All-in-One Method for Iterative Single-Cell Dielectrophoretic and Optical Characterization: Advancing the OpenDEP Platform.

An integrated platform for single-cell biophysical characterization is presented. Combining dielectrophoresis (DEP) and optical tweezers (OT) within a single experiment, this approach enables the extraction of both electric properties and optical trap stiffness from individual, living cells in suspension without the need for external calibration beads, labels, or adherence to surfaces. Unlike traditional population-based DEP methods, which average over large cell ensembles and obscure cellular heterogeneity, the presented single-cell approach provides precise DEP spectra and allows direct computation of electric parameters such as membrane conductivity, permittivity, and cytoplasmic conductivity. The method is compatible with structurally and optically complex particles, such as living cells, overcoming the limitations of calibration procedures designed for spherical, homogeneous particles. It supports repeated testing of the same cell, facilitating dynamic studies of cellular responses to chemical or physical perturbations. OT stiffness measurements are performed directly on nonadherent cells that otherwise would be excluded from surface-based assays. The system includes an open-source software for data acquisition, automated image-based analysis, and OT and DEP forces computing. It is compatible with various electrode geometries, making it broadly adaptable to different experimental designs. Overall, this platform offers a robust, label-free method for high-resolution, single-cell electric and optic profiling, expanding the capabilities of DEP and OT in fundamental research, diagnostics, and bioengineering applications.