This is a knockout-validated antibody summary, based on publications "Mitochondrial permeabilization engages NF-κB-dependent anti-tumour activity under caspase deficiency", as cited below [1], "A NIK-SIX Signalling Axis Controls Inflammation by Targeted Silencing of Non-Canonical NF-κB" for western blot (figure e5b) [2], and "KIX domain determines a selective tumor-promoting role for EP300 and its vulnerability in small cell lung cancer" for western blot knockout validation (figure 4b) [3]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Rabbit monoclonal IgG

Company: Cell Signalling

Antibody: NF-κB p65

Catalog number: 8242

Summary: Rabbit monoclonal IgG against a synthetic peptide corresponding to residues surrounding Glu498 of human NF-κB p65/RelA protein. Reacts with human, mouse, rat, hamster, monkey, and dog. Suitable for western blot, immunoprecipitation, immunohistochemistry (paraffin), immunofluorescence (immunocytochemistry), flow cytometry, chromatin IP, and chromatin IP-seq.

Western blot | Immunocytochemistry

Control and p65-deleted BCL-xL dependent SVEC cells.

WB: Cells were lysed in NP-40 lysis buffer (1% NP-40, 1mM EDTA, 150mM NaCl, 50 mM Tris pH 7.4, 1 mM PMSF and complete protease inhibitors (Roche)).

IC: Cells were fixed in 4% PFA/PBS for 10 minutes and then permeabilized with 0.2%

Triton/PBS for 10 minutes.

IC: 2% BSA/PBS for 1 hour at room temperature.

WB: 1/1000 dilution overnight in 4°C.

IC: 1/400 dilution in blocking buffer overnight at 4°C.

WB: HRP-linked secondary antibodies.

IC: 1/300 dilution Alexa Fluor 594 goat anti-rabbit (A11037, Invitrogen).

WB: ECL.

IC: Nikon A1R confocal microscope.

Cell lysates from control and p65-deleted BCL-xL dependent SVEC cells were immunoblotted for p65 and actin. Control or p65-deleted BCL-xL dependent SVEC cells were treated with ABT-737 (10 uM) and Q-VD-OPh (30 uM) for 6 h and immunostained for nuclear p65. Scale bar represents 30 uM. Please see Suppl. Figure 4e,f in the article [1].