WO2016106688A1 - 一种有核红细胞报警方法、装置及流式细胞分析仪 - Google Patents

一种有核红细胞报警方法、装置及流式细胞分析仪 Download PDF

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WO2016106688A1
WO2016106688A1 PCT/CN2014/095905 CN2014095905W WO2016106688A1 WO 2016106688 A1 WO2016106688 A1 WO 2016106688A1 CN 2014095905 W CN2014095905 W CN 2014095905W WO 2016106688 A1 WO2016106688 A1 WO 2016106688A1
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Prior art keywords
scattered light
cells
fluorescence
information
blood sample
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PCT/CN2014/095905
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English (en)
French (fr)
Inventor
叶波
祁欢
钱程
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深圳迈瑞生物医疗电子股份有限公司
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Priority to PCT/CN2014/095905 priority Critical patent/WO2016106688A1/zh
Priority to CN201480081573.XA priority patent/CN106687810B/zh
Publication of WO2016106688A1 publication Critical patent/WO2016106688A1/zh
Priority to US15/638,902 priority patent/US11105742B2/en

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N15/1456—Electro-optical investigation, e.g. flow cytometers without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals
    • G01N15/1459—Electro-optical investigation, e.g. flow cytometers without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals the analysis being performed on a sample stream
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/483—Physical analysis of biological material
    • G01N33/487—Physical analysis of biological material of liquid biological material
    • G01N33/49—Blood
    • G01N2015/014—

Definitions

  • the present application relates to the field of medical devices, and in particular, to a nucleated red blood cell alarm method, device and flow cytometer.
  • Nucleated red blood cells i.e., immature red blood cells
  • NRBC Nucleated red blood cells
  • normal human blood samples blood other than bone marrow
  • nucleated red blood cells The presence of nucleated red blood cells in blood samples is due to the release of naive bone marrow erythroid cells into blood samples, and its appearance in blood samples is associated with blood diseases. Therefore, it is clinically necessary to check whether there are nucleated red blood cells in the blood.
  • the principle of the flow cytometer is to generate a photoelectric signal by irradiating the laser with the receiving laser, and then, the photoelectric signal generated by the laser irradiation of the cell is expressed in the form of, for example, a scattergram for subsequent analysis.
  • the scattered light signal and the fluorescent signal reflect the physicochemical characteristics of the cells, such as the size of the cells, the granularity, and the expression of the antigen molecules.
  • the classification and counting of white blood cells are usually performed by an electrical impedance method or a laser scattering method. These methods usually require first dissolving the red blood cells in the blood sample, and then passing the test sample through the detector, and using various white cell electrical signals or optical signal differences to classify them into several types, for example, can be divided into neutrophils, lymphocytes, Monocytes, eosinophils or basophils.
  • the presence of nucleated red blood cells often interferes with white blood cell measurements.
  • the signal obtained by the detection of nucleated red blood cells is similar to that of lymphocytes, and is difficult to distinguish. This will first affect the classification of white blood cells; in addition, more importantly, if it is not possible to suggest that there may be nucleated red blood cells in the sample, it is easy to cause missed diagnosis and delay the optimal treatment of the disease.
  • the present application proposes a nucleated red blood cell alarm method, a device and a flow cell analyzer, which can realize alarming of nucleated red blood cells while classifying and detecting white blood cells.
  • an aspect of the embodiments of the present application provides a nucleated red blood cell alarm method for alarming whether or not nucleated red blood cells are present in a blood sample, including the following steps:
  • the white blood cells are at least divided into four groups of lymphocyte population, monocyte population, eosinophil cell population, neutrophil and basophil population;
  • the step of counting the number of the characterization cells in the predetermined feature area, determining whether the number of the characterization cells exceeds a threshold, and performing an alarm process when the determination result is YES includes:
  • the number of characterized cells is greater than the predetermined threshold, it is determined that nuclear red blood cells are present in the blood sample and an alarm process is performed.
  • the step of acquiring the predetermined feature region of the forward scattered light-fluorescence scattergram includes:
  • the forward scattered light-fluorescence scattergram is analyzed, and an area on the left side of the white blood cell group region and the upper side of the blood shadow region in the forward scattered light-fluorescence scattergram is determined as a predetermined feature region.
  • the alarm processing is: performing an alarm prompt by using a text, a sound, a light, or a pop-up window.
  • nucleated red blood cell alarm device for alerting the presence or absence of nucleated red blood cells in a blood sample, including:
  • a sample information acquiring unit configured to acquire forward scattered light information, side scattered light information, and fluorescence information when the cells in the blood sample pass through the detection area of the flow cytometer;
  • a scattergram generating unit configured to generate a side scattered light-fluorescence scattergram according to the side scattered light information and the fluorescence information, and distinguish at least a white blood cell into a lymphocyte group, a monocyte group, and an eosinophilic particle a four-class classification of a cell population, a neutrophil and a basophil population, and a forward scattered light-fluorescence scattergram of the blood sample according to the forward scattered light information and the fluorescence information, wherein
  • the forward scattered light-fluorescence scattergram includes a white blood cell population region;
  • An analysis alarm unit configured to acquire a predetermined feature region of the forward scattered light-fluorescence scattergram, count the number of characterization cells in the predetermined feature region, determine whether the number of the characterization cells exceeds a threshold, and determine The result is an alarm processing when it is YES, wherein the predetermined feature area is located on the left side of the white blood cell group area.
  • the analysis alarm unit further includes:
  • a feature region determining unit configured to analyze the forward scattered light-fluorescence scattergram, and determine an area of the forward scattered light-fluorescence scattergram located on a left side of the white blood cell group region and an upper side of the blood shadow region as Predetermined feature area.
  • the analysis alarm unit comprises:
  • a counting unit configured to count the number of characterization cells in the predetermined feature region
  • a comparing unit configured to compare the number of the characterized cells with a predetermined threshold
  • a determining unit configured to determine that nucleated red blood cells are present in the blood sample when the number of characterization cells is greater than the predetermined threshold
  • the alarm processing unit is configured to perform an alarm process after determining that the red blood cells are present in the blood sample.
  • the sample information acquiring unit acquires forward scattered light information, side scattered light information, and fluorescence information of cells in the blood sample from measurement data of the same channel.
  • Another aspect of the present application also provides a flow cytometer comprising:
  • a sampling device for sucking a blood sample, the blood sample containing at least white blood cells
  • a pretreatment device for pretreating the blood sample to obtain a processed blood sample, the pretreatment comprising fluorescently labeling cells in the blood sample;
  • a detecting device configured to pass the cells in the processed blood sample one by one through the detection area, and detect forward scattered light information, side scattered light information, and fluorescence information of the cells in the blood sample;
  • An analyzing device generates a forward scattered light-fluorescence scattergram of the blood sample according to forward scattered light information, side scattered light information, and fluorescence information detected by the detecting device, and acquires the forward scattered light a predetermined feature area of the fluorescent scattergram, and counting the number of characterization cells in the predetermined feature area, determining whether the number of the characterization cells exceeds a threshold, and performing an alarm process when the determination result is YES, wherein
  • the forward scattered light-fluorescence scattergram includes a white blood cell population region that is located to the left of the white blood cell population region.
  • analysis device comprises:
  • a sample information acquiring unit configured to acquire forward scattered light information, side scattered light information, and fluorescence information when the cells in the blood sample pass through the detection area of the flow cytometer;
  • a scattergram generating unit configured to generate a side scatter light-fluorescence scattergram according to the side scatter light information and the fluorescence information acquired by the sample information acquiring unit, and distinguish at least the white blood cells into a lymphocyte group, a monocyte group, and a hobby Four classifications of acid granulocyte cell population, neutrophil and basophil population, and forward scattered light information and fluorescence information acquired according to sample information acquisition unit, generating forward scattered light-fluorescence of the blood sample a scattergram, wherein the forward scattered light-fluorescence scattergram includes a white blood cell population region;
  • An analysis alarm unit configured to acquire a predetermined feature region of the forward scattered light-fluorescence scattergram, count the number of characterization cells in the predetermined feature region, determine whether the number of the characterization cells exceeds a threshold, and determine The result is an alarm processing when it is YES, wherein the predetermined feature area is located on the left side of the white blood cell group area.
  • the analysis alarm unit further includes:
  • a feature region determining unit configured to analyze the forward scattered light-fluorescence scattergram, and determine an area of the forward scattered light-fluorescence scattergram located on a left side of the white blood cell group region and an upper side of the blood shadow region as Predetermined feature area.
  • the detecting device detects and collects forward scattered light information, side scattered light information and fluorescent information of cells in the blood sample in the same channel.
  • a nucleated red blood cell alarm method for alerting the presence or absence of nucleated red blood cells in a blood sample, comprising the following steps:
  • the pretreatment comprising at least fluorescent labeling of cells in the blood sample
  • the processed blood sample is injected into a flow cytometer to collect forward scattered light information, side scattered light information and fluorescence information of the cells;
  • the white blood cells are at least divided into four groups of lymphocyte population, monocyte population, eosinophil cell population, neutrophil and basophil population;
  • the forward scattered light-fluorescence scattergram including a white blood cell group region
  • the step of counting the number of the characterization cells in the predetermined feature area, determining whether the number of the characterization cells exceeds a threshold, and performing an alarm process when the determination result is YES includes:
  • the number of characterized cells is greater than the predetermined threshold, it is determined that nuclear red blood cells are present in the blood sample and an alarm process is performed.
  • the step of injecting the processed blood sample into the flow cytometer to collect the forward scattered light information, the side scattered light information and the fluorescence information of the cells in the blood sample is specifically as follows:
  • the forward scattered light information, the side scattered light information, and the fluorescence information of the cells in the blood sample are simultaneously collected.
  • the step of acquiring the predetermined feature region of the forward scattered light-fluorescence scattergram includes:
  • the forward scattered light-fluorescence scattergram is analyzed, and an area on the left side of the white blood cell group region and the upper side of the blood shadow region in the forward scattered light-fluorescence scattergram is determined as a predetermined feature region.
  • the step of acquiring the predetermined feature region of the forward scattered light-fluorescence scattergram includes:
  • the normal blood sample containing at least white blood cells, the abnormal blood sample containing at least white blood cells and nucleated red blood cells;
  • the pretreatment including at least fluorescent labeling of cells in the blood sample;
  • the processed normal blood sample and the treated abnormal blood sample are respectively injected into a flow cytometer to collect forward scattered light information, side scattered light information and fluorescence information of the cells;
  • the white blood cells are at least divided into four groups of lymphocyte population, monocyte population, eosinophil cell population, neutrophil and basophil population. Generating a forward scattered light-fluorescence scattergram of the normal blood sample and a forward scattered light-fluorescence scattergram of the abnormal blood sample according to the forward scattered light information and the fluorescence information;
  • the leukocyte population region of the forward scattered light-fluorescence scatter plot of the abnormal blood sample A particle group on the left side and a region of the white blood cell group region on the left side of the leukocyte group region of the normal blood sample, which is a forward-scattering light-fluorescence scattergram, is determined as a predetermined feature region.
  • each cell in the blood sample is detected using a flow cytometer to obtain side scattered light, forward scattered light, and fluorescence information of the cell.
  • the white blood cells in the blood sample are divided into four categories by forming a side scattered light-fluorescence scattergram; by forming a forward scattered light-fluorescence scattergram, the scatter plot is located in the left region of the white blood cell group Characterization of specific regions on the side of the cells for counting analysis can achieve alarms for nucleated red blood cells, that is, the function of white blood cell four classification and nucleated red blood cell alarm can be realized in one channel measurement.
  • the white blood cells when the dedicated nucleated red blood cell reagent is not needed, and the number of nucleated red blood cells is not determined, the white blood cells can be classified and detected, and a more accurate nucleated red blood cell alarm can be provided, so that the measurement is not increased.
  • important information can be provided quickly and easily for clinical screening.
  • FIG. 1 is a schematic diagram of a main flow of an embodiment of a nucleated red blood cell alarm method provided by the present application
  • 2A is a schematic diagram of a side scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing no erythrocytes when the first reagent is used in the present application;
  • 2B is a schematic diagram of a forward scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing no erythrocytes when the first reagent is used in the present application;
  • 3A is a schematic diagram of a side scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing nucleated red blood cells when the first reagent is used in the present application;
  • 3B is a schematic diagram of a forward scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing nucleated red blood cells when the first reagent is used in the present application;
  • FIG. 4 is a schematic diagram of a three-dimensional scattergram corresponding to FIG. 3A and FIG. 3B;
  • 5A is a schematic diagram of a side scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing no nucleated red blood cells when the second reagent is used in the present application;
  • 5B is a schematic diagram of a forward scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing no erythrocytes when the second reagent is used in the present application;
  • 6A is a schematic diagram of a side scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing nucleated red blood cells when a second reagent is used in the present application;
  • 6B is a schematic diagram of a forward scattered light-fluorescence scatter plot obtained by analyzing a blood sample containing nucleated red blood cells when a second reagent is used in the present application;
  • Figure 7 is a schematic diagram of a three-dimensional scatter plot corresponding to Figures 6A and 6B;
  • FIG. 8 is a schematic structural view of an embodiment of a nucleated red blood cell alarm device provided by the present application.
  • Figure 9 is a schematic structural view of the analysis alarm unit of Figure 8.
  • FIG. 10 is a schematic structural view of an embodiment of a flow cytometer provided by the present application.
  • Figure 11 is a schematic view showing a specific structure of the detecting device of Figure 10;
  • Figure 12 is a schematic illustration of an embodiment of obtaining a predetermined feature area by dynamic adjustment in the present application.
  • a scatter plot is a two-dimensional map generated by a flow cytometer, on which two-dimensional feature information of a plurality of particles is distributed, wherein the X coordinate axis and the Y coordinate axis of the scatter plot image represent each particle a property, such as in a scatter plot, the X coordinate axis characterizes the forward scattered light intensity, and the Y coordinate axis characterizes the fluorescence intensity;
  • a population of cells distributed in a region of a scattergram composed of a plurality of particles having the same characteristics, such as a white blood cell population, and a neutrophil population, a lymphocyte population, a monocyte population, and a hobby in white blood cells. Acidic granulocyte population or basophilic granulocyte group.
  • FIG. 1 a schematic diagram of a main flow of an embodiment of a nucleated red blood cell alarm method provided by the present application is shown; in this embodiment, the nucleated red blood cell alarm method is used for the presence or absence of a blood sample.
  • Nucleated red blood cells for alarms including the following steps:
  • a flow cytometer and a blood sample containing white blood cells are provided, and then the blood sample is pretreated to obtain a processed blood sample.
  • the blood sample and the reagent having the fluorescent dye and the hemolysis component are mixed in a certain ratio to form a processed blood sample.
  • the reagent is used to dissolve the red blood cells in the blood sample so as not to interfere with the counting of white blood cells and nucleated red blood cells, and the fluorescent dye in the reagent binds the nucleic acids in the white blood cells and the nucleated red blood cells to label the cells, because each type of cells binds the fluorescent dye. The ability to differ will produce different fluorescence information.
  • Step S12 injecting the processed blood sample into a flow cytometer, allowing each cell in the blood sample to pass through a detection area of the flow cytometer, detecting and collecting forward scattered light information of each cell in the blood sample, Side scattered light information and fluorescence information; in this step, the forward scattered light of the cells in the blood sample is obtained from the measurement data of the same channel of the flow cytometer Information, side scattered light information, and fluorescence information; it will be appreciated that in various embodiments, forward scattered light information, side scattered light information, and fluorescence of cells in the blood sample can be simultaneously obtained in one detection step.
  • the forward scattered light information, the side scattered light information, and the fluorescence information of the cells in the blood sample are simultaneously collected.
  • the forward scattered light information, the side scattered light information, and the fluorescence information of the cells in the blood sample may also be obtained in a plurality of detecting steps.
  • the processed blood sample may be divided into two parts of the same amount, and the first The blood sample is injected into the flow cytometer, and each cell in the blood sample is passed through a detection area of the flow cytometer to detect and collect forward scattered light information and fluorescence information of each cell in the blood sample; The second blood sample is injected into the flow cytometer, and each cell in the blood sample is passed through a detection area of the flow cytometer to detect and collect side scatter light information and fluorescence information of each cell in the blood sample.
  • Step S14 generating a side scattered light-fluorescence information scattergram of the blood sample according to the collected side scattered light information and fluorescence information, and distinguishing the lymphocyte group, the monocyte group, and the eosinophil cell population from the white blood cells. , four classifications of neutrophils and basophils.
  • Step S16 generating a forward scattered light-fluorescence information scattergram of the blood sample according to the collected forward scattered light information and fluorescence information, where the scatter plot includes a white blood cell group region.
  • Step S18 acquiring a predetermined feature region of the forward scattered light-fluorescence scattergram, wherein the predetermined feature region is located on the left side of the white blood cell group region, and then counting the forward scattered light-fluorescence scatter generated in step S16
  • the number of characterization cells i.e., cell particles
  • the number of characterization cells in the predetermined feature region in the figure determines whether the number of characterization cells exceeds a threshold value, and performs an alarm process when the judgment result is YES.
  • the position and size of the predetermined feature region may be fixed (ie, predetermined), or may be determined according to a certain position of the white blood cell group and the blood shadow region in the forward scattered light-fluorescence scattergram.
  • the rule can be dynamically adjusted, and the normal blood sample containing no nucleated red blood cells and the abnormal blood sample containing nucleated red blood cells can be separately measured in the same detection system, and the forward scattered light-fluorescence of the two can be obtained by the above method.
  • a scatter plot by contrast, will find a region on the left side of the white blood cell population, a particle swarm in this region of the forward scattered light-fluorescence scatter plot of the abnormal blood sample, and forward scattered light from the normal blood sample -
  • the region of the particle swarm does not appear in this region of the fluorescent scattergram, and the region can be determined as a predetermined feature region.
  • the step S18 includes:
  • the predetermined threshold may be an empirical value obtained according to experimental statistics, and when it is understood, the predetermined threshold may be adjusted according to actual conditions;
  • the alarm processing includes: prompting an alarm by means of text, sound, illumination, or pop-up window.
  • the step S18 further includes: before counting the number of characterization cells in the predetermined feature area:
  • the forward scattered light-fluorescence scattergram is analyzed, and an area on the left side of the white blood cell group region and the upper side of the blood shadow region in the forward scattered light-fluorescence scattergram is determined as a predetermined feature region.
  • FIG. 12 an example of obtaining a predetermined feature area by dynamic adjustment is shown. It can be seen from the forward scattered light-fluorescence scattergram in Fig. 12 that the upper boundary of the white blood cell group region, the left boundary of the white blood cell group region b, and the blood shadow can be obtained according to the position data of the white blood cell group region and the position data of the blood shadow region.
  • the upper boundary of the white blood cell group region may be the upper boundary of the predetermined feature region, or the upper limit d of the fluorescence intensity may be the upper boundary of the predetermined feature region; the left boundary b of the white blood cell group region may be used as a predetermined feature.
  • the right boundary of the region, the line e obtained by shifting the left boundary b of the white blood cell group region by a certain distance may be used as the right boundary of the predetermined feature region; the upper boundary b of the blood shadow region may be used as the lower boundary of the predetermined feature region, or may be
  • the boundary f obtained by shifting the upper boundary c of the blood shadow region by a certain distance is taken as the lower boundary of the predetermined feature region; and the vertical axis g is set as the left boundary of the predetermined feature region.
  • an area surrounded by the boundaries a (or d), b (or e), c (or f), and the vertical axis g may be used as the predetermined feature area.
  • the certain distance from the left shift may be the same as or different from the upward shift, and the value may be determined in advance according to requirements.
  • a feature area may be pre-stored in the flow cytometer and stored with position data of the corresponding white blood cell group area and position data of the blood shadow area.
  • detecting position data of the white blood cell group region and position data of the blood shadow region in the forward scattered light-fluorescence scattergram when the position data and the flow If the corresponding data pre-stored by the cytometer is not the same (exceeding a certain error range), then The current white blood cell group region position data and the blood shadow region position data determine the position of the nucleated red blood cell alarm feature region (ie, the predetermined feature region).
  • the current nucleated red blood cell alarm feature area may be determined according to the manner described in the previous paragraph.
  • the preset area is circular or elliptical, the shape may be determined by other methods (such as central displacement).
  • the predetermined feature area position pre-stored in the system is used as the forward scattered light of the blood sample. - The position of the predetermined feature area of the fluoroscopic map, so that complicated calculations can be avoided.
  • the "scatter plot" used in this article can be either a visual graph or a collection of unvisualized data, as long as the above method can be implemented.
  • the nucleated red blood cell alarm method used in the present application mainly uses the characterization of the cell population for the alarm of nucleated red blood cells.
  • the applicant analyzed a large number of data collected from the classification of forward scattered light, side scattered light and fluorescence information during white blood cell classification, and found that when using side scattered light information and fluorescence information to achieve four classifications of white blood cells , a relatively fixed region on the forward scattered light-fluorescence scatter plot obtained by the corresponding detection, a good correlation between a cell population within the blood sample and whether the blood sample is clinically nucleated red blood cell sample.
  • the cell population is a characterized cell population of nucleated red blood cells.
  • the characterization of the cell population can be used to perform nucleated red blood cell alarms while the white blood cells are classified.
  • these red blood cells are present in blood samples because of the presence of nucleated red blood cells in blood samples from patients with severe diseases such as hemolytic anemia, malignancy, erythroleukemia, and myelofibrosis. There may be some degree of abnormality, such as variations in cell membranes. These anomalies have certain anti-hemolytic properties when the reagents are applied. Therefore, these red blood cells generate weak forward scattered light signals when passing through the flow cytometer.
  • red blood cells such as cytochromes and nuclei Flavin and the like may non-specifically bind to fluorescent dyes, so they also exhibit weak fluorescent signals.
  • the scatters generated by these abnormal red blood cells through the flow cytometer will appear in a specific region of the forward scattered light-fluorescence scatter plot, so it can be detected by the forward scattered light-fluorescence scatter plot. Whether a characteristic cell population is present in a particular region is used to determine whether the blood sample has clinically significant nucleated red blood cells.
  • the characteristic particle group can be well distinguished from white blood cells. For ease of understanding, two embodiments will be described below.
  • a first reagent for counting leukocyte counts is prepared as follows, the first reagent consisting of the following components:
  • Fluorescent dye of structural formula A 0.5ppm Mercapto bromide isoquinoline 0.4g/L Dodecyl alcohol polyoxyethylene (23) ether 1.3g/L Methanol 50g/L Sodium dihydrogen phosphate 3g/L Disodium phosphate 4.8g/L
  • step S12 white blood cells are detected by a laser flow method (light source: red semiconductor laser, wavelength 640 nm).
  • the fluorescence intensity information of the cells was measured by side fluorescence with a measurement angle of 90 degrees, and the side scattered light intensity information of the cells was measured by using side-scattering light with an angle of 90 degrees, and the forward direction of the measurement angle was 2-8 degrees.
  • the scattered light measures the forward scattered light information of the cells, thereby obtaining forward scattered light information, side scattered light information, and fluorescence information for each of the two blood samples.
  • a side-scattered light-fluorescence scatter plot of normal blood samples and nucleated red blood cell blood samples (shown in Figures 2A and 3A) is generated, respectively, and white blood cells are differentiated from lymphocytes.
  • Figure 4 is a forward scattered light-fluorescence three-dimensional scattergram of a nucleated red blood cell blood sample showing the presence of a characteristic cell population associated with nucleated red blood cells in a particular region.
  • a second reagent is formulated in this embodiment using a fluorescent dye B of a different structure, the second reagent consisting of the following components:
  • Fluorescent dye of structural formula B 0.5ppm Tetradedyltrimethylammonium chloride 0.3g/L Dodecyl alcohol polyoxyethylene (23) ether 1.3g/L Phthalate 3.0g/L Tris 2.4/L The pH was adjusted to 7.0 with hydrochloric acid.
  • step S10 20 ⁇ l of each of the normal blood sample and the anticoagulant known as the nucleated red blood cell blood sample are separately added to the above 1 ml of the second reagent, and the temperature is maintained at 25 ° C. Under the conditions, mix for 25 seconds.
  • step S12 white blood cells are detected by a laser flow method (light source: red semiconductor laser, wavelength 640 nm).
  • the fluorescence intensity information of the cells was measured by side fluorescence with a measurement angle of 90 degrees, and the side scattered light intensity information of the cells was measured by using side-scattering light with an angle of 90 degrees, and the forward direction of the measurement angle was 2-8 degrees.
  • the scattered light measures the forward scattered light information of the cells, thereby obtaining forward scattered light information, side scattered light information, and fluorescence information for each of the two blood samples.
  • a side-scattering light-fluorescence scatter plot of normal blood samples and nucleated red blood cell blood samples is generated (as shown in FIG. 5A and FIG. 6A), and white blood cells are differentiated from lymphocytes.
  • Figure 7 is a forward scattered light-fluorescence three-dimensional scattergram of a nucleated red blood cell blood sample showing the presence of a characteristic cell population associated with nucleated red blood cells in a particular region.
  • the method of the embodiment of the present application can be implemented by a flow cytometer and its host, and more specifically, it can be implemented by running software.
  • the method provided by the embodiment of the present application is not directly related to the reagent used, as long as the white blood cell four classification can be clearly realized in the side scattered light-fluorescence scatter diagram, according to the forward scattered light at this time. Such a characteristic region appears in the forward scattered light-fluorescence scattergram generated by the information and fluorescence information above the blood shadow or to the left of the white blood cell region.
  • the applicant conducted a series of comparative experiments.
  • the applicant collected 1229 blood samples and used the H20-A2 method recommended by CLSI (Clinical and Laboratory Standards Institute) for artificial microscopy.
  • the results of the microscopic examination were based on the microscopic examination by the International Hematology Reexamination Experts Group.
  • the positive standard is a positive result when the smear of the blood smear is NRBC ⁇ 1%, otherwise it is a negative result.
  • 131 were positive samples (ie, the ratio of nucleated red blood cells to white blood cells was greater than or equal to 1%), and 1098 samples were negative samples.
  • the 1229 blood samples were detected by a flow cytometry analyzer, and the samples which may contain nucleated red blood cells were alarmed by the method provided in the examples of the present application, and the alarm information was given to 180 samples, of which 1049 samples were obtained. No alarm information is given;
  • each sample in the alarm result is compared with the artificial microscopic examination result.
  • 117 are positive samples and 63 are negative samples; in 1049 unalarmed samples Among them, 14 were positive samples and 1035 were negative samples.
  • the specific comparison results are shown in Table 1.
  • the true positive rate, the true negative rate, the false negative rate and the false positive rate are obtained by the following formula:
  • False positive rate false positive number / (true negative number + false positive number) * 100%
  • False negative rate false negative number / (true positive number + false negative number) * 100%.
  • a nucleated red blood cell alarm device is also provided. As shown in FIGS. 8 to 9, an embodiment of a nucleated red blood cell alarm device is shown. In this embodiment, the nucleated red blood cell alarm device 1 is used to alarm whether or not nucleated red blood cells are present in a blood sample. , which includes:
  • the sample information acquiring unit 12 is configured to acquire forward scattered light information, side scattered light information, and fluorescence information when the cells in the blood sample pass through the detection area of the flow cytometer; specifically, the sample information acquiring unit 12 is The forward scattered light information, the side scattered light information, and the fluorescence information of the cells in the blood sample are obtained from the measurement data of the same channel.
  • the scattergram generating unit 14 is configured to generate a side scattered light-fluorescence scattergram according to the side scattered light information and the fluorescence information acquired by the sample information acquiring unit 12, and distinguish the white blood cells into at least a lymphocyte group and a monocyte Four groups of groups, eosinophils, neutrophils, and basophils, and forward scattered light information and fluorescence information acquired by the sample information acquiring unit 12 to generate forward scattering of blood samples a light-fluorescence scattergram, wherein the forward scattered light-fluorescence scattergram includes a white blood cell population region;
  • the analysis alarm unit 16 is configured to obtain a predetermined feature region of the forward scattered light-fluorescence scattergram to count the number of characterization cells in the predetermined feature region, determine whether the number of characterization cells exceeds a threshold, and determine that the result is The alarm processing is performed, wherein the predetermined feature area is located on the left side of the white blood cell group.
  • the analysis alarm unit 16 includes:
  • the feature region determining unit 160 is configured to analyze the forward scattered light-fluorescence scattergram, and determine a region on the left side of the white blood cell group region and the upper side of the blood shadow region in the forward scattered light-fluorescence scattergram as a predetermined feature region. ;
  • a counting unit 162 configured to count the number of characterization cells in the predetermined feature area
  • Comparing unit 164 configured to compare the number of characterization cells with a predetermined threshold
  • the determining unit 166 is configured to determine that nucleated red blood cells are present in the blood sample when the number of characterization cells is greater than a predetermined threshold;
  • the alarm processing unit 168 is configured to perform an alarm process after determining that the nucleated red blood cells are present in the blood sample, and the alarm processing includes: prompting an alarm by using a text, a sound, a light, or a pop-up window.
  • the analysis alarm unit 16 may not include the feature area determining unit 160.
  • the embodiment of the present application provides a schematic structural diagram of a detection device in a flow cytometer, and the flow cytometer includes:
  • the pretreatment device 30 is configured to pretreat the blood sample to obtain the processed blood sample, wherein the pretreatment comprises at least fluorescent labeling the cells in the blood sample;
  • the detecting device 20 is configured to pass the cells in the processed blood sample one by one through the detection area, and detect forward scattered light information, side scattered light information and fluorescence information of the cells in the blood sample. Specifically, the detecting device 20 is in the same Detecting and collecting forward scattered light information, side scattered light information, and fluorescence information of cells in the blood sample in one channel;
  • the analyzing device 40 is configured to generate a forward scattered light-fluorescence scattergram of the blood sample according to the forward scattered light information, the side scattered light information and the fluorescence information detected by the detecting device, and obtain the forward scattered light- a predetermined feature area of the fluorescent scattergram, and counting the number of characterization cells in the predetermined feature area, determining whether the number of characterization cells exceeds a threshold, and performing an alarm process when the determination result is YES, wherein the forward scattered light-fluorescence
  • the scattergram includes a white blood cell group region, the predetermined feature region being located on the left side of the white blood cell group region, the position and size of the predetermined feature region being fixed or dynamically set.
  • the analysis device 40 can be implemented in a host comprising the nucleated red blood cell alarm device 1 as described above with reference to Figures 8-9.
  • the analysis device 40 can be implemented in a host comprising the nucleated red blood cell alarm device 1 as described above with reference to Figures 8-9.
  • the detecting device 20 includes a light source 1025, a flow chamber 1022 as a detection area, a forward scattered light collecting device 1023 disposed on the optical axis, and a side scattered light collecting device 1026 disposed at the side of the optical axis. And a lateral fluorescent scattered light collecting device 1027.
  • the cells of the 30-treated blood sample are sequentially passed through the conduit 3 into the flow chamber 1022 (detection zone), wherein the forward scattered light collecting means 1023, the side scattered light collecting means 1026 and the lateral fluorescence scattering in the same channel
  • the light collecting device 1027 sequentially detects and collects the forward scattered light, the side scattered light, and the side fluorescent scattered light information of each cell, and transmits the information to the analyzing device 40 via the communication interface 4.
  • each cell in the blood sample is detected using a flow cytometer to obtain side scattered light, forward scattered light, and fluorescence information of the cell.
  • the white blood cells in the blood sample are divided into four categories by forming a side scattered light-fluorescence scattergram; by forming a forward scattered light-fluorescence scattergram, the scatter plot is located in the left region of the white blood cell group Characterization of specific regions on the side of the cells for counting analysis can achieve alarms for nucleated red blood cells, that is, the function of white blood cell four classification and nucleated red blood cell alarm can be realized in one channel measurement.
  • the white blood cells when the dedicated nucleated red blood cell reagent is not needed, and the number of nucleated red blood cells is not determined, the white blood cells can be classified and detected, and a more accurate nucleated red blood cell alarm can be provided, so that the measurement is not increased.
  • important information can be provided quickly and easily for clinical screening.
  • the storage medium may be a magnetic disk, an optical disk, a read-only memory (ROM), or a random access memory (RAM).

Abstract

一种有核红细胞报警装置、流式细胞分析仪及相应的有核红细胞报警方法，用于对血液样本中是否存在有核红细胞进行报警，该报警装置包括：样本信息获取单元(12)，用于获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；散点图生成单元(14)，用于根据所述侧向散射光信息和所述荧光信息，生成侧向散射光-荧光散点图，将白细胞至少区分为四分类，以及根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；分析报警单元(16)，用于获取位于所述白细胞群区域的左侧的预定特征区域，统计所述预定特征区域内的表征细胞的数量，在表征细胞的数量超过阈值时进行报警处理。该有核红细胞报警装置、报警方法以及流式细胞分析仪可以在对白细胞进行分类检测的同时实现对有核红细胞的报警。

Description

一种有核红细胞报警方法、装置及流式细胞分析仪 技术领域
本申请涉及医疗设备领域，尤其涉及一种有核红细胞报警方法、装置及流式细胞分析仪。
背景技术
有核红细胞(nucleated red blood cell，NRBC),即幼稚红细胞，正常情况下存在于骨髓中，因此正常人血液样本(除骨髓之外的血液)不会出现有核红细胞。血液样本中出现有核红细胞是由于幼稚的骨髓红系细胞释放入血液样本所致，它在血液样本中的出现与血液疾病相关。因此，临床上需要检查血液中是否存在有核红细胞。
近年来，应用流式细胞计数原理对血液细胞进行计数和分类的流式细胞分析仪相继问世。由于其操作简便、快速、结果准确及精密度高，大大促进了血液检验技术的发展并为临床医学提供了更多的诊断信息。流式细胞分析仪的原理是通过接收激光照射细胞产生光电信号，然后，将激光照射细胞产生的光电信号通过例如散点图等的形式表现出来，供后续分析。其散射光信号和荧光信号反映细胞的物理化学特征，如细胞的大小、颗粒度和抗原分子的表达情况等。
在现有流式细胞分析仪中，对白细胞分类和计数通常采用电阻抗法、激光散射法等方法。这些方法通常需要先对血液样本中的红细胞进行溶解处理，然后将检测样品通过检测器，利用各个白细胞电信号或者光学信号的差异将其分成几类，例如可以分成中性粒细胞，淋巴细胞，单核细胞，嗜酸性细胞或嗜碱性细胞等。
在测定白细胞的过程中，有核红细胞的存在常常会对白细胞测量结果造成干扰。在如前述的测量方法中，对有核红细胞检测获到的信号和淋巴细胞比较相似，不易区分。这样首先会对白细胞的计数分类造成影响；另外，更为重要的是，如果不能提示样本中可能出现了有核红细胞，也容易造成漏诊，贻误疾病的最佳治疗时机。
在现有的技术中，为了实现对有核红细胞的检测，一般需要采用有核红细胞的专用检测试剂和方法。因为需要使用专用的试剂体系和检测装置对有 核红细胞进行计数。这种方法虽然可以实现有核红细胞的计数，但是使得检测仪器的体积和复杂度增加，也会增加临床检验的成本。
发明内容
为了消除现有技术的上述缺陷，本申请提出了一种有核红细胞报警方法、装置及流式细胞分析仪，可以在对白细胞进行分类检测的同时实现对有核红细胞进行报警。
为了解决上述技术问题，本申请实施例的一方面提供了一种有核红细胞报警方法，用于对血液样本中是否存在有核红细胞进行报警，包括如下步骤：
获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；
根据所述侧向散射光信息和所述荧光信息，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类；
根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于所述白细胞群区域的左侧。
其中，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理的步骤包括：
计数所述预定特征区域中的表征细胞数量；
将所述表征细胞数量与一预定阈值进行比较；
如果所述表征细胞数量大于所述预定阈值，则判定所述血液样本中存在有核红细胞，并进行报警处理。
其中，获取所述前向散射光-荧光散点图的预定特征区域的步骤包括：
分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
其中，在获取所述血液样本中细胞经过流式细胞仪检测区时的前向散射光信息、侧向散射光信息和荧光信息的步骤中，从同一个通道的测定数据中 获取所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
其中，所述报警处理为：通过文本、声音、发光或弹窗方式进行报警提示。
相应地，本申请的另一方面还提供一种有核红细胞报警装置，用于对血液样本中是否存在有核红细胞进行报警，包括：
样本信息获取单元，用于获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；
散点图生成单元，用于根据所述侧向散射光信息和所述荧光信息，生成侧向散射光-荧光散点图，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，以及根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
分析报警单元，用于获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于所述白细胞群区域的左侧。
其中，所述分析报警单元还包括：
特征区域确定单元，用于分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
其中，所述分析报警单元包括：
计数单元，用于计数所述预定特征区域中的表征细胞数量；
比较单元，用于将所述表征细胞数量与一预定阈值进行比较；
判定单元，用于在所述表征细胞数量大于所述预定阈值时，判定所述血液样本中存在有核红细胞；
报警处理单元，用于在判定所述血液样本中存在有核红细胞后，进行报警处理。
其中，所述样本信息获取单元从同一个通道的测定数据中获取所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
相应地，本申请的另一方面还提供一种流式细胞分析仪，包括：
采样装置，用于吸取血液样本，所述血液样本中至少含有白细胞；
预处理装置，用于对所述血液样本进行预处理，获得处理后的血液样本，所述预处理包括对所述血液样本中的细胞进行荧光标记；
检测装置，用于使所述处理后的血液样本中的细胞逐个通过检测区，检测所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息；
分析装置，根据所述检测装置所检测到的前向散射光信息、侧向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，获取所述前向散射光-荧光散点图的预定特征区域，并统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述前向散射光-荧光散点图包括白细胞群区域，所述预定特征区域位于所述白细胞群区域的左侧。
其中，所述分析装置包括：
样本信息获取单元，用于获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；
散点图生成单元，用于根据样本信息获取单元获取的侧向散射光信息和荧光信息，生成侧向散射光-荧光散点图，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，以及根据样本信息获取单元获取的前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
分析报警单元，用于获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于所述白细胞群区域的左侧。
其中，所述分析报警单元还包括：
特征区域确定单元，用于分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
其中，所述检测装置在同一个通道中检测并收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
相应地，本申请的另一方面，还提供一种有核红细胞报警方法，用于对血液样本中是否存在有核红细胞进行报警，包括如下步骤：
提供流式细胞分析仪和含有白细胞的血液样本；
对所述血液样本进行预处理，获得处理后的血液样本，所述预处理至少包括对血液样本中的细胞进行荧光标记；
将所述处理后的血液样本注入流式细胞分析仪，收集细胞的前向散射光信息、侧向散射光信息和荧光信息；
根据所述侧向散射光信息和所述荧光信息，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类；
根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，所述前向散射光-荧光散点图包括白细胞群区域；
获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理；其中，所述预定特征区域位于所述白细胞群区域的左侧。
其中，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理的步骤包括：
计数所述预定特征区域中的表征细胞数量；
将所述表征细胞数量与一预定阈值进行比较；
如果所述表征细胞数量大于所述预定阈值，则判定所述血液样本中存在有核红细胞，并进行报警处理。
其中，将处理后血液样本注入流式细胞分析仪，收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息的步骤具体为：
当细胞通过流式细胞仪检测区时，同时收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
其中，获取所述前向散射光-荧光散点图的预定特征区域的步骤包括：
分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
其中，获取所述前向散射光-荧光散点图的预定特征区域的步骤包括：
提供正常血液样本和异常血液样本，所述正常血液样本中至少含有白细胞，所述异常血液样本中至少含有白细胞和有核红细胞；
对所述正常血液样本和异常血液样本进行预处理，获得处理后的正常血液样本和处理后的异常血液样本，所述预处理至少包括对血液样本中的细胞进行荧光标记；
将所述处理后的正常血液样本和处理后的异常血液样本分别注入流式细胞分析仪，收集细胞的前向散射光信息、侧向散射光信息和荧光信息；
根据所述侧向散射光信息和所述荧光信息，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，根据所述前向散射光信息和荧光信息，生成所述正常血液样本的前向散射光-荧光散点图，和异常血液样本的前向散射光-荧光散点图；
比较所述正常血液样本的前向散射光-荧光散点图和异常血液样本的前向散射光-荧光散点图，将在异常血液样本的前向散射光-荧光散点图的白细胞群区域左侧出现粒子群且正常血液样本的前向散射光-荧光散点图的白细胞群区域左侧未出现粒子群的区域，确定为预定特征区域。
实施本申请的实施例，具有如下有益效果：
在本申请实施例中，通过将血液样本使用试剂进行预处理，使用流式细胞分析仪对血液样本中的各细胞进行检测，获得细胞的侧向散射光、前向散射光和荧光信息。通过形成侧向散射光-荧光散点图，将所述血液样本中的白细胞区分为四个类别；通过形成前向散射光-荧光散点图，对该散点图中的位于白细胞群区域左侧的特定区域的表征细胞进行计数分析，可以实现对有核红细胞的报警，也就是在一个通道的测量中可以实现白细胞四分类和有核红细胞报警的功能。
在本申请实施例中，在无需采用专用的有核红细胞试剂，不用测定有核红细胞数量的情况下，可以在对白细胞进行分类检测的同时提供较为准确的有核红细胞报警，从而在不增加测量复杂度和成本的情况下，可以快速简便地为临床筛检提供了重要信息。
附图说明
为了更清楚地说明本申请实施例或现有技术中的技术方案，下面将对实施例或现有技术描述中所需要使用的附图作简单地介绍，显而易见地，下面 描述中的附图仅仅是本申请的一些实施例，对于本领域普通技术人员来讲，在不付出创造性劳动的前提下，还可以根据这些附图获得其他的附图。
图1是本申请提供的一种有核红细胞报警方法的一个实施例的主流程示意图；
图2A为本申请中采用第一种试剂时对一个不含有核红细胞的血液样本进行分析获得的侧向散射光-荧光散点图示意图；
图2B为本申请中采用第一种试剂时对一个不含有核红细胞的血液样本进行分析获得的前向散射光-荧光散点图示意图；
图3A为本申请中采用第一种试剂时对一个含有核红细胞的血液样本进行分析获得的侧向散射光-荧光散点图示意图；
图3B为本申请中采用第一种试剂时对一个含有核红细胞的血液样本进行分析获得的前向散射光-荧光散点图示意图；
图4为对应图3A和图3B的三维散点图示意图；
图5A为本申请中采用第二种试剂时对一个不含有核红细胞的血液样本进行分析获得的侧向散射光-荧光散点图示意图；
图5B为本申请中采用第二种试剂时对一个不含有核红细胞的血液样本进行分析获得的前向散射光-荧光散点图示意图；
图6A为本申请中采用第二种试剂时对一个含有核红细胞的血液样本进行分析获得的侧向散射光-荧光散点图示意图；
图6B为本申请中采用第二种试剂时对一个含有核红细胞的血液样本进行分析获得的前向散射光-荧光散点图示意图；
图7为对应图6A和图6B的三维散点图示意图；
图8是本申请提供的一种有核红细胞报警装置的一个实施例的结构示意图；
图9是图8中分析报警单元的结构示意图；
图10是本申请提供的一种流式细胞分析仪的一个实施例的结构示意图；
图11是图10中检测装置的一个具体结构示意图；
图12是本申请中通过动态调整获得预定特征区域的实施例的示意图。
具体实施方式
下面将结合本申请实施例中的附图，对本申请实施例中的技术方案进行 清楚、完整地描述，显然，所描述的实施例仅仅是本申请一部分实施例，而不是全部的实施例。基于本申请中的实施例，本领域普通技术人员在没有作出创造性劳动前提下所获得的所有其他实施例，都属于本申请保护的范围。
下面参考附图对本申请的优选实施例进行描述。
为便于后续说明的方便，首先需要对下文中所涉及的一些术语进行简要说明如下：
散点图，是由流式细胞分析仪生成的一种二维图，其上分布有多个粒子的二维特征信息，其中散点图的X坐标轴和Y坐标轴均表征每个粒子的一种特性，例如在一个散点图中，X坐标轴表征前向散射光强度，而Y坐标轴表征荧光强度；
细胞群，分布在散点图的某一区域，由具有相同特性的多个粒子形成的粒子团，例如白细胞群，以及白细胞中的中性粒细胞群、淋巴细胞群、单核细胞群、嗜酸性粒细胞群或嗜碱性粒细胞群等。
如图1所示，示出了本申请提供的一种有核红细胞报警方法的一个实施例的主流程示意图；在该实施例中，该有核红细胞报警方法，用于对血液样本中是否存在有核红细胞进行报警，包括如下步骤：
步骤S10，提供流式细胞分析仪和含有白细胞的血液样本，然后对血液样本进行预处理，获得处理后的血液样本。具体地，将所述血液样本与具有荧光染料与溶血成份的试剂按一定比例进行混合，形成处理后的血液样本。其中，采用试剂溶解血液样本中的红细胞，使之不干扰白细胞和有核红细胞的计数，同时试剂中的荧光染料对白细胞和有核红细胞中的核酸结合以标记细胞，由于各类型细胞结合荧光染料的能力有差异，会产生不同的荧光信息。另外，由于各类型细胞的大小不同，会产生不同的前向散射光信息，由于细胞内形态或复杂程度不同，会产生不同的侧向散射光信息。可以理解的是，通过利用流式细胞仪，让细胞逐个通过检测区，收集这些信息，然后通过分析，可以在散点图上区分各类型细胞。
步骤S12，将处理后的血液样本注入流式细胞分析仪，让该血液样本中各细胞经过流式细胞分析仪的检测区，检测并收集该血液样本中每一个细胞的前向散射光信息、侧向散射光信息和荧光信息；在该步骤中，是从流式细胞分析仪的同一个通道的测定数据中获取该血液样本中细胞的前向散射光 信息、侧向散射光信息和荧光信息；可以理解的是，在不同的实施例中，可以在一次检测步骤中同时获得该血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息，即当细胞通过流式细胞仪检测区时，同时收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。也可以在多次检测步骤中获得该血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息，例如，可以将经处理后的血液样本分成相同份量的两份，将第一份血液样本注入流式细胞分析仪，让该血液样本中各细胞经过流式细胞分析仪的检测区，检测并收集该血液样本中每一个细胞的前向散射光信息以及荧光信息；而随后将第二份血液样本注入流式细胞分析仪，让该血液样本中各细胞经过流式细胞分析仪的检测区，检测并收集该血液样本中每一个细胞的侧向散射光信息以及荧光信息。
步骤S14，根据收集到的侧向散射光信息和荧光信息，生成该血液样本的侧向散射光-荧光信息散点图，将白细胞区分淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类。
步骤S16，根据收集到的前向散射光信息和荧光信息，生成该血液样本的前向散射光-荧光信息散点图，该散点图上包括白细胞群区域。
步骤S18，获取所述前向散射光-荧光散点图的预定特征区域，其中，所述预定特征区域位于白细胞群区域的左侧，然后统计步骤S16中生成的前向散射光-荧光散点图中的该预定特征区域内的表征细胞(即细胞粒子)的数量，判断表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理。
可以理解的是，该预定特征区域的位置与大小可以是固定的(即预先确定好的)，也可以是随前向散射光-荧光散点图中白细胞群、血影区的位置根据一定的规则动态调整获得，也可以在同一个检测体系中分别测定不含有核红细胞的正常血液样本和已知含有有核红细胞的异常血液样本，通过上述的方式，获得两者的前向散射光-荧光散点图，通过对比，将会在白细胞群的左侧找到一个区域，在异常血液样本的前向散射光-荧光散点图的该区域出现粒子群，而正常血液样本的前向散射光-荧光散点图的该区域未出现粒子群的区域，该区域可确定为预定特征区域。
具体地，当该预定特征区域的位置与大小是固定情形时，则该步骤S18包括：
计数该预定特征区域中的表征细胞数量；
将表征细胞数量与一预定阈值进行比较，其中该预定阈值可以是根据实验统计获得的经验值，当可以理解的，该预定阈值可以根据实际情况进行调整；
如果该表征细胞数量大于预定阈值，则确定血液样本中存在有核红细胞，并进行报警处理，该报警处理包括：通过文本、声音、发光或弹窗等方式进行报警提示。
当该预定特征区域的位置与大小是动态调整获得的情形时，则该步骤S18在计数该预定特征区域中的表征细胞数量之前进一步包括：
分析该前向散射光-荧光散点图，将该前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
具体地，可以参见图12所示，示出了一种通过动态调整获得预定特征区域的例子。从图12中的前向散射光-荧光散点图可以看出，根据白细胞群区域的位置数据和血影区域的位置数据可以获得白细胞群区域上边界a、白细胞群区域左边界b、血影区域上边界c。在一个例子中，可以将该白细胞群区域上边界a作为预定特征区域的上边界，也可以将是荧光强度的上限d作为预定特征区域的上边界；可以将白细胞群区域左边界b作为预定特征区域的右边界，也可以将该白细胞群区域左边界b左移一定距离获得的线e作为预定特征区域的右边界；可以将血影区域上边界c作为预定特征区域的下边界，也可以将该血影区域上边界c上移一定距离获得的边界f作为预定特征区域的下边界；而将纵轴g设为预定特征区域的左边界。从而根据预定特征区域的上边界、下边界、左边界以及右边界动态来确定预定特征区域的大小与位置。在如图12中，由边界a(或d)、b(或e)、c(或f)以及纵轴g所围成的区域可以作为预定特征区域。其中，左移的一定距离与上移的一定距离可以相同，也可以不同，其数值预先根据需要确定。
具体地，在一些实施例中，可以在流式细胞计数仪中预存一个特征区域，且存储有对应的白细胞群区域的位置数据和血影区域的位置数据。在生成当前血液样本的前向散射光-荧光散点图时，检测该前向散射光-荧光散点图中的白细胞群区域的位置数据和血影区域的位置数据，当这些位置数据与流式细胞计数仪预存的相应的数据不同时(超出一定的误差范围)，则根据 当前白细胞群区域位置数据和血影区域位置数据，确定有核红细胞报警特征区域(即预定特征区域)的位置。当预设区域为矩形时，可以按上段介绍的方式确定当前有核红细胞报警特征区域，当预设区域为圆形或椭圆形等其他形状时，可以用其他方式(例如中心位移等方法)确定当前有核红细胞报警特征区域。当检测到的位置数据与流式细胞计数仪预存的相应的数据基本相同时(例如在一定的误差范围之内)，则将系统中预存的预定特征区域位置作为该血液样本的前向散射光-荧光散点图的预定特征区域的位置，这样可以避免复杂的计算。
可以理解的是，以上的步骤的标号仅为表述方便，并不限定先后顺序。
本文中使用的“散点图”即可以是可视化的图形，也可以是未可视化的数据的集合，只要能实现以上方法即可。
本申请所采用的有核红细胞报警方法，主要是利用表征细胞群来进行有核红细胞的报警。申请人经过深入的研究，分析了大量白细胞分类时的收集到的前向散射光、侧向散射光和荧光信息的数据，发现当利用侧向散射光信息和荧光信息对白细胞实现四分类的时候，在相应检测获得的前向散射光-荧光的散点图上的一个相对固定的区域，其内的一个细胞群与该血液样本是否临床意义上的有核红细胞样本之间具有良好的相关性，申请人称该细胞群为有核红细胞的表征细胞群。因此，可以在白细胞分类的同时利用该表征细胞群进行有核红细胞的报警。虽然不希望受理论约束，但申请人猜测，由于有核红细胞常出现在溶血性贫血、恶性肿瘤、红白血病、骨髓纤维化等严重疾病的患者的血液样本中，这些存在于血液样本中的红细胞可能存在一定程度的异常，如细胞膜的变异等。这些异常在试剂作用时会出现一定的抗溶血特性，因此这些红细胞在经过流式细胞分析仪时会产生微弱的前向散射光的信号；同时，这些红细胞内部的某些成分如细胞色素、核黄素等会与荧光染料发生非特异性结合，故它们也会出现微弱的荧光信号。这样，这些异常红细胞经过流式细胞分析仪所产生的散点就会出现在前向散射光-荧光散点图的一个特定区域内，故可以通过检测在前向散射光-荧光散点图的特定区域是否出现表征细胞群，来判断该血液样本是否存在临床意义上的有核红细胞。
具体来说，在本申请的一个具体实施例方式中，当侧向散射光-荧光散 点图上将白细胞能够区分成四分群(淋巴细胞群、单核细胞群、嗜酸性粒细胞群、中性粒细胞加嗜碱性粒细胞群)时，切换至前向散射光-荧光散点图，则该特征粒子群就能和白细胞很好的区分。为了便于理解，下述采用两个实施例来进行说明。
实施例一
采用本申请前述描述的流程步骤，其中在步骤S10的预处理中，采用如下配制用于白细胞分类计数的第一试剂，该第一试剂由以下成分组成：
结构式A的荧光染料 0.5ppm
癸基溴化异喹啉 0.4g/L
十二烷基醇聚氧乙烯(23)醚 1.3g/L
甲醇 50g/L
磷酸二氢钠 3g/L
磷酸氢二钠 4.8g/L
Figure PCTCN2014095905-appb-000001
将正常血液样本和已知为有核红细胞血液样本的抗凝血各20μl分别加入到上述的1ml的第一试剂中，在温度保持25℃的条件下，混合25秒。
然后在步骤S12中，用激光流式法(光源：红色半导体激光，波长640nm)检测白细胞。采用测定角度为90度的侧向荧光测定细胞的荧光强度信息，并采用测定角度为90度的侧向散射光测定细胞的侧向散射光强度信息，采用测定角度为2-8度的前向散射光测定细胞的前向散射光信息，从而获得两份血液样本中每一个细胞的前向散射光信息、侧向散射光信息和荧光信息。
根据收集到的侧向散射光信息和荧光信息，分别生成正常血液样本和有核红细胞血液样本的侧向散射光-荧光散点图(如图2A和图3A所示)，将白细胞区分淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类；将收集到的前向散射光信息和荧光信息，生成两份 血液样本的前向散射光-荧光散点图(如图2B和图3B所示)，从已知为有核红细胞血液样本的前向散射光-荧光信息散点图(图3B)的特定的区域出现表征细胞群(认为其是与有核红细胞相关的粒子群)。图4是有核红细胞血液样本的前向散射光-荧光三维散点图，可见在特定区域出现了与有核红细胞相关的表征细胞群。
实施例二
在该实施例中使用不同结构的荧光染料B来配制第二试剂，该第二试剂由以下成分组成：
结构式B的荧光染料 0.5ppm
十四基三甲基氯化铵 0.3g/L
十二烷基醇聚氧乙烯(23)醚 1.3g/L
邻苯二甲酸 3.0g/L
Tris 2.4/L
用盐酸将pH调节到7.0。  
Figure PCTCN2014095905-appb-000002
采用本申请前述描述的流程步骤，在步骤S10中，将正常血液样本和已知为有核红细胞血液样本的抗凝血各20μl分别加入到上述的1ml第二试剂中，在温度保持25℃的条件下，混合25秒。
然后在步骤S12中，用激光流式法(光源：红色半导体激光，波长640nm)检测白细胞。采用测定角度为90度的侧向荧光测定细胞的荧光强度信息，并采用测定角度为90度的侧向散射光测定细胞的侧向散射光强度信息，采用测定角度为2-8度的前向散射光测定细胞的前向散射光信息，从而获得两份血液样本中每一个细胞的前向散射光信息、侧向散射光信息和荧光信息。
根据收集到的侧向散射光信息和荧光信息，分别生成正常血液样本和有核红细胞血液样本的侧向散射光-荧光散点图(如图5A和图6A所示)，将白细胞区分淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类；将收集到的前向散射光信息和荧光信息，生成两份 血液样本的前向散射光-荧光散点图(如图5B和图6B所示)，从已知为有核红细胞血液样本的前向散射光-荧光信息散点图(图6B)的特定的区域出现表征细胞群(认为其是与有核红细胞相关的粒子群)。图7是有核红细胞血液样本的前向散射光-荧光三维散点图，可见在特定区域出现了与有核红细胞相关的表征细胞群。
从上述两个实施例可以看出，在预处理步骤采用不同的试剂，最后均可以在特定区域中出现表征细胞群。
可以理解的，本申请实施例的方法可以通过流式细胞分析仪及其主机实现，更具体地，其可以采用运行软件的方式来实现。另外，可以理解的是，本申请实施例提供方法与所采用的试剂无直接关联，只要在侧向散射光-荧光散点图中能明显实现白细胞四分类时，根据此时的前向散散光信息和荧光信息生成的前向散射光-荧光散点图中，在其血影上方或、及白细胞区左方，就会出现这样的特征区域。
为了进一步地确认本申请实施例所提供的有核红细胞报警方法的效果，申请人进行了一系列的对比实验。申请人收集了1229例血液样本，采用CLSI(Clinical and Laboratory Standards Institute，临床实验室标准委员会)推荐的H20-A2方法进行人工镜检，镜检结果参考国际血液学复检专家组制定的镜检阳性标准，当血涂片人工镜检NRBC≥1％时作为阳性结果，否则作为阴性结果。在这1229个样本中，其中有131个样本为阳性样本(即其所含有的有核红细胞与白细胞的比例大于或等于1％)，1098个样本为阴性样本。
利用流式细胞分析仪对该1229例血液样本进行检测，采用本申请实施例提供的方法对其中可能含有核红细胞的样本进行报警，结果对其中180个样本给出报警信息，对其中1049个样本未给出报警信息；
然后将报警结果中的每个样本与人工镜检结果进行比对，最终统计出，在180个报警的样本中，有117个为阳性样本、63个为阴性样本；在1049个未报警的样本中，有14个为阳性样本、1035个为阴性样本。具体对比结果如表1所示。
表1，采用本申请实施例提供的方法报警结果和人工镜检结果对比
Figure PCTCN2014095905-appb-000003
Figure PCTCN2014095905-appb-000004
其中，真阳率、真阴率、假阴率和假阳率采用如下的计算公式获得：
真阳率＝真阳个数/(真阳个数+假阴个数)*100％；
真阴率＝真阴个数/(真阴个数+假阳个数)*100％；
假阳率＝假阳个数/(真阴个数+假阳个数)*100％；
假阴率＝假阴个数/(真阳个数+假阴个数)*100％。
从上述的结果显示，采用本申请实施所提供的方法对血液样本中进行有核红细胞的检测具有较高的报警灵敏度和特异度，报警准确性较好。
相应地，本申请实施的另一方面，还提供了一种有核红细胞报警装置。如图8至图9所示，示出了一种有核红细胞报警装置的一个实施例，在该实施例中，该有核红细胞报警装置1用于对血液样本中是否存在有核红细胞进行报警，其包括：
样本信息获取单元12，用于获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；具体地，该样本信息获取单元12，是从同一个通道的测定数据中获取血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
散点图生成单元14，用于根据样本信息获取单元12所获取的侧向散射光信息和荧光信息，生成侧向散射光-荧光散点图，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，以及根据样本信息获取单元12所获取的前向散射光信息和荧光信息，生成血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
分析报警单元16，用于获取所述前向散射光-荧光散点图的预定特征区域统计该预定特征区域内的表征细胞的数量，判断表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于白细胞群的左侧。
具体地，分析报警单元16包括：
特征区域确定单元160，用于分析前向散射光-荧光散点图，将前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域；
计数单元162，用于计数预定特征区域中的表征细胞数量；
比较单元164，用于将表征细胞数量与一预定阈值进行比较；
判定单元166，用于在表征细胞数量大于预定阈值时，则判定血液样本中存在有核红细胞；
报警处理单元168，用于在判定血液样本中存在有核红细胞后，进行报警处理，该报警处理包括：通过文本、声音、发光或弹窗方式进行报警提示。
可以理解的是，如果该预定特征区域的位置与大小是固定的，则在分析报警单元16可以不包括特征区域确定单元160。
更多的细节，可以一并参照前述对图1至图7的说明，在此不进行赘述。
如图10所示，是本申请实施例提供一种流式细胞分析仪中的检测装的结构示意图，该流式细胞分析仪包括：
采样装置10，用于吸取包含有白细胞的血液样本；
预处理装置30，用于对血液样本进行预处理，获得处理后的血液样本，其中，该预处理至少包括对血液样本中的细胞进行荧光标记；
检测装置20，用于使处理后的血液样本中的细胞逐个通过检测区，检测血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息，具体地，该检测装置20在同一个通道中检测并收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息；
分析装置40，用于根据检测装置所检测到的前向散射光信息、侧向散射光信息和荧光信息，生成血液样本的前向散射光-荧光散点图，获取所述前向散射光-荧光散点图的预定特征区域，并统计该预定特征区域内的表征细胞的数量，判断表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，前向散射光-荧光散点图中包括白细胞群区域，该预定特征区域位于白细胞群区域的左侧，该预定特征区域的位置与大小是固定的，或者是动态设定的。
其中，该分析装置40可以在一主机中实现，其包括前述如图8-9所描述的有核红细胞报警装置1，更多的细节可参照前述对图8-图9的描述。
具体地，如图11所示，示出了图10中的一个检测装置20的具体结构示意图。从中可以看出，该检测装置20包括：光源1025、作为检测区域的流动室1022、设置在光轴上的前向散射光收集装置1023、设置在光轴侧边的侧向散射光收集装置1026和侧向荧光散射光收集装置1027。经预处理装 置30处理后的血液样本的细胞依次经管道3进入流动室1022(检测区)，其中，在同一个通道中的前向散射光收集装置1023、侧向散射光收集装置1026和侧向荧光散射光收集装置1027依次检测并收集每一细胞的前向散射光、侧向散射光和侧向荧光散射光信息，并经通信接口4传送给分析装置40。
更多的细节，可以一并参照前述对图1至图9的说明，在此不进行赘述。
实施本申请的实施例，具有如下有益效果：
在本申请实施中，通过将血液样本使用试剂进行预处理，使用流式细胞分析仪对血液样本中的各细胞进行检测，获得细胞的侧向散射光、前向散射光和荧光信息。通过形成侧向散射光-荧光散点图，将所述血液样本中的白细胞区分为四个类别；通过形成前向散射光-荧光散点图，对该散点图中的位于白细胞群区域左侧的特定区域的表征细胞进行计数分析，可以实现对有核红细胞的报警，也就是在一个通道的测量中可以实现白细胞四分类和有核红细胞报警的功能。
在本申请实施例中，在无需采用专用的有核红细胞试剂，不用测定有核红细胞数量的情况下，可以在对白细胞进行分类检测的同时提供较为准确的有核红细胞报警，从而在不增加测量复杂度和成本的情况下，可以快速简便地为临床筛检提供了重要信息。
可以理解的是，本领域普通技术人员可以理解实现上述实施例方法中的全部或部分流程，是可以通过计算机程序来指令相关的硬件来完成，的程序可存储于一计算机可读取存储介质中，该程序在执行时，可包括如上述各方法的实施例的流程。其中，的存储介质可为磁碟、光盘、只读存储记忆体(Read-Only Memory，ROM)或随机存储记忆体(Random Access Memory，RAM)等。
本发明实施例中描述的技术特征或操作步骤可以按照任何合适的方式进行组合。本领域内普通技术人员容易理解，本发明实施例描述的方法中的步骤或动作的顺序是可以改变的。因此，除非另有说明要求一定的顺序，在附图或者详细描述中的任何顺序只是为了用作说明的目的，而不是必须的顺序。
以上所揭露的仅为本申请一种较佳实施例而已，当然不能以此来限定本申请之权利范围，因此依本申请权利要求所作的等同变化，仍属本申请所涵盖的范围。

Claims (18)

  1. 一种有核红细胞报警方法，用于对血液样本中是否存在有核红细胞进行报警，其特征在于，包括如下步骤：
    获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；
    根据所述侧向散射光信息和所述荧光信息，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类；
    根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
    获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于所述白细胞群区域的左侧。
  2. 如权利要求1所述的一种有核红细胞报警方法，其特征在于，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理的步骤包括：
    计数所述预定特征区域中的表征细胞数量；
    将所述表征细胞数量与一预定阈值进行比较；
    如果所述表征细胞数量大于所述预定阈值，则判定所述血液样本中存在有核红细胞，并进行报警处理。
  3. 如权利要求1所述的一种有核红细胞报警方法，其特征在于，
    获取所述前向散射光-荧光散点图的预定特征区域的步骤包括：
    分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
  4. 如权利要求1所述的一种有核红细胞报警方法，其特征在于，在获取所述血液样本中细胞经过流式细胞仪检测区时的前向散射光信息、侧向散射光信息和荧光信息的步骤中，从同一个通道的测定数据中获取所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
  5. 如权利要求1所述的一种有核红细胞报警方法，其特征在于，所述报警处理为：通过文本、声音、发光或弹窗方式进行报警提示。
  6. 一种有核红细胞报警装置，用于对血液样本中是否存在有核红细胞进行报警，其特征在于，包括：
    样本信息获取单元，用于获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；
    散点图生成单元，用于根据所述侧向散射光信息和所述荧光信息，生成侧向散射光-荧光散点图，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，以及根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
    分析报警单元，用于获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于所述白细胞群区域的左侧。
  7. 如权利要求6所述的一种有核红细胞报警装置，其特征在于，所述分析报警单元还包括：
    特征区域确定单元，用于分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
  8. 如权要求6所述的一种有核红细胞报警装置，其特征在于，所述分析报警单元包括：
    计数单元，用于计数所述预定特征区域中的表征细胞数量；
    比较单元，用于将所述表征细胞数量与一预定阈值进行比较；
    判定单元，用于在所述表征细胞数量大于所述预定阈值时，判定所述血液样本中存在有核红细胞；
    报警处理单元，用于在判定所述血液样本中存在有核红细胞后，进行报警处理。
  9. 如权利要求6所述的一种有核红细胞报警装置，其特征在于，所述 样本信息获取单元从同一个通道的测定数据中获取所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
  10. 一种流式细胞分析仪，其特征在于，包括：
    采样装置，用于吸取血液样本，所述血液样本中至少含有白细胞；
    预处理装置，用于对所述血液样本进行预处理，获得处理后的血液样本，所述预处理包括对所述血液样本中的细胞进行荧光标记；
    检测装置，用于使所述处理后的血液样本中的细胞逐个通过检测区，检测所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息；
    分析装置，根据所述检测装置所检测到的前向散射光信息、侧向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，获取所述前向散射光-荧光散点图的预定特征区域，并统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理；其中，所述前向散射光-荧光散点图包括白细胞群区域，所述预定特征区域位于所述白细胞群区域的左侧。
  11. 如权利要求10所述的一种流式细胞分析仪，其特征在于，所述分析装置包括：
    样本信息获取单元，用于获取血液样本中的细胞经过流式细胞分析仪检测区时的前向散射光信息、侧向散射光信息和荧光信息；
    散点图生成单元，用于根据样本信息获取单元获取的侧向散射光信息和荧光信息，生成侧向散射光-荧光散点图，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，以及根据样本信息获取单元获取的前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，其中，所述前向散射光-荧光散点图包括白细胞群区域；
    分析报警单元，用于获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理，其中，所述预定特征区域位于所述白细胞群区域的左侧。
  12. 如权利要求11所述的一种流式细胞分析仪，其特征在于，所述分 析报警单元还包括：
    特征区域确定单元，用于分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
  13. 如权利要求10所述的一种流式细胞分析仪，其特征在于，所述检测装置在同一个通道中检测并收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
  14. 一种有核红细胞报警方法，用于对血液样本中是否存在有核红细胞进行报警，其特征在于，包括如下步骤：
    提供流式细胞分析仪和含有白细胞的血液样本；
    对所述血液样本进行预处理，获得处理后的血液样本，所述预处理至少包括对血液样本中的细胞进行荧光标记；
    将所述处理后的血液样本注入流式细胞分析仪，收集细胞的前向散射光信息、侧向散射光信息和荧光信息；
    根据所述侧向散射光信息和所述荧光信息，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类；
    根据所述前向散射光信息和荧光信息，生成所述血液样本的前向散射光-荧光散点图，所述前向散射光-荧光散点图包括白细胞群区域；
    获取所述前向散射光-荧光散点图的预定特征区域，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理；其中，所述预定特征区域位于所述白细胞群区域的左侧。
  15. 如权利要求14所述的一种有核红细胞报警方法，其特征在于，统计所述预定特征区域内的表征细胞的数量，判断所述表征细胞的数量是否超过阈值，并在判断结果为是时进行报警处理的步骤包括：
    计数所述预定特征区域中的表征细胞数量；
    将所述表征细胞数量与一预定阈值进行比较；
    如果所述表征细胞数量大于所述预定阈值，则确定所述血液样本中存在 有核红细胞，并进行报警处理。
  16. 如权利要求14所述一种有核红细胞报警方法，其特征在于，将处理后血液样本注入流式细胞分析仪，收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息的步骤具体为：
    当细胞通过流式细胞仪检测区时，同时收集所述血液样本中细胞的前向散射光信息、侧向散射光信息和荧光信息。
  17. 如权利要求14散射的一种有核红细胞报警方法，其特征在于，获取所述前向散射光-荧光散点图的预定特征区域的步骤包括：
    分析所述前向散射光-荧光散点图，将所述前向散射光-荧光散点图中位于白细胞群区域左侧及血影区域上侧的一个区域确定为预定特征区域。
  18. 如权利要求14所述的一种有核红细胞报警方法，其特征在于，获取所述前向散射光-荧光散点图的预定特征区域的步骤包括：
    提供正常血液样本和异常血液样本，所述正常血液样本中至少含有白细胞，所述异常血液样本中至少含有白细胞和有核红细胞；
    对所述正常血液样本和异常血液样本进行预处理，获得处理后的正常血液样本和处理后的异常血液样本，所述预处理至少包括对血液样本中的细胞进行荧光标记；
    将所述处理后的正常血液样本和处理后的异常血液样本分别注入流式细胞分析仪，收集细胞的前向散射光信息、侧向散射光信息和荧光信息；
    根据所述侧向散射光信息和所述荧光信息，将白细胞至少区分为淋巴细胞群、单核细胞群、嗜酸性粒细胞细胞群、中性粒细胞与嗜碱性粒细胞群的四分类，根据所述前向散射光信息和荧光信息，生成所述正常血液样本的前向散射光-荧光散点图，和异常血液样本的前向散射光-荧光散点图；
    比较所述正常血液样本的前向散射光-荧光散点图和异常血液样本的前向散射光-荧光散点图，将在异常血液样本的前向散射光-荧光散点图的白细胞群区域左侧出现粒子群且正常血液样本的前向散射光-荧光散点图的白细胞群区域左侧未出现粒子群的区域，确定为预定特征区域。
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