Real-time tagless monitoring of cell viability using patch-clamp microchips is reported and validated by using fluorescence imaging techniques for the first time. Specifically, four human breast cancer cell lines (MDA-MB231, MDA-MB231-brain metastatic subline (abbreviated as MB231-BR), MB231-BR over-expressing HER2 gene (MB231-BR-HER2), and MB231-BR-vector control for the HER2 (MB231-BR-vector)) have been used for these studies. Systematic experiments on these cells found that the seal impedance/resistance of cells captured by the micro-pipettes always decreases during the process when the cell loses its viability, and therefore it is a valid indicator of live or dead cells. Systematic experiments also found that the Mega-seal of patch-clamp microchip is sufficient for monitoring cell viability. Given its simplicity of direct electrical measurement of cells without fluorescence labeling, this technology may provide an efficient technical platform to monitor the drug effects on cells, thereby significantly benefiting high throughput drug screening and discovery process.