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. 2002 Aug 19;158(4):751–760. doi: 10.1083/jcb.200203088

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Copyright © 2002, The Rockefeller University Press

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Figure 1. — Syntaxin 1A binds to endogenous SNAP-25 on inside-out sheets of plasma membrane derived from PC12 cells. (A and B) Membrane sheets were reacted for 50 min at 37°C with syntaxin 1A(1–262)–Alexa594 (red channel) and then washed, fixed, and immunostained for SNAP-25 (green channel). (C and D) Magnified views from A and B. (E) Overlap from C and D. Circles indicate identical pixel locations. (F) On membrane sheets such as shown in A and B the staining intensity of random images in the green channel was measured and plotted against the staining intensity in the red channel, showing a linear correlation. (G) Effect of SNAP-25 cleavage by 2 μM of BoNT/E and/or Cl 71.1 (antibody specific for SNAP-25) on syntaxin binding, measured as above. Dilution of the antibody as indicated. (H) Effect of antibodies against rab3 (Cl 42.1), synaptobrevin 2 (Cl 69.1), and SNAP-25 (Cl 71.1) on syntaxin binding. (I) Binding of syntaxin in the presence of 2 μM BoNT/C1 (cleaving syntaxin) and 2 μM TeNT (cleaving synaptobrevin).