A novel method of cell culture, enabling growth rate control of sessile Gram-negative populations, has been employed to assess the sensitivity of Escherichia coli towards the aminoglycoside antibiotic, tobramycin. Changes in sensitivity, dependent on the growth rate, were compared with those for suspended populations grown in a chemostat and also those for newly-formed daughter cells shed from the biofilm during its growth and development. At specific growth rates up to 0.3 h-1 the susceptibility both of the resuspended biofilm cells and of their planktonic, chemostat grown controls increased in proportion to the growth rate. As the growth rate was increased further (up to 0.7h-1), the susceptibility of the resuspended biofilm cells remained high, whilst that of the planktonic controls decreased. Newly-formed daughter cells, dislodged from the biofilm, demonstrated a uniformly high sensitivity to the antibiotic at all growth rates. This sensitivity corresponded to that of the fastest-growing cells resuspended from biofilms. Lack of growth rate dependency of killing for the newly-formed daughter cells and their high sensitivity to tobramycin suggested that tobramycin activity might vary during the cellular division cycle. Indeed, when synchronous populations were exposed to tobramycin at various times during their division cycle, sensitivity decreased markedly 20 min before the onset of septation, and increased as septation began. Regulation of the cellular division cycle might therefore account, at least partly, for the observed effects of growth rate on susceptibility.