Budding-driven localization of lipid domains at prescribed positions in confined giant unilamellar vesicles

Giant unilamellar vesicles (GUVs) are widely used minimal cell models where essential biological features can be reproduced, isolated and studied. Although precise spatio-temporal distribution of membrane domains is a process of crucial importance in living cells, it is still highly challenging to generate anisotropic GUVs with domains at user-defined positions. Here we describe a novel and robust method to control the spatial position of lipid domains of liquid-ordered (Lo) / liquid-disordered (Ld) phase in giant unilamellar vesicles (GUVs). Our strategy consists in confining Lo/Ld phase-separated GUVs in microfluidic channels to define free budding regions where the minority phase lipid domains localize and grow by decreasing the line tension between the co-existing phases. We show that this process is governed by the respective fraction of the two phases, and not by the chemical nature of the lipids involved. The spatial position and number of domains are controlled by the design of the confining microchannel and could result in polarized GUVs with a controllable number of poles. The developed method is versatile and user-friendly, while allowing to perform multiple single-vesicle experiments in parallel.