Organotin compounds (OTCs) have been excessively used in antifouling paint on ship hulls and remain in the environment at toxicologically relevant levels. Electrothermal vaporization coupled with inductively coupled plasma-mass spectrometry (ETV/ICP-MS) shows potential as a rapid and chemical-saving screening tool to complement chromatographic approaches for the analysis of metal-based contaminants in environmental samples. In this study, a quantification method based on on-line isotope dilution analysis has been developed to overcome matrix effects. Using an optimized multi-step ETV temperature program, the separation of an OTC fraction from inorganic tin, and major matrix components could be achieved and applied to real-world samples.