微量淋巴细胞毒试剂盒中阳性血清的研制与鉴定  

The Preparation and Identification of Positive Serum in CDC Kit

作　　者：封青[1] 胡巧侠[1] 李研[1] 孙丽君[1] 孙晶莹[1] 梁导艳[1] 郭春艳[1] FENG Qing;HU Qiaoxia;LI Yan;SUN Lijun;SUN Jingying;LIANG Daoyan;GUO Chunyan(Shaanxi Provincial People’s Hospital,Xi’an 710068,China)

机构地区：[1]陕西省人民医院中心实验室,陕西西安710068

出　　处：《标记免疫分析与临床》2022年第1期153-157,共5页Labeled Immunoassays and Clinical Medicine

基　　金：国家重点研发计划项目(编号:2016YFD0500700);陕西省重点研发计划项目(编号:2021ZDLSF01-03);陕西省人民医院孵化基金项目(编号:2019YXQ-12)。

摘　　要：目的研制微量淋巴细胞毒(CDC)试剂盒中阳性血清,为临床检测节约成本。方法收集近年来临床检测人类白细胞抗原(HLA)配型的供者和受者外周血,用淋巴细胞分离液分离人外周血单个核细胞,通过多次免疫新西兰大白兔得到抗人淋巴细胞的抗体,即为试剂盒中的阳性血清,提取兔血清进行效价测定和微量淋巴细胞毒实验,与试剂盒中自带的阳性血清作对比分析;并对兔血清进行加速稳定性实验,采用过度升高和降低温度的条件来增加抗体降解的速度,来验证血清的稳定性。结果免疫的两只兔子中,在做微量淋巴细胞毒实验中,1号兔子在免疫第十一次后实验组中死亡细胞比例已接近阳性对照组,在免疫第十二次后实验组中死亡细胞比例高于阳性对照组,2号兔子的实验组中死亡细胞比例最高只达50%,经验证对比1号兔子的血清比试剂盒中的阳性血清效果更佳,且效价达10^(7),获取总量达60mL,并对其做加速稳定性实验,具有很好的稳定性。结论此研究方法适宜于研发部门参考文中成熟的研究方法,开发微量淋巴细胞毒试剂盒为其提供优质的实验材料,并可大批量制备后投放于市场,由于其制备成本低,也较易得到,其大大降低了试剂盒的制作成本,具有很好的实际应用价值。Objective In order to save the cost of clinical test,the positive serum in CDC Kit was prepared.Methods Peripheral blood of donor and recipient of clinical test human leukocyte antigen in recent years was collected for the study.Human peripheral blood mononuclear cells were isolated with lymphocyte,and antibodies against human lymphocyte were obtained by immunizing New Zealand rabbits multiple times,which were the positive serum in the kit.The rabbit serum was extracted and tested for titer and lymphocyte,and compared with the positive serum in the kit.The rabbit serum was then tested for accelerated stability,in order to verify the stability.The condition of over-raising and decreasing temperature was used to increase the rate of antibody degradation.Results Among the immunized rabbits,the percentage of dead cells in the experimental group was similar with that of the positive control group after the 11th dose of lymphocyte,while the percentage of dead cells in the experimental group was higher than that in the positive control group after the 12th immunization,and the highest percentage of dead cells was only 50%in the experimental group of No.2 rabbit.Compared with the positive serum in the kit,the serum of rabbit No.1 had a better effect,and the titer was 107,with the total amount of 60mL,suggesting a good stability.Conclusion The developed method is suitable for the research and development department to develop the lymphocyte kit to provide high quality experimental materials,and can be prepared in large quantities for the market because of its low preparation cost.It is also easy to obtain,which greatly reduces the production cost of the kit and has a good practical application value.

关 键 词：微量淋巴细胞毒检测 群体反应性抗体 人类白细胞抗原配型 阳性血清 稳定性 器官移植 

分 类 号：R392-33[医药卫生—免疫学]