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. Author manuscript; available in PMC: 2015 May 1.
Published in final edited form as: Traffic. 2014 Mar 7;15(5):516–530. doi: 10.1111/tra.12159

Figure 6. Exocytic SNARE complex formation is impaired in E. coli exposed mast cells.

Figure 6

(A) RBLs were cultured with or without (control) E. coli for 2h at an MOI of 10,000. The cells were harvested and stimulated for 30min with 1μM PMA/1μM Ionomycin at a concentration of 2×106 cells/ml. Cells were treated with 1mM NEM for 15min. Next, NEM was inactivated with 2mM DTT for 15min on ice. NEM-treated cells were washed and lysed. 1mg of total protein was immunoprecipitated using anti-SNAP23 antibody. Samples were analyzed by Western blot using anti-SNAP23, -Syntaxin4, -VAMP8, and –VAMP2 antibodies. Each blot is representative of at least 3 independent experiments. (B) Quantification of the mean band intensity ratio’s of the indicated protein to SNAP23 ± the standard deviation of at least 3 independent experiments. The protein:SNAP23 ratio of the control population was considered 100%. Asterisks denote statistically significant p values, where * ≤ 0.05 and ** ≤ 0.01. (C) 40μg of total protein from exposed or control whole cell lysates was analyzed by Western blot as described in A.