A novel method using capillary electrophoresis (CE) coupled with inductively coupled plasma mass spectrometry (ICP-MS) for the separation of four organotin compounds has been developed. Semi-permanent coating of dioctadecyl dimethyl ammonium chloride (DDAC) was applied and investigated to enhance separation efficiency of trimethyltin (TMT) chloride, tributyltin (TBT) chloride, dibutyltin (DBT) dichloride and monobutyltin (MBT) trichloride. The electrolyte composition and ionic strength were optimized. Running buffer containing 0.608 mol L⁻¹ HAc and 7 mmol L⁻¹ NaAc was selected for the separation. The detection limits of the four organotin compounds ranged from 0.037 to 0.112 mg L⁻¹ (as Sn) and the values of the relative standard deviation (RSD) were below 3.1% for migration time, less than 7.8% for peak height and not more than 8.4% for peak area. The developed method was successfully applied for speciation analysis of four organotin compounds in one certified reference material (CRM 477, mussel tissue), and environmental samples including water samples, grape wine and biological samples (Mya arenaria Linnaeus). The determined values (as organotin compounds) of CRM 477 (1.81 ± 0.13 (TBT), 1.36 ± 0.16 (DBT), 1.17 ± 0.18 (MBT)) were in good agreement with the certified values (2.20 ± 0.19 (TBT), 1.54 ± 0.12 (DBT), 1.50 ± 0.28 (MBT)). 0.325 mg L⁻¹ MBT and 1.02 μg g⁻¹ (as Sn) TBT were found in grape wine and Mya arenaria Linnaeus samples.