Spectral phase interferometry for direct electric field reconstruction (SPIDER),as a fast and accurate technique,is usually used in spectral phase measurement of femtosecond optical pulses. The effects of pulse replicas separation on the spectral phase retrieval of femtosecond optical pulses are investigated by experiments. The results show that the difference of reconstructed pulse width (FWHM) is within 2.76% in the replicas separation range from 0.22 to 1.82 ps. This research proves that spectral phases can be precisely retrieved in a broad range of pulse replica separation,which reduces the demand of high resolution spectrometer and facilitates the optimal parameter of pulse replicas separation in experimental measurement.