Abstract : In this paper, we present an approach to calculate the wavefront in the back pupil plane of an objective in a fluorescent MACROscope. We use the three-dimensional image of a fluorescent bead because it contains potential pupil information in the ‘far' out-of-focus planes for sensing the wavefront at the back focal plane of the objective. Wavefront sensing by phase retrieval technique is needed for several reasons. Firstly, the point-spread function of the imaging system can be calculated from the estimated pupil phase and used for image restoration. Secondly, the aberrations in the optics of the objective can be determined by studying this phase. Finally, the estimated wavefront can be used to correct the aberrated optical path with- out a wavefront sensor. In this paper, we estimate the wavefront of a MACROscope optical system by using Bayesian inferencing and derive the Gerchberg-Saxton algorithm as a special case.