A sensitive and fast method for the quantification of the illegal dye chrysoidine in aquaculture products with ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) is presented. Muscle tissues were made alkaline with sodium hydroxide and extracted with ethyl acetate. After evaporation and subsequent defatting with n-hexane, extracts were directly injected onto the UPLC-column. Chromatography was performed on a C(18) column using 0.1% formic acid in water and an acetonitrile gradient within 6 min. Mass spectrometric analysis was performed in the positive electrospray MS/MS mode. The limit of quantification was 0.25 ng/g, which was 30 times lower than the only previously published method with gas chromatographic detection. A complete validation according to the scientific literature and as defined by the European Union was performed. The applicability of the method was shown in the analysis of more than 50 unknown samples in the framework of a monitoring program.