Integrated transcriptomic and bioelectrical profiling of stem-like cellular states in a colorectal cancer using SdFFF and UHF-DEP

Cancer stem-like cells (CSCs) enrichment and characterization remain an analytical challenge, as conventional marker-based approaches are often time-consuming and can alter cellular properties. Here, we develop a complete label-free protocol for the enrichment and characterization of cellular states associated to stemness properties in colorectal cancer model, based on the coupling adjustment of sedimentation field-flow fractionation (SdFFF) and ultra-high-frequency dielectrophoresis (UHF-DEP). To this end, the operating conditions of SdFFF were adapted to a low conductivity dielectrophoresis buffer while maintaining the hyperlayer elution regime. Fractionated cellular subpopulation exhibits phenotypic differences in proliferative status, clonogenic potential, stem-cell related protein levels and UHF-DEP highlights significantly different high-frequency crossover distributions. Transcriptome sequencing revealed dysregulation of pathways associated with stemness, cell adhesion, membrane organization, calcium signaling and ion-channel activity, that can be linked to dielectric signatures. Overall, UHF-DEP coupling to SdFFF permit to enrich, characterize and quantify stem cell-like subpopulation within heterogeneous population of cancer cells.