微孔板法筛查RhCE抗原关键参数及RHCE基因多态性分析  

Screening RhCE antigen key parameters by microplate method and RHCE gene polymorphism analysis

作　　者：杨红梅[1] 陈敏洁 邹昕 马思飞 熊玉琪 邹书英 YANG Hongmei;CHEN Minjie;ZOU Xin;MA Sifei;XIONG Yuqi;ZOU Shuying(Changzhou Blood Center/Changzhou Key Laboratory of Clinical Blood Transfusion,Jiangsu 213004)

机构地区：[1]常州市中心血站/常州市临床输血重点专科实验室,江苏213004

出　　处：《交通医学》2025年第1期13-17,共5页Medical Journal of Communications

基　　金：常州市中心血站站级课题(xz202304,xz202402);常州市科技局课题项目(CJ20245046,CJ20241124)。

摘　　要：目的:研究适合微孔板法检测RhCE抗原的最优条件,分析常州地区献血人群RHCE基因型及表型特征。方法:选取2558例无偿献血者血标本,以试管法为标准,凝集试验为原理,研究微孔板法不同抗体容量、红细胞浓度及振荡频率、时间对反应结果的影响,建立试验反应和关键参数。对RhCE抗原弱表达样本进行RHCE基因10个外显子测序。结果:选取待检红细胞浓度为4%、单克隆试剂抗体容量比例为1∶1的反应体系,按1100 r/min频率震荡120 s、450 r/min频率震荡900 s、1100r/min震荡60 s和30 r/min震荡180 s 4次,读取的结果与手工试管法一致率达99.96%。2558例中表型CCDee占43.47%,CcDEe占35.49%,ccDEE占8.37%,CcDee占8.21%,ccDEe占3.05%,CCDEe占0.71%,ccDee占0.39%,CcDEE占0.31%,未见CCDEE表型。10例RHCE抗原弱表达基因多态性为RHCE*01/*03和RHCE*04/*03,发现1例RHCE*04/*02*new。结论:RhCE抗原在献血人群中具有基因多态性特征,存在稀有基因型,建立微孔板法筛查RhCE抗原试验关键参数为大规模筛查献血人群中RHCE基因多态性提供试验方法。新核苷酸突变位点为RHCE变异型发生机制研究提供样本来源,为RHCE精准输血提供基础。Objective:To study the optimal conditions suitable for the detection of RhCE antigen by microplate method,and to analyze the genotype and phenotypic characteristics of RHCE in the blood donation population in Changzhou.Methods:Samples from 2558 blood donors were selected,and the effects of antibody dosage,different concentrations of red blood cells,oscillation frequency and time on the reaction results were studied by taking the test-tube method as the result standard and the agglutination test as the principle,so as to establish the test response and key parameters.In the experiment,10 exons of RHCE gene were sequenced on the samples with weak expression of RhCE antigen.Results:The reaction system with 4%red blood cell concentration and 1∶1 monoclonal reagent antibody volume ratio was selected and the reaction system was shaken at 1100 r/min for 120 s,450 r/min for 900 s,1100 r/min for 60 s and 30 r/min for 180 s,and the coincidence rate of reading results with manual test tube method was 99.96%.The phenotypes of 2558 blood donors were CCDee(43.47%),CcDEe(35.49%),ccDEE(8.37%),CcDee(8.21%),ccDEe(3.05%),CCDEe(0.71%),ccDee(0.39%),CcDEE(0.31%),but CCDEE phenotype was not observed.The polymorphisms of weakly expressed genes of RHCE antigen in 10 cases were RHCE*01/*03 and RHCE*04/*03,additionally a case with RHCE*04/*02*new was found.Conclusion:RhCE antigen has the characteristics of gene polymorphisms and rare genotypes in the blood donation population,and the key parameters of the RhCE antigen screening test by microplate method are established to provide a test method for large-scale screening of RHCE gene polymorphisms in the blood donation population.The new nucleotide mutation sites provide a sample source for the study of the pathogenesis mechanism of RHCE variants and provide a basis for RHCE precision blood transfusion.

关 键 词：微孔板法 关键参数 抗原弱表达 RhCE抗原筛查 

分 类 号：R392.1[医药卫生—免疫学]