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Technology

Split peroxidase reporters, such as split horseradish peroxidase (HRP) and split enhanced ascorbate peroxidase 2 (APEX2), overcome these limitations because once the full-length peroxidase is reconstituted, it is a versatile catalyst that generates signal for a wide array of detection modalities, including fluorescence, colorimetric readouts, and chemiluminescence. Furthermore, the peroxidases can generate contrast through a reaction with diaminobenzidine (DAB), generating a polymer that can be visualized with nanometer resolution using electron microscopy, or they can be utilized for spatially restricted proteomic mapping through proximity tagging with the probe biotin phenol.

This technology is based on the generation of a split-peroxidase system, where the peroxidase enzyme is split into two components that, alone, are catalytically inactive. Proteins of interest can be tagged with these peroxidase components. When distinct proteins of interest interact, the two complementary components of the peroxidase enzyme are brought into proximity. These components associate to form the complete, active enzyme that can catalyze a wide array of reactions, enabling diverse detection modalities. Split peroxidases can be expressed in mammalian, yeast, and bacterial cells.