An antibody induced in rabbits against a gentamicin-bovine serum albumin conjugate cross-reacted with the chemically related aminoglycoside, sisomicin, facilitating the development of a sisomicin radioimmunoassay. Sisomicin was labeled with 125 I using an acylating agent [3-(4-hydroxyphenyl)propionic acid- N -hydroxy-succinimide ester]. The resulting assay produced a linear standard curve on a logit-log plot with a sensitivity of 140 pg. Comparison with a microbiological assay showed no significant difference ( P < 0.001) in the measurement of sisomicin by these two methods.