The killing and postantibiotic effects (PAE) of cefotaxime, ciprofloxacin, and gentamicin against Escherichia coli were determined in Mueller-Hinton broth (MHB) and pooled human cerebrospinal fluid (CSF). MICs performed in MHB and CSF were within one dilution for all antimicrobial agent-organism combinations. At two times the MIC, CSF significantly (P less than 0.05) increased the duration of the PAE compared with MHB when cefotaxime, ciprofloxacin, and gentamicin were used against all strains tested. This effect occurred despite similar reductions in bacterial growth in both fluids after the 2-h antimicrobial agent exposure. We conclude that pooled human CSF markedly increases the PAE of cefotaxime, ciprofloxacin, and gentamicin against E. coli compared with MHB, without affecting bacterial killing.