Department of Biomedical Engineering, School of Engineering, King Mongkut's Institute of Technology, Ladkrabang, Bangkok 10520, Thailand.
College of Biomedical Engineering, Rangsit University, Pathum Thani 12000, Thailand.
Sensors (Basel). 2021 Aug 2;21(15):5230. doi: 10.3390/s21155230.
Surface plasmon microscopy has been of interest to the science and engineering community and has been utilized in broad aspects of applications and studies, including biochemical sensing and biomolecular binding kinetics. The benefits of surface plasmon microscopy include label-free detection, high sensitivity, and quantitative measurements. Here, a theoretical framework to analyze and compare several non-interferometric surface plasmon microscopes is proposed. The scope of the study is to (1) identify the strengths and weaknesses in each surface plasmon microscopes reported in the literature; (2) quantify their performance in terms of spatial imaging resolution, imaging contrast, sensitivity, and measurement accuracy for quantitative and non-quantitative imaging modes of the microscopes. Six types of non-interferometric microscopes were included in this study: annulus aperture scanning, half annulus aperture scanning, single-point scanning, double-point scanning, single-point scanning, at 45 degrees azimuthal angle, and double-point scanning at 45 degrees azimuthal angle. For non-quantitative imaging, there is a substantial tradeoff between the image contrast and the spatial resolution. For the quantitative imaging, the half annulus aperture provided the highest sensitivity of 127.058 rad/μm RIU, followed by the full annulus aperture of 126.318 rad/μm RIU. There is a clear tradeoff between spatial resolution and sensitivity. The annulus aperture and half annulus aperture had an optimal resolution, sensitivity, and crosstalk compared to the other non-interferometric surface plasmon resonance microscopes. The resolution depends strongly on the propagation length of the surface plasmons rather than the numerical aperture of the objective lens. For imaging and sensing purposes, the recommended microfluidic channel size and protein stamping size for surface plasmon resonance experiments is at least 25 μm for accurate plasmonic measurements.
表面等离子体显微镜一直受到科学界和工程界的关注，并在包括生化传感和生物分子结合动力学在内的广泛应用和研究领域得到了应用。表面等离子体显微镜的优点包括无标记检测、高灵敏度和定量测量。在这里，提出了一种分析和比较几种非干涉表面等离子体显微镜的理论框架。研究的范围是：(1)确定文献中报道的每一种表面等离子体显微镜的优缺点；(2)根据空间成像分辨率、成像对比度、灵敏度以及显微镜的定量和非定量成像模式的测量精度，对其性能进行量化。本研究包括六种非干涉显微镜：环空孔径扫描、半环空孔径扫描、单点扫描、双点扫描、45 度方位角单点扫描和 45 度方位角双点扫描。对于非定量成像，图像对比度和空间分辨率之间存在很大的权衡。对于定量成像，半环空孔径提供了 127.058 rad/μm RIU 的最高灵敏度，其次是全环空孔径的 126.318 rad/μm RIU。空间分辨率和灵敏度之间存在明显的权衡。与其他非干涉表面等离子体共振显微镜相比，环空孔径和半环空孔径具有最佳的分辨率、灵敏度和串扰。分辨率强烈依赖于表面等离子体的传播长度，而不是物镜的数值孔径。对于成像和传感目的，建议用于表面等离子体共振实验的微流道尺寸和蛋白质印迹尺寸至少为 25 µm，以进行准确的等离子体测量。
