Zinc is an essential trace element effective at very low concentrations, but it is also an important environmental pollutant dangerous after excessive intake or exposure. The aim of this study was to evaluate in vitro nephrotoxicity of zinc sulfate heptahydrate ZnSO₄ × 7H₂O (1, 10, 50, 100, 200 mg/l) using rabbit epithelial kidney cells RK13 as the model cell line. The xCELLigence system (RTCA) for real-time monitoring of cell response and the end-point assays for determining metabolic activity (MTT test), cytotoxicity (LDH test), proliferation (BrdU test) and cell cycle analysis were compared. Exposure to zinc sulfate produced dose-dependent cytotoxicity. The inhibition concentration IC50 value for xCELLigence monitoring was 101.8 mg/l, for MTT test 135.9 mg/l and 197.4 mg/l for BrdU test. There was a significant correlation between used assays (p≤0.05) except for the LDH test. Based on mean of IC50 values, the effect of zinc sulfate at 150 mg/l on cell cycle was evaluated. We observed the accumulation of cells in S phase accompanied with the reduction of cells in the G0/G1 phase. In conclusion, the mean of our IC50 values for kidney cells is relatively high in comparison to the recommended as well as therapeutic daily doses.