A surface plasmon resonance imaging (SPRI) system was developed for the discrimination of proteins on a gold surface. As a label-free and high-throughput technique, SPRI enables simultaneously monitoring of the biomolecular interactions at low concentrations. We used SPRI as a label-free and parallel method to detect different proteins based on protein microarray. Bovine Serum Albumin (BSA), Casein and Immunoglobulin G (IgG) were immobilized onto the Au surface of a gold-coated glass chip as spots forming a 6×6 matrix. These proteins can be discriminated directly by changing the incident angle of light. Excellent reproducibility for label-free detection of protein molecules was achieved. This SPRI platform represents a simple and robust method for performing high-sensitivity detection of protein microarray.