Dark dyes–bright complexes: fluorogenic protein labeling

Abstract

Complexes formed between organic dyes and genetically encoded proteins combine the advantages of stable and tunable fluorescent molecules and targetable, biologically integrated labels. To overcome the challenges imposed by labeling with bright fluorescent dyes, a number of approaches now exploit chemical or environmental changes to control the properties of a bound dye, converting dyes from a weakly fluorescent state to a bright, easily detectable complex. Optimized, such approaches avoid the need for removal of unbound dyes, facilitate rapid and simple assays in cultured cells and enable hybrid labeling to function more robustly in living model organisms.

Fluorogenic protein labeling using a genetically encoded unstrained alkene

“…1–4 Fluorogenicity, which leads to good signal-to-noise ratio, is highly desirable for protein labeling in a complex biological environment. Fluorogenic bioorthogonal reactions, where the removal of unreacted reagents is not necessary, could simplify and, in some situations, enable real-time imaging experiments in live cells.…”

Circularly Permuted Fluorogenic Proteins for the Design of Modular Biosensors

“…[1][2][3][4] Chemical-genetic fluorogenic reporters provide an attractive alternative to FPs as they display many of the same advantages, while capitalizing on the specific interaction of a fluorogenic chromophore (so-called fluorogen) with its cognate genetically encoded receptor in order to generate fluorescence. [5][6][7] Fluorogens are chromophores that display no fluorescence until they interact with their cognate tag, resulting in systems with low background signal and little to no maturation time. The modular nature of these reporters opens exciting prospects for the design of genetically encoded biosensors in which fluorogen binding, and thus fluorescence, is conditioned to the recognition of a given analyte.…”

A general method to fine-tune fluorophores for live-cell andin vivoimaging

“…Importantly, the new HaloTag ligands derived from JF 585 and JF 635 show a high degree of chromogenicity and fluorogenicity ( Fig. 3c-j), a critical parameter in advanced imaging experiments 33 . Of particular interest is the ability to deliver these two dyes to neural tissue in explants (Fig.…”