Lu-Walther Hui-Wen, Kielhorn Martin, Förster Ronny, Jost Aurélie, Wicker Kai, Heintzmann Rainer
Leibniz Institute of Photonic Technology, Jena, Germany.
Methods Appl Fluoresc. 2015 Jan 16;3(1):014001. doi: 10.1088/2050-6120/3/1/014001.
A significant improvement in acquisition speed of structured illumination microscopy (SIM) opens a new field of applications to this already well-established super-resolution method towards 3D scanning real-time imaging of living cells. We demonstrate a method of increased acquisition speed on a two-beam SIM fluorescence microscope with a lateral resolution of ~100 nm at a maximum raw data acquisition rate of 162 frames per second (fps) with a region of interest of 16.5 × 16.5 µm, free of mechanically moving components. We use a programmable spatial light modulator (ferroelectric LCOS) which promises precise and rapid control of the excitation pattern in the sample plane. A passive Fourier filter and a segmented azimuthally patterned polarizer are used to perform structured illumination with maximum contrast. Furthermore, the free running mode in a modern sCMOS camera helps to achieve faster data acquisition.
结构照明显微镜（SIM）采集速度的显著提高为这种已成熟的超分辨率方法开辟了新的应用领域，可用于活细胞的三维扫描实时成像。我们展示了一种在双光束SIM荧光显微镜上提高采集速度的方法，其横向分辨率约为100纳米，在感兴趣区域为16.5×16.5微米时，最大原始数据采集速率为每秒162帧（fps），且无机械移动部件。我们使用了可编程空间光调制器（铁电液晶硅），它能够精确快速地控制样品平面中的激发图案。采用无源傅里叶滤波器和分段方位图案化偏振器来实现具有最大对比度的结构照明。此外，现代科学互补金属氧化物半导体（sCMOS）相机的自由运行模式有助于实现更快的数据采集。
