This is a knockout-validated antibody summary, based on the publications "STING-dependent sensing of self-DNA drives silica-induced lung inflammation", as cited below [1], "Autophagy regulates inflammatory programmed cell death via turnover of RHIM-domain proteins" for western blot knockout validation (see [2] figure 2s1b) and "An NLRP3 inflammasome-triggered cytokine storm contributes to Streptococcal toxic shock-like syndrome (STSLS)" for western blot knockout validation (figure 2b) [3]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Figure 1. Immunoblots of the indicated proteins in lungs from WT, STING-/- and NLRP3-/- mice. From [1].

Rabbit monoclonal IgG

Company: Cell Signaling

Antibody: NLRP3

Catalog number: 15101

Summary: Rabbit monoclonal IgG against a synthetic peptide corresponding to residues surrounding Ala306 of mouse NLRP3 protein. Reacts with human, and mouse. It is predicted to react based on 100% sequence homology with hamster. Suitable for western blot and immunoprecipitation.

Western blot

Lung tissues from WT, STING-/- and NLRP3-/- mice. Tissues were lysed with 0.5 ml of 1X RIPA lysis buffer (50 mMTris pH 7.5, 150 mM sodium chloride, 0.5% sodium deoxycholate, and 1% NP-40) containing 1X complete EDTA-free Protease inhibitor cocktail tablets (Roche Diagnostics), 1X Phosphatase Inhibitor Cocktail 2 (Sigma-Aldrich), 1 mM sodium orthovanadate (Sigma-Aldrich), 25 mM sodium fluoride (Sigma-Aldrich), and 1 mM phenylmethylsulfonyl fluoride (Sigma-Aldrich).

10% non-fat milk in 1X TBS-T (20 mM Tris Base, 150 mM sodium chloride, and 0.05% Tween-20 pH 7.6).

1/1000 dilution.

1/4000 dilution goat anti-rabbit-IgG-HRP-conjugated (#ab97057; Abcam) in 5% non-fat milk in TBS-T for 2 h at RT.

Amersham ECL Prime Western Blotting Detection Reagent (Thermofischer) was used to develop the blots on film (PXi gel doc system®).