The Xikuangshan (XKS) mine in central China is the largest antimony (Sb) mine in the world. The mining activity has seriously contaminated the waters in the area. To determine the sources, migration and transformation of Sb contamination, 32 samples from groundwater (aquifer water), surface water and mine water were collected for water chemistry, trace element and SSO4 and Sr stable isotope analyses. The results showed that the groundwater and surface water were in an oxidized environment. The SSO4 and Sr isotope compositions in the water indicated that dissolved Sb and SO42 originated from sulfide mineral (Sb2S3) oxidation, whereas radiogenic Sr may have been sourced from silicified limestone and stibnite in the Shetianqiao aquifer. Furthermore, a positive correlation between δ34SSO4 and δ87Sr values revealed that the Sr, S and Sb in the waters had a common contamination source, i.e., silicified limestone and stibnite, whereas the Sr, S and Sb in rock and ore were sourced from Proterozoic basement clastics. The analysis also indicated that the isotope composition of dissolved SO42− had been influenced by slight bacterial SO4 reduction in the Magunao aquifer. Mining or rock collapse may have caused Shetianqiao aquifer water to contaminate the Magunao aquifer water via mixing. This study has demonstrated that the stable isotopes of 34SSO4 and 87Sr, combined with hydrochemical methods, are effective in tracking the sources, migration and transformation of Sb contamination.