Objective To compare the application value of filter membrane method and smear method on the evaluation of disinfection effect of flexible endoscope.Methods 242 flexible endoscopes after cleaning and disinfection from January 2022 to November 2022 were collected as the study subjects, and the disinfection effect of all endoscopes was tested by filter membrane method and smear method. The qualified rate, pathogen detection rate, colony detection and use cost of the two detection methods were compared.Results Among the 242 flexible endoscopes, the total qualified rate of filter membrane method was lower than that of smear method while the total detection rate of pathogenic bacteria and the number of bacterial colony were significantly higher than those of smear method (P < 0.05). In the two evaluation methods, the colony count detected by gastroscopy was mainly ≤ 100 cfu/n, and the colony count detected by enteroscopy was mainly ≥ 100 cfu/n (P > 0.05). The economic cost and time cost of filter membrane method were higher than those of smear method (P < 0.05), and filter membrane method increased the annual depreciation cost of the suction filtration system of 976 yuan.Conclusion Compared with smear method, filter membrane method is more effective in evaluating the disinfection of flexible endoscope, but its economic cost and time cost are also higher. However, in order to build a green environment-friendly hospital, it is necessary to increase the investment of endoscopy center, thus it is recommended to apply filter membrane method.