A mine dump aquifer in the Lusatian lignite mining district, Germany, is contaminated with acid mine drainage (AMD). The only natural process that can counteract the effects of the contamination is bacterial sulphate reduction. The technical measures chosen to handle the contamination include the injection of glycerol into the aquifer to supply electron donors and to accelerate the growth and activity of sulphate-reducing bacteria. An initial assessment of the hydrochemical conditions in the aquifer showed that sulphate concentrations are subject to alteration due to flow-related processes. Consequently, the decision whether sulphate reduction is occurring in the investigated aquifer section was based on the stable isotopic composition of dissolved sulphate and sulphide, which were used in combination with sulphate concentrations. The significant enrichment of both heavy sulphur and heavy oxygen in the remaining sulphate pool and a characteristic isotope fractionation pattern are a clear evidence for the activity of sulphate-reducing bacteria utilising the injected glycerol as an electron donor. This activity seemed to intensify over the observation period. The spatial distribution of sulphate reduction activity, however, appeared to be highly inhomogeneous. Rather than occurring ubiquitously, sulphate reduction activity seemed to concentrate in a defined reaction zone. Regardless of the inhomogeneous distribution, the overall turnover of sulphate during the period of investigation proves the applicability of this enhanced natural attenuation method to handle the restoration of aquifers contaminated with AMD.