Mouse Caspase-11/Caspase-4 Antibody

Detection of Mouse Caspase-11/Caspase-4 by Western Blot. Western blot shows lysates of mouse splenocytes untreated (-) or treated (+) with LPS. PVDF membrane was probed with 1 µg/mL of Rabbit Anti-Mouse Caspase-11/Caspase-4 Monoclonal Antibody (Catalog # MAB8648) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). Specific bands were detected for Caspase-11/Caspase-4 at approximately 36, 44, and 46 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of Mouse Caspase-11/Caspase-4 by Western Blot Pro-casp-1, casp-11, NLRP3, gasdermin D (GSDMD) expression were assessed by western blot. Statistical significance was determined by using Student’s t-test or one-way ANOVA. Asterisks indicate p-values *=p<0.05, **=p<0.01, and ***=p<0.001, only significant values are shown. All data depicted in this figure are provided as source data. Image collected and cropped by CiteAb from the following open publication (https://elifesciences.org/articles/88686), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Caspase-11/Caspase-4 by Western Blot DENV NS1-induced inflammasome activation is NLPR3-independent.(A) WT and Nlrp3-/- BMDMs were primed with PAM3CSK4 (1μg/mL) for 17h and then treated with DENV2 NS1 at indicated concentrations, nigericin (5μM), or medium (PAM only). IL-1 beta levels in supernatant 2h (nigericin) or 24h (NS1 and PAM only) were measured by ELISA. Statistical significance was determined using two-way ANOVA with Holm-Sidak’s multiple comparisons test. (B) Representative Western blots of cell lysates from WT and Nlrp3-/- BMDMs after priming with PAM3CSK4 (1μg/mL) for 17h and treatment with DENV2 NS1 (10 or 5 μg/mL), treatment with nigericin (5μM), or no treatment for 24h. (C) BMDMs were primed with PAM3CSK4 (1μg/mL) for 17h and then pre-treated with MCC950 at the indicated concentrations before addition of DENV2 NS1 (10μg/mL), nigericin (5μM), or medium (Inhibitor only). IL-1 beta levels in the supernatant after 2h (Nigericin) or 24h (NS1 and PAM only) were measured by ELISA. (D) Representative Western blots of cell lysates from BMDMs nucleofected with Cas9-gRNA ribonuclear protein complexes to knock out the indicated genes. Two gRNAs per gene were used per nucleofection. NTG = non-targeting guide. (E) Knockout BMDMs from (D) were primed with PAM3CSK4 (1μg/mL) for 17h and treated with DENV2 NS1 (10μg/mL) or left untreated for 48h. Statistical significance was determined using two-way ANOVA followed by with Holm-Sidak’s multiple comparisons test. The data are shown as the mean ± SD of 3 biological replicates (A,C), a representative image taken from 2 biological replicates (B,D), or data pooled from 8 independent experiments with at least 3 biological replicates per guide (E). *p<0.05, **p<0.01, *** p< 0.001, ****p<0.0001, ns (not significant), p> 0.05. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.ppat.1012167), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Caspase-11/Caspase-4 by Western Blot NAD+ specifically inhibits the non-canonical inflammasome by targeting caspase-11.Bone marrow was isolated from mice and bone marrow-derived macrophages (BMDMs) were differentiated in vitro. Subsequently, BMDMs were cultured in the presence of NAD+ or PBS. BMDMs were then primed with either Pam3CSK4 or lipopolysaccharide (LPS) O111:B4. Next primed BMDMs were stimulated with ATP or LPS and cholera toxin B (CTB). (A) Pro-casp-1, pro-casp-11, casp-11, NLRP3, casp-1, IL1 beta, and gasdermin D (GSDMD) expression were determined using western blot and (B) IL-1 beta secretion and LDH release were assessed in the supernatant. Column plots display mean with standard deviation (n=5-8). Image collected and cropped by CiteAb from the following open publication (https://elifesciences.org/articles/88686), licensed under a CC-BY license. Not internally tested by R&D Systems.