牛奶体细胞数量标准物质定值方法研究与不确定度评定  

Research on Valuation Method and Uncertainty Evaluation for Milk Somatic Cell Count Reference Materials

作　　者：刘瑛颖[1] 周玉虎 王梓权 LIU Yingying;ZHOU Yuhu;WANG Ziquan(National Institute of Metrology,Beijing 100029,China;Harold(Beijing)Technology Co.,Ltd,Beijing 100083,China)

机构地区：[1]中国计量科学研究院,北京100029 [2]哈罗德(北京)科技有限公司,北京100083

出　　处：《计量科学与技术》2024年第7期17-22,48,共7页Metrology Science and Technology

基　　金：中国计量科学研究院基本业务费重点项目(31-AKYZD2110-1)。

摘　　要：建立了基于显微成像流式细胞技术对牛奶体细胞数量标准物质中的牛奶体细胞浓度进行测量的方法。该方法通过细胞核荧光染料溴化乙锭(Ethidium Bromide)对牛奶体细胞进行标记,显微成像流式细胞技术提供的单细胞多参数显微图像可有效辅助流式细胞分析方法中对目标细胞的圈门识别,从而提高圈门策略的正确性和重复性。通过多份不同体细胞浓度的牛奶体细胞标准物质样本的多次测量分析证明,该方法具有良好的室内重复性和日间重复性。建立了基于显微成像流式技术的牛奶体细胞数量标准物质中牛奶体细胞浓度测量方法的数学模型,分析确定了该测量方法不确定度主要来源于测量重复性、样本前处理、试剂加样、圈门操作和仪器测量体积偏差等。综合评定,针对研究制备的牛奶体细胞数量标准物质候选物V级(~1.0×10^(6)/mL),基于显微成像流式细胞技术的牛奶体细胞浓度测量方法的测量相对不确定度为4.84%。A method based on microscopic imaging flow cytometry was established to measure the concentration of milk somatic cells in reference materials for milk somatic cell count.This method labels milk somatic cells using the fluorescent dye ethidium bromide.The single-cell multi-parameter microscopic images provided by microscopic imaging flow cytometry can effectively assist in identifying target cells in flow cytometric analysis,thereby improving the accuracy and repeatability of gating strategies.Through multiple measurements and analyses of milk somatic cell reference material samples with different somatic cell concentrations,it has been proven that this method has good intra-laboratory and interday repeatability.A mathematical model for measuring the concentration of milk somatic cells in reference materials for milk somatic cell count based on microscopic imaging flow cytometry was established.The analysis determined that the main sources of uncertainty in this measurement method come from measurement repeatability,sample pretreatment,reagent addition,gating operation,and instrument measurement volume deviation.Comprehensive evaluation targeting the candidate Grade V(~1.0×10^(6) cells/mL)reference material for milk somatic cell count prepared in this study shows that the relative uncertainty of the measurement method for milk somatic cell concentration based on microscopic imaging flow cytometry is 4.84%.

关 键 词：计量学 牛奶体细胞 细胞浓度 不确定度 显微成像流式细胞技术 

分 类 号：TB99[一般工业技术—计量学]