Imaging ofXenopus laevisOocyte Plasma Membrane in Physiological-Like Conditions by Atomic Force Microscopy

Abstract Xenopus laevisoocytes are an interesting model for the study of many developmental mechanisms because of their dimensions and the ease with which they can be manipulated. In addition, they are widely employed systems for the expression and functional study of heterologous proteins, which can be expressed with high efficiency on their plasma membrane. Here we applied atomic force microscopy (AFM) to the study of the plasma membrane ofX. laevisoocytes. In particular, we developed and optimized a new sample preparation protocol, based on the purification of plasma membranes by ultracentrifugation on a sucrose gradient, to perform a high-resolution AFM imaging ofX. laevisoocyte plasma membrane in physiological-like conditions. Reproducible AFM topographs allowed visualization and dimensional characterization of membrane patches, whose height corresponds to a single lipid bilayer, as well as the presence of nanometer structures embedded in the plasma membrane and identified as native membrane proteins. The described method appears to be an applicable tool for performing high-resolution AFM imaging ofX. laevisoocyte plasma membrane in a physiological-like environment, thus opening promising perspectives for studyingin situcloned membrane proteins of relevant biomedical/pharmacological interest expressed in this biological system.