CD14反向辅助设门法在淋巴细胞亚群检测中的应用  

Application of CD14 reverse assisted gate method in detection of lymphocyte subsets

作　　者：景蓝汀 朱梦婷 杨振坤 罗园园 邹健 殷莹 JING Lanting;ZHU Mengting;YANG Zhenkun;LUO Yuanyuan;ZOU Jian;YIN Ying(Department of Clinical Laboratory,Wuxi People's Hospital,Nanjing Medical University,Wuxi 214023,Jiangsu,China;School of Medicine,Jiangsu University,Zhenjiang 212013,Jiangsu,China)

机构地区：[1]南京医科大学附属无锡人民医院医学检验科,江苏无锡214023 [2]江苏大学医学院,江苏镇江212013

出　　处：《检验医学》2024年第3期227-236,共10页Laboratory Medicine

摘　　要：目的建立CD14反向辅助设门法(简称CD14反向法),并探讨其在淋巴细胞亚群检测中的应用。方法在常规6色流式细胞术检测方案(常规法)中加入CD14-BV421抗体,建立CD14反向法。采用常规法和CD14反向法分别检测质控品、不同放置时间(4、24、48、72 h)的样本和不同疾病(系统性红斑狼疮、病毒性肺炎和肾移植术后)患者的淋巴细胞亚群,比较2种方法检测结果的差异和偏移情况。结果CD14反向法检测细胞低值质控品的淋巴细胞百分比(LYMPH%)和自然杀伤(NK)细胞百分比(NK%)低于常规法(P<0.05);细胞中值质控品2种方法之间所有项目差异均无统计学意义(P>0.05)。与放置4 h比较,CD14^(+)单核细胞与淋巴细胞的区分度随样本放置时间的延长而变小。与全血样本室温放置4 h比较,放置24 h后CD14反向法和常规法各项目的相对偏移均<5%,差异无统计学意义(P>0.05);放置48 h,CD14反向法各项指标相对偏移均<5%,常规法细胞毒性T淋巴细胞(CTL)(CD3^(+)CD8^(+))、B细胞(CD3-CD19^(+))百分比及其绝对数的相对偏移均>5%;放置72 h,2种方法所有项目的相对偏移均>10%,为临床不可接受。采用CD14反向法检测系统性红斑狼疮、病毒性肺炎、肾移植术后患者外周血淋巴细胞亚群,可较好地区分淋巴细胞和单核细胞,同时减少非典型单核细胞对NK%和NK细胞绝对数的影响。结论CD14反向法可减少流式细胞术检测中淋巴细胞门的误圈,有助于提高检测结果的准确性。Objective To establish CD14 reverse assisted gate method and investigate its application in the detection of lymphocyte subsets.Methods CD14-BV421 antibody was added to the conventional 6-color flow cytometry assay(conventional flow cytometry),and CD14 reverse assisted gate method was established.The lymphocyte subsets of quality control products,samples with different placement times(4,24,48 and 72 h)and patients with different diseases(systemic lupus erythematosus,viral pneumonia and kidney transplantation)were detected by conventional flow cytometry and CD14 reverse assisted gate method,and the difference and deviation of the detection results of the 2 methods were compared.Results The percentage of lymphocytes(LYMPH%)and the percentage of natural killer(NK)cells(NK%)detected by CD14 reverse assisted gate method were lower than those by conventional flow cytometry method(P<0.05),while there was no statistical significance in all the items between the 2 methods(P>0.05).Compared with 4 h,the differentiation between CD14^(+)monocytes and lymphocytes was decreased with the extension of the sample placement time.Compared with whole blood samples at room temperature for 4 h,the relative deviations of CD14 reverse assisted gate method and conventional flow cytometry were<5%after 24 h(P>0.05).After 48 h of sample placement,the relative deviations of CD14 reverse assisted gate method were all<5%,and the relative deviations of cytotoxic T lymphocyte(CTL)(CD3^(+)CD8^(+))and B cells(CD3-CD19^(+))percentages and their absolute values were all>5%by conventional flow cytometry.After 72 h of sample placement,the relative deviation of all the items by the both methods was>10%,which was clinically unacceptable.CD14 reverse assisted gate method can be used to detect lymphocyte subsets in peripheral blood of patients with systemic kidney erythematosus,viral pneumonia and kidney transplantation,which could better distinguish lymphocytes from monocytes and reduce the influence of atypical monocytes on NK%and the absolute value of N

关 键 词：CD14 反向辅助设门 淋巴细胞亚群 流式细胞术 

分 类 号：R446.1[医药卫生—诊断学]