Narang U, Anderson G P, Ligler F S, Burans J
Naval Research Laboratory, Center for Bio/Molecular Science and Engineering, Washington, DC 20375-5348, USA.
Biosens Bioelectron. 1997;12(9-10):937-45. doi: 10.1016/s0956-5663(97)00027-4.
We report on an evanescent wave fiber-optic biosensor for detecting a potently toxic protein, ricin, in the picograms per milliliter range. A sandwich immunoassay scheme was used to detect ricin. First, an anti-ricin IgG was immobilized onto the surface of an optical fiber in two different ways. In the first method, the antibody was directly coated to the silanized fiber using a crosslinker. Second, avidin-coated fibers were incubated with biotinylated anti-ricin IgG to immobilize the antibody using an avidin-biotin bridge. The assay using the avidin-biotin linked antibody demonstrated higher sensitivity and wider linear dynamic range than the assay using antibody directly conjugated to the surface. The linear dynamic range of detection for ricin in buffer using the avidin-biotin chemistry is 100 pg/ml-250 ng/ml. The limits of detection for ricin in buffer solution and river water are 100 pg/ml and 1 ng/ml, respectively. At higher concentrations of ricin (> 50 ng/ml), we observe a strong interaction of ricin with the avidin coated on the surface of the fibers. We have demonstrated that this interaction is primarily due to the lectin activity of ricin and is significantly reduced using fibers coated with neutravidin or by adding galactose to the ricin samples.
我们报道了一种用于检测皮克/毫升范围内剧毒蛋白蓖麻毒素的倏逝波光导纤维生物传感器。采用夹心免疫分析方案检测蓖麻毒素。首先,以两种不同方式将抗蓖麻毒素IgG固定在光纤表面。第一种方法是使用交联剂将抗体直接包被到硅烷化的光纤上。第二种方法是将抗生物素蛋白包被的光纤与生物素化的抗蓖麻毒素IgG孵育,通过抗生物素蛋白-生物素桥固定抗体。与使用直接偶联到表面的抗体的分析相比,使用抗生物素蛋白-生物素连接抗体的分析显示出更高的灵敏度和更宽的线性动态范围。使用抗生物素蛋白-生物素化学方法在缓冲液中检测蓖麻毒素的线性动态范围为100 pg/ml - 250 ng/ml。在缓冲溶液和河水中蓖麻毒素的检测限分别为100 pg/ml和1 ng/ml。在较高浓度的蓖麻毒素(>50 ng/ml)下,我们观察到蓖麻毒素与包被在光纤表面的抗生物素蛋白有强烈相互作用。我们已经证明这种相互作用主要归因于蓖麻毒素的凝集素活性,并且使用中性抗生物素蛋白包被的光纤或向蓖麻毒素样品中添加半乳糖可显著降低这种相互作用。