MINFLUX Nanoscopy: A “Brilliant” Technique Promising Major Breakthrough

MINFLUX nanoscopy relies on the localization of single fluorophores with expected ~ 2 nm precision in 3D mapping, roughly one order of magnitude better than standard stimulated emission depletion microscopy or stochastic optical reconstruction microscopy. This “brilliant” technique takes advantage of specialized localization principles and algorithms that require only dim fluorescence signals with a minimum flux of photons; hence the name follows. With this level of performance, MINFLUX imaging and tracking should allow for the routine study of biological processes down to the molecular scale, revealing previously unresolved details in cell structures, such as the organization of calcium channels in muscle cells or the clustering of receptors in synapses. Whereas the high localization precision is definitely a strength of the MINFLUX technique, limitations and challenges also exist, especially in the labeling procedures aiming at appropriate density and on/off switching kinetics. This primer presents some significant results achieved with MINFLUX so far and highlights specific operational procedures crucial for this technique.