Flesch Julia, Kappen Marie, Drees Christoph, You Changjiang, Piehler Jacob
Department of Biology/Chemistry, University of Osnabrück, Barbarastr. 11, 49076, Osnabrück, Germany.
Center for Cellular Nanoanalytics (CellNanOs), University of Osnabrück, Barbarastr. 11, 49076, Osnabrück, Germany.
Anal Bioanal Chem. 2020 May;412(14):3413-3422. doi: 10.1007/s00216-020-02551-6. Epub 2020 Mar 21.
Localized surface plasmon resonance (LSPR) detection offers highly sensitive label-free detection of biomolecular interactions. Simple and robust surface architectures compatible with real-time detection in a flow-through system are required for broad application in quantitative interaction analysis. Here, we established self-assembly of a functionalized gold nanoparticle (AuNP) monolayer on a glass substrate for stable, yet reversible immobilization of Histidine-tagged proteins. To this end, one-step coating of glass substrates with poly-L-lysine graft poly(ethylene glycol) functionalized with ortho-pyridyl disulfide (PLL-PEG-OPSS) was employed as a reactive, yet biocompatible monolayer to self-assemble AuNP into a LSPR active monolayer. Site-specific, reversible immobilization of His-tagged proteins was accomplished by coating the AuNP monolayer with tris-nitrilotriacetic acid (trisNTA) PEG disulfide. LSPR spectroscopy detection of protein binding on these biocompatible functionalized AuNP monolayers confirms high stability under various harsh analytical conditions. These features were successfully employed to demonstrate unbiased kinetic analysis of cytokine-receptor interactions. Graphical abstract.
局域表面等离子体共振（LSPR）检测为生物分子相互作用提供了高灵敏度的无标记检测。在定量相互作用分析的广泛应用中，需要简单且稳健的表面结构，以便在流通系统中进行实时检测。在此，我们在玻璃基板上建立了功能化金纳米颗粒（AuNP）单层的自组装，用于稳定但可逆地固定组氨酸标签蛋白。为此，用邻吡啶基二硫化物（PLL-PEG-OPSS）功能化的聚-L-赖氨酸接枝聚（乙二醇）对玻璃基板进行一步涂层，作为一种具有反应性但生物相容性的单层，将AuNP自组装成LSPR活性单层。通过用三（次氮基三乙酸）（trisNTA）聚乙二醇二硫化物涂覆AuNP单层，实现了组氨酸标签蛋白的位点特异性、可逆固定。LSPR光谱法检测这些生物相容性功能化AuNP单层上的蛋白质结合，证实了在各种苛刻分析条件下的高稳定性。这些特性已成功用于证明细胞因子-受体相互作用的无偏动力学分析。图形摘要。
