Turbo-refrigerator based on reverse Brayton cycle is an attractive solution for providing refrigeration at ultra-low temperatures below −60 °C for deep freezing. Employing a motor-driven turboexpander compressor (MTEC) not only allows the turbo-refrigerator to achieve high compactness, reliability and flexibility, but also leads to new operating characteristics especially in a closed cycle. This paper aims to investigate the operating characteristics of a closed-cycle single-stage turbo-refrigerator with a MTEC. A high-speed turbo-refrigerator system with the design refrigeration temperature of −86 °C was manufactured and then tested under variable rotating speeds and refrigeration conditions. Test results show that a refrigeration temperature below −96.4 °C was obtained in a pull-down progress. Compared with a fixed-speed operation, variable-speed operation enables the independent control of the refrigeration temperature and the cooling capacity. With the decrease in the refrigeration temperature (or in the cooling capacity), the rotating speed has a greater effect on the COP than that on the cooling capacity (or on the refrigeration temperature). The mass flow rate along with the compression and expansion work shows a decreasing trend as the refrigeration temperature decreases at a fixed speed of the MTEC. The advanced exergy analysis shows a total exergy efficiency of 0.42 (with a COP of 0.62 at refrigeration temperature of −86 °C) could be expected with component improvement in the near future, and it is of the priority to increase the efficiency of the turboexpander.