Superresolution Localization Methods

Superresolution localization microscopy methods produce nanoscale images via a combination of intermittently active fluorescent probes and algorithms that can precisely determine the positions of these probes from single-molecule or few-molecule images. These algorithms vary widely in their underlying principles, complexity, and accuracy. In this review, we begin by surveying the principles of localization microscopy and describing the fundamental limits to localization precision. We then examine several different families of fluorophore localization algorithms, comparing their complexity, performance, and range of applicability (e.g., whether they require particular types of experimental information, are optimized for specific situations, or are more general). Whereas our focus is on the localization of single isotropic emitters in two dimensions, we also consider oriented dipoles, three-dimensional localization, and algorithms that can handle overlapping images of several nearby fluorophores. Throughout the review, we try to highlight practical advice for users of fluorophore localization algorithms, as well as open questions.