We tracked and documented the time of positivity of blood cultures by using the BACTEC 9120 (Becton Dickinson Diagnostic Instrument Systems) blood culture system over a 5-year study period. A 7-day protocol of the incubation period was selected, and a total of 11,156 blood cultures were evaluated. The clinically significant microorganisms (32.95%) were isolated in 3,676 specimens. Gram-positive and -negative bacterial isolation rates were found to be 41.07 and 44.88%, respectively. Yeasts were found in 14.03% of all pathogens. Both the false-positivity and -negativity rates were very low (0.1 and 0.3%, respectively). The mean detection times for all of the pathogens were determined to be 19.45 h. Yeasts, nonfermentative gram-negative bacteria, and Brucella melitensis strains were isolated within 5 days. By taking these data into account, we decided to establish a 5-day-incubation protocol in our laboratory instead of the 7 days that are commonly used.