In this short Letter, we have synthesized the BSA protected Au-25 nanoclusters and studied their two photon luminescence behavior. We demonstrate that BSA Au-25 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 Au-25 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 Au-25 can be potentially used as a probe for deep tissue imaging. (C) 2013 Elsevier B. V. All rights reserved.