HaloTag

HaloTag is a genetically encoded protein tag used to label, visualize, and manipulate fusion proteins in vitro and in live cells. It is based on an engineered haloalkane dehalogenase that reacts specifically and covalently with synthetic chloroalkane ligands. The tag is typically expressed as a fusion to a protein of interest and is labeled by adding cell-permeable or impermeable ligands carrying fluorescent dyes, affinity handles, or other chemical functionalities.

The covalent bond between HaloTag and its ligand provides stable, wash-resistant labeling and enables applications that require persistent attachment, such as long-term imaging, pulse-chase experiments, protein immobilization, and single-molecule studies. A wide range of ligands are available, including fluorescent probes for live-cell and super-resolution microscopy, biotin or affinity tags for purification, and linkers for surface attachment or protein–protein interaction assays.

Advantages of HaloTag include rapid and specific labeling, modularity of ligand chemistry, and compatibility with many experimental formats. Limitations include the tag’s size (approximately 33 kDa), which can affect folding or function of fused proteins, and the need to introduce exogenous ligands. HaloTag is commonly used alongside other self-labeling systems such as SNAP- and CLIP-tag, offering orthogonal labeling strategies for multicolor experiments. The system has been commercialized and widely adopted in cell biology, biochemistry, and microscopy.