Laboratorio Nacional de Microscopía Avanzada, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico.
Centro de Investigación en Ciencias, Instituto de Investigación en Ciencias Básicas y Aplicadas, Universidad Autónoma del Estado de Morelos, Cuernavaca, Morelos, Mexico.
J Microsc. 2022 Dec;288(3):218-241. doi: 10.1111/jmi.13135. Epub 2022 Aug 9.
Due to the wave nature of light, optical microscopy has a lower-bound lateral resolution limit of approximately half of the wavelength of visible light, that is, within the range of 200 to 350 nm. Fluorescence fluctuation-based super-resolution microscopy (FF-SRM) is a term used to encompass a collection of image analysis techniques that rely on the statistical processing of temporal variations of the fluorescence signal. FF-SRM aims to reduce the uncertainty of the location of fluorophores within an image, often improving spatial resolution by several tens of nanometers. FF-SRM is suitable for live-cell imaging due to its compatibility with most fluorescent probes and relatively simple instrumental and experimental requirements, which are mostly camera-based epifluorescence instruments. Each FF-SRM approach has strengths and weaknesses, which depend directly on the underlying statistical principles through which enhanced spatial resolution is achieved. In this review, the basic concepts and principles behind a range of FF-SRM methods published to date are described. Their operational parameters are explained and guidance for their selection is provided.
由于光的波动性，光学显微镜的横向分辨率下限约为可见光波长的一半，即在 200 至 350nm 范围内。基于荧光波动的超分辨率显微镜（FF-SRM）是一个术语，涵盖了一系列依赖于荧光信号时间变化的统计处理的图像分析技术。FF-SRM 的目的是降低图像中荧光团位置的不确定性，通常可将空间分辨率提高几十纳米。FF-SRM 适用于活细胞成像，因为它与大多数荧光探针兼容，并且仪器和实验要求相对简单，主要是基于相机的落射荧光仪器。每种 FF-SRM 方法都有其优缺点，这直接取决于实现增强空间分辨率的基础统计原理。在本文中，描述了迄今为止发表的一系列 FF-SRM 方法背后的基本概念和原理。解释了它们的操作参数，并提供了选择它们的指导。
