应用FITC系统建立非均衡竞争雌二醇(E2)化学发光法

Construction of non-equilibrium competitive chemiluminesce immunoassay using FITC system for the detection of estradiol

目的建立能广泛应用于雌二醇(E2)检测的方法。方法制备辣根过氧化物酶(HRP)标记兔抗人E2多克隆抗体及异硫氰酸荧光素(FITC)标记E2类似物。以抗FITC抗体包被发光板,FITC-E2类似物与抗FITC抗体结合形成固相抗原,固相抗原与血清中E2竞争结合抗E2抗体-HRP,建立化学发光法(CLIA)血清E2检测系统(FITC系统),进行方法学评价,并与E2-牛血清清蛋白直接包被系统(非FITC系统)和罗氏公司Elecsys2010系统进行比较。结果 FITC系统检测线性范围为10～3000pg/mL,灵敏度为10pg/mL;批内、批间变异系数均小于5%,优于非FITC系统;与Elecsys2010系统检测结果的相关系数为0.9780,测定结果差异无统计学意义(P>0.05)。结论成功建立基于FITC系统的非均衡竞争E2检测CLIA系统,精密度、灵敏度等指标均符合临床要求,可用于临床标本检测。

Objective To construct a method for the detection of estradiol（E2）.Methods Anti-E2 polyclonal antibody was labeled with horse radish peroxidase（HRP）,and fluorescein isothiocyanate（FITC） was conjugated to E2-analogue.Microwell was coated by anti-FITC antibody,FITC-E2-analogue was combined with anti-FITC to form solid phase antigen,and solid phage antigen and E2 in sample would combine to anti-E2-HRP competively（FITC system）.FITC system was evaluated and compared with non-FITC system,based on E2-bovine serum albumin,and Roche Elecsys2010 system.Results The constructed FITC system was with linear range of 10-3 000 pg/mL,sensitivity of 10 pg/mL,and coefficients of variability within and between-run under 5%,which was better than non-FITC system.Compared with Elecsys2010 system,the correlation coefficient was 0.979 0,and with no statistical difference of detection results（P0.05）.Conclusion The non-equilibrium competitive CLIA based on FITC system was successfully constructed,with satisfying precision and sensitivity,and could be applied for the detection of clinical samples.