In the present study, potential of polymeric microspheres for treatment of allergic conjunctivitis was investigated. Azelastine hydrochloride loaded Eudragit RL100 microspheres were prepared by solvent evaporation technique. The change in drug-polymer ratio on the particle size, zeta potential, entrapment efficiency and in vitro drug release was investigated. As Eudragit concentration ranged from 40 to 80 mg/ml the size range obtained was 4.18-7.36 mm with positive zeta potential. With the increase in drug polymer ratio, the entrapment efficiency was increased with maximum 14.56%. In vitro release studies demonstrated prolonged release of the drug over the period of 6 hr. Scanning electron micrographs showed that microspheres were spherical with distinct solid dense structure. Fourier transform infrared and differential scanning calorimetry studies concluded slight change in peak intensities of drug in microspheres. In vivo studies in rat model indicated that reduction in eosinophil count number was more pronounced in azelastine hydrochloride microspheres than marketed formulation, Azelast (R).