Three distinct permeabilization procedures affect strongly detection of surface csA-SA. Cells were treated following a classical immunofluorescence procedure (upper panel) or a two-step procedure (lower panel). Cells were permeabilized using methanol at −20°C (A, B) triton X-100 (C, D) or saponin (E, F). All the pictures presented in this figure were taken sequentially with the same microscope and with identical settings. Scale bar: 5 μm.