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. Author manuscript; available in PMC: 2014 Mar 26.
Published in final edited form as: Biochem J. 2013 Feb 1;449(3):613–621. doi: 10.1042/BJ20121198

Figure 5. ASC CARD is required for foci formation between NLRC4 CARD and Caspase-1 CARD and for NLRC4 inflammasome activation upon Salmonella infection.

Figure 5

A. BiFC bridge assay. HEK293T cells were transfected with VC-NLRC4 CARD and VNCaspase-1 CARD. Addition of WT ASC full-length led to a change from diffuse to punctate staining, whereas ASC full-length mutants E130R and D134R were incapable of inducing foci formation (left). Immunostaining for ASC and ß-actin showing proper expression levels for WT ASC and mutants (right). B. RAW264.7 cells stably expressing WT ASC, mutant ASC or empty vector were infected with Salmonella (MOI 20) for 3 hours. Supernatants were analyzed for IL-1ß production using ELISA (mean±SD; n=3) and immunoblotting using anti-IL-1ß antibody. Cell lysates were also analyzed by immunoblotting, showing equivalent expression levels of ASC WT and ASC mutants. Cell lysates were probed for ß-actin as a loading control.