment, the biological sample analysis device of the present disclosure can be configured as a biological particle sorting device, such as a cell sorting device. The biological particle sorting device can be a device that analyzes and / or sorts biological particles in a microchip without forming droplets. The biological particle sorting device can be configured to perform the above verification process. The following will refer to Figure 10 and Figure 11 to describe this embodiment.

[0257] Figure 10 FIG. shows a schematic diagram of a configuration example of a biological particle sorting microchip and a configuration example of a biological particle analyzer including the microchip. Figure 11 FIG. shows an example of a flowchart of a biological particle sorting operation performed by a biological particle analyzer.

[0258] In Figure 10 the biological particle sorting microchip 150 shown includes a sample liquid flow channel 152 and a sheath liquid flow channel 154 joined to the sample liquid flow channel 152 at a junction 162. The biological particle sorting microchip 150 further includes a sample liquid inlet 151 and a sheath liquid inlet 153.

[0259] It should be noted that in Figure 10 a part of the sheath liquid flow channel 154 is represented by a dashed line. The part represented by the dashed line is located at a position lower than the position of the sample liquid flow channel 152 represented by a solid line (a position shifted in the optical axis direction described later), and the flow channel represented by the dashed line and the flow channel represented by the solid line are not connected to each other at the position where the flow channels cross. Further, in Figure 10 the sample liquid flow channel 152 is shown to bend twice between the sample liquid inlet 151 and the junction 162, which is beneficial for differentiating the sample liquid flow channel 152 from the sheath liquid flow channel 154. The sample liquid flow channel 152 can be linearly formed between the sample liquid inlet 151 and the junction 162 without bending in this way.

[0260] In the biological particle sorting operation, a sample liquid containing biological particles is introduced from the sample liquid inlet 151 into the sample liquid flow channel 152, and a sheath liquid not containing biological particles is introduced from the sheath liquid inlet 153 into the sheath liquid flow channel 154.

[0261] The biological particle sorting microchip 150 includes a joining flow channel 155 having a junction 162 at one end.

[0262] The sample liquid and the sheath liquid are joined at the junction 162 and then flow through the joined flow channel 155 toward the particle sorting unit 157. Specifically, the sample liquid and the sheath liquid are combined at the junction 162 to form, for example, a laminar flow in which the sample liquid is surrounded by the sheath liquid. Preferably, in the laminar flow, the biological particles are substantially arranged in a line. The sample liquid flow channel 152 and the two sheath liquid flow channels 154 are joined at the junction 162 and include a flow channel structure having a joined flow channel 155 with one end serving as the junction 162 to form a laminar flow including biological particles flowing substantially in a line. Therefore, when light is irradiated onto the detection region 156 described later, it is possible to easily distinguish the light generated by irradiating one biological particle with light and the light generated by irradiating another biological particle with light.

[0263] The biological particle sorting microchip 150 further includes a particle sorting unit 157 located at the other end of the joined flow channel 155. Figure 12 is an enlarged view of the particle sorting unit 157. As Figure 12 shown in A of, at the other end, the joined flow channel 155 is connected to the biological particle recovery flow channel 159 via a connection flow channel 170. As Figure 12 shown in A of, the joined flow channel 155, the connection flow channel 170, and the biological particle recovery flow channel 159 can be coaxial with each other.

[0264] When the target particle to be recovered flows into the particle sorting unit 157, as Figure 12 shown in B of, a flow is formed from the connected flow channel 155 through the connection flow channel 170 into the biological particle recovery flow channel 159, and the target particle to be recovered is recovered into the biological particle recovery flow channel 159. In this way, the target particle to be recovered flows through the connection flow channel 170 into the biological particle recovery flow channel 159.

[0265] When biological particles that are not the target particle to be recovered flow into the particle sorting unit 157, the biological particles that are not the target particle to be recovered flow into the branch flow channel 158, as Figure 12 shown in C of. In this case, no flow into the biological particle recovery flow channel 159 is formed.

[0266] As Figure 10 shown, the biological particle recovery flow channel 159 is formed to linearly extend from the particle sorting unit 157, make a U-turn, and then reach the same surface as the surface where the sample liquid inlet 151 and the sheath liquid inlet 153 are formed. The liquid flowing through the biological particle recovery flow channel 159 is discharged out of the chip from the recovery flow channel terminal 163.

[0267] As Figure 10As shown, the two branch flow channels 158 are also formed to linearly extend from the particle sorting unit 157, make a U-turn, and then reach the same surface as the surface where the sample liquid inlet 151 and the sheath liquid inlet 153 are formed. The liquid flowing in the branch flow channels 158 is discharged out of the chip from the branch flow channel terminals 166.

[0268] In Figure 10 , the display method of the biological particle recovery flow channel 159 changes from a solid line to a dotted line at the U-turn. This change indicates that the position in the optical axis direction changes midway. By changing the position in the optical axis direction in this way, the biological particle recovery flow channel 159 does not communicate with the branch flow channel 158 in the portion where they cross.

[0269] The recovery flow channel terminal 163 and the two branch flow channel terminals 166 are both formed on the surface where the sample liquid inlet 151 and the sheath liquid inlet 153 are formed. In addition, the introduction flow channel inlet 164 for introducing the liquid into the introduction flow channel 161 is also formed on the surface. In this way, the biological particle sorting microchip 150 has all the inlets from which the liquid is introduced and the outlets from which the liquid is discharged formed on one surface. This helps to attach the chip to the biological particle analyzer 100. For example, compared with the case where the inlets and / or outlets are formed on more than two surfaces, the connection between the flow channels provided in the biological particle analyzer 100 and the flow channels of the biological particle sorting microchip 150 becomes easier.

[0270] As Figure 10 and Figure 12 shown, the biological particle sorting microchip 150 includes an introduction flow channel 161 for introducing the liquid into the connection flow channel 170.

[0271] By introducing the liquid from the introduction flow channel 161 into the connection flow channel 170, the connection flow channel 170 is filled with the liquid. Therefore, unwanted biological particles can be prevented from entering the biological particle recovery flow channel 159.

[0272] The biological particle sorting microchip 150 includes two branch flow channels 158 connected to the joining flow channel 155 at the other end of the joining flow channel 155. In this way, in the biological particle sorting microchip used in the present technology, the connected flow channel can be branched into a connection flow channel and at least one branch flow channel.

[0273] Biological particles other than the target particles to be recovered flow into either of the two branch flow channels 158 without entering the biological particle recovery flow channel 159.

[0274] In addition, as Figure 10As shown, the biological particle sorting microchip 150 forms part of the biological particle analyzer 100, which includes a light irradiation unit 101, a detection unit 102, and a control unit 103 in addition to the microchip. The light irradiation unit 101, the detection unit 102, and the control unit 103 respectively correspond to the light irradiation unit 6101, the detection unit 6102, and the information processing unit 6103 described in (2) above, and their descriptions also apply to this configuration example. As Figure 13 shown, the control unit 103 of the biological particle analyzer 100 may include a signal processing unit 104, a determination unit 105, and a sorting control unit 106.

[0275] As Figure 11 shown, the biological particle sorting operation using the biological particle sorting microchip 150 includes a flow step S101 of allowing a liquid containing biological particles to flow through the joined flow channel 155, a determination step S102 of determining whether the biological particles flowing through the joined flow channel 155 are target particles to be recovered, and a recovery step S103 of recovering the target particles to the biological particle recovery flow channel 159. Each step will be described below.

[0276] (5-1) Flow step

[0277] In the flow step S101, a sample liquid containing biological particles and a sheath liquid not containing biological particles are respectively introduced into the sample liquid flow channel 152 and the sheath liquid flow channel 154 from the sample liquid inlet 151 and the sheath liquid inlet 153.

[0278] The sample liquid and the sheath liquid are combined at the junction 162 to form, for example, a laminar flow in which the sample liquid is surrounded by the sheath liquid. Preferably, in the laminar flow, the biological particles are substantially arranged in a line. That is, in the flow step S101, a laminar flow containing biological particles flowing substantially in a line can be formed.

[0279] In this way, in the flow step S101, the liquid containing biological particles is allowed to flow through the joined flow channel 155, specifically as a laminar flow. The liquid flows through the joined flow channel 155 from the junction 162 toward the particle sorting unit 157.

[0280] (5-2) Determination step

[0281] In the determination step S102, it is determined whether the biological particles flowing through the joined flow channel 155 are target particles to be recovered. This determination can be performed by the determination unit 105. The determination unit 105 can make a determination based on the light generated when the light irradiation unit 101 irradiates the biological particles with light.

[0282] The signal processing unit 104 included in the control unit 103 can process the waveform of the digital electrical signal obtained by the detection unit 102 to generate information (data) regarding the characteristics of the light to be determined by the determination unit 105. As the information regarding the characteristics of the light, the signal processing unit 104 can obtain, for example, one, two, or all of the width of the waveform, the height of the waveform, and the area of the waveform from the waveform of the digital electrical signal. In addition, the information regarding the characteristics of the light can include, for example, the time when the light is detected. Particularly in embodiments for detecting scattered light and / or fluorescence, the processing of the signal processing unit 104 described above can be performed.

[0283] Based on the light generated by irradiating biological particles flowing through the flow channel with light, the determination unit 105 included in the control unit 103 determines whether the biological particles are the target particles to be recovered.

[0284] (5-3) Recovery step

[0285] In the recovery step S103, the biological particles determined to be the target particles to be recovered in the determination step S102 are recovered into the biological particle recovery flow channel 159. The recovery step S103 is performed in the particle sorting unit 157 in the microchip 150. In the particle sorting unit 157, the laminar flows flowing through the connected flow channel 155 flow into the two branch flow channels 158 respectively. Figure 10 The particle sorting unit 157 shown in Figure 10 includes two branch flow channels 158, but the number of branch flow channels is not limited to two. The particle sorting unit 157 can be provided with, for example, one or more (e.g., two, three, or four) branch flow channels. The branch flow channels can be formed to branch into a Y shape on a plane as

[0286] In the recovery step S103, due to the pressure fluctuations in the biological particle recovery flow channel 159, the target particles to be recovered are recovered into the biological particle recovery flow channel through the connection flow channel. For example, as described above, the recovery can be performed by generating a negative pressure within the biological particle recovery flow channel 159. For example, when the wall defining the biological particle recovery flow channel 159 is deformed by an actuator 107 (specifically, a piezoelectric actuator) attached to the outer side of the microchip 150, a negative pressure can be generated. The negative pressure can form a flow into the biological particle recovery flow channel 159. To generate the negative pressure, the actuator 107 can be attached to the outside of the microchip 150 such that, for example, the wall of the biological particle recovery flow channel 159 can be deformed. The negative pressure can be generated by changing the deformation of the wall of the internal space of the biological particle recovery flow channel 159. The actuator 107 can be, for example, a piezoelectric actuator. When the target particles to be recovered are sucked into the biological particle recovery flow channel 159, the sample liquid forming a laminar flow or the sample liquid and the sheath liquid forming a laminar flow can also flow into the biological particle recovery flow channel 159. In this way, the target particles to be recovered are sorted in the particle sorting unit 157 and are recovered into the biological particle recovery flow channel 159.

[0287] The connection flow channel 170 is provided with an introduction flow channel 161 to prevent biological particles that are not the target particles to be recovered from entering the biological particle recovery flow channel 159 through the connection flow channel 170. Liquid is introduced into the connection flow channel 170 from the introduction flow channel 161. By introducing the liquid, the connection flow channel 170 is filled with liquid. In addition, since the flow from the connection flow channel 170 to the joining flow channel 155 is formed by a part of the liquid, biological particles other than the target particles to be recovered can be prevented from entering the biological particle recovery flow channel 159. The liquid forming the flow from the connection flow channel 170 to the joining flow channel 155 flows to the branch flow channel 158 through the liquid flowing through the joining flow channel 155 in a manner similar to the liquid not flowing through the joining flow channel 155.

[0288] It should be noted that the remaining part of the liquid introduced into the connection flow channel 170 flows to the biological particle recovery flow channel 159. Therefore, the biological particle recovery flow channel 159 can be filled with liquid.

[0289] The flow to the branch flow channel 158 can be discharged from the microchip at the branch flow channel terminal 160. In addition, the target particles to be recovered and recovered into the biological particle recovery flow channel 159 can be discharged from the microchip at the recovery flow channel terminal 163. A container can be connected to the recovery flow channel terminal 163 via a flow channel such as a tube. The target particles to be recovered can be recovered into the container.

[0290] 2. Second Embodiment (Biological Sample Analysis System)

[0291] The present disclosure also provides a biological sample analysis system configured to perform the verification process described in 1 above. That is, the system includes: an information processing unit configured to perform information processing on light generated by irradiating a flow channel through which particles flow with light; wherein, the information processing unit may be configured to perform a data replacement step that replaces the signal intensity data of light generated by irradiating any type of particle population in a data set of signal intensity data of light generated by irradiating light on a sample containing multiple types of particle populations, the multiple types of particle populations having different fluorescence intensity levels, the sample having signal intensity data obtained by irradiation, and in the verification process of the state of the verification device, using light, the flow channel through which no particles flow.

[0292] In addition to the information processing unit, the biological sample analysis system may include the light irradiation unit and the detection unit described in 1 above. A sorting unit may also be included. These components may be provided in one device or may be provided in multiple devices in a distributed manner. For example, the biological sample analysis system may include an information processing device configured as the information processing unit. The biological sample analysis system may include an analyzer that includes a light irradiation unit and a detection unit (and a sorting unit) in addition to the information processing device. These devices may be connected to each other in a wired or wireless manner. In addition, these devices may be connected to each other via a network.

[0293] For example, the system may perform the following verification process according to the present disclosure.

[0294] In the data set acquisition step S11 of the flowchart described in 1 above Figure 3 the analyzer (specifically, the light irradiation unit and the detection unit) constituting the biological sample analysis system performs light irradiation and detection of the light generated by the light irradiation, and the information processing device acquires a signal intensity data set regarding the light.

[0295] In the trigger piece setting step S12, the information processing device performs trigger piece setting. The information processing device sends the set trigger piece to the analyzer.

[0296] In the background data acquisition step S13, the analyzer uses the trigger piece to perform light irradiation and light detection for background data acquisition, and sends the background data detected by the light detection to the information processing device. The information processing device receives the background data.

[0297] The information processing device uses the acquired signal intensity data set and the background data to perform a data separation step S14, a data replacement step S15, and an evaluation value calculation step S16.

[0298] 3. Third Embodiment (Verification Method for Biological Sample Analysis Device and Program for Executing Verification Method)

[0299] The present disclosure further provides a verification method for verifying the state of a biological sample analysis device, including performing the verification process described in 1 above. That is, the verification method includes: a data replacement step of flowing a sample containing a plurality of types of particle groups with different fluorescence intensity levels into a flow channel, and replacing the signal intensity data of the light generated by irradiating any one type of particle group in the data set regarding the signal intensity data of the light generated by irradiating the particles flowing through the flow channel with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light. For example, referring to Figure 3 , the verification method can be performed as described in 1 above, and this description also applies to the verification method.

[0300] The present disclosure also provides a program for causing a biological sample analysis device or an information processing device to execute the verification method. The program can be stored, for example, in a biological sample analysis device or an information processing device (specifically, a storage unit), or can be stored in an information storage medium. The information storage medium can be, for example, an SD card, a micro SD card, a CD, a DVD, a flash memory, or a magnetic recording medium.

[0301] It should be noted that the present disclosure may also have the following configurations. [1]

[0303] A biological sample analysis device, including:

[0304] An information processing unit configured to perform information processing on the light generated by irradiating the flow channel through which particles flow using signal intensity data, where

[0305] the information processing unit performs a data replacement step of replacing, in the verification process of verifying the state of the device, the signal intensity data of the light generated by irradiating any one type of particle group in the data set of the signal intensity data with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light, the signal intensity data being regarding the light generated by irradiating a sample containing a plurality of types of particle groups, and the plurality of types of particle groups having varying fluorescence intensity levels. [2]

[0307] The biological sample analysis device according to [1], wherein the information processing unit calculates one or more evaluation values for verifying the state of the device using the data set obtained after the data replacement process. [3]

[0309] The biological sample analysis device according to [2], wherein one or more evaluation values include at least any one of the accuracy evaluation value of the device, the linearity evaluation value of the device, or the sensitivity evaluation value of the device. [4]

[0311] The biological sample analysis device according to any one of [1] to [3], wherein, in the data replacement step, the signal intensity data of the particle group having the lowest signal intensity among multiple types of particle groups is replaced. [5]

[0313] The biological sample analysis device according to any one of [1] to [4], wherein

[0314] The sample containing multiple types of particle groups contains one type of non-fluorescent particle group and one or more types of fluorescent particle groups, and

[0315] In the data replacement step, the signal intensity data of one type of non-fluorescent particle group is replaced. [6]

[0317] The biological sample analysis device according to any one of [1] to [5], configured to perform a trigger piece setting step of setting a trigger piece using a data set. [7]

[0319] The biological sample analysis device according to [6], configured to obtain signal intensity data of light generated by irradiating a flow channel through which no particles flow with light using a trigger piece. [8]

[0321] The biological sample analysis device according to any one of [1] to [7], configured to perform a background data acquisition step of acquiring signal intensity data of light generated by irradiating a flow channel through which no particles flow with light. [9]

[0323] The biological sample analysis device according to [8], wherein the acquisition of the signal intensity data in the background data acquisition step is performed in a state where the sample does not flow into the flow channel but only the sheath fluid flows into the flow channel.

[10]

[0325] The biological sample analysis device according to [8] or [9], wherein the acquisition of the signal intensity data in the background data acquisition step is performed at a predetermined time interval.

[11]

[0327] A biological sample analysis device according to any one of [8] to

[10] , wherein the number of signal intensity data obtained in the background data acquisition step is the same as the number set based on the number of event data in the dataset and the number of peaks formed by the sample.

[12]

[0329] A biological sample analysis device according to any one of [8] to

[11] , wherein the number of signal intensity data obtained in the background acquisition step is the same as the number set based on the number of signal intensity data replaced in the data replacement step.

[13]

[0331] A biological sample analysis device according to any one of [1] to

[12] , configured to further perform a data separation step that separates the dataset into data corresponding to each of a plurality of types of particle populations.

[14]

[0333] The biological sample analysis device according to

[13] , configured to perform data separation through aggregation processing in the data separation step.

[15]

[0335] A biological sample analysis device according to any one of [1] to

[14] ,

[0336] The biological sample analysis device is configured to, in the data replacement step, replace the signal intensity data of the particle population with the lowest fluorescence level among multiple types of particle populations with the signal intensity data obtained by irradiating a flow channel through which no particles flow with light.

[16]

[0338] A biological sample analysis device according to any one of [1] to

[15] , wherein,

[0339] A sample containing multiple types of particle populations contains one type of non-fluorescent particle population and one or more types of fluorescent particle populations,

[0340] The biological sample analysis device is configured to, in the data replacement step, replace the signal intensity data of one type of non-fluorescent particle population with the signal intensity data obtained by irradiating a flow channel through which no particles flow with light.

[17]

[0342] A biological sample analysis device according to any one of [1] to

[16] , used as a flow cytometer.

[18]

[0344] A biological sample analysis system, comprising:

[0345] An information processing unit configured to perform information processing on light generated by irradiating a flow channel through which particles flow with light, using signal intensity data, wherein

[0346] The information processing unit performs a data replacement step, in a verification process for verifying the state of a verification device, of replacing signal intensity data regarding light generated by irradiating a particle group of any one type in a dataset of signal intensity data regarding light generated by irradiating a sample containing multiple types of particle groups with light, with signal intensity data obtained by irradiating a flow channel through which no particles flow with light. The multiple types of particle groups have varying fluorescence intensity levels.

[19]

[0348] A method for verifying the state of a biological sample analysis device, the method comprising:

[0349] A data replacement step of causing a sample containing multiple types of particle groups having different fluorescence intensity levels to flow into a flow channel, and replacing signal intensity data regarding light generated by irradiating a particle group of any one type in a dataset of signal intensity data regarding light generated by irradiating particles flowing through the flow channel with light, with signal intensity data obtained by irradiating a flow channel through which no particles flow with light.

[0350] List of reference symbols

[0351] 6100 Biological sample analysis device

[0352] 6101 Light irradiation unit

[0353] 6102 Detection unit

[0354] 6103 Information processing unit

Claims

1. A biological sample analysis device, comprising: An information processing unit configured to perform information processing on light generated by irradiating a flow channel through which particles flow with light, wherein, In a verification process of verifying the state of the device, the information processing unit performs a data replacement step, and for a data set of signal intensity data of light generated by irradiating a sample containing multiple types of particle groups with varying fluorescence intensity levels with light, replaces the signal intensity data of light generated by irradiating any one type of particle group in the data set with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light.

2. The biological sample analysis device according to claim 1, wherein, The information processing unit uses the data set obtained after the data replacement process to calculate one or more evaluation values for verifying the state of the device.

3. The biological sample analysis device according to claim 2, wherein, The one or more evaluation values include at least any one of an accuracy evaluation value of the device, a linearity evaluation value of the device, or a sensitivity evaluation value of the device.

4. The biological sample analysis device according to claim 1, wherein, In the data replacement step, the signal intensity data of the particle group with the lowest signal intensity among the multiple types of particle groups is replaced.

5. The biological sample analysis device according to claim 1, wherein, The sample containing the multiple types of particle groups contains one type of non-fluorescent particle group and one or more types of fluorescent particle groups, and In the data replacement step, the signal intensity data of the one type of non-fluorescent particle group is replaced.

6. The biological sample analysis device according to claim 1, configured to perform a trigger piece setting step of setting a trigger piece using the data set.

7. The biological sample analysis device according to claim 6, configured to obtain the signal intensity data of light generated by irradiating the flow channel through which no particles flow with light using the trigger piece.

8. The biological sample analysis device according to claim 1, configured to perform a background data acquisition step of obtaining the signal intensity data of light generated by irradiating the flow channel through which no particles flow with light.

9. The biological sample analysis device according to claim 8, wherein, The acquisition of the signal intensity data in the background data acquisition step is performed in a state where the sample does not flow into the flow channel, but only the sheath fluid flows into the flow channel.

10. The biological sample analysis device according to claim 8, wherein, The acquisition of the signal intensity data in the background data acquisition step is performed at a predetermined time interval.

11. The biological sample analysis device according to claim 8, wherein, The number of signal intensity data obtained in the background data acquisition step is the same as the number set based on the number of event data in the data set and the number of peaks formed by the sample.

12. The biological sample analysis device according to claim 8, wherein, The number of signal intensity data obtained in the background data acquisition step is the same as the number set based on the number of signal intensity data replaced in the data replacement step.

13. The biological sample analysis device according to claim 1, configured to further perform a data separation step of separating the data set into data corresponding to each of the multiple types of particle groups.

14. The biological sample analysis device according to claim 13, configured to perform the data separation through clustering processing in the data separation step.

15. The biological sample analysis device according to claim 1, wherein the biological sample analysis device is configured to, in the data replacement step, replace the signal intensity data of the particle group with the lowest fluorescence level among the multiple types of particle groups with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light.

16. The biological sample analysis device according to claim 1, wherein, the sample containing the multiple types of particle groups contains one type of non-fluorescent particle group and one or more types of fluorescent particle groups, and the biological sample analysis device is configured to, in the data replacement step, replace the signal intensity data of the one type of non-fluorescent particle group with the signal intensity data obtained by irradiating the flow channel through which no particles flow with light.

17. The biological sample analysis device according to claim 1, used as a flow cytometer.

18. A biological sample analysis system, comprising: an information processing unit configured to perform information processing on light generated by irradiating a flow channel through which particles flow with light using signal intensity data, wherein in a verification process for verifying the state of the device, the information processing unit performs a data replacement step on a data set of signal intensity data of light generated by irradiating a sample containing multiple types of particle groups with varying fluorescence intensity levels, and replaces the signal intensity data of light generated by irradiating any one type of particle group in the data set with the signal intensity data obtained by irradiating a flow channel through which no particles flow with light.

19. A method for verifying the state of a biological sample analysis device, the method comprising: a data replacement step of flowing a sample containing multiple types of particle groups with varying fluorescence intensity levels into a flow channel, and for a data set of signal intensity data of light generated by irradiating the particles flowing through the flow channel, replacing the signal intensity data of light generated by irradiating any one type of particle group in the data set with the signal intensity data obtained by irradiating a flow channel through which no particles flow with light.

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