A cell-microelectronic sensing technique is developed for profiling chemical cytotoxicity and is used to study different cytotoxic effects of the same class chemicals using nitrosamines as examples. This technique uses three human cell lines (T24 bladder, HepG2 liver, and A549 lung carcinoma cells) and Chinese hamster ovary (CHO-K1) cells in parallel as the living components of the sensors of a real-time cell electronic sensing (RT-CES) method for dynamic monitoring of chemical toxicity. The RT-CES technique measures changes in the impedance of individual microelectronic wells that is correlated linearly with changes in cell numbers during t log phase of cell growth, thus allowing determination of cytotoxicity. Four nitrosamines, N-nitrosodimethylamine (NDMA), N-nitrosodiphenylamine (NDPhA), N-nitrosopiperidine (NPip), and N-nitrosopyrrolidine (NPyr), were examined and unique cytotoxicity profiles were detected for each nitrosamine. In vitro cytotoxicity values (IC(50)) for NDPhA (ranging from 0.6 to 1.9 mM) were significantly lower than the IC(50) values for the well-known carcinogen NDMA (15-95 mM) in all four cell lines. T24 cells were the most sensitive to nitrosamine exposure among the four cell lines tested (T24>CHO>A549>HepG2), suggesting that T24 may serve as a new sensitive model for cytotoxicity screening. Cell staining results confirmed that administration of the IC(50) concentration from the RT-CES experiments inhibited cell growth by 50% compared to the controls, indicating that the RT-CES method provides reliable measures of IC(50). Staining and cell-cycle analysis confirmed that NDPhA caused cell-cycle arrest at the G0/G1 phase, whereas NDMA did not disrupt the cell cycle but induced cell death, thus explaining the different cytotoxicity profiles detected by the RT-CES method. The parallel cytotoxicity profiling of nitrosamines on the four cell lines by the RT-CES method led to the discovery of the unique cytotoxicity of NDPhA causing cell-cycle arrest. This study demonstrates a new approach to comprehensive testing of chemical toxicity.