This study compared the bacterial removal performance of ultramicrofibre cloths and mops (UMF) moistened with water (UMF+water), with those moistened with a novel copper-based biocide (UMF+CuWB50, 300ppm) in several working hospital environments, specifically accident and emergency (A&E) and three other wards. A total of 13 defined sampling sites (10 sites per ward) were sampled in order to retrieve, culture, and enumerate total viable (bacterial) counts (TVC) for each site. We sampled 1h before, and 1 and 4h after, cleaning three times per week. The trial ran for 7 weeks. Two wards were cleaned with UMF+water for 3 weeks, and UMF+CuWB50 for 4 weeks. The reverse applied to the other two wards in a cross-over design fashion, to eliminate ward- and time-specific bias. Multivariate statistical analyses were used to establish extent and significance of any perceived differences, and to eliminate the effects of potential confounders. Cleaning with UMF+water reduced TVC on the test surfaces by 30%, whereas cleaning with TVC+CuWB50 reduced TVC by 56%. CuWB50 had two separate effects; a direct antibacterial effect (evident shortly after cleaning), and a residual antibacterial effect that lasted approximately 2 weeks. The residual effect requires regular application of CuWB50 if it is to persist. This ‘real life’ hospital implementation study demonstrates encouraging microbiological cleaning performance for UMF, which is further enhanced with CuWB50.