摘要
目的探讨隐匿性HBV感染(OBI)相关S基因突变对多种抗-HBs及HBsAg临床检测试剂的反应性。方法9种代表性S基因突变株(M1–M9,其中M1–M5为本课题组首次报道)分离自1例OBI患者和3例OBI献血人员血清。分别采用野生型及突变型S基因重组质粒转染中国仓鼠卵巢细胞,分析9种突变株HBsAg表达及其突变蛋白对抗体和检测试剂反应的影响。结果采用HBsAg罗氏诊断试剂Elecsys定量检测9种突变株胞内HBsAg水平,均较野生株的水平明显下降;相同样品用抗-His抗体检测显示仅M1、M6和M7的HBsAg-His融合蛋白表达水平与野生株比较明显降低。6种抗-HBs与各突变HBsAg的反应性结果(S/CO值)显示,与野生株相比,多数突变株与6种抗-HBs的反应性明显降低,M1、M4、M7和M9产生的HBsAg与抗体4及M9产生的HBsAg与抗体6反应性最差(S/CO<1)。6种商品化HBsAg临床试剂检测结果显示,多数突变株与6种HBsAg试剂反应性较野生株明显降低(P<0.05),ELISA试剂D、E和F漏检率分别为11.1%、22.2%和55.6%。结论本实验涉及的突变HBsAg与抗-HBs的结合力降低是引起OBI表现的主要原因之一。当前临床常用HBsAg检测试剂对突变HBsAg的检测能力存在明显不足,亟待提高。
Objective To determine the reactivity of occult HBV infection（OBI）-related S-gene mutations to various anti-HBs antibodies（Abs） and HBs Ag detection reagents. Methods Nine representative S-gene mutations M1–M9（including 5 novel mutations M1–M5） from 1 OBI patient and 3 OBI blood donors were investigated. S-gene recombinant plasmids harboring 9 mutants or wild-type sequences constructed before were transfected into Chinese hamster ovar y（CHO） cells, respectively. HBs Ag expression levels and reactivity with various anti-HBs Abs and regular HBs Ag detection reagents of all the mutants were analyzed. Results Compared to wild-type strain, intracellular HBs Ag levels of the 9 mutants were obviously reduced upon Roche quantitative Elecsys assay. In contrast, analysis of the same samples using anti-His-tag Ab showed that the levels of HBs Ag-Histag fusion protein were significantly reduced only in mutants M1, M6, and M7. The results of reactivity of mutant HBs Ag against 6 anti-HBs Abs（S/CO values） showed that poor reactivity was observed for most mutants. Specifically, M1-, M4-, M7-, and M9-produced HBs Ag with Ab4, and M9-produced HBs Ag with Ab6 had the worst reactivity（S/CO〈1） compared to the wild-type. The results of reactivity of mutant HBs Ag with 6 commercial HBs Ag detection reagents showed that the reactivity for most mutants was significantly lower than that of the wild-type（P〈0.05）. The miss rates of ELISA reagents D, E, and F were 11.1%, 22.2%, and 55.6%, respectively. Conclusions Lower-affinity of studied mutant HBs Ag with anti-HBs is one of the major causes of OBI presentation. The regular HBs Ag detection reagents used currently in clinic show a significant deficiency in detection of the mutant HBs Ag and thus need to be improved.
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2017年第10期865-869,共5页
Medical Journal of Chinese People's Liberation Army
基金
国家自然科学基金(81572010
81373136)
首都卫生发展科研专项项目(2016-2-5032)
北京市自然科学基金(7172206)~~
