A real-time system structure for multiple sound sources segmentation and separation using Multiple Signal Classification algorithm is proposed in this thesis. Using a calibrated array manifold vector, the proposed calibration method improves the accuracy of the MUSIC algorithm for wide-band detections, hence providing high accuracy source segmentation and separation results. And system structure using the Multiple Signal Classification algorithm to detect and estimate the localization of sound source’s spectrum distribution. And then using probability decision method to determine the direction of sound sources. Finally, multiple sources were extracted from array signals by using beamforming method. This proposed method can track and separate multiple sources at the same time and maintain high detection rate.