G145R变异重组HBsAg ELISA检测质控参照品制备的初步研究  

The preparation for the reference sample for quality control of G145R mutation of recombinant hepatitis B surface antigen

作　　者：张波[1] 陈妍[2] 龚劲松[2] 张春燕[2] 黄永国[2] 张小燕[2] 田拥军[2] 杨东亮[2] 李方和[2] 

机构地区：[1]华中科技大学同济医学院附属同济医院 [2]华中科技大学同济医学院附属同济医院实验医学研究中心,武汉市430030

出　　处：《实用医学杂志》2007年第12期1775-1778,共4页The Journal of Practical Medicine

基　　金：国家十五科技攻关项目(编号:2001BA705B05)

摘　　要：目的:实验性制备G145R变异重组乙型肝炎表面抗原(rHBsAg)质控参照品。方法:收集含G145R变异rHBsAg的细胞培养上清,采用50%饱和硫酸铵盐析,根据其在D12-ELISA中的检测信号,以卫生部颁布的HBsAgELISA质控血清精确标定盐析物中靶物质的含量,适量稀释分装,置-20℃冻存。采用不同方法对制备物的特异性与稳定性进行考核,并将其试用于对几种市售试剂盒检测变异能力的评价。结果:制备物中靶物质含量在0.50～32.00ng/mLwcHBsAg之间,在D12-ELISA检测中其信号的线性关系与质控血清有很好的一致性;反复冻融对其稳定性无显著影响;采用不同市售ELISA试剂检测时其反应信号强度各不相同。结论:本研究制备的G145R变异rHBsAgELISA质控参照品具有较高的稳定性、特异性与实用性;其研制为乙肝病毒免疫逃逸现象在基础、临床、诊断试剂研发,以及流行病学调查等方面研究的开展提供了重要的物质基础。To prepare the reference sample for quality control of G145R mutation of recombinant hepatitis B surface antigen （rHBsAg）. Methods The culture supernatant of mutate cell line expressing G145R rHBsAg was collected. The G145R rHBsAg was salted out by 50% saturated ammonium sulfate. The content of target protein in the precipitate was precisely detected by the reactive signal of D12-ELISA, then diluted properly, packaged and stored at -20℃. The specificity and stability of the reference sample were detected by different methods. Immunoreaction of the reference sample was assayed by several HBsAg ELISA diagnostic kits on the market. Results The content of target protein prepared was among 0.50 ～32.00 ng/mL wcHBsAg. The reactive signal of the reference sample detected by D12-ELISA was coherent with that of the national standard. Repeat freezing had no effects on its stability. Different reactive signal levels occurred when detected by different ELISA reagent kits. Conclusions This reference sample for quality control has higher stability, specificity, and availability. It provides the material foundation for investigating HBV mutations laboratorially, clinically, epidemiologically, and for developing new diagnostic reagents.

关 键 词：肝炎表面抗原 乙型 酶联免疫吸附测定 G145R变异株 质控参照品 

分 类 号：R446.6[医药卫生—诊断学]