A two-photon fluorescent probe (QNO) for nitric oxide is reported. The probe is designed with a photoinduced electron transfer (PeT) mechanism and shows 12-fold fluorescence enhancement toward NO. Adopting a quinoline derivative as the fluorophore, QNO has a large two-photon action cross section value of 52 GM and long-wavelength emission. It also features high selectivity, low cytotoxicity, and pH insensitivity. By utilizing two-photon microscopy (TPM), QNO can detect NO in live cells and live tissues at a depth of 90-180 μm.