Calibration of the diffuse reflectance spectrum for instrument response and time-dependent fluctuation as well as interdevice variations is complicated, time consuming, and potentially inaccurate. We describe a novel fiber optic probe with a real-time self-calibration capability that can be used for tissue optical spectroscopy. The probe was tested in a number of liquid phantoms over a relevant range of tissue optical properties. Absorption and scattering coefficients are extracted with an average absolute error and standard deviation of 6.9%+/-7.2% and 3.5%+/-1.5%, respectively.