Opstad Ida S, Ströhl Florian, Fantham Marcus, Hockings Colin, Vanderpoorten Oliver, van Tartwijk Francesca W, Lin Julie Qiaojin, Tinguely Jean-Claude, Dullo Firehun T, Kaminski-Schierle Gabriele S, Ahluwalia Balpreet S, Kaminski Clemens F
Department of Physics and Technology, UiT The Arctic University of Norway, Tromsø, Norway.
Department of Chemical Engineering and Biotechnology, University of Cambridge, Cambridge, UK.
J Biophotonics. 2020 Jun;13(6):e201960222. doi: 10.1002/jbio.201960222. Epub 2020 Mar 16.
Large fields of view (FOVs) in total internal reflection fluorescence microscopy (TIRFM) via waveguides have been shown to be highly beneficial for single molecule localisation microscopy on fixed cells [1,2] and have also been demonstrated for short-term live-imaging of robust cell types [3-5], but not yet for delicate primary neurons nor over extended periods of time. Here, we present a waveguide-based TIRFM set-up for live-cell imaging of demanding samples. Using the developed microscope, referred to as the ChipScope, we demonstrate successful culturing and imaging of fibroblasts, primary rat hippocampal neurons and axons of Xenopus retinal ganglion cells (RGCs). The high contrast and gentle illumination mode provided by TIRFM coupled with the exceptionally large excitation areas and superior illumination homogeneity offered by photonic waveguides have potential for a wide application span in neuroscience applications.
通过波导实现的全内反射荧光显微镜（TIRFM）中的大视野（FOV）已被证明对固定细胞上的单分子定位显微镜非常有益[1,2]，并且也已在强健细胞类型的短期活细胞成像中得到证实[3-5]，但尚未用于脆弱的原代神经元或长时间成像。在这里，我们展示了一种基于波导的TIRFM装置，用于对要求苛刻的样品进行活细胞成像。使用我们开发的称为ChipScope的显微镜，我们展示了成纤维细胞、原代大鼠海马神经元和非洲爪蟾视网膜神经节细胞（RGC）轴突的成功培养和成像。TIRFM提供的高对比度和柔和照明模式，再加上光子波导提供的超大激发区域和卓越的照明均匀性，在神经科学应用中具有广泛的应用潜力。
