In this article, a new approach to chiral resolution based on enantioselective crystallization on chiral polymeric microspheres is presented. To demonstrate the approach, a new synthetic method is developed for the preparation of porous and hollow chiral polymeric microspheres. Chiral microspheres based on poly(N-vinyl α-L-phenylalanine) (PV-L-Phe) are synthesized from uniform polystyrene (PS) microsphere templates by a single-step swelling process. The chiral microspheres display a narrow size distribution, and their properties, especially the porosity, can be controlled by varying the synthesis conditions. The chiral discrimination ability of these chiral microspheres is studied for DL-valine crystallization, as a model system for chiral racemic crystallization. X-ray diffraction (XRD) and differential scanning calorimetry (DSC) provide evidence for the crystallization of enantiopure crystals on the chiral polymeric microspheres. Optical rotation measurements during crystallization show an enatiomeric excess of ca. 25% for L-valine in the chiral microspheres. Although this excess is not yet very high, the basic principle of chiral discrimination by enantioselective crystallization on chiral microspheres is demonstrated.