We compared the Difco ESP 384 blood culture system with the pediatric Septi-Chek system for the detection of bloodstream infections in pediatric patients. A total of 10,762 blood culture sets included an ESP aerobic bottle and a Septi-Chek bottle. From these cultures, a total of 278 organisms classified as probable pathogens were isolated, including 237 from ESP bottles and 221 from Septi-Chek bottles. This difference was not statistically significant. More organisms classified as possible contaminants were also isolated from ESP bottles (for ESP, 480 bottles; for Septi-Chek, 418 bottles; P < 0.01). The time to detection was shorter for probable pathogens isolated from ESP bottles (median times for organisms isolated from both systems: ESP, 14.0 h; Septi-Chek, 34.5 h; P < 0.001). The proportions of all probable pathogens detected by 24 and 48 h after inoculation were 78 and 96%, respectively, for ESP compared with 31 and 74%, respectively, for Septi- Chek. The time to final identification was also shorter for organisms grown in ESP bottles (median times for organisms isolated from both systems: ESP, 48.8 h; Septi-Chek, 58.5 h; P ≤ 0.001). A subset of 4,442 cultures also included an ESP anaerobic bottle in addition to an ESP aerobic bottle and a Septi-Chek bottle. There were no significant differences in the recovery of probable pathogens by any of the possible two bottle combinations, but five anaerobic pathogens were recovered only in the anaerobic bottle. We conclude that the ESP 384 is an excellent system for culturing pediatric blood samples and that it provides for the very rapid detection of bloodstream pathogens.