A rapid, specific, and reliable technique has been developed for the determination of Amikacin in biological tissues by high performance liquid chromatography. The technique involves the use of reverse phase liquid chromatography employing methanol, water (69 : 21, v=v), and tripotassium EDTA (2.2 g=L) as the mobile phase and emission and excitation wavelengths of 360 nm and 435 nm, respectively; the range of concentrations used being 0.25-25 mg=mL.The techniques afford excellent linearity, accuracy, and within-and between-day precision for the concentration range studied, allowing it to be used in daily practice for the determination of Amikacin at the laboratory.