In this short Letter, we have synthesized the BSA protected Au25 nanoclusters and studied their two photon luminescence behavior. We demonstrate that BSA Au25 nanoclusters can be used as a probe with two photon excitation capability. Our results show a quadratic relation between excitation power and emission intensity whereas with one photon excitation shows a linear dependence. The emission spectrum of BSA Au25 nanoclusters with one photon and two photon excitation shows no appreciable change. Due to its long wavelength emission (650 nm) and two photon excitation, BSA Au25 can be potentially used as a probe for deep tissue imaging.