A colorimetric surface plasmon resonance (SPR) imaging biosensor array based on polarization orientation rotation is presented in this paper. It measures the spectral characteristic variations caused by the steep phase difference between the p- and s-polarization occurring at surface plasmon excitation. It provides one-order of magnitude sensor resolution improvement comparing to existing phase-sensitive SPR imaging sensors and the two-dimensional (2D) sensing capability of the imaging sensor enables multiplex, high throughput array based simultaneous detection for a range of different bio-molecular interactions. Experiments on the binding interactions detection between anti-bovine serum albumin (anti-BSA) and BSA antigen have been performed. All binding interactions occurred at 5×4 protein array were real-time monitored simultaneously. A sensor resolution of 8.26ng/ml (125pM) has been demonstrated, which is one-order of magnitude (12 times) better than the detection limit reported by existing phase-sensitive SPR imaging sensors in the literature, while no time-consuming phase modulation and phase extraction processes are required. Furthermore, the optical colorimetric image read-out of the sensor is easy to be identified by the end users comparing to conventional intensity or phase information. The colorimetric SPR imaging biosensor array can find promising potential applications in high throughput clinical disease diagnosis, protein biomarkers screening and drug screening.