Opt Express. 2021 Oct 11;29(21):34097-34108. doi: 10.1364/OE.433945.
Total internal reflection fluorescence (TIRF) microscopy is an important imaging tool for the investigation of biological structures, especially the study on cellular events near the plasma membrane. Imaging at cryogenic temperatures not only enables observing structures in a near-native and fixed state but also suppresses irreversible photo-bleaching rates, resulting in increased photo-stability of fluorophores. Traditional TIRF microscopes produce an evanescent field based on high numerical aperture immersion objective lenses with high magnification, which results in a limited field of view and is incompatible with cryogenic conditions. Here, we present a waveguide-based TIRF microscope, which is able to generate a uniform evanescent field using high refractive index waveguides on photonic chips and to obtain cellular observation at cryogenic temperatures. Our method provides an inexpensive way to achieve total-internal-reflection fluorescence imaging under cryogenic conditions.
全内反射荧光（TIRF）显微镜是研究生物结构的重要成像工具，特别是在研究细胞膜附近的细胞事件方面。在低温下成像不仅可以观察到接近天然状态和固定状态的结构，还可以抑制不可逆的光漂白率，从而提高荧光团的光稳定性。传统的 TIRF 显微镜利用高数值孔径浸没物镜产生消逝场，具有高放大率，这导致视场有限，并且与低温条件不兼容。在这里，我们提出了一种基于波导的 TIRF 显微镜，它可以利用光子芯片上的高折射率波导产生均匀的消逝场，并在低温下获得细胞观察。我们的方法提供了一种在低温条件下实现全内反射荧光成像的廉价方法。
