Objective: To evaluate the correlation between microbiological culture, ATP tests by bioluminescence and visual inspection for monitoring the surface cleaning and disinfection in an emergency room. Methods: This is a prospective, analytical study with a quantitative approach. Data analysis was guided by the following tests: Spearman’s correlation, Fisher ‘s exact test and receiver operating characteristic (ROC) curve. Results: There was a correlation between ATP quantification methods and microbial count for the women’s bathroom door handle (ρ = 0.526; p= 0.008). In this study, considering the ROC curve, the ATP value below 20 RLU is suggested to classify surfaces as approved in the emergency room. Conclusion: Using different methods of monitoring the cleaning and disinfection process is essential, considering that each method has a different purpose.