Laboratoire Nanotechnologies Nanosystèmes (LN2), CNRS UMI-3463, Université de Sherbrooke, Sherbrooke, J1K 0A5, Canada; Institut Interdisciplinaire d'Innovation Technologique (3IT), Université de Sherbrooke, Sherbrooke, J1K 0A5, Canada; Laboratoire Charles Fabry (LCF), Institut d'Optique Graduate School, Université Paris-Saclay, CNRS, Palaiseau, 91127, France.
Laboratoire Charles Fabry (LCF), Institut d'Optique Graduate School, Université Paris-Saclay, CNRS, Palaiseau, 91127, France.
Biosens Bioelectron. 2019 Sep 15;141:111478. doi: 10.1016/j.bios.2019.111478. Epub 2019 Jun 25.
Surface plasmon resonance imaging (SPRI) is a powerful label-free imaging modality for the analysis of morphological dynamics in cell monolayers. However, classical plasmonic imaging systems have relatively poor spatial resolution along one axis due to the plasmon mode attenuation distance (tens of μm, typically), which significantly limits their ability to resolve subcellular structures. We address this limitation by adding an array of nanostructures onto the metal sensing surface (25 nm thick, 200 nm width, 400 nm period grating) to couple localized plasmons with propagating plasmons, thereby reducing attenuation length and commensurately increasing spatial imaging resolution, without significant loss of sensitivity or image contrast. In this work, experimental results obtained with both conventional unstructured and nanostructured gold film SPRI sensor chips show a clear gain in spatial resolution achieved with surface nanostructuring. The work demonstrates the ability of the nanostructured SPRI chips to resolve fine morphological detail (intercellular gaps) in experiments monitoring changes in endothelial cell monolayer integrity following the activation of the cell surface protease-activated receptor 1 (PAR1) by thrombin. In particular, the nanostructured chips reveal the persistence of small intercellular gaps (<5 μm) well after apparent recovery of cell monolayer integrity as determined by conventional unstructured surface based SPRI. This new high spatial resolution plasmonic imaging technique uses low-cost and reusable patterned substrates and is likely to find applications in cell biology and pharmacology by allowing label-free quantification of minute cell morphological activities associated with receptor dependent intracellular signaling activity.
表面等离子体共振成像（SPRI）是一种强大的无标记成像方式，可用于分析细胞单层中的形态动力学。然而，由于等离子体模式的衰减距离（通常为数十微米），传统的等离子体成像系统在一个轴向上的空间分辨率相对较差，这极大地限制了它们解析亚细胞结构的能力。为了解决这个限制，我们在金属传感表面上添加了纳米结构阵列（25nm 厚、200nm 宽、400nm 周期光栅），以将局域等离子体与传播等离子体耦合，从而减少衰减长度并相应地提高空间成像分辨率，而不会显著降低灵敏度或图像对比度。在这项工作中，使用传统的无结构和纳米结构化金膜 SPRI 传感器芯片获得的实验结果清楚地表明，表面纳米结构化可实现明显的空间分辨率提高。该工作证明了纳米结构化 SPRI 芯片能够分辨出细微的形态细节（细胞间间隙），在实验中监测凝血酶激活细胞表面蛋白酶激活受体 1（PAR1）后内皮细胞单层完整性的变化。特别是，纳米结构化芯片揭示了小细胞间间隙（<5μm）的持续存在，而传统的无结构表面基于 SPRI 确定的细胞单层完整性似乎已经恢复。这种新的高空间分辨率等离子体成像技术使用低成本且可重复使用的图案化衬底，有望通过无标记定量与受体依赖性细胞内信号活性相关的微小细胞形态活动，在细胞生物学和药理学中得到应用。
