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The visualization and manipulation of proteins in live cells are critical for studying complex biological processes. Self-labeling proteins do so by enabling the specific and covalent attachment of synthetic probes, offering unprecedented flexibility in the chemical labeling of proteins in live cells and in vivo. By combining the excellent photophysical properties of synthetic dyes with genetic targetability, these tags provide a modular and innovative toolbox for live-cell and high-resolution fluorescence imaging. In this review, we explore the development and diverse applications of the key self-labeling protein technologies, HaloTag7, SNAP-tag, and CLIP-tag, as well as the covalent trimethoprim (TMP)-tag. We discuss recent innovations in both protein engineering and substrate design that have introduced new functionalities to enable multiplexed imaging, super-resolution microscopy, and the design of novel biosensors and recorders.
Porzberg et al. (Wed,) studied this question.