基于纳米金标记的阳极溶出伏安免疫分析用于免疫球蛋白A的测定

Anodic stripping immunoassay based on gold label for determination of immunoglobulin A

提出了一种基于纳米金标记的阳极溶出伏安免疫分析方法。免疫反应在聚苯乙烯微孔板中以夹心分析模式进行,通过物理吸附将兔抗人免疫球蛋白 A(IgA)抗体固定于微孔板上,与相应抗原 IgA 发生免疫反应后,再通过夹心模式捕获相应的纳米金标记的羊抗人 IgA 抗体,然后再与金标羊抗人 IgA 抗体和金标兔抗羊二抗形成的免疫复合物反应,在微孔板上引入大量的纳米金,将金溶解后,在碳糊电极上用阳极溶出伏安法(ASV)对金离子进行检测,溶出峰电流的大小间接与待分析物 IgA 的浓度成正比。对免疫分析的一些实验条件进行了优化。阳极溶出峰电流与 IgA 的浓度在0.018 mg/L～181 mg/L 范围内呈良好的线性关系,检测下限为10μg/L。将该分析方法应用于人血清中 IgA 浓度的测定,取得了满意结果。

An anodic stripping immunoassay based on gold label has been developed. The immunoreaction is performed in a polystyrene microwell using the sandwich format. Rabbit anti - human immunoglobulin A （IgA） antibodies are adsorbed passively on the walls of a polystyrene microwell. The IgA analyte is first captured by the rabbit anti - human antibody and then sandwiched by a colloidal gold - labeled goat anti - human IgA antibody. The subsequent interaction with the immunocomplex consisting of the colloidal gold - labeled goat anti - human IgA antibody and the colloidal gold - labeled rabbit anti - goat secondary antibody result in the adsorption of a large amount of colloidal gold onto the walls of a polystyrene microwell which, after gold metal dissolution in an appropriate solution, is determined by anodic stripping voltammetry （ASV） at a carbon - paste electrode. The influence of some immunoassay conditions upon the anodic stripping peak current is examined and optimized. The anodic stripping peak current depended linearly on the IgA concentration over the range of 0. 018 mg/L ～181 mg/L and a detection limit as low as 10 μg/L is achieved. The anodic stripping immunoassay is applied to the determination of IgA concentration in human serum with satisfactory results.