To optimize the process of aseptic preparation and maintenance for BD FACSAria cell sorter to maintain long-term stable sterile sorting state, the cost of aseptic preparation, operation process and system stability are optimized. The method of monitoring the aseptic status of the fluidics system with LB medium is explored, and the aseptic sorting status and long-term culture of HCT116 cells are observed. The optimized aseptic preparation methods simplify the operation process, save the cost, and can keep the aseptic state of the instrument liquid flow pipeline for a long time. HCT116 colorectal cancer cells are sorted and cultured in antibiotic-free medium for a long time without contamination. The optimized aseptic preparation methods and maintenance procedure of BD FACSAria sorter provide a simple and feasible scheme for the operation and management of the instrument, and provide high-quality and stable aseptic sorting services for scientific research.