Li Shaoheng, Tamura Ryo, Banzai Kota, Kamiyama Daichi, Kner Peter
School of Electrical and Computer Engineering, University of Georgia, Athens, GA, USA.
Department of Cellular Biology, University of Georgia, Athens, GA, USA.
bioRxiv. 2025 May 15:2025.05.11.653285. doi: 10.1101/2025.05.11.653285.
Superresolution microscopy enables imaging of subcellular structures and dynamics with nanoscale detail. Among the various superresolution techniques, structured illumination microscopy (SIM) stands out for its compatibility with live-cell imaging. Linear SIM is restricted to a resolution improvement of a factor of two, improving the resolution to about 100 nm. Nonlinear SIM (NSIM) utilizes reversibly switchable fluorescent proteins to generate a nonlinear response, allowing for the collection of higher spatial frequency information and theoretically extending the resolution without limit. By employing rsEGFP2 and patterned depletion illumination (PD) to generate the desired nonlinearity in the fluorescent response, we have successfully achieved 2D PD-NSIM imaging of actin in live U2OS cells with sub-80 nm resolution.
超分辨率显微镜能够以纳米级细节对亚细胞结构和动态进行成像。在各种超分辨率技术中，结构光照明显微镜（SIM）因其与活细胞成像的兼容性而脱颖而出。线性SIM的分辨率提高仅限于两倍，将分辨率提高到约100纳米。非线性SIM（NSIM）利用可逆转换的荧光蛋白产生非线性响应，从而能够收集更高空间频率的信息，并且理论上可以无限制地扩展分辨率。通过使用rsEGFP2和图案化耗尽照明（PD）在荧光响应中产生所需的非线性，我们成功地在活的U2OS细胞中实现了肌动蛋白的二维PD-NSIM成像，分辨率低于80纳米。
