The separation of protein in a cross-flow microfiltration of microbe/protein binary suspension is studied. The yeast cells and albumin bovine serum (BSA) are served as typical samples for preparing binary suspensions. Effects of cross-flow velocity and filtration pressure on the cake properties, the filtration flux and the BSA rejection are discussed. An increase in cross-flow velocity leads to a more compact cake with a higher specific filtration resistance; however, the thinner cake thickness results in a higher filtration flux. On the other hand, the cake thickness, the filtration resistance as well as the filtration flux increase with the increase of filtration pressure. The filter cake plays a major role on the BSA rejection. An increase in cross-flow velocity or filtration pressure causes the observed BSA rejection to become higher. A theoretical model based on the resistance-in-series model, the force balance model for particle deposition, the concentration polarization model and the standard capture equation for depth filtration is derived for predicting the filtration flux and the observed BSA rejection directly from operating conditions. The agreements between calculated results and experimental data demonstrate the reliability of the proposed model. The optimum operating condition for highest BSA separation efficiency is also suggested.