Supplementary Figure 12: Multicolor 4Pi-SMS images of the ER and contact site proteins in COS-7 cells.

(a) Two-color image of the ER membrane (overexpressed GFP-Sec61β labeled with anti-GFP antibody; shown in the upper right half) and ORP5 (overexpressed mCherry-ORP5 labeled with anti-mCherry antibody; shown in the lower left half). (b) y-z view of the boxed region (3 × 1 μm) in (a). (c) Axial intensity profile averaged across the dashed box in (b). (d) Two-color image of the ER membrane (overexpressed GFP-Sec61β labeled with anti-GFP antibody; shown in the upper right half) and E-Syt2 (overexpressed mCherry-E-Syt2 labeled with anti-mCherry antibody; shown in the lower left half). (e) y-z view of the boxed region (3 × 1 μm) in (d). (f) Axial intensity profile averaged across the dashed box in (e). (g) Three-color image of Sec61β, ORP5 and WGA (Supplementary Video 7, part I). y-z view of the boxed region in (g) is shown in (Fig. 4h). (h) Three-color image of Sec61β, E-Syt2 and WGA (Supplementary Video 7, part II). y-z view of the boxed region in (h) is shown in (Fig. 4j). Representative images of five (a-b), three (d-e) or two (g-h) independent experiments are shown.