The in-vitro activity of tosufloxacin (A-61827) was compared with that of temafloxacin, ciprofloxacin and selected members of other groups of antimicrobial agents, against 684 recent distinct clinical isolates and strains with known mechanisms of resistance. Against members of the Enterobacteriaceae, ciprofloxacin was slightly more active than tosufloxacin, which was more active than temafloxacin. The MIC90 of tosufloxacin for all species of Enterobacteriaceae, Pseudomonas aeruginosa and Acinetobacter spp. was less than or equal to 1 mg/L. Tosufloxacin was slightly more active than temafloxacin, and four to eight fold more active than ciprofloxacin, against the Gram-positive species tested. The MIC90 of tosufloxacin for Staphylococcus aureus was 0.12 mg/L, and for Streptococcus pneumoniae was 0.5 mg/L. All strains of Neisseria spp., Haemophilus influenzae and Moraxella catarrhalis were inhibited by tosufloxacin at a concentration of less than or equal to 0.12 mg/L. Tosufloxacin was the most active quinolone against the anaerobic organisms tested. Cross resistance between quinolones was seen, but not between quinolones and other groups of antimicrobials. The protein binding of tosufloxacin across a range of concentrations averaged 60%. Human serum at a concentration of 70% decreased the bactericidal activity of tosufloxacin by about four-fold.