BACKGROUND: Blood culture is important in the diagnosis of blood infections and the identification of treatment strategies. Increased contamination in blood culture is a reduction in quality of care. This retrospective observational study, set in an emergency department in Japan, aims to elucidate the contamination rate before and after the introduction of contamination prevention activities. METHODS: We compared the contamination rates before and one year after contamination prevention activities. Specifically, these activities included a novel and specific use of a blood culture cart and adherence to a checklist. The checklist included noting the actual site of collection, the method of disinfection, and whether gloves were worn. RESULTS: There were 1184 and 1129 blood culture samples before and after the preventive activities, respectively. The rates of contamination were 2.03% (95% Cl = 1.30% to 3.00%) and 1.24% (95% Cl = 0.68% to 2.07%), respectively. CONCLUSION: Prevention activities including specific use of a blood culture cart and careful adherence to a checklist were not associated with a significant decrease in contamination rate in our hospital. Further studies based in hospitals with greater rates of contamination may see higher rates of reduction.