A calibration apparatus and method for supporting a Relay Station (RS) in a multiple antenna communication system are provided. The calibration method in the multiple antenna communication system which supports the RS includes, when a modem stage sends a downlink calibration signal, receiving, at a calibrator, the downlink calibration signal and estimating a downlink path channel, when the calibrator sends an uplink calibration signal, receiving, at the modem stage, the uplink calibration signal and estimating an uplink path channel, determining, at the modem stage, calibration coefficients per path not to generate a phase and amplitude difference with respect to the estimated uplink path channel and the estimated downlink path channel, and applying the calibration coefficients per path to corresponding antennas respectively.