Protocol to study internalization and localization dynamics of exogenously added proteins in detergent-permeabilized human cells

Cell-permeabilization and resealing assays are essential tools for studying the localization and functional roles of exogenous factors in detergent-permeabilized cells. We describe a protocol for studying the internalization and localization of exogenous proteins in detergent-permeabilized human cells. It includes detailed steps for culturing, permeabilizing, and preparing ghost cells, followed by lysate incubation and fixation. Subsequent immunofluorescence staining and imaging allow visualization of subcellular protein localization. This assay can be adapted to suit a variety of experimental conditions and research objectives. For complete details on the use and execution of this protocol, please refer to Ray et al.¹.