NSE时间分辨荧光免疫分析法的建立

采用夹心法建立神经元特异烯醇化酶（NSE）时间分辨荧光免疫分析技术（TRFIA）来测定血清中NSE的含量。以NSE单克隆抗体E1包被板,双功能螯合剂异氰酸苄基二乙烯三胺四乙酸络合Eu3＋及标记NSE单克隆抗体E7,发光增强系统为以β-二酮体为主的增强液。采用平衡饱和法建立NSE-TRFIA,数据采用双对数函数数据处理程序处理。本方法的连续批内和批间CV分别为2.4%和4.8%,平均回收率为102.6%,灵敏度为0.31ng/mL,可测范围为0.31～320ng/mL,ED20、ED50和ED80分别为20.72ng/mL、57.23ng/mL和157.25ng/mL。本方法与AFP、CEA均无交叉反应。Eu3＋标记抗体-20℃保存8个月免疫反应基本无损失,同批试剂连续8个月应用分析结果稳定。临床应用检测结果与E170所测值高度相关。实验结果表明,本文所建立的NSE-TRFIA的灵敏度、特异性、准确度等均符合临床应用要求。

Establishment of Time-resolved Fluoroimmunoassay for NSE

To establish a two site time-resolved fluoroimmunoassay（TRFIA） for NSE based on the direct sandwich technique.One monoclonal antibody E1 against a specific antigenic site on the NSE was coated to plates,another monoclonal antibody E7 was conjugated with EDTA and labeled with Eu3＋.The detection range of NSE was from 0.31 to 320 ng/mL.The intra-assay and inter-assay CV of were 2.4% and 4.8% respectively.The mean recovery rate was 102.6%,and the sensitivity of NSE was 0.31 ng/mL.The 80%,50% and 20% inhibition binding effect dose（ED80,ED50,ED20） of NSE were 157.25,57.23 and 20.72ng/mL,respectively.There was no cross reaction with AFP and CEA.The TRFIA for NSE established in this study was suitable for clinical application with high sensitivity,specificity and accuracy.