Analysis of Fluorescent Proteins with a Nanoparticle Probe

This Letter presents the first application of high-energy, single nanoparticle probes (e.g., 520 keV Au₄₀₀ 2 nm NP) in the characterization of surfaces containing fluorescent proteins (e.g., GFP variants) by their coemitted photon, electron and secondary ion signals. NP-induced protein luminescence increases with the NP incident energy, is originated by the NP impact, and is transferred to the protein fluorophor via electronic energy transfer. Multielectron emission is observed per single NP impacts, and their distributions are specific to the target morphology and composition. Fragment ions of protein subunits consisting of 2–7 amino acid peptides are observed under individual NP impacts that can be correlated to the random protein orientation relative to the impact site (e.g., outer layer or “skin” of the protein).