This review describes the use of vancomycin in neonates over the last three decades. Given the relation of late-onset neonatal septicaemia to outcome and the increase in coagulase-negative staphylococcal infection as causative organism, vancomycin remains an important antibacterial in the neonatal intensive care unit. The pharmacokinetic behaviour of vancomycin in neonates can be adequately described by a one- or two-compartment model and is mainly determined by postconceptional age and renal function. In neonates, a patent ductus arteriosus as well as treatment with indomethacin or extracorporeal membrane oxygenation (ECMO) leads to an increase in volume of distribution and a decrease in clearance. Microbiological studies in vitro have shown that an increase in vancomycin concentrations above the minimum inhibitory concentration does not result in more effective killing. The microbiological and clinical efficacy of vancomycin in neonates has only been studied explicitly in a restricted number of patients. There are no definitive data relating serum concentrations to effect in this patient group. Vancomycin-related nephrotoxicity and ototoxicity in neonates is rare, and no clear relation to serum concentrations has been demonstrated. Based on the pharmacokinetic profile of vancomycin in neonates, several administration regimens have been constructed. Recent guidelines have suggested that dosage can be independent of gestational age or postconceptional age in neonates without renal failure. In patients with renal failure, therapy can be adequately tailored by using a regimen based on serum creatinine. The usefulness of routine monitoring of peak serum concentrations is doubtful based on the current literature. Recent research demonstrates a shift towards taking only routine trough serum concentrations in order to optimise efficacy. Patients with renal failure and other special subpopulations, such as patients exposed to ECMO or indomethacin, need to be monitored more closely.