APEX peroxidase is a versatile tool for proximity labeling and electron microscopy (EM) but its requirement for toxic H2O2 is a liability. We now report LaccID, a multicopper oxidase evolved from an ancestral fungal laccase that oxidizes aromatic substrates using non-toxic O2 instead of H2O2. We use LaccID for EM imaging of cultured cells and fly brains, and for proteomic mapping of the changing surface composition of T cells that engage with tumor cells via antigen-specific T cell receptors.