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. Author manuscript; available in PMC: 2016 Jan 31.
Published in final edited form as: J Immunol. 2014 Dec 29;194(3):1274–1284. doi: 10.4049/jimmunol.1402388

Figure 9. CRAMP binds to CpG DNA.

Figure 9

A and B, RAW-TLR9-HA cells transfected with mCAP18 or control were stimulated with or without 1 μM CpG-C for 90 min. Cells were washed and lysed. Cell lysates were immunoprecipitated with anti-CRAMP and immunoblotted with anti-HA (A) or immunoprecipitated with anti-HA and immunoblotted with anti-CRAMP (B). Total cell lysates were immunoblotted with anti-CRAMP or β-actin for control. C, 10 μM mCRAMP was incubated with 1 μM CpG-C or 1 μM biotinylated-CpG-A, -CpG-B or -CpG-C for 1 h. Biotinylated-CpG was recovered by streptavidin beads, and then binding of CRAMP to CpG was examined by immunoblot with anti-CRAMP. D, Various concentration of mCRAMP were incubated with 1 μM CpG-C or 1 μM biotinylated-CpG-C for 1 h. Biotinylated-CpG was recovered by streptavidin beads, and then binding of CRAMP to CpG was examined by immunoblot with anti-CRAMP. E and F, RAW-TLR9-HA cells were transfected with mCAP18, CRAMP or control. Cells were lysed and immunoprecipitated with anti-CRAMP and immunoblotted with anti-HA, or immunoprecipitated with anti-HA and immunoblotted with anti-CRAMP. G, WT and Camp−/− GM-CSF BMDCs were stimulated with 1 μM CpG-C or CpG-C-biotin for 1 h. Cells were washed and lysed. Cell lysates were incubated with streptavidin beads. CpG-C-biotin-bound CRAMP was immunoblotted with anti-CRAMP. Data are representative of three independent experiments.