Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, Pushchino, Moscow region, Russia.
Bull Exp Biol Med. 2020 Oct;169(6):840-843. doi: 10.1007/s10517-020-04988-2. Epub 2020 Oct 24.
The article presents a new method of immunoblotting for simple, rapid, and highly sensitive detection of proteins. Electrophoretic separation of sample is carried out under non-denaturing conditions in a thin conductive layer between cellulose membranes without polyacrylamide gel. The membrane surface is preliminarily modified with azidophenyl groups to photochemically immobilize proteins in situ. For visualization of protein bands, the membranes are treated with magnetic beads coated with specific antibodies, unbound particles are then removed with a magnet. The detection limit in the model system with biotinylated BSA and magnetic beads coated with streptavidin reaches 10 fg or about 10 molecules, while the total blotting time does not exceed 5 min. The method was applied for detection of IgA in a sample of human exhaled air. The method can be used for the analysis of various complex biological samples containing low amounts of the analyte.
本文提出了一种新的免疫印迹方法，用于简单、快速、高度灵敏地检测蛋白质。在纤维素膜之间的薄导电层中，在非变性条件下进行样品的电泳分离，而无需聚丙烯酰胺凝胶。膜表面预先用叠氮苯基基团修饰，以光化学方式将蛋白质原位固定。为了可视化蛋白质条带，将膜用涂有特异性抗体的磁性珠处理，然后用磁铁去除未结合的颗粒。在带有生物素化 BSA 和涂有链霉亲和素的磁性珠的模型系统中的检测限达到 10 fg 或约 10 个分子，而总印迹时间不超过 5 分钟。该方法用于检测人呼出空气中的 IgA。该方法可用于分析含有低浓度分析物的各种复杂生物样品。
