Nitrogen-doped graphene quantum dot (N-GQD) nanomaterials conjugated with polyethylenimine (PEI)-polystyrene sulfonate (PSS)-anti-epidermal growth factor receptor (AbEGFR) antibody (N-GQD-PEI-PSS-AbEGFR) demonstrated impressive two-photon properties and stability, signifying that they can serve as an effective two-photon contrast agent in two-photon bioimaging. Furthermore, they provided high intensity, brightness, and signal-to-noise ratios at an ultra-low two-photon excitation (TPE) power level in an observation extending to a deep, three-dimensional depth.