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Fluorescence fluctuation‐based super‐resolution microscopy: Basic concepts for an easy start
Author(s) -
Alva Alma,
BritoAlarcón Eduardo,
Linares Alejandro,
TorresGarcía Esley,
Hernández Haydee O.,
PintoCámara Raúl,
Martínez Damián,
HernándezHerrera Paul,
D'Antuono Rocco,
Wood Christopher,
Guerrero Adán
Publication year - 2022
Publication title -
journal of microscopy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.569
H-Index - 111
eISSN - 1365-2818
pISSN - 0022-2720
DOI - 10.1111/jmi.13135
Subject(s) - microscopy , resolution (logic) , optics , image resolution , fluorescence microscope , fluorescence , wavelength , fluorescence lifetime imaging microscopy , microscope , computer science , superresolution , temporal resolution , biological system , materials science , physics , computer vision , artificial intelligence , image (mathematics) , biology
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.