Fluorophore-labeled nanocapsules displaying IgG Fc-binding domains for the simultaneous detection of multiple antigens
Fluorophore-labeled nanocapsules displaying IgG Fc-binding domains for the simultaneous detection of multiple antigens
复制标题
DOI：
10.1016/j.biomaterials.2011.08.012
复制
发表时间：
2011-12-01
期刊：
影响因子：
14
通讯作者：
Kuroda, Shun'ichi
中科院分区：
文献类型：
--
作者：
Iijima, Masumi;Matsuzaki, Takashi;Kuroda, Shun'ichi
通过常规免疫学方法同时检测多种抗原受到一抗来源的限制。每种抗体应来源于不同的宿主物种（或免疫球蛋白（IG）的亚类），以抑制二抗的交叉反应。在这里，我们描述了一种创新的方法，使用类似于30 nm的生物纳米胶囊（BNC）展示来自金黄色葡萄球菌蛋白A（ZZ-BNC）的IgG Fc结合Z结构域，同时，快速，灵敏地检测多种抗原。用Cy 2标记的ZZ-BNC（Cy 2-ZZ-BNC）代替Cy 2标记的二抗进行Western blot分析，灵敏度和信号强度均显著提高。Cy 5-ZZ-BNC和小鼠IgG 2a的复合物（其显示对Z结构域的中等亲和力）不解离，即使在8倍过量的游离小鼠IgG 2a存在下也是如此。此外，用BS（3）（双磺基琥珀酰亚胺辛二酸酯）交联有效地稳定了相互作用。用不同的Cy染料标记的ZZ-BNC便于使用来自同一宿主物种的一抗通过蛋白质印迹分析、免疫细胞化学和流式细胞术同时检测多种抗原，这可以扩展各种免疫荧光技术中的生物成像探针的可能性。(C)2011爱思唯尔有限公司保留所有权利。
Simultaneous detection of multiple antigens by conventional immunological methods has been limited by the source of primary antibodies. Each antibody should be derived from a different host species (or subclass of immunoglobulin (Ig)) for suppressing the cross-reactions of secondary antibodies. Here we describe an innovative method for simultaneous, rapid, and sensitive detection of multiple antigens using similar to 30-nm bio-nanocapsules (BNCs) displaying IgG Fc-binding Z domains derived from Staphylococcus aureus protein A (ZZ-BNC). When Cy2-labeled ZZ-BNC (Cy2-ZZ-BNC) was used instead of Cy2-labeled secondary antibody in western blot analysis, both sensitivity and signal intensity were significantly increased. The complex of Cy5-ZZ-BNC and mouse IgG2a (which shows moderate affinity to the Z domain) was not dissociated, even in the presence of 8-fold excess of free mouse IgG2a. In addition, crosslinking with BS(3) (bis-sulfosuccinimidyl suberate) efficiently stabilized the interaction. The ZZ-BNCs labeled with various Cy dyes facilitated the simultaneous detection of multiple antigens using primary antibodies derived from the same host species, by western blot analysis, immunocytochemistry and flow cytometry, which could expand the possibility of bio-imaging probes in various immunofluorescence techniques. (C) 2011 Elsevier Ltd. All rights reserved.