Solid immersion lens interferometric scattering microscopy for improved sensitivity and mass-resolution

固体浸没透镜干涉散射显微镜可提高灵敏度和质量分辨率

• 批准号: 2124837
• 金额: --
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2018
• 资助国家: 英国
• 起止时间: 2018 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2124837
• 关键词: Solid; immersion; lens; interferometric; scattering

Interferometric scattering microscopy (iSCAT) is an optical imaging technique in which scattered and reflected light interfere at a detector to obtain an image. iSCAT can be used for single molecule label-free detection, single-molecule tracking and quantitative mass imaging of proteins in solution. Alternative optical imaging techniques such as fluorescence microscopy are not suitable for quantitative mass imaging due to labelling efficiency and dynamic range problems. Recently, a considerable improvement in the performance of fluorescence microscopy was obtained through the incorporation of a super-hemispherical solid immersion lens into the experimental set-up. In this project, the effect of incorporating a super-hemispherical solid immersion lens in an iSCAT experimental set-up will be analysed. The potential impact of this project is to improve the sensitivity and mass-resolution of current iSCAT experimental set-ups. This will improve the technique's ability to study many relevant biological and chemical problems in the academic field, ranging from protein self-assembly to drug screening. The aims and objectives of this project is to improve the performance of iSCAT through replacing the currently used oil immersion objective with a solid immersion lens. Use of a solid immersion medium with higher refractive index than immersion oil (n = 1.515) will lead to a considerably greater effective numerical aperture, enabling collection of more scattered photons and a higher spatial resolution. Furthermore, removing the microscope immersion oil and replacing it with solid components will improve the mechanical stability of the experimental set-up, reducing drift of the sample relative to the detector. This could bring about improvements in sensitivity since, while averaging images collected over time reduces the shot noise induced fluctuations, after ~100 ms sample drift (rather than number of photons) becomes the limiting factor. This work will entail testing different high refractive index materials for the solid immersion lens, including cubic zirconia, moissanite and various types of optical glass. For each material, the extent of monochromatic aberration and image quality will be analysed to determine the ideal material. As well as this, the effect of replacing an air objective with an aspheric lens will be probed. Replacing the air objective is advantageous because it will reduce the cost of the experimental set-up, making iSCAT more accessible to the wider academic community. The research methodology is novel as it aims to push the current boundaries of a ground-breaking technique beyond what is currently possible. Currently, all iSCAT experimental set-ups consist of an oil-immersion objective and therefore incorporating a solid immersion lens is a novel approach. It is expected that the performance of iSCAT will be improved through the research completed in this project. This project falls within the EPSRC Biophysics and soft matter physics research area, which is under the Physical Sciences theme. No companies or collaborators are involved in the project.

干涉散射显微镜(ISCAT)是一种光学成像技术，在该技术中，散射光和反射光在探测器上干涉以获得图像。ISCAT可用于溶液中蛋白质的单分子无标记检测、单分子示踪和定量质量成像。替代的光学成像技术，如荧光显微镜，由于标记效率和动态范围问题，不适合定量质量成像。最近，通过将超半球固体浸没透镜引入到实验装置中，荧光显微镜的性能得到了相当大的改善。在这个项目中，将分析在iSCAT实验装置中加入超半球固体浸没透镜的效果。该项目的潜在影响是提高当前iSCAT实验装置的灵敏度和质量分辨率。这将提高该技术研究从蛋白质自组装到药物筛选等学术领域许多相关生物和化学问题的能力。该项目的目的和目标是通过用固体浸没透镜取代目前使用的油浸物镜来提高iSCAT的性能。使用折射率高于浸油(n=1.515)的固体浸没介质将产生相当大的有效数值孔径，从而能够收集更多的散射光子和更高的空间分辨率。此外，去除显微镜浸油并将其替换为固体组件将提高实验装置的机械稳定性，减少样品相对于检测器的漂移。这可以带来灵敏度的提高，因为在约100毫秒的样本漂移(而不是光子数量)成为限制因素后，随着时间的推移收集的图像的平均值减少了散粒噪声引起的波动。这项工作将需要测试用于固体浸没透镜的不同高折射率材料，包括立方氧化锆、三氧化二铝和各种类型的光学玻璃。对于每种材料，将分析单色像差的程度和图像质量，以确定理想的材料。此外，还将探讨用非球面透镜代替空气物镜的效果。更换AIR物镜是有利的，因为它将降低实验设置的成本，使更广泛的学术界更容易接触到ISCAT。这一研究方法是新颖的，因为它的目标是将一项开创性技术的现有边界推向目前可能的范围。目前，所有的iSCAT实验装置都包括一个油浸物镜，因此结合固体浸没透镜是一种新的方法。预计通过本项目的研究，综合监控系统的性能将得到进一步的提高。该项目属于EPSRC生物物理学和软物质物理研究领域，该领域属于物理科学主题。没有任何公司或合作者参与该项目。