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In view of the lack of a unified and reliable calibration method for existing micro spectrometer, a calibration method for micro spectrometer based on moving least square (MLS) method is proposed. First, the standard spectrograms of mercury-argon lamp and neon lamp were obtained by spectrometer. Then, the original spectral image containing high frequency noise was de-noised by wavelet. After that, the characteristic peak was found through the peak positioning algorithm. The corresponding pixel number of the characteristic peak that needed to participate in the fitting was selected. Finally, a certain number of standard points were selected and fitted by moving least square method. A certain number of characteristic peaks not involved in the calibration were selected and substituted into the fitting function for accuracy verification. The experimental results show that the standard deviation of calibration set is 0.136 nm and the standard deviation of test set is 0.192 nm after calibration based on moving least square method, which is higher than traditional least square method. This method can realize fast and accurate calibration of micro spectrometer, and it has important guiding significance in practical engineering application.