The multilongitudinal mode (MLM) high-spectral-resolution lidar (HSRL) based on the Mach–Zehnder interferometer (MZI) is constructed in Xi’an for accurate measurements of aerosol optical properties. The critical requirement of the optimal match between the free spectral range of MZI and the longitudinal mode interval of the MLM laser is influenced by the laboratory temperature, pressure, and vibration. To realize the optimal separation of aerosol Mie scattering signals and molecular Rayleigh scattering signals excited by the MLM laser, a self-tuning technique to dynamically adjust the optical path difference (OPD) of the MZI is proposed, which utilizes the maximum ratio between the received power of the Mie channel and Rayleigh channel as the criterion of the OPD displacement. The preliminary experiments show the feasibility of the MLM-HSRL with self-tuning MZI and the stable performance in the separation of aerosol Mie scattering signals and molecular Rayleigh scattering signals.