Super-Resolution Imaging and Quantitative Analysis of Membrane Protein/Lipid Raft Clustering Mediated by Cell-Surface Self-Assembly of Hybrid Nanoconjugates

Super-resolution imaging was used to quantify organizational changes in the plasma membrane after treatment with hybrid nanoconjugates. The nanoconjugates crosslinked CD20 on the surface of malignant B cells, thereby inducing apoptosis. Super-resolution images were analyzed by using pair-correlation analysis to determine cluster size and to count the average number of molecules in the clusters. The role of lipid rafts was investigated by pre-treating cells with a cholesterol chelator and actin destabilizer to prevent lipid raft formation. Lipid raft cluster size correlated with apoptosis induction after treatment with the nanoconjugates. Lipid raft clusters had radii of similar to 200 nm in cells treated with the hybrid nanoconjugates. Super-resolution images provided precise molecule location coordinates that could be used to determine density of bound conjugates, cluster size, and number of molecules per cluster.