Impedance flow cytometry (IFC) was evaluated as a possible alternative to fluorescence-based methods for on-line quality monitoring of hybridoma cells. Hybridoma cells were cultured at different cell densities and viability was estimated by means of IFC and fluorescence-based flow cytometry (FCM). Cell death was determined by measuring the impedance phase value at high frequency in low conductivity buffer. IFC data correlate well with reference FCM measurements using AnnexinV and 7-AAD staining. Hybridoma cells growing at different densities in cell culture revealed a density-dependent subpopulation pattern. Living cells of high density cultures show reduced impedance amplitudes, indicating particular cellular changes. Dead cell subpopulations become evident in cultures with increasing cell densities. In addition, a novel intermediate subpopulation, which most probably represents apoptotic cells, was identified. These results emphasize the extraordinary sensitivity of high frequency impedance measurements and their suitability for hybridoma cell culture quality control.