Fig. 2. Formation of reticular and fibrillar laminin patterns. (a-b) Schwann cells were incubated with fragments and stained with antibodies specific for the corresponding fragments following washing and fixation. Only E3 was found to bind to cell surfaces, appearing in a punctate pattern (a). Fragments E8, E4 and E1' had levels corresponding to background (b). (c-h) Schwann cells were pre-incubated for 20 minutes with blocking or activating reagents followed by the addition of 2 µg/ml of laminin-1. The laminin distribution was detected 8 hours after incubation with EHS laminin-specific antibody (c,d,e,h) or fragment E4-specific antibody (f,g). Laminin-1 (c) formed both fibrillar (arrows) and reticular patterns (box, with corresponding twofold magnification inset) without blocking reagents. In the presence of ß1-integrin blocking antibody Ha2/5 (10 µg/ml), laminin assembled almost exclusively into reticular patterns. In the presence of -dystroglycan blocking antibody IIH6 (e), laminin was detected in fibrillar structures (arrows) with only few small surviving reticular arrays. Coincubation of laminin with E3 (50 µg/ml) substantially decreased formation of reticular, but not fibrillar (arrows) arrays. Addition of both E3 fragment and anti ß1-integrin blocking antibody inhibited formation of both reticular and fibrillar structures, with surviving laminin largely in a punctate distribution (g). Treatment with 0.5 mM MnSO₄ increased the ratio of the area occupied by fibrillar structures compared with reticular structures (h). All images were taken at the same exposure settings and are shown at the same magnification (bar, 10 µm). (i) Laminin fragment map. (j) Relative surface area covered by laminin determined for the conditions described in c-h.