Gentian violet (GV) is a triphenylmethane dye antiseptic with potential for illegal use in livestock production, especially aquaculture where the related malachite green has been widely used. This potential misuse has regulatory importance because of the observed rodent carcinogenicity of GV. This report describes the use of online LC-APCyMS for confirmation of incurred GV residues, and those of its principal metabolite, LGV, in caffish muscle following treatment of live caffish with GV under putative use conditions. LC with APCYMS detection provided sensitive analysis of GV and LGV with estimated detection limits of e l pg observed for both compounds. Fragmentation of GV and LGV via in-source CID was effected by varying the sampling coneskimmer voltage. Ion intensity data were collected using a rapid cone voltage switching procedure that permits selected ion acquisition under optimal conditions for the parent molecule and several selected fragment ions. For GV, four ions including the ionized molecule were used and for LGV, six ions including the protonated molecule were used. The levels of GV and LGV in muscle from fish dosed with 10 pg/l in aquarium water were determined by LCNIS to be 0.5 and 44 ppb, respectively. Analysis of these samples yielded ion intensity ratios that agreed precisely between injections (4%) and accurately with those generated by a comparable amount of authentic GV and LGV ( ~1 0 % deviation). These results show the utility of on-line LC-APCYMS to do both sensitive confirmatory analyses of incurred drug residues for use in monitoring the food supply.