Combining Live Cell Imaging with Cellular Impedance to Monitor Apoptotic Cell Death in Real Time

Abstract

Although apoptosis can be tracked using various biomarker‐based assays, these typically require multiple manual handling steps and only yield endpoint measurements. Requiring just a cell seeding step and a drug addition step, we used the Agilent xCELLigence RTCA eSight to continuously monitor drug‐mediated apoptosis over the course of multiple days. Providing a direct and objective assessment of cell number, cell size, cell‐substrate attachment strength, and cell barrier function, impedance biosensors embedded within the base of eSight microplates quantitatively track early (cell shrinkage) to late (fragmentation) apoptotic events with high analytical sensitivity. Concurrently, eSight captures live cell images in brightfield and three fluorescence channels (red, green, and blue), providing an orthogonal readout of the apoptosis process. By combining the strengths of real‐time impedance monitoring (simplicity, analytical sensitivity, and objectivity) with that of live cell imaging (specificity of the readout), eSight increases the information richness of the apoptosis assay without increasing the workload. Importantly, the drug EC50 values determined using these two approaches are nearly identical, suggesting that eSight can simultaneously provide both the primary and secondary (confirmatory) readouts for apoptosis studies.Corroborating the impedance response with live cell images. Image panels demonstrate the progression of apoptosis 20 and 40 hours after treating A549‐Blue cells with 5.5 μm MG132. White arrows denote large membrane blebs that contain phosphatidylserine in their outer leaflet.Figure 1Multiplex monitoring of apoptosis in real‐time: combining readouts from cellular impedance and live cell imaging.Figure 2