Song Ki-Hee, Dong Biqin, Sun Cheng, Zhang Hao F
Department of Biomedical Engineering, Northwestern University, Evanston, Illinois 60208, USA.
Department of Mechanical Engineering, Northwestern University, Evanston, Illinois 60208, USA.
Rev Sci Instrum. 2018 Dec;89(12):123703. doi: 10.1063/1.5054144.
Spectroscopic single-molecule localization microscopy (sSMLM) is a novel super-resolution imaging technology, which simultaneously records the nanoscopic location and the corresponding full emission spectrum of every stochastic single-molecule emission event. This spectroscopic imaging capability of sSMLM necessitates the establishment of a theoretical foundation of the newly introduced spectral precision and to guide the system design and optimization. Based on numerical simulation and analytical solution, we introduced such a theoretical model to analyze spectral precision by considering the main system parameters, including signal and background shot noises, readout noise, and the spectral calibration procedure. Using this model, we demonstrated the delicate balance among these parameters in achieving the optimal spectral precision and discovered that the best spectral precision can only be achieved at a particular system spectral dispersion. For example, with a given signal of 3000 photons and a readout noise of 2 e-, a system spectral dispersion of 1.6 nm/pixel is required for sSMLM to achieve the highest spectral precision of 1.31 nm.
光谱单分子定位显微镜（sSMLM）是一种新型的超分辨率成像技术，它能同时记录每个随机单分子发射事件的纳米级位置和相应的完整发射光谱。sSMLM的这种光谱成像能力需要建立新引入的光谱精度的理论基础，以指导系统设计和优化。基于数值模拟和解析解，我们引入了这样一个理论模型，通过考虑信号和背景散粒噪声、读出噪声以及光谱校准程序等主要系统参数来分析光谱精度。使用这个模型，我们展示了这些参数在实现最佳光谱精度方面的微妙平衡，并发现只有在特定的系统光谱色散下才能实现最佳光谱精度。例如，在给定3000个光子的信号和2 e-的读出噪声的情况下，sSMLM要实现1.31 nm的最高光谱精度需要1.6 nm/像素的系统光谱色散。
