Therapeutic drug monitoring (TDM) has become standard clinical practice for gentamicin, amikacin, and vancomycin to optimize efficacy and reduce toxicity. TDM after the first dose of antibiotic was adopted in our institution. This study aims to evaluate if target therapeutic drug concentrations could be achieved more rapidly in patients with TDM performed after the first dose of gentamicin, amikacin, or vancomycin compared to TDM at steady state. A single‐center retrospective cohort study was conducted at KK Women's and Children's Hospital, Singapore. Patients aged 1 month to 18 years old who received amikacin, gentamicin, or vancomycin between October 2012 to March 2016 and had at least 2 serum drug concentrations done within the same dosing interval were included. The primary objective was to compare the time taken to achieve target serum drug concentrations between first‐dose and steady‐state TDM. A total of 334 patients on amikacin, 211 patients on gentamicin, and 140 patients on vancomycin were included. Using Kaplan‐Meier analysis, the median number of days to optimize therapy was significantly shorter after first‐dose TDM was performed for amikacin (first dose, 1.51 [95% confidence interval (CI), 1.44‐1.89] days vs steady state, 2.85 [95%CI, 2.50‐3.25] days; P < .01] and gentamicin (first dose, 1.66 [95%CI, 1.25‐2.08] days vs steady state, 3.54 [95%CI, 2.40‐4.75] days; P < .01] but not vancomycin (first dose, 1.34 [95%CI, 1.06‐1.52] days vs steady state, 1.42 [95%CI, 1.26‐1.59] days; P = .99]. First‐dose TDM for gentamicin and amikacin resulted in faster attainment of target serum concentrations but did not for vancomycin. Further validation of its impact on actual clinical outcomes may be required.