A sensitive and selective colorimetric biosensor for determination of gentamicin, amikacin and tobramycin was proposed with the unmodified gold nanoparticles (GNPs) as the sensing element. Gentamicin, amikacin and tobramycin can rapidly induce the aggregation of gold nanoparticles and is accompanied by a color change from red to blue. The concentration of gentamicin, ami-kacin and tobramycin can be determined by using UV-Vis spectrometer. The experimental parameters were optimized with regard to pH, incubation time and the concentration of the GNPs. Under optimal experimental conditions, the linear range of the colorimetric sensor for gentamicin/amikacin/tobramycin were 2.67--33.93 ngmL-1, 13.33--66.67 ngmL-1 and 20--ISOngmL-1, respectively. The corresponding limit of detection (3er) was 0.354ngmL-1, 0.999 ngmL-1 and 0.579ng inL -1, respectively. This assay was simple and used to detect ami-noglycoside antibiotics in milk and medicine products. [ABSTRACT FROM AUTHOR]