磁微球标记物载体在高灵敏电化学发光免疫检测中的应用

A New Electrochemiluminescence Immunoassay Based on Magnetic Microbeads as Carrier of Labels

报道了一种基于磁性微球作为发光标记物载体的高灵敏相思子毒素电化学发光免疫检测方法。首先在酶标板上吸附固定相思子毒素多抗,与相思子毒素作用后再与生物素化的单抗结合,形成双抗体夹心免疫复合物,然后通过生物素-链霉亲和素相互作用连接三联吡啶钌标记的亲和素包被磁性微球形成磁性微球免疫复合物,将复合物从酶标板上解离后进行电化学发光检测。利用此方法检测相思子毒素,浓度在2～200ng/L范围内与电化学发光强度呈良好的对数线性关系,拟合方程为lgY=1.295lgX＋0.201（R=0.995,n=7,p〈0.0001）,检出限达2ng/L。方法特异性高、重现性好,可用于痕量毒素的高灵敏检测,在临床诊断、环境监测、生物防护等领域有较好的应用前景。

An ultrasensitive elctrochemiluminescense （ECL） immunoassay was proposed by using magnetic microbeads as the cartier of ECL labels for ECL signal amplification. First, polyclonal antibody （pcAb） of abrin was coated on microplate. Then, abrin was bound to pcAb. Next, biotinylated monoclonal antibody （mcAb） was conjugated to abrin, yielding the sandwich immunocomplex on the plate. After that, Ru（bpy） 23＋ labeled magnetic microbeads was conjugated on the plate through specific biotin-streptavidin interaction. Finally, Ru（bpy） 23＋ labeled magnetic microbeads was released from the plate and ECL of Ru （bpy）3^2＋ was measured for abrin determination. A logarithmic linear relationship between ECL intensity and the concentration of abrin in the range of 2- 200μg/L was obtained, the linear regression equation was lgY= 1. 2951gX＋0. 201 （R=0. 995, n=7 ,p〈0. 0001 ） and the detection limit was 2 μg/L. This new approach holds great promise for highly sensitive detection of target proteins in various fields such as biodiagnostics, environmental monitoring and biodefense.