The aim of this study was to evaluate the utility of a volume-modified blood culture system to diagnose bacteremia in newborns and infants. A total of 793 paired blood cultures, obtained from 464 patients (173 newborns and 291 infants), were analyzed. Vacutainer tubes containing 18 ml supplemented peptone broth sodium-polyanethol-sulfonate were used as the gold standard, in comparison with a blood micro-culture system containing 1.8 ml of the broth. Prior to antibiotic treatment, 2.2 ml of blood was obtained from each patient; 0.2 ml was inoculated in a blood micro-culture tube and 2 ml in a routine tube. Sensitivity, specificity and predictive values were calculated. Microorganisms were isolated in 153 standard blood culture tubes and 151 blood micro-culture tubes. The sensitivity of the blood micro-culture system was 95%, specificity 99% and positive and negative predictive values 96% and 99% respectively. The sensitivity and specificity of blood micro-culture in neonates and infants is high. We recommend that this system be used for the diagnosis of bacteremia in newborns and infants in laboratories where manual systems are still in use.