胃蛋白酶原Ⅰ时间分辨荧光免疫分析法的建立  

Time-resolved fluoroimmunoassay of pepsinogen Ⅰ

作　　者：黄飚[1] 肖华龙[1] 张祥瑞[1] 朱岚[1] 蒋叶华[1] 王翌[1] 蒋孟军[1] 王铁生[1] 

机构地区：[1]江苏省原子医学研究所,无锡214063

出　　处：《中华微生物学和免疫学杂志》2004年第6期492-495,共4页Chinese Journal of Microbiology and Immunology

基　　金：江苏省自然科学基金资助项目 (BK0 2 0 13 3 )

摘　　要：目的　采用时间分辨荧光免疫分析技术建立高灵敏的胃蛋白酶原Ⅰ的快速全自动检测方法。方法　以PGⅠ单克隆抗体 80 0 3#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PGⅠ单克隆抗体 80 16 # ,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PGⅠ时间分辨荧光免疫分析 ,数据采用Log Logit法函数和四参数Logitc函数数据处理程序处理。结果　方法的批内和批间CV分别为 1.9%和 4 .7% ,平均回收率为 10 2 .6 5 % ,灵敏度为 0 .0 5 μg L ,可测范围为 3.5～ 32 8μg L ,ED2 0 、ED50 和ED80 分别为 11.34μg L、38.73μg L和 132 .3μg L。Eu3+ 标记抗体 2 0℃保存 8个月免疫反应性基本无损失 ,同批试剂连续 8个月应用分析结果稳定 ,临床结果与免疫放射分析法的结果相符。结论　胃蛋白酶原Ⅰ时间分辨荧光免疫分析法是目前胃蛋白酶原Ⅰ检测中最灵敏的方法 ,该分析法稳定性好 ,具有很好的应用前景。Objective To establish a two-site time-resolved fluoroimmunoassay (TRFIA) of pepsinogen Ⅰ (PGⅠ) based on the direct sandwich technique. PGⅠ-TRFIA was used to detect serum PGⅠ concentration. Methods In the equilibrium procedure, the monoclonal antibodies (McAb) against a specific antigenic site on the PGⅠ was 8016#, the europium-labelled McAb was prepared by europium-chelate of N-(p-isothiocyanatobenzyl)- diethylenetriamine-N, N, N, N-tetraacetic acid directed against a different antigenic site on the PGⅠ molecule was 8003#. The luminescent enhancement system was enhancement solution containing 2-naphthoyltrifluoroacetone. PGⅠ in sera from patients and healthy controls were examined by PGⅠ-TRFIA with auto DELFIA1235 system. Results The intra- and inter-assay CV of the PGⅠ-TRFIA were 1.9% and 4.7%, respectively, and the recovery rate was 102.65%, the sensitivity was 0.05μg/L. The McAb provided a linear response from 3.5 to 328μg/L with ED 50 of 38.73μg/L. The cross-reacting rate with pepsinogen Ⅱ was negligible. The europium-labelled 8003# was stable for at lest eight months at -20℃ and the results of the TRFIA with same reagents were stable for eight months. Conclusion The newly developed PGⅠ-TRFIA was high sensitivity and very stable. The method can be used in serum immunoassay such as gastric cancer detection. The detection of PGⅠ in sera will show patient an early warning of developing gastric cancer in future.

关 键 词：胃蛋白酶原 时间分辨荧光免疫分析法 PG 胃癌 抗体 抗原 

分 类 号：R341[医药卫生—基础医学]