Kumar S, Aaron J, Sokolov K
Department of Biomedical Engineering, University of Texas at Austin, Austin, Texas 78712, USA.
Nat Protoc. 2008;3(2):314-20. doi: 10.1038/nprot.2008.1.
Molecular optical imaging has shown promise in visualizing molecular biomarkers with subcellular resolution both noninvasively and in real-time. Here, we use gold nanoparticles as optical probes to provide meaningful signal in the presence of targeted biomarkers. We present a novel conjugation technique to control the binding orientation of antibodies on the surface of gold nanoparticles to maximize antibody functionality. Briefly, a heterobifunctional linker, hydrazide-polyethylene glycol-dithiol, is used to directionally attach the Fc, or nonbinding region of the antibody, to the gold nanoparticle surface. The conjugation strategy allows for multiplexing various glycosylated antibodies on a single nanoparticle. We present a method to prepare multifunctional nanoparticles by incorporating targeting and delivery moieties on the same nanoparticle that addresses the challenge of imaging intracellular biomarkers. The time estimate for the entire protocol is approximately 6 h.
分子光学成像已显示出在以亚细胞分辨率非侵入性且实时地可视化分子生物标志物方面的前景。在此，我们使用金纳米颗粒作为光学探针，以便在存在靶向生物标志物时提供有意义的信号。我们提出了一种新颖的偶联技术，以控制抗体在金纳米颗粒表面的结合方向，从而使抗体功能最大化。简而言之，一种异双功能连接子，即酰肼 - 聚乙二醇 - 二硫醇，用于将抗体的Fc或非结合区域定向连接到金纳米颗粒表面。这种偶联策略允许在单个纳米颗粒上复用各种糖基化抗体。我们提出了一种通过在同一纳米颗粒上整合靶向和递送部分来制备多功能纳米颗粒的方法，该方法解决了对细胞内生物标志物进行成像的挑战。整个方案的时间估计约为6小时。
