Nanoparticles are now utilized in many diverse biological and medical applications. Despite this, it remains challenging to tailor their surface for specific molecular targeting while maintaining high biocompatibility. To address this problem, we evaluate a phytochelatin-related peptide surface coating to produce functional and biocompatible nanoparticles (NPs) based on fluorescent InP/ZnS and CdSe/ZnS or superparamagnetic FePt and Fe(3)O(4). Using a combination of transmission electron microscopy, size-exclusion chromatography and gel electrophoresis (GE), we demonstrate the excellent colloidal properties of the peptide-coated NPs (pNPs) and the compact nature of the coating ( approximately 4 nm thickness). We develop a simple protocol for the monofunctionalization of the pNPs with targeting biomolecules, by combining covalent conjugation with GE purification. We then employ functionalized InP/ZnS pNPs in a live-cell, single-molecule imaging application to specifically target and detect individual proteins in the cell membrane. These findings showcase the versatility of the peptides for preparing compact NPs of various compositions and sizes, which are easily functionalized, and suitable for a broad range of biomedical applications.