WO2018148942A1 - 血液细胞分析方法及血液细胞分析仪 - Google Patents

血液细胞分析方法及血液细胞分析仪 Download PDF

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WO2018148942A1
WO2018148942A1 PCT/CN2017/073971 CN2017073971W WO2018148942A1 WO 2018148942 A1 WO2018148942 A1 WO 2018148942A1 CN 2017073971 W CN2017073971 W CN 2017073971W WO 2018148942 A1 WO2018148942 A1 WO 2018148942A1
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blood cell
information
white blood
platelet
scattergram
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PCT/CN2017/073971
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English (en)
French (fr)
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叶波
胡长松
祁欢
郑文波
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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 CN201780085853.1A priority Critical patent/CN110249223B/zh
Priority to PCT/CN2017/073971 priority patent/WO2018148942A1/zh
Publication of WO2018148942A1 publication Critical patent/WO2018148942A1/zh
Priority to US16/542,267 priority patent/US11501852B2/en
Priority to US17/986,726 priority patent/US20230071301A1/en

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    • GPHYSICS
    • G16INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR SPECIFIC APPLICATION FIELDS
    • G16BBIOINFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR GENETIC OR PROTEIN-RELATED DATA PROCESSING IN COMPUTATIONAL MOLECULAR BIOLOGY
    • G16B45/00ICT specially adapted for bioinformatics-related data visualisation, e.g. displaying of maps or networks
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/02Devices for withdrawing samples
    • G01N1/10Devices for withdrawing samples in the liquid or fluent state
    • G01N1/14Suction devices, e.g. pumps; Ejector devices
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • G01N1/31Apparatus therefor
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N15/14Optical investigation techniques, e.g. flow cytometry
    • G01N15/1456Optical investigation techniques, e.g. flow cytometry without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals
    • G01N15/1459Optical investigation techniques, e.g. flow cytometry 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
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/483Physical analysis of biological material
    • G01N33/487Physical analysis of biological material of liquid biological material
    • G01N33/49Blood
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5005Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
    • G01N33/5094Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for blood cell populations
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/02Devices for withdrawing samples
    • G01N1/10Devices for withdrawing samples in the liquid or fluent state
    • G01N1/14Suction devices, e.g. pumps; Ejector devices
    • G01N2001/1418Depression, aspiration
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/01Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials specially adapted for biological cells, e.g. blood cells
    • G01N2015/016White blood cells
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/01Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials specially adapted for biological cells, e.g. blood cells
    • G01N2015/018Platelets
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N2015/1006Investigating individual particles for cytology
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N15/14Optical investigation techniques, e.g. flow cytometry
    • G01N2015/1402Data analysis by thresholding or gating operations performed on the acquired signals or stored data
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N15/14Optical investigation techniques, e.g. flow cytometry
    • G01N2015/1486Counting the particles

Definitions

  • a blood cell analyzer comprising:
  • An analyzing device configured to generate a first scattergram according to the forward scattered light information and the fluorescence information detected by the detecting device, and generate a second scattergram according to the side scattered light information and the fluorescence information, at the first scatter
  • the figure is obtained by characterizing platelet information, and the white blood cell fragment information is obtained in the second scattergram, and the platelet count value is obtained from the characterization of the platelet information and the characterization of the white blood cell fragment information.
  • Figure 4 is a side scatter light-fluorescence scatter plot of a normal blood sample without leukocyte debris.
  • Figure 5 is a forward scattered light-fluorescence scatter plot of a second blood sample containing leukocyte debris.
  • FIG. 9 is a schematic structural diagram of a detection area of a blood cell analyzer according to an embodiment of the present invention.
  • the sizes are relatively close, and since the forward scattered light characterizes the volume of the particles in the platelet optical counting method, the forward scattered light information formed by the small volume of white blood cell fragments is indistinguishable from the forward scattered light information formed by the platelets. Since white blood cell debris carries nucleic acid substances, it generates relatively strong fluorescence information.
  • a blood sample is provided.
  • the blood sample is subjected to a certain treatment, for example, spheroidizing the red blood cells in the blood sample, and, for example, mixing the fluorescent dye reagent with the blood sample in a certain ratio.
  • a certain treatment for example, spheroidizing the red blood cells in the blood sample, and, for example, mixing the fluorescent dye reagent with the blood sample in a certain ratio.
  • the above-mentioned spheroidization treatment of red blood cells in a blood sample means that the integrity of the cell membrane of the red blood cells is substantially maintained, and the red blood cells are not dissolved and broken.
  • Step 103 Generate a first scattergram according to the forward scattered light information and the fluorescence information to obtain characterization of platelet information.
  • the first scattergram generated according to the forward scattered light information and the fluorescence information can divide the blood cells into at least white blood cells, red blood cells, and platelets. class.
  • the platelet count is characterized by direct statistical characterization of the platelet information
  • the characterization of the platelet count value is not necessarily an accurate platelet count, as the characterization of the platelet count may contain a count of white blood cell debris.
  • a reticulocyte count value is also obtained in the first scatter plot.
  • Step 104 Generate a second scattergram according to the side scattered light information and the fluorescence information, to obtain characterization of white blood cell fragment information. It has been found that the particle group appearing in a specific region of the two-dimensional scattergram obtained from the fluorescence information and the side scattered light information has a strong correlation with the appearance of white blood cell debris, and thus the second scattergram is utilized. The specific region is determined in the second scattergram, and information on the white blood cell fragments can be obtained, thereby distinguishing the white blood cell fragments from the platelets, and obtaining the count of the white blood cell fragments.
  • a true platelet count value can be obtained by a certain algorithm or process.
  • the blood cell analysis method further includes providing a display device for displaying the obtained scattergram. More specifically, in the display device, according to the characterization of the white blood cell fragment information obtained in step 104, the platelets and white blood cell fragments are visualized on the first scattergram, for example, scatters displayed as different colors/shapes, or each is drawn The boundaries or contours allow visual differentiation of platelets and white blood cell debris on the first scatter plot.
  • platelets and leukocyte fragments can be distinguished, but platelets and red blood cells are not well distinguished, and thus a predetermined feature region can be determined in Fig. 6, and a scatter indicating white blood cell fragment information can be identified in the predetermined feature region.
  • the white blood cell fragment count value is then removed from the scatter points corresponding to these white blood cell fragments in Fig. 5, and a forward scattered light-fluorescence scattergram after removing the white blood cell debris as shown in Fig. 7 can be obtained.
  • mature red blood cells, reticulocytes, white blood cells, and platelet counts can be identified.
  • the existing blood cell analyzer has a detection channel for detecting reticulocytes, and uses forward scattered light and fluorescence to classify and count reticulocytes and other cells in a blood sample, and can also classify and count platelets and other cells, but As mentioned above, platelet classification and counting may be affected by the interference of white blood cell debris in the blood sample, which affects the accuracy of the measurement results. With the method of the present invention, it is possible to accurately report reticulocyte counts and platelet counts in the reticulocyte detection channel without substantially increasing the cost.
  • the characterization cells in the predetermined feature region may also be marked, removed or alarmed.
  • the labeled characterization cell can also be observed in other two-dimensional or three-dimensional images generated from the detected forward scattered light information, side scattered light information, and fluorescence information; Similarly, alerts for alarm processing can appear in other 2D or 3D plots.
  • the characterization of the cells is removed, the influence of the white blood cell debris on the accuracy of the platelet number is avoided, and when the detected forward scattered light information, the side scatter light information, and the fluorescence information are used to generate various types of two-dimensional or three-dimensional images, Accurately distinguish each type of cell on a scatter plot.
  • the platelet value obtained by the BriCyte E6 flow cytometer was 141 (10 ⁇ 9/L).
  • the sampling device 100 is configured to suck a predetermined amount of blood samples.
  • a sample preparation device 200 is for pretreating the blood sample to obtain a processed blood sample, the pretreatment comprising fluorescently staining cells in the blood sample and spheroidizing red blood cells in the blood sample.
  • the spheroidization of red blood cells can substantially maintain the integrity of the cell membrane of red blood cells.
  • the detecting device 300 is configured to pass the cells in the pre-treated blood sample one by one through the detection region, and detect forward scattered light information, side scattered light information, and fluorescence information of each of the cells in the blood sample.
  • the detecting device 300 is provided with a flow chamber 301 through which cells pass one by one, a laser 302 emits laser light, is concentrated by the focusing lens 303, and is aligned with the flow chamber 301, and the cells flow through the flow.
  • a photo-electric signal is generated by laser irradiation, wherein a forward scattered light signal is generated along the laser irradiation direction, and side scattered light and a fluorescent signal are generated from the laser irradiation direction (generally 90 degrees).
  • the analyzing device 400 generates a first scattergram according to the forward scattered light information and the fluorescence information detected by the detecting device 300, and generates a second scattergram according to the side scattered light information and the fluorescence information, in the first scattergram.
  • Platelet information is characterized, and white blood cell debris information is obtained in a second scatter plot, and platelet count values are obtained from the characterized platelet information and the white blood cell debris information.
  • the analyzing device 400 includes a determining unit 401 and an alarm unit 402.
  • the determining unit 401 obtains a white blood cell fragment count value according to the white blood cell fragment information, and determines whether the white blood cell fragment count value exceeds a threshold.
  • the alarm unit 402 is determined according to the determining unit.
  • the judgment result of 401 determines whether or not to give an alarm prompt. For example, when the judging unit 401 judges that the white blood cell debris count value exceeds the threshold value, the alarm unit 402 performs an alarm prompt.
  • the blood cell analyzer further includes a display device 500, and the display device 500 The resulting scatter plot is displayed.
  • Display device 500 can visually distinguish between displaying platelets and white blood cell debris on a first scatter plot, such as scatters displayed as different colors/shapes, or drawing respective boundaries or contours to distinguish the two based on characterization of white blood cell debris information.
  • FIG. 10 is a schematic view showing the appearance of a blood cell analyzer provided by an embodiment.
  • the blood cell analyzer has a housing that is substantially cubic in structure.
  • the sampling device 100 is exposed to the housing to facilitate the suction of the blood sample. More specifically, the housing may also be provided with a sample placement table, and the sampling device 100 may absorb the blood sample placed on the sample placement table; the sample preparation device 200, The detecting device 300 and the analyzing device 400 are both housed in the casing; the display device 500 is located on the surface of the casing for displaying information such as the detection result of the blood cell analyzer.
  • the above blood cell analyzer has a neat appearance and is convenient for operation.

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Abstract

一种血液细胞分析方法及血液细胞分析仪。该方法和仪器中根据血液细胞的侧向散射光信息和荧光信息,得到表征白细胞碎片信息,根据前向散射光信息和荧光信息,得到表征血小板信息,再由表征血小板信息和表征白细胞碎片信息,得到血小板计数值,可避免白细胞碎片对血小板的计数影响,在不提高成本的前提下,保障血小板的计数精确度。

Description

血液细胞分析方法及血液细胞分析仪 技术领域
本发明涉及医疗设备领域,特别是涉及一种血液细胞分析方法及血液细胞分析仪。
背景技术
近年来,应用流式细胞技术对血液细胞进行计数和分类的血液细胞分析仪相继问世。传统的血液细胞分析仪利用库尔特原理(亦称电阻法)进行血小板的计数,但是易受到小红细胞和红细胞碎片的影响。近几年来兴起了使用荧光染色激光散射的方法计数血小板,也称为血小板光学计数法,其原理是通过激光照射经荧光试剂处理过的细胞产生光电信号,因前向散射光信号和荧光信号等不同的光电信号可以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,从而可以区分不同种类的细胞,在一定程度上避免了小红细胞和红细胞碎片对血小板计数的影响。但是,随着研究的深入,发现白细胞碎片会对血小板计数值造成影响而导致血小板计数不准。在上述血小板光学计数法中,虽可通过建立新的通道和添加新的试剂来避免白细胞碎片对血小板的计数造成的影响,但是明显增加了设备和时间等成本。
发明内容
基于此,有必要针对上述现有技术中的不足,提供一种血液细胞分析方法和血液细胞分析仪,能够减少可能存在的白细胞碎片对血小板计数的干扰。
一种血液细胞分析方法,包括:
提供血液样本;
使血液样本中的细胞逐个通过血液细胞分析仪的检测区,获取每个细胞的前向散射光信息、侧向散射光信息和荧光信息;
根据前向散射光信息和荧光信息生成第一散点图,得到表征血小板信息;
根据侧向散射光信息和荧光信息生成第二散点图,得到表征白细胞碎片信息;
根据表征血小板信息和表征白细胞碎片信息,得到血小板计数值。
一种血液细胞分析仪,包括:
采样装置,用于吸取预设量的血液样本;
制样装置,用于将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染色以及对所述血液样本中的红细胞球形化;
检测装置,用于使所述预处理后的血液样本中的细胞逐个通过检测区,检测每个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,用于根据所述检测装置所检测到的前向散射光信息和荧光信息生成第一散点图,根据侧向散射光信息和荧光信息生成第二散点图,在第一散点图得到表征血小板信息,在第二散点图中得到表征白细胞碎片信息,并由所述表征血小板信息和表征白细胞碎片信息得到血小板计数值。
上述方法和仪器中通过在荧光-侧向散射散点图中来区分血小板和白细胞碎片,避免白细胞碎片对血小板的计数影响,在不提高成本的前提下,可保障血小板的计数精确度。
附图说明
图1为本发明一实施例提供的血液细胞分析方法的流程示意图。
图2为由含白细胞碎片的第一血液样本的前向散射光-荧光散点图。
图3为所述第一血液样本的侧向散射光-荧光散点图。
图4为不含白细胞碎片的正常血液样本的侧向散射光-荧光散点图。
图5为含白细胞碎片的第二血液样本的前向散射光-荧光散点图。
图6为含白细胞碎片的第二血液样本的侧向散射光-荧光散点图。
图7为第二血液样本中去除白细胞碎片散点后得到的前向散射光-荧光散点图。
图8为本发明一实施例提供的血液细胞分析仪的结构框图。
图9为本发明一实施例提供的血液细胞分析仪的检测区的结构示意图。
图10为本发明一实施例提供的血液细胞分析仪的外观结构示意图。
具体实施方式
白细胞碎片,也被称为破碎白细胞,是细胞膜完全或者部分破裂的白细胞,白细胞破碎释放出的颗粒以及核内容物呈散乱分布,这些物质大小不等,体积较小的白细胞碎片与血小板的体积大小较为接近。
血液样本在经过试剂处理后,使用流式细胞技术逐个检测细胞的前向散射光信息、侧向散射光信息和荧光信息,可以获得散点图。散点图是由采用流式细胞技术的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,Y坐标轴表征前向散射光强度,而X坐标轴表征荧光强度。利用前向散射光-荧光散点图,可以将血液细胞分为至少三类:白细胞、红细胞、血小板。
经过深入的研究,分析了含有白细胞碎片的血液样本和正常血液样本(基本不含白细胞碎片),意外发现由荧光信息和侧向散射光信息得到的二维散点图的特定区域出现的粒子群与白细胞碎片的出现具有较强的相关性。
虽然不希望受理论约束,发明人猜测,白细胞碎片的细胞膜完全或者部分破裂,细胞破碎释放出的颗粒以及核内容物呈散乱分布,这些物质大小不等,体积较小的白细胞碎片与血小板的体积大小较为接近,由于在血小板光学计数法中,前向散射光表征颗粒的体积,因而小体积白细胞碎片形成的前向散射光信息与血小板形成的前向散射光信息难以区分。由于白细胞碎片带有核酸物质,所以会产生较为强烈的荧光信息,在血小板光学法计数时,荧光信息表征的是颗粒核酸含量,因而白细胞碎片产生的荧光信息强度与血小板,特别是大血小板,产生的荧光信息强度比较接近,白细胞碎片和血小板形成的荧光信息难以区分;而采用血小板光学法计数时,血小板由前向散射光和荧光共同确定,因而在前向散射光-荧光散点图中,血小板的计数会受到白细胞碎片的影响,导致计数不准确。通过研究分析发现,由于白细胞碎片为破碎的细胞,则表征细胞内容物的侧向散射光信息强度减弱,在荧光-侧向散射光散点图中可以较好的来区分血小板和白细胞碎片,因而本发明利用上述发现对血液细胞进行类别分析以及精确计数。
如图1所示,本发明一实施例提供的血液细胞分析方法,包括如下步骤:
步骤101,提供血液样本。具体地,血液样本需经一定处理,例如将血液样本中的红细胞进行球形化处理,再例如,将荧光染料试剂按一定比例与血液样本进行混合。上述所称将血液样本中的红细胞进行球形化处理,是指基本保持红细胞的细胞膜的完整性,不对红细胞进行溶解破碎。
步骤102,使血液样本中的细胞逐个通过血液细胞分析仪的检测区,获取每个细胞的前向散射光信息、侧向散射光信息和荧光信息。由于血液样本中的红细胞、白细胞、血小板结合荧光染料的能力有差异,会产生不同的荧光信息。另外,由于上述各类型细胞的大小不同,会产生不同的前向散射光信息, 而细胞内形态或复杂程度的不同,会产生不同的侧向散射光信息。因而让细胞逐个通过检测区,收集这些信息,就可以在散点图上区分上述各类型细胞。
步骤103,根据前向散射光信息和荧光信息生成第一散点图,得到表征血小板信息。在一实施例中,根据所述前向散射光信息和荧光信息生成的第一散点图,也即前向散射光-荧光散点图,可将血液细胞至少分为白细胞、红细胞、血小板三类。其中,通过直接统计表征血小板信息获得表征血小板计数值,该表征血小板计数值不一定就是准确的血小板计数,因为该表征血小板计数值中可能含有白细胞碎片的计数。在一些实施例中,在第一散点图中还可得到网织红细胞计数值。
步骤104,根据所述侧向散射光信息和荧光信息生成第二散点图,得到表征白细胞碎片信息。经研究发现,由荧光信息和侧向散射光信息得到的二维散点图的特定区域出现的粒子群与白细胞碎片的出现具有较强的相关性,因而利用所述第二散点图,在第二散点图中确定所述特定区域,可以得到表征白细胞碎片信息,从而将白细胞碎片与血小板进行区分,获得白细胞碎片的计数。
步骤105,根据上述两个步骤中获得的表征血小板信息和表征白细胞碎片信息,通过一定的算法或处理,即可得到真实的血小板计数值。
例如,可以根据第一散点图得到的表征血小板信息获得表征血小板计数值,根据第二散点图得到表征白细胞碎片信息获得表征白细胞碎片值,将表征血小板计数值减去表征白细胞碎片值,得到血小板计数值。
再例如,根据步骤104中获得的表征白细胞碎片信息,确定第一散点图中这些表示表征白细胞碎片信息的散点,并在第一散点图上,去除这些表征白细胞碎片的散点,从而使表征白细胞碎片信息从第一散点图中的表征血小板信 息中剥离,获得精确的血小板信息,根据血小板信息即可得到血小板计数值。在一些实施例中,上述血液细胞分析方法还包括,根据表征白细胞碎片信息获得白细胞碎片计数值,判断白细胞碎片计数值是否超过阈值,如果超过阈值,提示该血液样本存在白细胞碎片。提示的形式可以是文本、声音、发光或弹窗方式等类型。
在一些实施例中,上述血液细胞分析方法还包括提供显示装置,所述显示装置用于显示得到的散点图。更具体的,于显示装置中,根据步骤104中获得的表征白细胞碎片信息,在第一散点图上可视化地区分血小板和白细胞碎片,例如显示为不同的颜色/形状的散点,或绘制各自的边界或轮廓,从而可在第一散点图上直观地将血小板和白细胞碎片予以区分。
为了便于进一步理解本发明,下面采用两个实施例并结合其他附图来进行说明。
实施例1
提供含有白细胞碎片的第一血液样本,对该第一血液样本进行处理,使红细胞球形化,并基本保持红细胞的完整性,使细胞逐个通过血液细胞分析仪的检测区,检测每个细胞的前向散射光、侧向散射光和荧光三个信号。其中图2示出了该第一血液样本的前向散射光-荧光散点图(第一散点图),图3示出了该第一血液样本的侧向散射光-荧光散点图(第二散点图)。图2中,血小板和白细胞碎片区分不好,因而无法精确获得血小板计数值。在图3中,血小板和白细胞碎片可以区分,但血小板和红细胞区分不好,所以可以利用图3计算出白细胞碎片,再反馈到图2中,不但能够得到血小板计数值,还可以同时得到红细胞等其他类型细胞的计数值。具体的,将图3中白细胞碎片所在预定特征区域中的散点对应图2中相应的散点,并在图2中用不同颜色 标出,或者直接将图2中血小板+白细胞碎片对应的散点计数减去图3中预定特征区域的表征白细胞碎片的散点计数,而得到血小板计数值。在第二散点图中,将侧向散射光强度比红细胞小,荧光强度比血小板大的细胞所在区别确定为所述预定特征区域,并将预定特征区域内的散点识别为白细胞碎片。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),也可以是随侧向散射光-荧光散点图中血小板、白细胞碎片的位置根据一定的规则动态调整获得。例如,可以确定红细胞的下边界、血小板的右边界和白细胞的左边界,并将红细胞的下边界、血小板的右边界、白细胞的左边界和横轴分别作为预定特征区域的上边界、左边界、右边界和下边界。还可以将红细胞的下边界下移一定距离作为预定特征区域的上边界,也可以将血小板的右边界右移一定距离作为预定特征区域的左边界。
该预定特征区域的位置与大小也可以在同一个检测体系中分别测定不含有白细胞的正常血液样本和已知含有白细胞碎片的异常血液样本,通过上述的方式,获得两者的侧向散射光-荧光散点图,通过对比,将会在血小板所在区域的右侧找到一个区域,在异常血液样本的侧向散射光-荧光散点图的该区域出现粒子群(例如图3中所示),而正常血液样本的侧向散射光-荧光散点图的该区域未出现粒子群(例如图4所示)的区域,该区域可确定为预定特征区域。
由图4中可以看到,正常的血液样本在前向散射光-荧光散点图中,红细胞、白细胞、血小板三类细胞各自分布在散点图的某一区域,且各区域分隔明显,因而能用于较好的统计红细胞、白细胞、血小板三类细胞的数量。
实施例2
提供含有白细胞碎片的第二血液样本,对该第二血液样本进行处理,使红细 胞球形化,并保持红细胞的完整性,使细胞逐个通过血液细胞分析仪的检测区,检测每个细胞的前向散射光、侧向散射光和荧光三个信号。其中图5示出了该第二血液样本的前向散射光-荧光散点图,图6示出了该第二血液样本的侧向散射光-荧光散点图。图5中,血小板和白细胞碎片区分不好,因而无法精确获得血小板计数值。在图6中,血小板和白细胞碎片可以区分,但血小板和红细胞区分不好,因而可在图6中确定预设特征区域,并在该预设特征区域内识别表示白细胞碎片信息的散点,得到白细胞碎片计数值,然后在图5中将对应这些白细胞碎片的散点去除,可以得到如图7所示的去除白细胞碎片散点后的前向散射光-荧光散点图。在图7中,可以识别成熟红细胞、网织红细胞、白细胞、血小板计数。
通过上述方法可以避免白细胞碎片误统计为血小板数量的风险,从而更加精确的反映各类血液细胞的组成。
上述方法中,由于无需建立新的通道或者另外使用其他的试剂,可以在有效控制成本的前提下完成白细胞碎片与血小板的区分,以及血小板等细胞的数量统计。现有的血液细胞分析仪具有检测网织红细胞检测通道,利用前向散射光和荧光将血液样本中的网织红细胞和其他细胞分类并计数,也可以将血小板和其他细胞分类并计数,但如前所述,血小板分类计数时可能受到血液样本中白细胞碎片的干扰而影响测量结果的准确性。采用本发明的方法,能够基本不增加成本的基础上,在网织红细胞检测通道中,准确报告网织红细胞计数,和血小板计数。
上述方法中,还可对所述预定特征区域内的表征细胞进行标示、去除或作报警处理。当作出标示时,根据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的其他二维或三维图中也可观察到该标示后的表征细胞;类 似的,报警处理的提示可以在其他二维或三维图中出现。当去除所述表征细胞后,避免了白细胞碎片对血小板数量精度的影响,再利用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各类二维或三维图时,即可在散点图上准确区分各类型细胞。
以下将通过一个对比实施例对本发明提供的分析方法作进一步说明。
对于一例含白细胞碎片的血液样本,分别使用深圳迈瑞电子股份有限公司生产的BC-6800血液细胞分析仪和深圳迈瑞电子股份有限公司生产的BriCyte E6血液细胞分析仪(该仪器成本高,但可区分白细胞碎片和血小板,采用国际血液学标准化委员会(ICSH)推荐的流式RBC/PLT比值法获得的测试结果充当比对标准)进行测试,结果如下:
通过BC-6800血液细胞分析仪,未采用上述方法时,血小板的值为:179(10^9/L);采用上述方法去除白细胞碎片影响后,血小板的值为:144(10^9/L)。
通过BriCyte E6流式细胞分析仪得到的血小板的值为:141(10^9/L)。
可见使用上述方法对含有白细胞碎片的血液样本进行测试时,血小板的值更接近对比标准中的值,因而使用上述方法可采用相对低成本的设备完成更为精确的血小板计数。
参考图8,相应的,本发明还提供一种血液细胞分析仪,其包括采样装置100、制样装置200、检测装置300和分析装置400。
其中,采样装置100用于吸取预设量的血液样本。
制样装置200用于将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染色以及对所述血液样本中的红细胞球形化。红细胞球形化后能基本保持红细胞的细胞膜的完整性。
检测装置300用于使所述预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每个细胞的前向散射光信息、侧向散射光信息和荧光信息。同时参考图9,具体地,该检测装置300设有流动室301,细胞逐个通过所述流动室301,一激光器302发出激光,经聚焦透镜303聚集后对准流动室301,细胞在流经流动室301时,被激光照射而产生光电信号,其中沿着激光照射方向产生前向散射光信号,偏离激光照射方向(一般为90度)产生侧向散射光和荧光信号。在激光照射方向,依次设置光阑304、收集透镜305、消杂光阑306和前向散射光探测器307,前向散射光信号被前向散射光探测器307收集。偏离激光照射方向上,依次设置另一收集透镜308和二色镜309,通过二色镜309分离侧向散射光信号和荧光信号,并分别被侧向散射光探测器310和荧光探测器311收集。上述检测在同一个通道中进行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,无需建立新的通道、采用额外的预处理试剂。
分析装置400根据所述检测装置300所检测到的前向散射光信息和荧光信息生成第一散点图,根据侧向散射光信息和荧光信息生成第二散点图,在第一散点图得到表征血小板信息,在第二散点图中得到表征白细胞碎片信息,并由所述表征血小板信息和表征白细胞碎片信息得到血小板计数值。
进一步地,所述分析装置400包括判断单元401和报警单元402,所述判断单元401根据表征白细胞碎片信息获得白细胞碎片计数值,判断白细胞碎片计数值是否超过阈值,所述报警单元402根据判断单元401的判断结果决定是否进行报警提示。例如,当判断单元401判断白细胞碎片计数值超过阈值时,则报警单元402进行报警提示。
进一步地,所述血液细胞分析仪还包括显示装置500,所述显示装置500用 于显示得到的散点图。例如上述的第一散点图、第二散点图或其他二维、三维散点图。显示装置500可根据表征白细胞碎片信息,在第一散点图上可视化地区别显示血小板和白细胞碎片,例如显示为不同的颜色/形状的散点,或绘制各自的边界或轮廓将两者区分。
图10所示为一实施例提供的血液细胞分析仪的外观结构示意图。血液细胞分析仪具有大致为立方结构的壳体。其中,采样装置100外露于壳体,便于血液样本的吸取,更为具体地,壳体还可设置样本放置台,采样装置100可吸取放置在样本放置台上的血液样本;制样装置200、检测装置300和分析装置400均收容在壳体内;显示装置500位于壳体表面,用于显示血液细胞分析仪的检测结果等信息。上述血液细胞分析仪具有整洁的外观,为操作带来便利。
以上所述实施例的各技术特征可以进行任意的组合,为使描述简洁,未对上述实施例中的各个技术特征所有可能的组合都进行描述,然而,只要这些技术特征的组合不存在矛盾,都应当认为是本说明书记载的范围。
以上所述实施例仅表达了本发明的几种实施方式,其描述较为具体和详细,但并不能因此而理解为对发明专利范围的限制。应当指出的是,对于本领域的普通技术人员来说,在不脱离本发明构思的前提下,还可以做出若干变形和改进,这些都属于本发明的保护范围。因此,本发明专利的保护范围应以所附权利要求为准。

Claims (12)

  1. 一种血液细胞分析方法,其特征在于包括:
    提供血液样本;
    使血液样本中的细胞逐个通过血液细胞分析仪的检测区,获取每个细胞的前向散射光信息、侧向散射光信息和荧光信息;
    根据前向散射光信息和荧光信息生成第一散点图,得到表征血小板信息;
    根据侧向散射光信息和荧光信息生成第二散点图,得到表征白细胞碎片信息;
    根据表征血小板信息和表征白细胞碎片信息,得到血小板计数值。
  2. 如权利要求1所述的血液细胞分析方法,其特征在于,还包括,根据表征白细胞碎片信息获得白细胞碎片计数值,判断白细胞碎片计数值是否超过阈值,如果超过阈值,提示该血液样本存在白细胞碎片。
  3. 如权利要求2所述的血液细胞分析方法,其特征在于,所述提示通过文本、声音、发光或弹窗方式进行。
  4. 如权利要求1所述的血液细胞分析方法,其特征在于,根据第一散点图得到的表征血小板信息获得表征血小板计数值,根据第二散点图得到的表征白细胞碎片信息获得表征白细胞碎片值,将表征血小板计数值减去表征白细胞碎片值,得到血小板计数值。
  5. 如权利要求1所述的血液细胞分析方法,其特征在于,根据表征白细胞碎片信息,在第一散点图上,去除表征白细胞碎片的散点,获得血小板信息,根据血小板信息,得到血小板计数值。
  6. 如权利要求1所述的血液细胞分析方法,其特征在于,还包括提供显示装置,所述显示装置显示第一散点图,根据表征白细胞碎片信息,在第一散点图上可视化地区别显示血小板和白细胞碎片。
  7. 如权利要求6所述的血液细胞分析方法,其特征在于,所述在第一散点图上可视化地区别显示血小板和白细胞碎片为显示为不同的颜色/形状的散点,或绘制血小板和白细胞碎片的边界或轮廓。
  8. 如权利要求1-7任意一项所述的血液细胞分析方法,其特征在于,还包括根据第一散点图得到网织红细胞计数值和/或红细胞计数值。
  9. 如权利要求1-7任意一项所述的血液细胞分析方法,其特征在于,还包括在使血液样本中的细胞逐个通过血液细胞分析仪的检测区前,对血液样本进行处理,所述处理是将红细胞球形化,并基本保持红细胞的细胞膜的完整性。
  10. 一种血液细胞分析仪,其特征在于包括:
    采样装置,用于吸取预设量的血液样本;
    制样装置,用于将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染色以及对所述血液样本中的红细胞球形化;
    检测装置,用于使所述预处理后的血液样本中的细胞逐个通过检测区,检测每个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,用于根据所述检测装置所检测到的前向散射光信息和荧光信息生成第一散点图,根据侧向散射光信息和荧光信息生成第二散点图,在第一散点图得到表征血小板信息,在第二散点图中得到表征白细胞碎片信息,并由所述表征血小板信息和表征白细胞碎片信息得到血小板计数值。
  11. 如权利要求10所述的血液细胞分析仪,其特征在于,还包括显示装置,所述显示装置用于显示第一散点图,并根据表征白细胞碎片信息,在第一散点图上可视化区分血小板和白细胞碎片。
  12. 如权利要求10所述的血液细胞分析仪,其特征在于,所述分析装置包括 判断单元和报警单元,所述判断单元根据表征白细胞碎片信息获得白细胞碎片计数值,判断白细胞碎片计数值是否超过阈值,所述报警单元根据判断单元的判断结果决定是否进行报警提示。
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