Sample analyzer and sample analysis method

Abstract

A sample analyzer and a sample analysis method. The method comprises: suctioning a blood sample to be measured (S210); mixing at least a part of said blood sample, a hemolytic agent, a first dye, and a second dye to prepare a measurement sample (S220); in a single test, enabling particles in the measurement sample to pass through one by one an optical detection area radiated by excitation light, so as to acquire optical information, wherein the optical information comprises at least one piece of scattered light information, first fluorescence information corresponding to the first dye, and second fluorescence information corresponding to the second dye, and the scattered light information, the first fluorescence information, and the second fluorescence information are acquired in the single test (S230); and on the basis of the at least one piece of scattered light information, the first fluorescence information, and the second fluorescence information, obtaining a five-type classification result of white blood cells and a counting result of nucleated red blood cells of the measurement sample (S240). In this way, both a five-type classification result of white blood cells and a counting result of a nucleated red blood cell group can be obtained in a single test.

Description

Sample analyzer and sample analysis methods

[0001] This application claims priority to Chinese Patent Application No. 202410704288.4, filed on May 31, 2024, entitled “Sample Analyzer and Sample Analysis Method”; Chinese Patent Application No. 202410702597.8, filed on May 31, 2024, entitled “Sample Analyzer and Sample Analysis Method”; and Chinese Patent Application No. 202410703128.8, filed on May 31, 2024, entitled “Sample Analyzer and Sample Analysis Method”, the entire contents of which are incorporated herein by reference. Technical Field

[0002] This application relates to the field of blood cell detection, and in particular to a sample analyzer and a sample analysis method. Background Technology

[0003] Currently, laser scattering and fluorescence staining methods are commonly used to classify and count blood cells in blood samples. To obtain parameters such as red blood cells, white blood cells, reticulocytes, and nucleated red blood cells, multiple reaction and detection channels are often set up in the hematology analyzer. Each reaction and detection channel can only detect and analyze one parameter. Different fluorescent dyes are used to fluoresce the blood cells in different reaction and detection channels, and then a single exciter is used to excite the blood cells, collecting a single fluorescence signal. In this process, each dye can only specifically stain a single cell.

[0004] Existing blood cell analyzers based on fluorescence detection have the following drawbacks: white blood cells and red blood cells need to be detected in different reaction and detection channels, resulting in low detection efficiency and increased reagent costs; the dyes and reagents used in each reaction and detection channel are different, resulting in a complex liquid path structure of the instrument.

[0005] For example, in existing blood cell analyzers based on fluorescence detection, in order to obtain white blood cell classification parameters and nucleated red blood cell parameters, two reaction and detection channels need to be set up in the blood cell analyzer. In one reaction and detection channel, a test sample for white blood cell classification is prepared from the blood sample to be tested, a hemolysin, and a fluorescent dye, and the test sample is detected to obtain the white blood cell classification result. In the other reaction and detection channel, a test sample for identifying nucleated red blood cells is prepared from the blood sample to be tested, another hemolysin, and another fluorescent dye, and the test sample is detected to obtain the nucleated red blood cell count result.

[0006] However, this method of obtaining white blood cell classification parameters and nucleated red blood cell parameters is inefficient, requires a large amount of blood sample and reagents, and the corresponding instrument structure is relatively complex.

[0007] For example, in existing blood cell analyzers based on fluorescence detection, to obtain accurate white blood cell differential (including neutrophils, lymphocytes, monocytes, eosinophils, and basophils), two reaction and detection channels need to be set up in the blood cell analyzer. In one reaction and detection channel, a test sample for the four-part differential white blood cell count is prepared from the blood sample, a hemolysin, and a fluorescent dye, and this test sample is detected to obtain the four-part differential white blood cell count (including neutrophils, lymphocytes, monocytes, and eosinophils). In the other reaction and detection channel, a test sample for identifying basophils is prepared from the blood sample, another hemolysin, and another fluorescent dye, and this test sample is detected to obtain the basophil count. Combining the detection results from these two reaction and detection channels ultimately yields an accurate five-part differential white blood cell count.

[0008] However, this method of obtaining five-part differential white blood cell counts is inefficient, requires a large amount of blood sample and reagents, and the corresponding instrument structure is relatively complex. Summary of the Invention

[0009] One objective of this application is to obtain white blood cell differential and nucleated red blood cell count results through only one reaction and detection channel.

[0010] Another objective of this application is to obtain accurate five-part differential white blood cell results using only one reaction and detection channel.

[0011] To achieve the objectives of this application, the first aspect of this application relates to a sample analyzer, comprising:

[0012] A sampling device is used to collect blood samples for testing.

[0013] A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample;

[0014] An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated when particles in the test sample are illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye; and

[0015] The processor is configured to: generate a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtain the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, wherein the white blood cell five-part differential result includes the count result of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

[0016] To achieve the objectives of this application, a second aspect of this application relates to a corresponding sample analysis method, comprising:

[0017] Collect the blood sample to be tested;

[0018] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0019] In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by light. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye; and

[0020] A scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the scatter plot. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

[0021] To achieve the objectives of this application, a third aspect of this application also relates to a sample analysis method, including:

[0022] Collect the blood sample to be tested;

[0023] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0024] In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by light. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye; and

[0025] The white blood cell differential and nucleated red blood cell count results of the test sample are obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information. The white blood cell differential results include the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample.

[0026] To achieve the objectives of this application, a fourth aspect of this application relates to a sample analyzer, comprising:

[0027] A sampling device is used to collect blood samples for testing.

[0028] A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample;

[0029] An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated when particles in the test sample are illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light information detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye. The light emitting device includes a first light source and a second light source. The first light source emits a light beam of a first wavelength capable of exciting the first dye, and the second light source emits a light beam of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength.

[0030] The processor is configured to: generate a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtain the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, wherein the white blood cell four-part differential result includes the count result of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

[0031] To achieve the objectives of this application, the fifth aspect of this application relates to a corresponding sample analysis method, comprising:

[0032] Collect the blood sample to be tested;

[0033] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0034] In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles after irradiation. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. The excitation light includes light of a first wavelength capable of exciting the first dye and light of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength.

[0035] A scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the scatter plot. The white blood cell four-part differential result includes the count results of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

[0036] To achieve the objectives of this application, the sixth aspect of this application also relates to a sample analysis method, including:

[0037] Collect the blood sample to be tested;

[0038] At least a portion of the blood sample to be tested, a hemolytic agent, a first dye, and a second dye are mixed to prepare a test sample, wherein the first dye and the second dye have different maximum absorption wavelengths;

[0039] In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles after irradiation. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. The excitation light includes light of a first wavelength capable of exciting the first dye and light of a second wavelength capable of exciting the second dye.

[0040] The white blood cell differential and nucleated red blood cell count results of the test sample are obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information. The white blood cell differential results include the count results of neutrophils, lymphocytes, monocytes and eosinophils in the test sample.

[0041] In the technical solutions provided in the first to sixth aspects of this application, a blood sample to be tested, a hemolysin, and two dyes are mixed in a reaction and detection channel to prepare a test sample, and the test sample is optically measured in one test to obtain white blood cell classification results and nucleated red blood cell count results based on the optical information obtained in one test.

[0042] To achieve the objectives of this application, the seventh aspect of this application relates to a sample analyzer, comprising:

[0043] A sampling device is used to collect blood samples for testing.

[0044] A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample;

[0045] An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated by the particles in the test sample after being illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light information detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye. The light beam includes at least a first wavelength capable of exciting the first dye, wherein the first wavelength is between 315 nm and 490 nm.

[0046] The processor is configured as follows:

[0047] The white blood cell differential count of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell differential count includes the counts of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

[0048] The basophil count in the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information.

[0049] To achieve the objectives of this application, the eighth aspect of this application relates to a sample analyzer, comprising:

[0050] A sampling device is used to collect blood samples for testing.

[0051] A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample;

[0052] An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated by the particles in the test sample after being illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye; and

[0053] The processor is configured as follows:

[0054] The white blood cell differential count of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell differential count includes the counts of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

[0055] The basophil count is corrected based on at least the second fluorescence information to obtain a corrected basophil count.

[0056] To achieve the objectives of this application, the ninth aspect of this application relates to a sample analyzer, comprising:

[0057] A sampling device is used to collect blood samples for testing.

[0058] A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample;

[0059] An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated by the particles in the test sample after being illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye; and

[0060] The processor is configured as follows:

[0061] A three-dimensional scatter plot of the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information; and

[0062] Based on the three-dimensional scatter plot, the white blood cells in the test sample are classified into neutrophil population, lymphocyte population, monocyte population, eosinophil population, and basophil population.

[0063] To achieve the objectives of this application, the tenth aspect of this application relates to a sample analysis method, comprising:

[0064] Collect the blood sample to be tested;

[0065] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0066] In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. The excitation light includes light capable of exciting the first dye at a first wavelength, which is between 315 nm and 490 nm.

[0067] The white blood cell differential count of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell differential count includes the counts of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

[0068] The basophil count in the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information.

[0069] To achieve the objectives of this application, the eleventh aspect of this application relates to a sample analysis method, comprising:

[0070] Collect the blood sample to be tested;

[0071] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0072] In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one type of scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test.

[0073] The white blood cell differential count of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell differential count includes the counts of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

[0074] The basophil count is corrected based on at least the second fluorescence information to obtain a corrected basophil count.

[0075] To achieve the objectives of this application, the twelfth aspect of this application relates to a sample analysis method, comprising:

[0076] Collect the blood sample to be tested;

[0077] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0078] In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye.

[0079] A three-dimensional scatter plot of the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information; and

[0080] Based on the three-dimensional scatter plot, the white blood cells in the test sample are classified into neutrophil population, lymphocyte population, monocyte population, eosinophil population, and basophil population.

[0081] In the technical solutions provided in aspects seven to twelfth of this application, a blood sample to be tested, a hemolysin, and two dyes are mixed in a reaction and detection channel to prepare a test sample, and the test sample is optically measured in one test to obtain an accurate five-part differential white blood cell result based on the optical information obtained in one test. Attached Figure Description

[0082] Figure 1 is a schematic diagram of one embodiment of the sample analyzer according to this application.

[0083] Figure 2 is a schematic diagram of the emission spectra of two dyes according to an embodiment of this application.

[0084] Figure 3 is a schematic diagram of the emission and excitation spectra of a large Stokes shift dye according to an embodiment of this application.

[0085] Figure 4 is a schematic diagram of an embodiment of the optical detection device according to this application.

[0086] Figures 5 to 10 are schematic flowcharts of different embodiments of the sample analysis method according to this application.

[0087] Figures 11A and 11B are scatter plots obtained from the detection of blood samples to be tested according to the first embodiment of this application.

[0088] Figures 12A and 12B are scatter plots obtained from the detection of blood samples to be tested according to the second embodiment of this application.

[0089] Figures 13A and 13B are scatter plots obtained from the detection of blood samples to be tested according to the third embodiment of this application.

[0090] Figures 14A and 14B are scatter plots obtained from the detection of blood samples to be tested according to the fourth embodiment of this application.

[0091] Figures 15A and 15B are scatter plots obtained from the detection of blood samples to be tested according to the fifth embodiment of this application.

[0092] Figures 16A and 16B are scatter plots obtained from the detection of blood samples to be tested according to the fifth embodiment of this application.

[0093] Figures 17A to 17F are correlation curves showing the cell count results obtained from multiple blood samples tested according to this application and the prior art. Detailed Implementation

[0094] The embodiments of this application will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are merely a part of the embodiments of this application, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of this application without inventive effort are within the scope of protection of this application.

[0095] The serial numbers assigned to components in this document, such as "first" and "second," are used only to distinguish the described objects and have no sequential or technical meaning. The terms "connection" and "linkage" used in this application, unless otherwise specified, include both direct and indirect connections (linkages).

[0096] To facilitate subsequent explanations, a brief explanation of some terms used in this application is provided below:

[0097] 1) Scatter plot: A two-dimensional or three-dimensional graph generated by a blood cell analyzer, displaying two-dimensional or three-dimensional feature information of multiple particles. The X, Y, and Z axes of a scatter plot each represent a characteristic of each particle. For example, in a scatter plot, the X axis represents the intensity of forward scattered light, the Y axis represents the fluorescence intensity, and the Z axis represents the intensity of side scattered light.

[0098] The term "scatter plot" as used in this application refers not only to a distribution of at least two sets of data in a rectangular coordinate system in the form of data points, but also to data arrays, i.e., regardless of the form in which they are presented graphically.

[0099] 2) Cell clusters: A group of particles with the same characteristics distributed in a certain area of ​​a scatter plot. It is also called a "particle cluster" or "cell cluster". For example, white blood cell (including all types of white blood cells) clusters, as well as white blood cell subpopulations, such as neutrophil clusters, lymphocyte clusters, monocyte clusters, eosinophil clusters, or basophil clusters.

[0100] 3) Blood shadow: Fragment particles obtained by dissolving red blood cells and platelets in blood with a hemolytic agent.

[0101] The blood cell analyzer used in this application classifies and counts particles in blood samples using flow cytometry, which combines laser scattering and fluorescence staining methods. The principle of the blood cell analyzer in detecting blood samples can be illustrated as follows: First, a blood sample is drawn and treated with a hemolysin and a fluorescent dye. Red blood cells are destroyed and dissolved by the hemolysin, while white blood cells are not dissolved. However, the fluorescent dye can enter the nucleus of white blood cells with the help of the hemolysin and bind to nucleic acid substances in the nucleus. Next, each particle in the sample passes through a detection aperture illuminated by a laser beam. When the laser beam illuminates the particles, the characteristics of the particles themselves (such as volume, staining degree, size and content of cell contents, and nuclear density) can block or change the direction of the laser beam, thereby generating scattered light at various angles corresponding to the particle characteristics. This scattered light is received by a signal detector to obtain information about the particle structure and composition. Forward scatter (FS) reflects the number and volume of particles, side scatter (SS) reflects the complexity of the internal cell structure (such as intracellular particles or the cell nucleus), and fluorescence (FL) reflects the content of nucleic acid substances in the cell. This light information can be used to classify and count cells in blood samples.

[0102] Figure 1 is a schematic diagram of the structure of a sample analyzer according to some embodiments of this application. The sample analyzer 100 includes at least a sample aspiration device 110, a sample preparation device 120, an optical detection device 130, and a processor 140. The sample aspiration device 110 is used to aspirate a blood sample to be tested. The sample preparation device 120 is used to mix at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample. The optical detection device 130 includes a light emitting device, a flow cell, and a photodetector. The light emitting device emits a light beam to irradiate the flow cell, which is connected to the reaction cell and allows individual particles in the test sample to pass through. The photodetector detects the optical information generated when particles in the test sample are irradiated by the light beam as they pass through the flow cell in a single test. Here, the optical information includes at least one scattered light information detected in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye.

[0103] In some embodiments, the sampling device 110 has a sampling needle (not shown) for drawing up a blood sample to be tested. Furthermore, the sampling device 110 may also include, for example, a driving device for driving the sampling needle to quantitatively draw up the blood sample to be tested from the test tube through the tip of the sampling needle. Additionally, the sampling device 110 can also deliver the drawn-up blood sample to the sample preparation device 120.

[0104] In some embodiments, the sample preparation apparatus 120 may include at least one reaction cell and a reagent supply device (not shown). The at least one reaction cell is used to receive a blood sample to be tested drawn by the sampling device 110, and the reagent supply device provides processing reagents (including hemolysin, first dye, second dye, etc.) to the at least one reaction cell, thereby mixing the blood sample to be tested drawn by the sampling device 110 with the processing reagents provided by the reagent supply device in the reaction cell to prepare a test sample.

[0105] In this embodiment, the hemolytic agent can dissolve red blood cells and platelets in a blood sample, but can maintain the morphology of white blood cells essentially unchanged. The fragment particles obtained after the red blood cells and platelets in the blood sample are dissolved form a blood shadow, thereby distinguishing it from the white blood cells and nucleated red blood cells in the blood sample.

[0106] In some embodiments, the hemolytic agent may comprise one or more combinations of cationic surfactants, nonionic surfactants, anionic surfactants, amphiphilic surfactants, and buffer pairs. Cationic surfactants are, for example, selected from at least one or more combinations of dodecyltrimethylammonium chloride, octyltrimethylammonium bromide, and tetradecyltrimethylammonium chloride. Nonionic surfactants are, for example, selected from at least one or more combinations of long-chain fatty alcohol polyoxyethylene, alkylphenol polyoxyethylene ethers, fatty acid polyoxyethylene ethers, and fatty amine polyoxyethylene ethers. Buffer pairs are, for example, selected from at least one or more combinations of phosphates, citrates, and Tris-HCl. Anionic surfactants are, for example, selected from at least one or more combinations of dodecylbenzenesulfonic acid, sodium fatty alcohol acyl sulfate, ethoxylated fatty acid methyl ester sulfonate sodium, secondary alkyl sulfonate sodium, and alcohol ether carboxylates.

[0107] In other embodiments, the hemolytic agent may include at least one of alkyl glycosides, triterpenoid saponins, and steroidal saponins.

[0108] In the embodiments of this application, the dye may be, for example, a fluorescent dye. Because the nucleic acids in leukocytes and nucleated erythrocytes, as well as the cytochromes and riboflavin inside erythrocytes and platelets, have different binding abilities to the fluorescent dye, different fluorescent information will be generated.

[0109] In some embodiments, the absolute value of the wavelength difference corresponding to the peak values ​​of the emission spectra of the first dye and the second dye is greater than 30 nm and less than 80 nm. Alternatively or additionally, the overlap of the emission spectra of the first dye and the second dye is no greater than 50%. By selecting such a first dye and second dye, not only can the mutual detection interference between the first fluorescence information and the second fluorescence information be greatly reduced, i.e., the distinction between the first fluorescence information and the second fluorescence information can be greatly increased, but the size and complexity of the optical detection device will not be increased.

[0110] Figure 2 shows a schematic diagram of the emission spectra of the first dye and the second dye. The dashed line represents the emission spectrum 210 of the first dye, and the solid line represents the emission spectrum 220 of the second dye. The peak point of the emission spectrum 210 of the first dye is A, and the peak point of the emission spectrum 220 of the second dye is D. Here, the absolute difference between the x-coordinates of peak points A and D (i.e., the difference in wavelengths corresponding to the peaks) is greater than 30 nm and less than 80 nm. Furthermore, the overlap between the emission spectrum 210 of the first dye and the emission spectrum 220 of the second dye can be the ratio of the area of ​​the first polygon to the area of ​​the second polygon. The area of ​​the first polygon is equal to the area of ​​the curved polygon enclosed by points E, G, and C, while the area of ​​the second polygon is equal to the area of ​​the curved polygon enclosed by the emission spectrum 210 (or the emission spectrum 220 of the second dye) and the baseline 230. The baseline 230 is a dashed horizontal line parallel to the horizontal axis, as shown in Figure 2, located at 5% of the normalized peak value of the emission spectra 210 and 220 of the first and second dyes, respectively. Points E and F are the left and right intersection points of the emission spectrum 220 of the second dye and the baseline 230, respectively, and points B and C are the left and right intersection points of the emission spectrum 210 of the first dye and the baseline 230, respectively. Here, the overlap between the emission spectra 210 of the first dye and the emission spectrum 220 of the second dye is no greater than 50%.

[0111] Furthermore, the absolute value of the wavelength difference corresponding to the peak values ​​of the emission spectra of the first dye and the second dye is greater than 40 and less than 80 nanometers, preferably greater than 50 nanometers and less than 80 nanometers, and more preferably greater than 50 nanometers and less than 70 nanometers, thereby further reducing the mutual detection interference between the first fluorescence information and the second fluorescence information without increasing the size and complexity of the optical detection device.

[0112] Furthermore, it is advantageous that the overlap of the emission spectra of the first dye and the second dye is no more than 35%, preferably no more than 15%, which can further reduce the detection interference between the first fluorescence information and the second fluorescence information.

[0113] In some embodiments, at least one of the first dye and the second dye, particularly the first dye, may be a large Stokes shift dye. Here, a large Stokes shift dye refers to a dye in which the difference between the wavelengths corresponding to the peaks of the emission spectrum and the excitation spectrum is greater than a predetermined threshold.

[0114] Figure 3 is a schematic spectral diagram of a large Stokes shift dye. The excitation spectrum (also known as the absorption spectrum) 310 of the large Stokes shift dye is shown by a dashed line, and the emission spectrum 320 is shown by a solid line. The peak point of the excitation spectrum 310 is A1, and the peak point of the emission spectrum 320 is A2. The difference between the abscissas of peak points A2 and A1 (i.e., the difference in wavelengths corresponding to the peaks of the emission and excitation spectra) is greater than a predetermined threshold. This predetermined threshold can be, for example, greater than 30 nm and less than 150 nm, preferably greater than 50 nm and less than 100 nm.

[0115] By using at least one large Stokes shift dye, the detection interference between the first fluorescence information and the second fluorescence information can be reduced.

[0116] In some embodiments, the first dye may be a nucleic acid dye capable of being used for leukocyte classification, comprising compounds with the following general structural formula.

[0117] In this general formula:

[0118] X is C(CH3)2, O, S, or Se;

[0119] R1 and R2 are each independently selected from H and C. 1-18 Alkyl, OR6, or halogen;

[0120] R3 is selected from C 1-18 Alkyl, C 1-18 Carboxyl group, C 1-18 Hydroxyl group, C 1-18 NR5R6, benzyl and substituted benzyl, wherein the substituted benzyl is optionally substituted by the following groups: C 1-18 Alkyl, CN, COOH, NH2, NO2, OH, SH, C 1-6 Alkoxy, C 1-6 Alkylamino, C 1-6 Acylamino, halogen or C 1-6 Halogenated alkyl groups;

[0121] R4 is selected from C 1-18 Alkyl, C 1-18 Carboxyl group, C 1-18 hydroxyl, benzyl, and substituted benzyl groups, wherein the substituted benzyl group is optionally substituted by the following groups: C 1-18 Alkyl, CN, COOH, NH2, NO2, OH, SH, C 1-6 Alkoxy, C 1-6 Alkylamino, C 1-6 Acylamino, halogen or C 1-6 Halogenated alkyl groups.

[0122] As some embodiments, the compound used for the first dye has one of the following structures: A, B, C, and D:

[0123] In some other embodiments, the first dye may be acridine orange dye.

[0124] In some embodiments, the second dye may be a fluorescent dye capable of recognizing nucleated erythrocytes, such as a lysosomal dye, comprising compounds with the following general structural formula.

[0125] In this general formula, X is selected from -O-, -S-, and One of them; R1 and R2 are each independently selected from C3-C 20 alkenyl and C3-C 20 Alkynes, preferably R1 and R2, are each independently selected from C3-C4. 10 alkenyl and C3-C 10 Alkynes.

[0126] As some implementations, compounds used for the second dye have one of the following structures: I, II, III, and IV:

[0127] However, this application does not particularly limit the type of lysosomal dye. According to some embodiments, the lysosomal dye is a red light-exciteable lysosomal dye. The term "red light-exciteable" refers to a lysosomal dye that can be excited by light in the red spectrum, such as light in the wavelength range of 600 nm to 800 nm, for example, light with a wavelength of approximately 633 nm, and which binds to lysosomes. According to other embodiments, the lysosomal dye is a blue light-exciteable lysosomal dye. The term "blue light-exciteable" refers to a dye that can be excited by light in the blue spectrum, such as light in the wavelength range of 400 nm to 500 nm, for example, light with a wavelength of approximately 488 nm, and which subsequently emits fluorescence. Those skilled in the art can select a suitable lysosomal dye based on the light source equipped in the specific blood analyzer. For example, suitable lysosomal dyes may include commercially available Lyso-Tracker red, but are not limited to this.

[0128] In some preferred embodiments, the first dye is acridine orange dye and the second dye is a lysosomal dye. In some embodiments, the first dye and / or the second dye is stored in a water-soluble organic phase such as glycerol, glycol, or ethylene glycol.

[0129] In some embodiments, the first dye and / or the second dye may be stored separately or mixed with a hemolysin.

[0130] In some embodiments, the flow chamber in the optical detection device 130 refers to a chamber with a focused liquid flow suitable for detecting light scattering signals and fluorescence signals. When a particle, such as a blood cell, passes through the detection aperture of the flow chamber, the particle scatters the incident light beam from the light source, which is guided through the detection aperture, in various directions. Photodetectors can be positioned at one or more different angles relative to the incident light beam to detect the light scattered by the particle, thereby obtaining a light scattering pulse signal (also known as a light scattering signal). Since different particles have different light scattering characteristics, the light scattering signal can be used to distinguish different groups of particles.

[0131] Specifically, the light scattering signal detected near the incident light beam is generally referred to as the forward light scattering signal or the small-angle light scattering signal. In some embodiments, the forward light scattering signal can be detected at an angle of about 1° to about 10° with respect to the incident light beam. In other embodiments, the forward light scattering signal can be detected at an angle of about 2° to about 6° with respect to the incident light beam. The light scattering signal detected at about 90° with respect to the incident light beam is generally referred to as the side light scattering signal. In some embodiments, the side light scattering signal can be detected at an angle of about 65° to about 115° with respect to the incident light beam. Typically, the fluorescent signal emitted from blood cells stained with fluorescent dye is also generally detected at about 90° with respect to the incident light beam.

[0132] In some embodiments, the photodetector in the optical detection device 130 may include a scattering light detector for detecting scattered light pulse signals (also known as scattered light information), such as side-scattered light pulse signals (also known as side-scattered light information), and a first fluorescence detector for detecting a first fluorescence pulse signal (also known as first fluorescence information) and a second fluorescence detector for detecting a second fluorescence pulse signal (also known as second fluorescence information).

[0133] In some embodiments, the optical detection device 130 includes a forward-scattering detector for detecting forward-scattering light information or a side-scattering detector for detecting side-scattering light information. Preferably, the optical detection device 130 includes both a forward-scattering detector and a side-scattering detector.

[0134] Figure 4 shows a specific example of an optical detection device 130. This optical detection device 130 includes a light emitting device 131, a front light assembly 132, a flow chamber 133, a forward-scattering light detector 134, a first dichroic mirror 135, a side-scattering light detector 136, a second dichroic mirror 137, a first fluorescence detector 138, and a second fluorescence detector 139. The first fluorescence detector 138 detects a first fluorescence signal corresponding to a first dye generated by particles passing through the flow chamber 133 after being irradiated by a light beam. The second fluorescence detector 139 detects a second fluorescence signal corresponding to a second dye generated by particles passing through the flow chamber 133 after being irradiated by a light beam. Here, the light emitting device 131, the front light assembly 132, the flow chamber 133, and the forward-scattering light detector 134 are arranged sequentially along the optical axis. The front light assembly is configured to converge the light beam emitted by the light emitting device 131 into the detection area of ​​the flow chamber 133 in the particle flow direction, so that particles flowing through the detection area of ​​the flow chamber 133 can generate scattered light. On one side of the flow chamber 133, a first dichroic mirror 135 is arranged at a 45° angle to the optical axis. A portion of the lateral light generated when particles flow through the detection area of ​​the flow chamber 133 is reflected by the first dichroic mirror 135 and captured by the side-scattering light detector 136, while another portion of the lateral light passes through the first dichroic mirror 135 and reaches a second dichroic mirror 137, which is also arranged downstream of the first dichroic mirror 135 at a 45° angle to the optical axis. A portion of the lateral light passing through the first dichroic mirror 135 is reflected by the second dichroic mirror 137 and captured by the first fluorescence detector 138, while another portion passes through the second dichroic mirror 137 and is captured by the second fluorescence detector 139.

[0135] In some embodiments, the light emitting device includes a first light source and a second light source. The first light source is used to emit a light beam of a first wavelength capable of exciting the first dye, and the second light source is used to emit a light beam of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength.

[0136] In some implementations, the first wavelength is between 315 nm and 490 nm, and the second wavelength is between 610 nm and 750 nm. Preferably, the first wavelength is between 400 nm and 450 nm, and the second wavelength is between 620 nm and 700 nm.

[0137] In some alternative embodiments, the light emitting device is configured to emit a single-wavelength light beam capable of exciting the first and second dyes. That is, in the optical detection apparatus 130 of this application, the test sample in the flow chamber is irradiated with a single-wavelength excitation light; that is, the optical information is generated by each particle in the test sample being irradiated by the single-wavelength excitation light as it passes through the optical detection area of ​​the optical detection device. For example, the light emitting device may be a laser emitting blue-green or red light, such as a laser emitting light with a wavelength of 488 or 520 nanometers.

[0138] In some embodiments, the processor 140 is used to process and perform calculations on the data to obtain the desired results, such as generating a two-dimensional or three-dimensional scatter plot based on the collected optical information, and performing particle analysis on the scatter plot according to the gating method.

[0139] In some embodiments, the processor 140 may visualize intermediate or final processing results and then display them through the display device 150. For example, the display device 150 may include a user interface, and the processor 140 may display the processing results on the user interface of the display device 150.

[0140] In some embodiments, the processor 140 includes, but is not limited to, devices such as a central processing unit (CPU), a microcontroller unit (MCU), a field-programmable gate array (FPGA), and a digital signal processor (DSP) used to interpret computer instructions and process data in computer software. For example, the processor is used to execute various computer applications in a computer-readable storage medium, thereby enabling the sample analyzer 100 to execute corresponding detection procedures and analyze the scattered light information and fluorescence information detected by the optical detection device 130 in real time.

[0141] In addition, the sample analyzer 100 may also include a liquid path system (not shown) for connecting the sample aspiration device 110, the sample preparation device 120 and the optical detection device 130 to facilitate liquid transfer between these devices.

[0142] Furthermore, the sample analyzer 100 may also include a first housing 160 and a second housing 170. An optical detection device 130 and a processor 140 are disposed inside the second housing 170. A sample preparation device 120 is disposed, for example, inside the first housing 160, and a display device 150 is disposed, for example, on the outer surface of the first housing 160 and is used to display the detection results of the sample analyzer 100.

[0143] The following describes how to obtain the five-part differential white blood cell count and nucleated red blood cell count through a single reaction and detection channel.

[0144] In order to obtain the five-part differential white blood cell count and nucleated red blood cell count in a single test, this application proposes to process and detect the same blood sample using two fluorescent dyes under hemolytic conditions, and then obtain the results based on the optical information obtained from the same test on the same processed blood sample.

[0145] Therefore, in some embodiments of this application, the processor 140 is configured to:

[0146] A scatter plot is generated based on at least one scattered light information, a first fluorescence information, and a second fluorescence information detected in a single test of the test sample, and the white blood cell five-part differential result and the nucleated red blood cell (NRBC) count result of the test sample are obtained based on the scatter plot. The white blood cell five-part differential result includes the count results of neutrophils (neu), lymphocytes (lym), monocytes (mon), eosinophils (eos), and basophils (baso) in the test sample.

[0147] In the embodiments of this application, the generated scatter plot can be a two-dimensional scatter plot or a three-dimensional scatter plot.

[0148] In some embodiments, the light emitting device includes a first light source and a second light source. The first light source emits a light beam of a first wavelength capable of exciting a first dye, while the second light source emits a light beam of a second wavelength capable of exciting a second dye, wherein the second wavelength is greater than the first wavelength. Here, the first dye is acridine orange dye and the second dye is a lysosomal dye. This allows for more accurate white blood cell classification results and nucleated red blood cell count results.

[0149] As some implementation methods, the processor 140 generates a scatter plot based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, and obtains the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, including:

[0150] A first scatter plot is generated based on at least one type of scattered light information, such as side-scattered light information and first fluorescence information, such as the two-dimensional first scatter plot shown in Figure 11A, and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot.

[0151] A second scatter plot is generated based on at least one type of scattered light information, such as side-scattered light information and second fluorescence information. For example, a two-dimensional second scatter plot as shown in 11B is generated. Based on this second scatter plot, nucleated red blood cells in the test sample are identified, and the count result of the nucleated red blood cell population is obtained.

[0152] The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

[0153] In other words, according to the embodiments of this application, basophils and nucleated red blood cells in the test sample can be accurately distinguished, so that the white blood cell five-part differential result and nucleated red blood cell count result of the test sample can be obtained through a single reaction and detection channel.

[0154] In some embodiments, the scattered light information includes first lateral scattered light information generated by particles passing through the flow chamber after being irradiated by a beam of light from a first light source, such as blue-violet light (i.e., a first wavelength between 315 nm and 490 nm), and / or second lateral scattered light information generated by particles passing through the flow chamber after being irradiated by a beam of light from a second light source, such as red light (i.e., a second wavelength between 620 nm and 700 nm). Here, the generation of a scatter plot based on at least one scattered light information, first fluorescence information, and second fluorescence information, and the obtaining of the white blood cell five-part differential result and nucleated red blood cell count result of the test sample based on the scatter plot, executed by the processor 140, includes:

[0155] Based on first side-scattered light information, such as first side-scattered light information and first fluorescence information obtained by blue-violet light excitation, and / or based on second side-scattered light information, such as second side-scattered light information and first fluorescence information obtained by red light excitation, a first scatter plot, especially a two-dimensional first scatter plot, is generated, and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot;

[0156] Based on first lateral scattered light information, such as first scattered light information obtained from blue-violet light excitation and second fluorescence information, and / or based on second lateral scattered light information, such as second lateral scattered light information obtained from red light excitation and second fluorescence information, a second scatter plot, especially a two-dimensional second scatter plot, is generated, and nucleated red blood cells in the test sample are identified based on the second scatter plot to obtain the counting result of the nucleated red blood cell population; and

[0157] The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

[0158] In some embodiments, at least one type of scattered light information includes lateral scattered light information. In this case, the processor 140 performs the following steps: generating a scatter plot based on at least one type of scattered light information, first fluorescence information, and second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot. This includes:

[0159] The first scatter plot is generated based on the side-scattered light information and the first fluorescence information. A first characteristic cell group RG1, including basophilic granulocytes and nucleated erythrocytes, is divided in the first scatter plot, and the first particle information of the first characteristic cell group is obtained.

[0160] The second scatter plot is generated based on the side-scattered light information and the second fluorescence information. The second characteristic cell group RG2, which only includes nucleated erythrocytes, is divided in the second scatter plot and the second particle information of the second characteristic cell group is obtained.

[0161] The count of nucleated erythrocytes in the test sample was obtained based on the second particle information; and

[0162] The basophil population count in the test sample is obtained based on the first particle information and the second particle information.

[0163] Further, the first particle information may include a first cell count of a first characteristic cell population, and the second particle information may include a second cell count of a second characteristic cell population. In this case, the process executed by the processor 140 to obtain the basophil count in the test sample based on the first and second particle information may include: subtracting the second cell count from the first cell count to obtain the basophil count in the test sample, i.e., the basophil count in the test sample = first cell count - second cell count.

[0164] Here, the second cell count is the count of nucleated red blood cells in the test sample.

[0165] In other embodiments, at least one type of scattered light information includes side-scattered light information. In this case, the processor 140 performs the following steps: generating a scatter plot based on at least one type of scattered light information, first fluorescence information, and second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot.

[0166] The first scatter plot is generated based on the side-scattered light information and the first fluorescence information. This first scatter plot is used to delineate a first characteristic cell population, including basophils and nucleated erythrocytes.

[0167] The basophil count in the test sample is obtained based on at least second fluorescence information of cells in the first characteristic cell population, such as second fluorescence information only.

[0168] As other implementations, the processor 140 generates a scatter plot based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, and obtains the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, including:

[0169] At least one type of scattered light information, first fluorescence information, and second fluorescence information are used to generate a three-dimensional scatter plot, and based on the three-dimensional scatter plot, the five-part differential white blood cell count and the nucleated red blood cell count of the test sample are obtained.

[0170] This application also proposes a sample analysis method capable of obtaining white blood cell five-part differential and nucleated red blood cell count results through a single reaction and detection channel, which is implemented, in particular, by the sample analyzer described above or one of its embodiments.

[0171] The sample analysis method according to the embodiments of this application includes:

[0172] Collect the blood sample to be tested;

[0173] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0174] In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by light. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye; and

[0175] The white blood cell differential and nucleated red blood cell count results of the test sample are obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information. The white blood cell differential results include the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample.

[0176] As some implementation methods are shown in FIG5, the sample analysis method 200 according to the embodiment of this application includes the following steps S210, S220, S230, S240 and S250.

[0177] In step S210, a blood sample to be tested is collected or aspirated. For example, a portion of the blood sample to be tested is aspirated from a test tube containing the blood sample to be tested.

[0178] In step S220, at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare a test sample.

[0179] For example, in step S220, the hemolysin, the first dye, and the second dye can be added sequentially to the same blood sample to obtain a test sample. The test sample is then incubated to allow the dye to fully stain the particles in the test sample. Alternatively, in step S220, the first dye and the second dye can be pre-mixed with the hemolysin to obtain a mixed reagent. This mixed reagent is then mixed with the blood sample to be tested at a volume ratio of 250:1 to 1000:1. After thorough mixing, the resulting test sample is incubated at 25°C to 50°C for 10 seconds to 1 minute, preferably 20 seconds to 40 seconds.

[0180] Examples of the hemolytic agent and dye used in step S220 can be found in the above description and will not be repeated here.

[0181] In step S230, each particle in the test sample is passed through the optical detection area irradiated by the excitation light in a single test to obtain the optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test.

[0182] Here, the first fluorescence information includes the fluorescence signal generated by photoexcitation after the particles in the test sample bind to the first dye, while the second fluorescence information includes the fluorescence signal generated by photoexcitation after the particles in the test sample bind to the second dye.

[0183] In other words, in step S230, the scattered light information and fluorescence information of the test sample are obtained based on the principle of flow cytometry. In the embodiments of this application, the scattered light information includes the scattered light signal intensity, and the fluorescence information includes the fluorescence signal intensity.

[0184] In step S240, a scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the scatter plot. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

[0185] In some embodiments, the excitation light may include light capable of exciting a first dye at a first wavelength and light capable of exciting a second dye at a second wavelength, wherein the second wavelength is longer than the first wavelength. Preferably, the first dye is acridine orange dye and the second dye is a lysosomal dye. This allows for more accurate white blood cell classification results and nucleated red blood cell count results.

[0186] Furthermore, the first wavelength can be between 315 nm and 490 nm, and the second wavelength is between 620 nm and 700 nm. That is, the excitation light includes blue-violet light and red light.

[0187] In other embodiments, the excitation light is a single wavelength of light capable of exciting the first dye and the second dye.

[0188] As some implementations, step S240 may include:

[0189] A first scatter plot is generated based on at least one type of scattered light information, such as scattered light information and first fluorescence information, for example, a two-dimensional first scatter plot as shown in Figure 11A, and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot;

[0190] A second scatter plot is generated based on at least one type of scattered light information, such as scattered light information and second fluorescence information. For example, a two-dimensional second scatter plot as shown in Figure 11B is generated. Based on this second scatter plot, nucleated red blood cells in the test sample are identified, and the count result of the nucleated red blood cell population is obtained.

[0191] The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

[0192] In some embodiments, when the excitation light includes light of a first wavelength capable of exciting a first dye and light of a second wavelength capable of exciting a second dye, for example, when the excitation light includes blue-violet light and red light, the scattered light information includes first lateral scattered light information generated by particles passing through the optical detection area after being irradiated by light of the first wavelength, i.e., blue-violet light, and / or second lateral scattered light information generated by particles passing through the optical detection area after being irradiated by light of the second wavelength, i.e., red light, obtained in the single test.

[0193] Accordingly, step S240 may include:

[0194] Based on the first side-scattered light information and the first fluorescence information, and / or based on the second side-scattered light information and the first fluorescence information, a first scatter plot is generated and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot;

[0195] Based on the first lateral scattering light information and the second fluorescence information, and / or based on the second lateral scattering light information and the second fluorescence information, a second scatter plot is generated, and based on the second scatter plot, nucleated red blood cells in the test sample are identified, and the counting result of the nucleated red blood cell population is obtained; and

[0196] The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

[0197] In some embodiments, at least one type of scattered light information includes side-scattered light information. Accordingly, step S240 may include:

[0198] The first scatter plot is generated based on the side-scattered light information and the first fluorescence information. A first characteristic cell group RG1, including basophilic granulocytes and nucleated erythrocytes, is divided in the first scatter plot, and the first particle information of the first characteristic cell group is obtained.

[0199] The second scatter plot is generated based on the side-scattered light information and the second fluorescence information. The second characteristic cell group RG2, which only includes nucleated erythrocytes, is divided in the second scatter plot and the second particle information of the second characteristic cell group is obtained.

[0200] The count of nucleated erythrocytes in the test sample was obtained based on the second particle information; and

[0201] The basophil population count in the test sample is obtained based on the first particle information and the second particle information.

[0202] Further, the first particle information may include a first cell count of a first characteristic cell population, and the second particle information may include a second cell count of a second characteristic cell population. In this case, obtaining the basophil count in the test sample based on the first and second particle information may include subtracting the second cell count from the first cell count to obtain the basophil count in the test sample. Here, the second cell count is the count of nucleated erythrocytes in the test sample.

[0203] In other embodiments, at least one type of scattered light information includes side-scattered light information. In this case, step S240 may include:

[0204] The first scatter plot is generated based on the side-scattered light information and the first fluorescence information. This first scatter plot is used to delineate a first characteristic cell population, including basophils and nucleated erythrocytes.

[0205] The basophil count in the test sample is obtained based on at least second fluorescence information of cells in the first characteristic cell population, such as second fluorescence information only.

[0206] As in some other implementations, step S240 may include: generating a three-dimensional scatter plot based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the three-dimensional scatter plot.

[0207] The following describes how, in an embodiment of the light emitting device including a first light source and a second light source, white blood cell four-part differential and nucleated red blood cell count results are obtained through a reaction and detection channel.

[0208] In order to obtain white blood cell four-part differential and nucleated red blood cell count results in a single test, this application proposes to process and detect the same blood sample using two fluorescent dyes under hemolytic conditions, and then obtain the results based on the optical information obtained from the same test on the same processed blood sample.

[0209] Therefore, in some embodiments of this application, the processor 140 is configured to:

[0210] A scatter plot is generated based on at least one scattered light information, a first fluorescence information, and a second fluorescence information detected in a single test of the test sample, and the white blood cell four-part differential result and the nucleated red blood cell (NRBC) count result of the test sample are obtained based on the scatter plot, wherein the white blood cell four-part differential result includes the count results of neutrophils (neu), lymphocytes (lym), monocytes (mon), and eosinophils (eos) in the test sample.

[0211] In some embodiments, the light emitting device includes a first light source and a second light source. The first light source emits a light beam of a first wavelength capable of exciting a first dye, while the second light source emits a light beam of a second wavelength capable of exciting a second dye, wherein the second wavelength is greater than the first wavelength. Here, the first dye is acridine orange dye and the second dye is a lysosomal dye. This allows for more accurate white blood cell classification results and nucleated red blood cell count results.

[0212] As some implementation methods, the processor 140 generates a scatter plot based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, and obtains the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, including:

[0213] A first scatter plot is generated based on at least one type of scattered light information, such as side-scattered light information and first fluorescence information. For example, a two-dimensional first scatter plot as shown in Figure 14A is generated, and the white blood cell four-part differential result of the test sample is obtained based on the first scatter plot.

[0214] A second scatter plot is generated based on at least one type of scattered light information, such as side-scattered light information and second fluorescence information. For example, a two-dimensional second scatter plot as shown in 14B is generated, and the count result of the nucleated red blood cell population in the test sample is obtained based on the second scatter plot.

[0215] In some examples, the scattered light information includes first lateral scattered light information generated by particles passing through the flow chamber after being irradiated by a beam of light from a first light source, such as blue-violet light, and / or second lateral scattered light information generated by particles passing through the flow chamber after being irradiated by a beam of light from a second light source, such as red light, detected in the single test. Here, the generation of a scatter plot based on at least one scattered light information, first fluorescence information, and second fluorescence information, and the obtaining of the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, executed by the processor 140, includes:

[0216] Based on first side-scattered light information, such as first side-scattered light information and first fluorescence information obtained by blue-violet light excitation, and / or based on second side-scattered light information, such as second side-scattered light information and first fluorescence information obtained by red light excitation, a first scatter plot, especially a two-dimensional first scatter plot, is generated, and the white blood cell four-part differential result of the test sample is obtained based on the first scatter plot; and

[0217] Based on the first side-scattered light information, such as the first scattered light information and the second fluorescence information obtained by blue-violet light excitation, and / or based on the second side-scattered light information, such as the second side-scattered light information and the second fluorescence information obtained by red light excitation, a second scatter plot, especially a two-dimensional second scatter plot, is generated, and the count result of the nucleated red blood cell population in the test sample is obtained based on the second scatter plot.

[0218] As other implementations, the processor 140 generates a scatter plot based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, and obtains the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, including:

[0219] A three-dimensional scatter plot is generated based on at least one type of scattered light information, a first fluorescence information, and a second fluorescence information, and the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the three-dimensional scatter plot.

[0220] In some embodiments, the processor 140 may also be configured to obtain the basophil count in the test sample based on at least one scattered light information, a first fluorescence information, and a second fluorescence information. That is, according to the embodiments of this application, basophils and nucleated red blood cells in the test sample can be accurately distinguished, thereby enabling the acquisition of the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample through a single reaction and detection channel, as described above.

[0221] As some implementations, at least one type of scattered light information includes side-scattered light information. In this case, the processor 140 executes a process to obtain the count results of nucleated erythrocytes and basophils in the test sample based on at least one type of scattered light information, first fluorescence information, and second fluorescence information, including:

[0222] A first scatter plot is generated based on the side-scattered light information and the first fluorescence information, especially a two-dimensional first scatter plot as shown in Figure 11A. In the first scatter plot, a first characteristic cell group RG1, including basophilic granulocytes and nucleated erythrocytes, is divided and the first particle information of the first characteristic cell group is obtained.

[0223] A second scatter plot is generated based on the side-scattered light information and the second fluorescence information, especially a two-dimensional second scatter plot as shown in Figure 11B. In the second scatter plot, a second characteristic cell group RG2, which only includes nucleated erythrocytes, is divided and the second particle information of the second characteristic cell group is obtained.

[0224] The count of nucleated erythrocytes in the test sample was obtained based on the second particle information; and

[0225] The basophil population count in the test sample is obtained based on the first particle information and the second particle information.

[0226] Further, the first particle information may include a first cell count of a first characteristic cell population, and the second particle information may include a second cell count of a second characteristic cell population. In this case, the process executed by the processor 140 to obtain the basophil count in the test sample based on the first and second particle information may include: subtracting the second cell count from the first cell count to obtain the basophil count in the test sample, i.e., the basophil count in the test sample = first cell count - second cell count.

[0227] Here, the second cell count is the count of nucleated red blood cells in the test sample.

[0228] In some other implementations, at least one type of scattered light information includes side-scattered light information. In this case, the processor 140 executes a process to obtain the basophil population count in the test sample based on at least one type of scattered light information, first fluorescence information, and second fluorescence information, including:

[0229] A first scatter plot is generated based on lateral scattering light information and first fluorescence information. This first scatter plot is used to delineate a first characteristic cell population, including basophils and nucleated erythrocytes.

[0230] The basophil count in the test sample is obtained based on at least second fluorescence information of cells in the first characteristic cell population, such as second fluorescence information only.

[0231] This application also proposes a sample analysis method capable of obtaining white blood cell four-part differential and nucleated red blood cell count results through a single reaction and detection channel, which is implemented, in particular, by the sample analyzer described above or one of its embodiments.

[0232] The sample analysis method according to the embodiments of this application includes:

[0233] Collect the blood sample to be tested;

[0234] At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample;

[0235] In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles after irradiation. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. The excitation light includes light of a first wavelength capable of exciting the first dye and light of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength.

[0236] The white blood cell differential and nucleated red blood cell count results of the test sample are obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information. The white blood cell differential results include the count results of neutrophils, lymphocytes, monocytes and eosinophils in the test sample.

[0237] In some embodiments, the first dye is acridine orange dye and the second dye is a lysosomal dye. This allows for more accurate white blood cell classification results and nucleated red blood cell count results.

[0238] As some implementation methods are shown in FIG6, the sample analysis method 300 according to the embodiment of this application includes the following steps S310, S320, S330, S340 and S350.

[0239] In step S310, a blood sample to be tested is collected or aspirated. For example, a portion of the blood sample to be tested is aspirated from a test tube containing the blood sample to be tested.

[0240] In step S320, at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare a test sample.

[0241] For example, in step S320, the hemolysin, the first dye, and the second dye can be added sequentially to the same blood sample to obtain a test sample. The test sample is then incubated to allow the dye to fully stain the particles in the test sample. Alternatively, in step S320, the first dye and the second dye can be pre-mixed with the hemolysin to obtain a mixed reagent. This mixed reagent is then mixed with the blood sample to be tested at a volume ratio of 250:1 to 1000:1. After thorough mixing, the resulting test sample is incubated at 25°C to 50°C for 10 seconds to 1 minute, preferably 20 seconds to 40 seconds.

[0242] Examples of the hemolytic agent and dye used in step S320 can be found in the above description and will not be repeated here.

[0243] In step S330, in a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by excitation light. This optical information includes at least one type of scattered light information obtained in the single test, first fluorescence information corresponding to the first dye, and second fluorescence information corresponding to the second dye. Here, the excitation light includes light of a first wavelength capable of exciting the first dye and light of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength.

[0244] Here, the first fluorescence information includes the fluorescence signal generated by photoexcitation after the particles in the test sample bind to the first dye, while the second fluorescence information includes the fluorescence signal generated by photoexcitation after the particles in the test sample bind to the second dye.

[0245] Preferably, the first dye is acridine orange dye and the second dye is a lysosomal dye.

[0246] In other words, in step S330, the scattered light information and fluorescence information of the test sample are obtained based on the principle of flow cytometry. In the embodiments of this application, the scattered light information includes the scattered light signal intensity, and the fluorescence information includes the fluorescence signal intensity.

[0247] In step S340, a scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the scatter plot. The white blood cell four-part differential result includes the count result of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

[0248] As some implementations, step S340 may include:

[0249] A first scatter plot is generated based on at least one type of scattered light information, such as scattered light information and first fluorescence information, for example, a two-dimensional first scatter plot as shown in Figure 14A, and the white blood cell four-part differential result of the test sample is obtained based on the first scatter plot; and

[0250] A second scatter plot is generated based on at least one type of scattered light information, such as scattered light information and second fluorescence information, for example, a two-dimensional second scatter plot as shown in Figure 14B, and the count result of the nucleated red blood cell population in the test sample is obtained based on the second scatter plot.

[0251] In some examples, the first wavelength is between 315 nm and 490 nm, and the second wavelength is between 620 nm and 700 nm. That is, the laser light includes blue-violet light and red light. Here, the scattered light information includes first lateral scattered light information generated by particles passing through the optical detection area after being irradiated with light of the first wavelength, i.e., blue-violet light, and / or second lateral scattered light information generated by particles passing through the optical detection area after being irradiated with light of the second wavelength, i.e., red light, acquired in the single test.

[0252] Accordingly, step S340 may include:

[0253] Based on the first side-scattered light information and the first fluorescence information, and / or based on the second side-scattered light information and the first fluorescence information, a first scatter plot is generated, and the white blood cell four-part differential result of the test sample is obtained based on the first scatter plot; and

[0254] Based on the first lateral scattering light information and the second fluorescence information, and / or based on the second lateral scattering light information and the second fluorescence information, a second scatter plot is generated, and the count result of the nucleated red blood cell population in the test sample is obtained based on the second scatter plot.

[0255] As in some other implementations, step S340 may include: generating a three-dimensional scatter plot based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, and obtaining the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample based on the three-dimensional scatter plot.

[0256] Furthermore, the sample analysis method 300 may also include: obtaining the count result of the basophil population in the test sample based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information.

[0257] In some embodiments, at least one type of scattered light information includes side-scattered light information. Accordingly, obtaining the count results of nucleated erythrocytes and basophils in the test sample based on at least one type of scattered light information, first fluorescence information, and second fluorescence information may include:

[0258] A first scatter plot is generated based on the side-scattered light information and the first fluorescence information, especially a two-dimensional first scatter plot as shown in Figure 8A. In the first scatter plot, a first characteristic cell group RG1, including basophilic granulocytes and nucleated erythrocytes, is divided and the first particle information of the first characteristic cell group is obtained.

[0259] A second scatter plot, especially a two-dimensional second scatter plot as shown in Figure 8B, is generated based on the side-scattered light information and the second fluorescence information. In this second scatter plot, a second characteristic cell group RG2, which includes only nucleated red blood cells, is divided and the second particle information of this second characteristic cell group is obtained.

[0260] The count of nucleated erythrocytes in the test sample was obtained based on the second particle information; and

[0261] The basophil population count in the test sample is obtained based on the first particle information and the second particle information.

[0262] Further, the first particle information may include a first cell count of a first characteristic cell population, and the second particle information may include a second cell count of a second characteristic cell population. In this case, obtaining the basophil count in the test sample based on the first and second particle information may include subtracting the second cell count from the first cell count to obtain the basophil count in the test sample. Here, the second cell count is the count of nucleated erythrocytes in the test sample.

[0263] In other embodiments, at least one type of scattered light information includes side-scattered light information. In this case, obtaining the basophil population count in the test sample based on at least one type of scattered light information, a first fluorescence information, and a second fluorescence information includes:

[0264] A first scatter plot is generated based on lateral scattering light information and first fluorescence information. This first scatter plot is used to delineate a first characteristic cell population, including basophils and nucleated erythrocytes.

[0265] The basophil count in the test sample is obtained based on at least second fluorescence information of cells in the first characteristic cell population, such as second fluorescence information only.

[0266] The following describes how to obtain accurate five-part differential white blood cell count results through a single reaction and detection channel.

[0267] To achieve accurate five-part differential white blood cell count results in a single test, this application proposes processing and detecting the same blood sample with two fluorescent dyes under hemolytic conditions, and then obtaining the results based on optical information obtained from the same test on the same processed blood sample.

[0268] In a first embodiment of the sample analyzer according to this application, the light emitting device emits a light beam that includes at least a first wavelength capable of exciting the first dye, the first wavelength being between 315 nm and 490 nm, particularly between 400 nm and 450 nm. In this first embodiment, the processor 140 is configured to:

[0269] The white blood cell four-part differential result of the test sample is obtained based on at least one scattered light information and first fluorescence information detected in a single test of the test sample. This white blood cell four-part differential result includes the counts of neutrophils (neu), lymphocytes (lym), monocytes (mon), and eosinophils (eos) in the test sample.

[0270] The basophil population (baso) count in the test sample is obtained based on at least one scattered light information, a first fluorescence information, and a second fluorescence information detected in the single test.

[0271] In some embodiments, the processor 140 is further configured to acquire the count of nucleated erythrocyte populations (NRBCs) in the test sample based on at least one type of scattered light information and a second type of fluorescence information detected in the single test.

[0272] As some implementations, the processor 140 may be configured to: generate a first scatter plot, particularly a two-dimensional first scatter plot, based on at least one type of scattered light information, particularly side-scattered light information and first fluorescence information detected in a single test of the test sample, and perform a four-category classification of white blood cells based on the first scatter plot, that is, to identify and count the neutrophil population, lymphocyte population, monocyte population and eosinophil population in the test sample.

[0273] In some implementations, the light emitting device includes a first light source and a second light source. The first light source emits a light beam of a first wavelength capable of exciting the first dye, while the second light source emits a light beam of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength. In this case, the scattered light information includes first lateral scattered light information generated by particles passing through the flow chamber after being irradiated by the light beam of the first light source and / or second lateral scattered light information generated by particles passing through the flow chamber after being irradiated by the light beam of the second light source, as detected in the single test. In some embodiments, the first dye is acridine orange dye and the second dye is a lysosomal dye. This allows for more accurate white blood cell classification results. Here, the acquisition of the four-part white blood cell classification result of the test sample based on at least one scattered light information and first fluorescence information, executed by the processor 140, includes: obtaining the four-part white blood cell classification result of the test sample based on the first lateral scattered light information and the first fluorescence information, and / or based on the second lateral scattered light information and the first fluorescence information. For example, processor 140 generates a first scatter plot, particularly a two-dimensional first scatter plot, based on first side-scattered light information and first fluorescence information, or based on second side-scattered light information and first fluorescence information, and obtains the four-part differential white blood cell count of the test sample based on the first scatter plot, as shown in Figures 11A, 12A, and 13A. Furthermore, the process executed by processor 140 to obtain the basophil count in the test sample based on at least one scattered light information, first fluorescence information, and second fluorescence information includes: obtaining the basophil count in the test sample based on first side-scattered light information, first fluorescence information, and second fluorescence information, and / or, based on second side-scattered light information, first fluorescence information, and second fluorescence information.

[0274] The inventors of this application discovered through research that when nucleated red blood cells are present in the blood sample to be tested, it is difficult to distinguish between nucleated red blood cells and basophils using only scattered light information and first fluorescence information. In other words, an accurate basophil count cannot be obtained using only scattered light information and first fluorescence information. Furthermore, the inventors of this application discovered through research that nucleated red blood cells and basophils can be accurately distinguished using scattered light information, first fluorescence information, and third fluorescence information.

[0275] Therefore, as one implementation, the process executed by the processor 140 to obtain the counting result of the basophil population in the test sample based on at least one scattered light information, a first fluorescence information, and a second fluorescence information includes: distinguishing between the basophil population and the nucleated erythrocyte population in the test sample based on at least one scattered light information, a first fluorescence information, and a second fluorescence information, so as to identify the basophil population in the test sample and count the basophil population in the test sample, thereby obtaining the counting result of the basophil population in the test sample.

[0276] In some embodiments, at least one type of scattered light information includes side-scattered light information. In this case, the process executed by processor 140 to obtain the basophil population count result in the test sample based on at least one type of scattered light information, first fluorescence information, and second fluorescence information includes:

[0277] The first particle information of the first characteristic cell group is obtained based on the side-scattered light information and the first fluorescence information, for example as shown in Figures 11A, 12A and 13A. A two-dimensional first scatter plot is generated based on the side-scattered light information and the first fluorescence information, and the first particle information of the first characteristic cell group RG1 is obtained based on the first scatter plot.

[0278] The second particle information of the second characteristic cell population is obtained based on the side-scattered light information and the second fluorescence information, as shown in Figures 11B, 12B and 13B. A two-dimensional second scatter plot RG2 is generated based on the side-scattered light information and the second fluorescence information, and the first particle information of the second characteristic cell population is obtained based on the second scatter plot.

[0279] The basophil count in the test sample is obtained based on the first particle information and the second particle information.

[0280] Furthermore, the first particle information of the first characteristic cell group, executed by the processor 140 based on the side-scattered light information and the first fluorescence information, may include: dividing the first characteristic cell group, which includes basophils and nucleated erythrocytes, based on the side-scattered light information and the first fluorescence information, and obtaining the first particle information of the first characteristic cell group; and the second particle information of the second characteristic cell group, executed by the processor 140 based on the side-scattered light information and the second fluorescence information, may accordingly include: dividing the second characteristic cell group, which includes only nucleated erythrocytes, based on the side-scattered light information and the second fluorescence information, and obtaining the second particle information of the second characteristic cell group.

[0281] For example, the processor 140 generates a two-dimensional first scatter plot based on side-scattered light information and first fluorescence information, divides a first characteristic cell group RG1 including basophils and nucleated red blood cells in the first scatter plot and obtains the first particle information of the first characteristic cell group, and the processor 140 generates a second scatter plot based on side-scattered light information and second fluorescence information, divides a second characteristic cell group RG2 including only nucleated red blood cells in the second scatter plot and obtains the second particle information of the second characteristic cell group.

[0282] Further, the first particle information may include a first cell count of the first characteristic cell population, and the second particle information may include a second cell count of the second characteristic cell population. In this case, the process executed by the processor 140 to obtain the basophil count in the test sample based on the first and second particle information may include: subtracting the second cell count from the first cell count to obtain the basophil count in the test sample, i.e., the basophil count in the test sample = first cell count - second cell count.

[0283] Here, the second cell count is the count of nucleated red blood cells in the test sample.

[0284] In some alternative embodiments, at least one type of scattered light information includes side-scattered light information. In this case, the process executed by processor 140 to obtain the basophil population count result in the test sample based on at least one type of scattered light information, first fluorescence information, and second fluorescence information includes:

[0285] The first characteristic cell population is obtained based on the side-scattered light information and the first fluorescence information. For example, a two-dimensional first scatter plot is generated based on the side-scattered light information and the first fluorescence information, and the first characteristic cell population, especially the first characteristic cell population including basophils and nucleated erythrocytes, is divided in the first scatter plot; and

[0286] The basophil count in the test sample is obtained based on at least the second fluorescence information of cells in the first characteristic cell population.

[0287] In a second embodiment of the sample analyzer according to this application, the processor 140 is configured to:

[0288] The white blood cell five-part differential result of the test sample is obtained based on at least one scattered light information and first fluorescence information. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample.

[0289] The basophil count is corrected based on at least a second fluorescence information to obtain a corrected basophil count.

[0290] As described above, the inventors of this application discovered through research that nucleated red blood cells in the blood sample to be tested interfere with basophil counts.

[0291] Therefore, as some implementations, the correction of the basophil population counting results based on at least second fluorescence information performed by the processor 140 includes:

[0292] Based on the second fluorescence information, the nucleated erythrocyte population within the basophil population is identified and counted to obtain the nucleated erythrocyte population count result; and

[0293] The count of the basophil population is corrected based on the count of the nucleated erythrocyte population.

[0294] In one example, when at least one type of scattered light information includes side-scattered light information, the correction of the basophil population counting result based on at least a second fluorescence information performed by the processor 140 includes:

[0295] The first cell count of the basophil population in the test sample was obtained based on the side-scattered light information and the first fluorescence information;

[0296] The second cell count of nucleated erythrocytes in the test sample was obtained based on side-scattered light information and second fluorescence information; and

[0297] The second cell count is subtracted from the first cell count to obtain the corrected basophil population count.

[0298] In another example, when at least one type of scattered light information includes side-scattered light information, the correction of the basophil population count result based on at least a second fluorescence information performed by processor 140 includes: processor 140

[0299] The basophil population of the test sample was obtained based on side-scattered light information and first fluorescence information; and

[0300] Based on the second fluorescence information of the basophil population, the nucleated erythrocyte population within the basophil population is identified to obtain a corrected basophil population count.

[0301] In a third embodiment of the sample analyzer according to this application, the processor 140 is configured to:

[0302] A three-dimensional scatter plot of the test sample is obtained based on at least one type of scattered light information, first fluorescence information, and second fluorescence information; and

[0303] Based on the three-dimensional scatter plot, the white blood cells in the test sample are classified into neutrophil population, lymphocyte population, monocyte population, eosinophil population and basophil population, and optionally the corresponding count values ​​of these cell populations are obtained.

[0304] For example, in the case where the light emitting device described above includes a first light source and a second light source, the scattered light information includes first lateral scattered light information generated by particles passing through the flow chamber after being irradiated by the beam of the first light source, as detected in the single test, and / or second lateral scattered light information generated by particles passing through the flow chamber after being irradiated by the beam of the second light source, as detected in the single test. In this case, the processor 140 obtains a three-dimensional scatter plot of the test sample based on the first lateral scattered light information, the first fluorescence information, and the second fluorescence information, and / or based on the second lateral scattered light information, the first fluorescence information, and the second fluorescence information. Preferably, the first dye is acridine orange dye and the second dye is a lysosomal dye.

[0305] This application also proposes a sample analysis method capable of obtaining accurate five-part differential white blood cell results through a single reaction and detection channel, which is implemented, in particular, by the sample analyzer described above or one of its embodiments.

[0306] As shown in Figure 7, in the fourth embodiment of the sample analysis method according to this application, the sample analysis method 400 includes the following steps S410, S420, S430, S440 and S450.

[0307] In step S410, a blood sample to be tested is collected or aspirated. For example, a portion of the blood sample to be tested is aspirated from a test tube containing the blood sample to be tested.

[0308] In step S420, at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare a test sample.

[0309] For example, in step S420, the hemolysin, the first dye, and the second dye can be added sequentially to the same blood sample to obtain a test sample. The test sample is then incubated to allow the dye to fully stain the particles in the test sample. Alternatively, in step S420, the first dye and the second dye can be pre-mixed with the hemolysin to obtain a mixed reagent. This mixed reagent is then mixed with the blood sample to be tested at a volume ratio of 250:1 to 1000:1. After thorough mixing, the resulting test sample is incubated at 25°C to 50°C for 10 seconds to 1 minute, preferably 20 seconds to 40 seconds.

[0310] Examples of the hemolytic agent and dye used in step S420 can be found in the above description and will not be repeated here.

[0311] In step S430, in a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by excitation light. The optical information includes at least one scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. The excitation light includes light of a first wavelength capable of exciting the first dye, the first wavelength being between 315 nm and 490 nm, preferably between 400 nm and 450 nm.

[0312] Here, the first fluorescence information includes the fluorescence signal generated by photoexcitation after the particles in the test sample bind to the first dye, while the second fluorescence information includes the fluorescence signal generated by photoexcitation after the particles in the test sample bind to the second dye.

[0313] In other words, in step S430, the scattered light information and fluorescence information of the test sample are obtained based on the principle of flow cytometry. In the embodiments of this application, the scattered light information includes the scattered light signal intensity, and the fluorescence information includes the fluorescence signal intensity.

[0314] In step S440, the white blood cell four-part differential result of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell four-part differential result includes the count results of neutrophils, lymphocytes, monocytes and eosinophils in the test sample.

[0315] In step S450, the count result of the basophil population in the test sample is obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information.

[0316] Therefore, by combining the four-part differential white blood cell count obtained in step S440 and the basophil count obtained in step S450, an accurate five-part differential white blood cell count can be obtained.

[0317] Furthermore, the excitation light also includes light of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength. For example, the second wavelength is between 610 nm and 750 nm, preferably between 620 nm and 700 nm. In some embodiments, the first dye is acridine orange dye and the second dye is a lysosomal dye.

[0318] Here, the scattered light information includes the first lateral scattered light information generated by the particles passing through the optical detection area after being irradiated by light of the first wavelength, i.e., blue-violet light, and / or the second lateral scattered light information generated by the particles passing through the optical detection area after being irradiated by light of the second wavelength, i.e., red light, obtained in the single test.

[0319] Therefore, in some embodiments, step S440 may include: obtaining the white blood cell four-part differential result of the test sample based on the first side-scattered light information and the first fluorescence information, and / or, based on the second side-scattered light information and the first fluorescence information. Step S450 may include: obtaining the basophil count result in the test sample based on the first side-scattered light information, the first fluorescence information, and the second fluorescence information, and / or, based on the second side-scattered light information, the first fluorescence information, and the second fluorescence information.

[0320] In some embodiments, step S450 includes: distinguishing between basophilic granulocytes and nucleated erythrocytes in the test sample based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, so as to identify the basophilic granulocytes in the test sample and count the basophilic granulocytes in the test sample, thereby obtaining the count result of the basophilic granulocytes in the test sample.

[0321] In some embodiments, at least one of the scattered light information includes side-scattered light information. In this case, as shown in FIG8, step S450 includes:

[0322] S451, Based on the side-scattered light information and the first fluorescence information, the first particle information of the first characteristic cell population is obtained;

[0323] S452, based on the side-scattered light information and the second fluorescence information, the second particle information of the second characteristic cell population is obtained; and

[0324] S453, based on the first particle information and the second particle information, the count result of the basophil population in the test sample is obtained.

[0325] Further, step S451 may include: based on the side-scattered light information and the first fluorescence information, identifying a first characteristic cell group comprising basophils and nucleated erythrocytes, and obtaining first particle information for the first characteristic cell group. And step S452 may include: based on the side-scattered light information and the second fluorescence information, identifying a second characteristic cell group comprising only nucleated erythrocytes, and obtaining second particle information for the second characteristic cell group.

[0326] This eliminates the interference of nucleated red blood cells on basophil counts.

[0327] In one example, the first particle information includes a first cell count of the first characteristic cell population, while the second particle information includes a second cell count of the second characteristic cell population. In this case, step S453 may include subtracting the second cell count from the first cell count to obtain the basophil population count in the assay sample.

[0328] As alternative to the embodiment shown in FIG8, in other embodiments, step S450 may include: obtaining a first characteristic cell population based on the side-scattered light information and the first fluorescence information, for example, dividing a first characteristic cell population including basophils and nucleated erythrocytes based on the side-scattered light information and the first fluorescence information; and obtaining the count result of the basophil population in the test sample based on at least a second fluorescence information of the cells in the first characteristic cell population.

[0329] As shown in Figure 9, in the fifth embodiment of the sample analysis method according to this application, the sample analysis method 500 includes:

[0330] Step S510: Collect the blood sample to be tested.

[0331] Step S520: Mix at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye to prepare a test sample.

[0332] Step S530: In a single test, each particle in the test sample is passed through the optical detection area irradiated by the excitation light to obtain the optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test.

[0333] Step S540: Obtain the white blood cell five-part differential result of the test sample based on at least one of the scattered light information and the first fluorescence information. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample.

[0334] Step S550: The basophil count is corrected based on at least the second fluorescence information to obtain a corrected basophil count.

[0335] In some embodiments, step S550 may include:

[0336] Based on the second fluorescence information, the nucleated erythrocyte population within the basophil population is identified and counted to obtain the nucleated erythrocyte population count result; and

[0337] The count of the basophil population is corrected based on the count of the nucleated erythrocyte population.

[0338] This eliminates the interference of nucleated red blood cells on basophil counts.

[0339] In some embodiments, at least one of the scattered light information includes side-scattered light information. In this case, step S350 may include:

[0340] The first cell count of the basophil population of the test sample is obtained based on the side-scattered light information and the first fluorescence information.

[0341] The second cell count of nucleated red blood cells in the test sample is obtained based on the side-scattered light information and the second fluorescence information; and

[0342] The second cell count is subtracted from the first cell count to obtain a corrected basophil population count.

[0343] Alternatively, step S550 may include:

[0344] The basophil population of the test sample is obtained based on the side-scattered light information and the first fluorescence information; and

[0345] Based on the second fluorescence information of the basophil population, the nucleated erythrocyte population within the basophil population is identified to obtain a corrected basophil population count.

[0346] In some embodiments, the excitation light includes light of a first wavelength capable of exciting a first dye and light of a second wavelength capable of exciting a second dye, wherein the second wavelength is greater than the first wavelength.

[0347] For example, the first wavelength is between 315 nm and 490 nm, and the second wavelength is between 610 nm and 750 nm. Preferably, the first wavelength is between 400 nm and 450 nm, and the second wavelength is between 620 nm and 700 nm.

[0348] In some embodiments, the first dye is acridine orange dye and the second dye is a lysosomal dye.

[0349] As shown in Figure 10, in the sixth embodiment of the sample analysis method according to this application, the sample analysis method 600 includes:

[0350] Step S610: Collect the blood sample to be tested.

[0351] Step S620: Mix at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye to prepare a test sample.

[0352] Step S630: In a single test, each particle in the test sample is passed through the optical detection area irradiated by the excitation light to obtain the optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye.

[0353] Step S640: A three-dimensional scatter plot of the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information.

[0354] Step S650: Based on the three-dimensional scatter plot, classify the white blood cells in the test sample into neutrophil population, lymphocyte population, monocyte population, eosinophil population and basophil population.

[0355] In some embodiments, the excitation light includes light of a first wavelength capable of exciting a first dye and light of a second wavelength capable of exciting a second dye, wherein the second wavelength is greater than the first wavelength.

[0356] Here, the first wavelength is, for example, between 315 nm and 490 nm, and the second wavelength is, for example, between 610 nm and 750 nm. Preferably, the first wavelength is between 400 nm and 450 nm, and the second wavelength is between 620 nm and 700 nm.

[0357] In some embodiments, the first dye is acridine orange dye and the second dye is a lysosomal dye.

[0358] In some embodiments, the scattered light information includes first lateral scattered light information generated by particles passing through the optical detection area after being irradiated with light of the first wavelength, such as blue-violet light, acquired in the single test, and / or second lateral scattered light information generated by particles passing through the optical detection area after being irradiated with light of the second wavelength, such as red light, acquired in the single test. Here, step S640 may include: obtaining a three-dimensional scatter plot of the test sample based on the first lateral scattered light information, the first fluorescence information, and the second fluorescence information, and / or, based on the second lateral scattered light information, the first fluorescence information, and the second fluorescence information.

[0359] The present application is described below with reference to embodiments intended to illustrate the application and not limit it. Unless otherwise specified, the experiments and methods described in the embodiments are generally performed according to conventional methods well known in the art and described in various references. Furthermore, where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0360] Example 1

[0361] The reagent formula is as follows:

[0362] A: Fluorescent dye solution

[0363] B: Hemolytic agent

[0364] The blood sample to be tested was processed and detected using a sample analyzer equipped with a red laser (emitting a beam with a wavelength of approximately 633 nm) and a blue laser (emitting a beam with a wavelength of approximately 488 nm) according to this application: 20 μL of anticoagulated blood sample and reagent A were added to 1 mL of reagent B, and the mixture was incubated at 42°C for 20 seconds to form a test sample; the intensity of side-scattered light corresponding to blue light excitation, the intensity of first fluorescence corresponding to blue light excitation, and the intensity of second fluorescence corresponding to red light excitation were detected; a two-dimensional first scatter plot was generated based on the side-scattered light intensity and the first fluorescence intensity, as shown in Figure 11A, and the five-part differential white blood cell count was obtained based on the first scatter plot; a two-dimensional second scatter plot was generated based on the side-scattered light intensity and the second fluorescence intensity, as shown in Figure 11B; nucleated red blood cells in the test sample were identified based on the second scatter plot, and the nucleated red blood cell count was obtained; the basophil count in the five-part differential white blood cell count was corrected based on the nucleated red blood cells identified in the second scatter plot to obtain a corrected five-part differential white blood cell count.

[0365] Alternatively, the white blood cell four-part differential count can be obtained based on the first scatter plot, the basophil count can be obtained based on the first and second scatter plots, and the nucleated red blood cell count can be obtained based on the second scatter plot.

[0366] Example 2

[0367] The reagent formula is as follows:

[0368] A: Fluorescent dye solution

[0369] B: Hemolytic agent

[0370] The blood sample to be tested was processed and detected using a sample analyzer equipped with a red laser (emitting a beam with a wavelength of approximately 633 nm) and a blue laser (emitting a beam with a wavelength of approximately 488 nm) according to this application: 20 μL of anticoagulated blood sample and reagent A were added to 1 mL of reagent B, and the mixture was incubated at 42°C for 20 seconds to form a test sample; the intensity of side-scattered light corresponding to blue light excitation, the intensity of first fluorescence corresponding to blue light excitation, and the intensity of second fluorescence corresponding to red light excitation were detected; a two-dimensional first scatter plot was generated based on the side-scattered light intensity and the first fluorescence intensity, as shown in Figure 12A, and the five-part differential white blood cell count was obtained based on the first scatter plot; a two-dimensional second scatter plot was generated based on the side-scattered light intensity and the second fluorescence intensity, as shown in Figure 12B; nucleated red blood cells in the test sample were identified based on the second scatter plot, and the nucleated red blood cell count was obtained; the basophil count in the five-part differential white blood cell count was corrected based on the nucleated red blood cells identified in the second scatter plot to obtain a corrected five-part differential white blood cell count.

[0371] Alternatively, the white blood cell four-part differential count can be obtained based on the first scatter plot, the basophil count can be obtained based on the first and second scatter plots, and the nucleated red blood cell count can be obtained based on the second scatter plot.

[0372] Example 3

[0373] The reagent formulation of Example 2 was used.

[0374] The blood sample to be tested was processed and detected using a sample analyzer according to this application, which is equipped with a red laser (emitting a beam with a wavelength of approximately 633 nm) and a blue laser (emitting a beam with a wavelength of approximately 450 nm). Reagent A and 20 μL of anticoagulated blood sample were added to 1 mL of reagent B, and incubated at 42°C for 20 seconds to form the test sample. The intensity of the side-scattered light corresponding to blue light excitation, the first fluorescence intensity corresponding to blue light excitation, and the second fluorescence intensity corresponding to red light excitation were detected. A two-dimensional first scatter plot was generated based on the side-scattered light intensity and the first fluorescence intensity, as shown in Figure 13A, and the four-part differential white blood cell count was obtained based on the first scatter plot. A two-dimensional second scatter plot was generated based on the side-scattered light intensity and the second fluorescence intensity, as shown in Figure 13B, and the basophil count was obtained based on the first and second scatter plots. The nucleated red blood cell count was obtained based on the second scatter plot.

[0375] Example 4

[0376] The reagent formula is as follows:

[0377] A: Fluorescent dye solution

[0378] B: Hemolytic agent

[0379] The blood sample to be tested was processed and detected using a sample analyzer equipped with a red laser (emitting a beam with a wavelength of approximately 633 nm) and a blue laser (emitting a beam with a wavelength of approximately 488 nm) according to this application: 20 μL of anticoagulated blood sample and reagent A were added to 1 mL of reagent B, and the mixture was incubated at 42°C for 20 seconds to form the test sample; the intensity of the side-scattered light corresponding to blue light excitation, the first fluorescence intensity corresponding to blue light excitation, and the second fluorescence intensity corresponding to red light excitation were detected; a two-dimensional first scatter plot was generated based on the side-scattered light intensity and the first fluorescence intensity, as shown in Figure 14A, and the four-part differential white blood cell count was obtained based on the first scatter plot; and a two-dimensional second scatter plot was generated based on the side-scattered light intensity and the second fluorescence intensity, as shown in Figure 14B, and the nucleated red blood cell count was obtained based on the second scatter plot.

[0380] Example 5

[0381] The reagent formula is as follows:

[0382] A: Fluorescent dye solution

[0383] B: Hemolytic agent

[0384] The blood sample to be tested was processed and detected using a sample analyzer according to this application, which is equipped with a red laser (emitting a beam with a wavelength of approximately 633 nm) and a blue laser (emitting a beam with a wavelength of approximately 488 nm). Reagent A and 20 μL of anticoagulated blood sample were added to 1 mL of reagent B, and the mixture was incubated at 42°C for 20 seconds to form the test sample. The intensity of the side-scattered light corresponding to blue light excitation, the first fluorescence intensity corresponding to blue light excitation, and the second fluorescence intensity corresponding to red light excitation were detected. A two-dimensional first scatter plot was generated based on the side-scattered light intensity and the first fluorescence intensity, as shown in Figure 15A, and the four-part differential white blood cell count was obtained based on the first scatter plot. A two-dimensional second scatter plot was generated based on the side-scattered light intensity and the second fluorescence intensity, as shown in Figure 15B, and the nucleated red blood cell count was obtained based on the second scatter plot.

[0385] Example 6

[0386] The reagent formula is as follows:

[0387] A: Fluorescent dye solution

[0388] B: Hemolytic agent

[0389] The blood sample to be tested was processed and detected using a sample analyzer according to this application, which is equipped with a red laser (emitting a beam with a wavelength of approximately 633 nm) and a blue laser (emitting a beam with a wavelength of approximately 488 nm). Reagent A and 20 μL of anticoagulated blood sample were added to 1 mL of reagent B, and the mixture was incubated at 42°C for 20 seconds to form the test sample. The intensity of the side-scattered light corresponding to blue light excitation, the first fluorescence intensity corresponding to blue light excitation, and the second fluorescence intensity corresponding to red light excitation were detected. A two-dimensional first scatter plot was generated based on the side-scattered light intensity and the first fluorescence intensity, as shown in Figure 16A, and the four-part differential white blood cell count was obtained based on the first scatter plot. A two-dimensional second scatter plot was generated based on the side-scattered light intensity and the second fluorescence intensity, as shown in Figure 16B, and the basophil count was obtained based on the first and second scatter plots. The nucleated red blood cell count was obtained based on the second scatter plot.

[0390] Example 7

[0391] The reagent formulation of Example 2 was used to verify the proposed solution in this application.

[0392] Multiple blood samples were tested using the sample analyzer and method (dual dye, single channel) of Example 2 to obtain white blood cell differential and nucleated red blood cell counts. A Mindray hematology analyzer (single dye, dual channel) was also used to test the same multiple blood samples to obtain white blood cell differential and nucleated red blood cell counts. A correlation analysis was performed between the white blood cell differential and nucleated red blood cell counts obtained using this application and those obtained using the existing hematology analyzer, as shown in Figure 17. Figures 17A to 17F show the correlations between the neutrophil count (Neu%), lymphocyte count (Lym%), monocyte count (Mon%), eosinophil count (Eos%), basophil count (Baso%), and nucleated red blood cell count (Nrbc%) obtained from testing multiple blood samples according to this application and the prior art, respectively.

[0393] As shown in Figure 17, the white blood cell differential and nucleated red blood cell count results obtained through this application have a good correlation with the white blood cell differential and nucleated red blood cell count results obtained through existing blood cell analyzers.

[0394] All features or combinations of features mentioned above in the specification, drawings, and claims may be used in any combination or individually, provided they are meaningful within the scope of this application and do not contradict each other. The advantages and features described with reference to the sample analyzer provided in this application shall be applied accordingly to the sample analysis method provided in this application, and vice versa.

[0395] The above description is merely a preferred embodiment of this application and does not limit the scope of patent protection of this application. All equivalent modifications made based on the content of the specification and drawings of this application, or direct / indirect applications in other related technical fields, are included within the scope of patent protection of this application.

Claims

1. A sample analyzer, comprising: A sampling device is used to collect blood samples for testing. A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample; An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device is used to emit a light beam to illuminate the flow chamber. The flow chamber is connected to the reaction cell and is used to allow individual particles in the test sample to pass through. The photodetector is used to detect, in a single test, the optical information generated when particles in the test sample are illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light information detected in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. as well as The processor is configured to: generate a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtain the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, wherein the white blood cell five-part differential result includes the count result of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

2. The sample analyzer of claim 1, wherein, The light emitting device includes a first light source and a second light source. The first light source is used to emit a light beam of a first wavelength that can excite the first dye, and the second light source is used to emit a light beam of a second wavelength that can excite the second dye, wherein the second wavelength is greater than the first wavelength. Preferably, the first dye is acridine orange dye and the second dye is a lysosomal dye.

3. The sample analyzer of claim 1, wherein, The light emitting device is configured to emit light of a single wavelength capable of exciting the first dye and the second dye.

4. The sample analyzer of any one of claims 1 to 3, wherein, The process of generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: A first scatter plot is generated based on at least one of the scattered light information and the first fluorescence information, and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot; A second scatter plot is generated based on at least one of the scattered light information and the second fluorescence information, and nucleated red blood cells in the test sample are identified based on the second scatter plot to obtain the count result of the nucleated red blood cell population; and The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

5. The sample analyzer of claim 2 or 3, wherein, The first wavelength is between 315 nm and 490 nm, and the second wavelength is between 620 nm and 700 nm.

6. The sample analyzer of claim 5, wherein, The scattered light information includes first lateral scattered light information generated by particles passing through the flow chamber after being irradiated by the beam of the first light source and / or second lateral scattered light information generated by particles passing through the flow chamber after being irradiated by the beam of the second light source, as detected in the single test. The step of generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: Based on the first side-scattered light information and the first fluorescence information, and / or based on the second side-scattered light information and the first fluorescence information, a first scatter plot is generated and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot; Based on the first side-scattered light information and the second fluorescence information, and / or based on the second side-scattered light information and the second fluorescence information, a second scatter plot is generated, and nucleated red blood cells in the test sample are identified based on the second scatter plot to obtain the counting result of the nucleated red blood cell population; and The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

7. The sample analyzer of claim 4, wherein, At least one of the scattered light information includes side-scattered light information. The step of generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: The first scatter plot is generated based on the side-scattered light information and the first fluorescence information. A first characteristic cell group including basophils and nucleated erythrocytes is divided in the first scatter plot, and the first particle information of the first characteristic cell group is obtained. The second scatter plot is generated based on the side-scattered light information and the second fluorescence information. The second scatter plot is used to divide the second characteristic cell group, which includes only nucleated erythrocytes, and the second particle information of the second characteristic cell group is obtained. The count of nucleated erythrocytes in the test sample is obtained based on the second particle information; and The basophil count in the test sample is obtained based on the first particle information and the second particle information.

8. The sample analyzer of claim 7, wherein, The first particle information includes a first cell count of the first characteristic cell population, and the second particle information includes a second cell count of the second characteristic cell population; and The step of obtaining the basophil population count result in the test sample based on the first particle information and the second particle information includes: subtracting the second cell count from the first cell count to obtain the basophil population count result in the test sample.

9. The sample analyzer according to claim 4, characterized in that, At least one of the scattered light information includes side-scattered light information. The step of generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: The first scatter plot is generated based on the lateral scattered light information and the first fluorescence information, and a first characteristic cell population, including basophils and nucleated erythrocytes, is identified in the first scatter plot; and The basophil count in the test sample is obtained based on at least the second fluorescence information of the cells in the first characteristic cell population.

10. The sample analyzer according to any one of claims 1 to 3, characterized in that, The process of generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: A three-dimensional scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the three-dimensional scatter plot.

11. A sample analysis method, comprising: Collect the blood sample to be tested; At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample; In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by light. The optical information includes at least one scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test. as well as A scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the scatter plot. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils, and basophils in the test sample.

12. The sample analysis method according to claim 11, characterized in that, The excitation light includes light of a first wavelength capable of exciting the first dye and light of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength; Preferably, the first dye is acridine orange dye and the second dye is a lysosomal dye.

13. The sample analysis method according to claim 11, characterized in that, The excitation light is a single wavelength of light capable of exciting the first dye and the second dye.

14. The sample analysis method according to any one of claims 11 to 13, characterized in that, A scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell five-part differential and nucleated red blood cell count results of the test sample are obtained based on the scatter plot, including: A first scatter plot is generated based on at least one of the scattered light information and the first fluorescence information, and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot; A second scatter plot is generated based on at least one of the scattered light information and the second fluorescence information, and nucleated red blood cells in the test sample are identified based on the second scatter plot to obtain the count result of the nucleated red blood cell population; and The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

15. The sample analysis method according to claim 12, characterized in that, The first wavelength is between 315 nm and 490 nm, and the second wavelength is between 620 nm and 700 nm.

16. The sample analysis method according to claim 15, characterized in that, The scattered light information includes the first lateral scattered light information generated by the particles passing through the flow chamber after being irradiated by the first wavelength of light, obtained in the single test, and / or the second lateral scattered light information generated by the particles passing through the flow chamber after being irradiated by the second wavelength of light. The process of generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential and nucleated red blood cell count results of the test sample based on the scatter plot, includes: Based on the first side-scattered light information and the first fluorescence information, and / or based on the second side-scattered light information and the first fluorescence information, a first scatter plot is generated and the white blood cell five-part differential result of the test sample is obtained based on the first scatter plot; Based on the first side-scattered light information and the second fluorescence information, and / or based on the second side-scattered light information and the second fluorescence information, a second scatter plot is generated, and nucleated red blood cells in the test sample are identified based on the second scatter plot to obtain the counting result of the nucleated red blood cell population; and The count of basophils in the five-part differential white blood cell count is corrected based on the nucleated red blood cells identified in the second scatter plot to obtain the corrected five-part differential white blood cell count.

17. The sample analysis method according to claim 14, characterized in that, At least one of the scattered light information includes side-scattered light information, wherein generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: The first scatter plot is generated based on the side-scattered light information and the first fluorescence information. A first characteristic cell group including basophils and nucleated erythrocytes is divided in the first scatter plot, and the first particle information of the first characteristic cell group is obtained. The second scatter plot is generated based on the side-scattered light information and the second fluorescence information. The second scatter plot is used to divide the second characteristic cell group, which includes only nucleated erythrocytes, and the second particle information of the second characteristic cell group is obtained. The count of nucleated erythrocytes in the test sample is obtained based on the second particle information; and The basophil count in the test sample is obtained based on the first particle information and the second particle information.

18. The sample analysis method according to claim 17, characterized in that, The first particle information includes a first cell count of the first characteristic cell population, and the second particle information includes a second cell count of the second characteristic cell population; and Obtaining the basophil population count in the test sample based on the first particle information and the second particle information includes: subtracting the second cell count from the first cell count to obtain the basophil population count in the test sample.

19. The sample analysis method according to claim 14, characterized in that, At least one of the scattered light information includes side-scattered light information, wherein generating a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtaining the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, includes: The first scatter plot is generated based on the lateral scattered light information and the first fluorescence information, and a first characteristic cell population, including basophils and nucleated erythrocytes, is identified in the first scatter plot; and The basophil count in the test sample is obtained based on at least the second fluorescence information of the cells in the first characteristic cell population.

20. The sample analysis method according to any one of claims 11 to 13, characterized in that, A scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell five-part differential and nucleated red blood cell count results of the test sample are obtained based on the scatter plot, including: A three-dimensional scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell five-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the three-dimensional scatter plot.

21. A sample analysis method, comprising: Collect the blood sample to be tested; At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample; In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by light. The optical information includes at least one scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test. as well as The white blood cell differential and nucleated red blood cell count results of the test sample are obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information. The white blood cell differential results include the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample.

22. A sample analyzer, comprising: A sampling device is used to collect blood samples for testing. A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample; An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated when particles in the test sample are illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light information detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye. The light emitting device includes a first light source and a second light source. The first light source emits a light beam of a first wavelength capable of exciting the first dye, and the second light source emits a light beam of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength. The processor is configured to: generate a scatter plot based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and obtain the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample based on the scatter plot, wherein the white blood cell four-part differential result includes the count result of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

23. The sample analyzer according to claim 22, characterized in that, The first dye is acridine orange dye and the second dye is a lysosomal dye.

24. A sample analysis method, comprising: Collect the blood sample to be tested; At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample; In a single test, each particle in the test sample is passed one by one through an optical detection area irradiated by excitation light to obtain optical information generated by the particles after irradiation. This optical information includes at least one type of scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. The excitation light includes light of a first wavelength capable of exciting the first dye and light of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength. The white blood cell four-part differential and nucleated red blood cell count results of the test sample are obtained based on at least one of the scattered light information, the first fluorescence information and the second fluorescence information. The white blood cell four-part differential results include the count results of neutrophils, lymphocytes, monocytes and eosinophils in the test sample. Preferably, a scatter plot is generated based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information, and the white blood cell four-part differential result and the nucleated red blood cell count result of the test sample are obtained based on the scatter plot. The white blood cell four-part differential result includes the count results of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample.

25. A sample analyzer, comprising: A sampling device is used to collect blood samples for testing. A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample; An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device emits a light beam to illuminate the flow chamber, which is connected to a reaction cell and allows individual particles in the test sample to pass through. The photodetector detects, in a single test, the optical information generated by the particles in the test sample after being illuminated by the light beam as they pass through the flow chamber. The optical information includes at least one type of scattered light information detected in the single test, a first fluorescence information corresponding to a first dye, and a second fluorescence information corresponding to a second dye. The light beam includes at least a first wavelength capable of exciting the first dye, wherein the first wavelength is between 315 nm and 490 nm. The processor is configured as follows: The white blood cell four-part differential result of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell four-part differential result includes the count results of neutrophils, lymphocytes, monocytes and eosinophils in the test sample. and The basophil count in the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information.

26. The sample analyzer according to claim 25, characterized in that, The light beam further includes a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength; and The first dye is acridine orange dye and the second dye is a lysosomal dye.

27. A sample analyzer, comprising: A sampling device is used to collect blood samples for testing. A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample; An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device is used to emit a light beam to illuminate the flow chamber. The flow chamber is connected to the reaction cell and is used to allow individual particles in the test sample to pass through. The photodetector is used to detect optical information generated by the particles in the test sample after being illuminated by the light beam when passing through the flow chamber in a single test. The optical information includes at least one scattered light information detected in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. as well as The processor is configured as follows: The white blood cell five-part differential result of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample. and The basophil count is corrected based on at least the second fluorescence information to obtain a corrected basophil count.

28. The sample analyzer according to claim 27, characterized in that, The light emitting device includes a first light source and a second light source. The first light source is used to emit a light beam of a first wavelength capable of exciting the first dye, and the second light source is used to emit a light beam of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength; and The first dye is acridine orange dye and the second dye is a lysosomal dye.

29. A sample analyzer, comprising: A sampling device is used to collect blood samples for testing. A sample preparation apparatus for mixing at least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye in a reaction cell to prepare a test sample; An optical detection device includes a light emitting device, a flow chamber, and a photodetector. The light emitting device is used to emit a light beam to illuminate the flow chamber. The flow chamber is connected to the reaction cell and is used to allow individual particles in the test sample to pass through. The photodetector is used to detect optical information generated by the particles in the test sample after being illuminated by the light beam when passing through the flow chamber in a single test. The optical information includes at least one scattered light information detected in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. as well as The processor is configured as follows: A three-dimensional scatter plot of the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information; and Based on the three-dimensional scatter plot, the white blood cells in the test sample are classified into neutrophil population, lymphocyte population, monocyte population, eosinophil population, and basophil population.

30. The sample analyzer according to claim 29, characterized in that, 28. The sample analyzer according to claim 27, wherein the light emitting device comprises a first light source and a second light source, the first light source being used to emit a light beam of a first wavelength capable of exciting the first dye, and the second light source being used to emit a light beam of a second wavelength capable of exciting the second dye, wherein the second wavelength is greater than the first wavelength; and The first dye is acridine orange dye and the second dye is a lysosomal dye.

31. A sample analysis method, comprising: Collect the blood sample to be tested; At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample; In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test. The excitation light includes light capable of exciting the first dye at a first wavelength, which is between 315 nm and 490 nm. The white blood cell differential count of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell differential count includes the counts of neutrophils, lymphocytes, monocytes, and eosinophils in the test sample. The basophil count in the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information.

32. A sample analysis method, comprising: Collect the blood sample to be tested; At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample; In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one type of scattered light information, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye obtained in the single test. The white blood cell five-part differential result of the test sample is obtained based on at least one of the scattered light information and the first fluorescence information. The white blood cell five-part differential result includes the count results of neutrophils, lymphocytes, monocytes, eosinophils and basophils in the test sample. and The basophil count is corrected based on at least the second fluorescence information to obtain a corrected basophil count.

33. A sample analysis method, comprising: Collect the blood sample to be tested; At least a portion of the blood sample to be tested, a hemolysin, a first dye, and a second dye are mixed to prepare the test sample; In a single test, each particle in the test sample is passed through an optical detection area irradiated by excitation light to obtain optical information generated by the particles in the test sample after being irradiated by the excitation light. The optical information includes at least one scattered light information obtained in the single test, a first fluorescence information corresponding to the first dye, and a second fluorescence information corresponding to the second dye. A three-dimensional scatter plot of the test sample is obtained based on at least one of the scattered light information, the first fluorescence information, and the second fluorescence information; and Based on the three-dimensional scatter plot, the white blood cells in the test sample are classified into neutrophil population, lymphocyte population, monocyte population, eosinophil population, and basophil population.

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