The survival and multiplication of Enterobacter agglomerans, Klebsiella pneumoniae, Serratia marcescens, and Pseudomonas aeruginosa in 0.45- and 0.22-micrometer in-line filter sets during simulated infusions were studied to evaluate the ability of each filter type to prevent infusions of these bacteria into patients. Bacteria were found to proliferate in the upstream compartment of sets housing both filter porosities. None of the 0.22-micrometer in-line filters were penetrated by the test bacteria. In contrast, P. aeruginosa was observed to penetrate each 0.45-micrometer in-line filter examined within 12 h of continuous infusion. Tribe Klebsielleae organisms penetrated a proportion of the 0.45-micrometer filters usually between 48 and 72 h of infusion. In addition, the elution of endotoxin from gram-negative bacteria trapped in the filter set during infusion is reported. Collected infusion filtrate exhibited a trend of increasing endotoxin-like activity with an increasing duration of infusion. In the case of E. agglomerans, mean peak levels of approximately 65 pg of Escherichia coli endotoxin per ml were attained after 72 h. Other bacteria produced similar results, except mean peak levels ranged from 5 to 30 pg/ml. It was noted that endotoxin-like activity was not detected in filtrate eluted from contaminated filter sets during the initial 24 h of infusion. We conclude that to avoid potential hazards of bacterial penetration and endotoxin production during continuous use of in-line filter sets, the 0.22-micrometer filter type must be employed and replaced every 24 h.