The EMIT (Enzyme Multiplied Immunoassay Technique) for serum gentamicin determination was evaluated by a standard procedure. The precision, accuracy, and specificity were assessed and proved satisfactory. Comparison with a bioassay was done, and results for patient samples showed a correlation coefficient of 0.95 between the two methods. Advantages of the enzyme immunoassay system were the provision of results within 15 min of receipt of blood in the laboratory, a requirement of minimal technical expertise, and an applicability to both large and small workloads. The EMIT proved during evaluation to be a practical alternative to current bioassays for the determination of serum gentamicin concentrations.