Affinity ultrafiltration using a large stereospecific binding agent can be used to separate enantiomeric mixtures; however, the overall yield and purification factor have generally been inadequate for commercial separations. The objective of this study was to examine the performance of a multi-stage diafiltration process for chiral separations. Data were obtained for the separation Of D- and L-tryptophan using bovine serum albumin (BSA) as the affinity macroligand. Tangential flow filtration (TIFF) was conducted with laboratory scale modules that are linearly scalable to industrial operation. The two-stage system gave purification factors of more than 20 at greater than 90% yield. Theoretical calculations based on a two site competitive binding model were in good agreement with the experimental data. Purification-yield diagrams were used to examine the effects of the ligand concentration, number of stages, and stage volume on the overall separation. The results clearly demonstrate that multi-stage affinity ultrafiltration processes can provide high purification factors and yield for enantiomeric separations.