Optical absorption contrast, large imaging depth at ultrasonic resolution, and the potential of functional/quantitative imaging are the features driving the rapid development of photoacoustic (PA) imaging. For quantitative and functional PA imaging, the fluence distribution is required to transform a PA image to a map of absorption coefficients. A suitable method to estimate the fluence for in vivo applications must not require a priori knowledge of the medium optical properties, should work for various illumination conditions, and must be applicable to different PA imaging geometries. Existing methods of estimating fluence in tissue do not meet these requirements simultaneously. Here we present a method to measure the fluence distribution in tissue using acousto-optics (AO) that meets all the above requirements. We developed a PA and AO tomography system for small-animal imaging to investigate the potential and the feasibility of fluence-corrected PA imaging using our method in a single instrument. We performed experiments on phantoms, an ex vivo tissue sample, and freshly sacrificed mice. We demonstrate that the correction for spatial and spectral fluence variations in PA images establishes the direct relation between image value and optical absorption, which in turns improves the quantitative estimation of blood oxygen saturation.