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© 2020 WILEY-VCH Verlag GmbH Co. KGaA, Weinheim.Dispersed negatively charged silica nanoparticles segregate inside microfluidic water-in-oil (W/O) droplets that are coated with a positively charged lipid shell. We report a methodology for the quantitative analysis of this self-assembly process. By using real-time fluorescence microscopy and automated analysis of the recorded images, kinetic data are obtained that characterize the electrostatically-driven self-assembly. We demonstrate that the segregation rates can be controlled by