Zhang Yide, Benirschke David, Abdalsalam Ola, Howard Scott S
Department of Electrical Engineering, University of Notre Dame, Notre Dame, IN 46556, USA.
Biomed Opt Express. 2018 Aug 6;9(9):4077-4093. doi: 10.1364/BOE.9.004077. eCollection 2018 Sep 1.
We present a novel super-resolution fluorescence lifetime microscopy technique called generalized stepwise optical saturation (GSOS) that generalizes and extends the concept of the recently demonstrated stepwise optical saturation (SOS) super-resolution microscopy [Biomed. Opt. Express9, 1613 (2018)]. The theoretical basis of GSOS is developed based on exploring the dynamics of a two-level fluorophore model and using perturbation theory. We show that although both SOS and GSOS utilize the linear combination of raw images to increase the imaging resolution by a factor of , SOS is a special and the simplest case of GSOS. The super-resolution capability is demonstrated with theoretical analysis and numerical simulations for GSOS with sinusoidal and pulse-train modulations. Using GSOS with pulse-train modulation, super-resolution and fluorescence lifetime imaging microscopy (FLIM) images can be obtained simultaneously. The super-resolution FLIM capability is experimentally demonstrated with a cell sample on a custom-built two-photon frequency-domain (FD) FLIM system based on radio frequency analog signal processing. To our knowledge, this is the first implementation of super-resolution imaging in FD-FLIM.
我们提出了一种名为广义逐步光学饱和（GSOS）的新型超分辨率荧光寿命显微镜技术，该技术对最近展示的逐步光学饱和（SOS）超分辨率显微镜[《生物医学光学快报》9, 1613 (2018)]的概念进行了推广和扩展。GSOS的理论基础是基于探索双能级荧光团模型的动力学并使用微扰理论而发展起来的。我们表明，尽管SOS和GSOS都利用原始图像的线性组合将成像分辨率提高了 倍，但SOS是GSOS的一种特殊且最简单的情况。通过对具有正弦调制和脉冲序列调制的GSOS进行理论分析和数值模拟，证明了其超分辨率能力。使用具有脉冲序列调制的GSOS，可以同时获得超分辨率和荧光寿命成像显微镜（FLIM）图像。基于射频模拟信号处理的定制双光子频域（FD）FLIM系统上的细胞样本通过实验证明了超分辨率FLIM能力。据我们所知，这是FD - FLIM中首次实现超分辨率成像。
