直接化学发光免疫分析法定量检测鳞状细胞癌抗原SCC-Ag试剂研究

Quantitative Detection of Squamous Cell Carcinoma Antigen SCC-Ag by Direct Chemiluminescence Immunoassay

目的:研制可定量检测人血清中鳞状细胞癌抗原SCC-Ag的直接化学发光免疫分析法试剂。方法:采用表面羧基修饰的磁性微球作为固相载体包被SCC-Ab1,以吖啶磺酰胺为信号标记物标记SCC Ab2;待测物SCC-Ag与试剂混合后,形成双抗体夹心复合物结合在磁珠上,清洗分离,复合物在激发液的作用下发光,相对发光强度(RLU)与样本中的SCC-Ag浓度呈正相关。结果:试剂性能分析显示,最低检出限不高于0.1 ng/mL;批内变异系数≤5%、批间变异系数≤10%;线性范围是1.2~200 ng/mL,线性相关系数R≥0.999;试剂热稳定性变化幅度≤10%、长期稳定性变化幅度≤15%;平均加样回收率98.7%;检测100例临床标本与罗氏SCC-Ag检测试剂盒对比,相关性系数为R2=0.9894。结论:该试剂具有灵敏度高、线性范围广、稳定性好、特异性高、准确度高等特点,为临床人血清鳞状细胞癌抗原SCC-Ag定量检测提供坚实的基础。

Objective:To develop a direct chemiluminescence immunoassay reagent which can be used to the quantitative detection of squamous cell carcinoma antigen SCC-Ag in human serum.Methods:SCC-Ab1 was coated with carboxyl-modified magnetic microspheres as a solid phase carrier,and SCC Ab2 was labeled with acridine sulfonamide as a signal marker.After the SCC-Ag is mixed with the reagent,a double antibody sandwich complex is formed and bound to the magnetic beads.With cleaning and separation,the compound glows under the action of excitation liquid.The relative luminescence intensity(RLU)is positively correlated with the SCC-Ag concentration in the sample.Result:Reagent performance analysis display the limit of detection were below 0.1 ng/mL;coefficient of variation of intra-assay≤5%;coefficient of variation of inter-assay≤10%;The linear ranges were from 1.2 to 200 ng/mL,the correlation coefficient were greater than or equal to 0.999;The heating stability of the reagents were less than or equal to 10%;the long-term stability were less than or equal to 15%;the average recovery was 98.7%.The relativity of Roche SCC-Ag detection kit from the experiment(100 clinical serum samples)were 0.9894.Conclusion:The reagent has the characteristics of high sensitivity,wide linear range,good stability,excellent specificity and high accuracy.It provides a solid foundation for the quantitative detection of SCC-Ag in clinical human serum.