Objective To evaluate the detection efficiency of different sampling methods to detect bacterial contamination on the surface of different materials, and explore a sampling method that not only conforms to national standards, but also can simply, quickly and accurately reflect the actual microbial contamination on object surface. Methods Surface of sterilized stainless steel plate and sterilized non-woven fabric were evenly coated with Staphylococcus saprophyticus, after natural drying, object surface were taken samples and detected the total number of bacterial colonies by adenosine triphosphate (ATP) bioluminescence assay, cotton swab smearing method, carrier imprinting method, and membrane filtration method, differences in different sampling methods were compared. Data were analyzed by SPSS 18.0 software. Results Under different contamination concentrations, stainless steel plates and non-woven fabrics had the highest number of bacteria detected by carrier imprinting method, followed by membrane filtration method, while cotton swab smearing method had the lowest number of detected bacteria. When bacterial colonies of non-woven fabric was 10⁶ CFU/mL, the qualified rate of carrier imprinting method (30.0%) was lower than that of cotton swab smearing method (80.0%), membrane filtration method (80.0%) and ATP bioluminescence assay (90.0%), differences were all statistically significant (all P < 0.05). ATP bioluminescence assay was prone to get false negative results. When bacterial concentration was 10⁷ CFU/mL, Pearson correlation coefficient between cotton swab smearing method and ATP bioluminescence assay was 0.838 (P=0.002), detection results of two sampling methods had good correlation. Conclusion Carrier imprinting method is more sensitive than cotton swab smearing method, membrane filtration method and ATP bioluminescence assay for the detection of bacterial colonies contaminated on object surface, carrier imprinting method can be used for the monitoring of bacteria on environmental object surface and hands in medical institutions; ATP bioluminescence assay and cotton swab smearing method have a good correlation to the detection results of bacterial load on artificially contaminated non-woven fabric surface.