IKKα antibody | knockout validation | Santa Cruz sc-7606

This is a knockout-validated antibody summary, based on the publication "Priming Phosphorylation of TANK-Binding Kinase 1 by IκB Kinase β Is Essential in Toll-Like Receptor 3/4 Signaling", as cited below [1]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Antibody information

Mouse monoclonal IgG2b κ

Company: Santa Cruz

Antibody: IKKα

Catalog number: sc-7606

Summary: Mouse monoclonal IgG2b κ against amino acids 1-745 representing full length human IKKα. Recommended for detection of IKKα of human origin by western blot, immunoprecipitation, immunofluorescence, immunohistochemistry (paraffin), flow cytometry and ELISA.

Validation Method

Western blot

Sample

WT, IKKα-KO, and IKKα/β-DKO 293T cells.

Blocking agent

Tris-buffered saline with 0.1% Tween 20 (TBST) containing 5% skim milk for 30 min at room temperature.

Primary incubation

Diluted in the blocking buffer overnight at 4°C.

Secondary incubation

Horseradish peroxidase (HRP)-conjugated secondary antibody in TBST for 1 h at room temperature.

Detection

LAS-4000 instrument (Fujifilm) by chemiluminescence using Chemi-lumi One Super (Nacalai Tesque).

Figure: WT 293T cells and IKKα, IKKβ, and IKKα/β KO 293T cells were transfected with expression constructs for HA-tagged TLR3 and Flag-tagged IRF-3 S386A. Twenty-four hours later, cells were stimulated with 50 μg/ml of poly(I·C) for 2 h. Cell lysates were subjected to immunoprecipitation with anti-HA, followed by immunoblotting. Please see Figure 4g in the article [1].

References
  1. Abe H, Satoh J, Shirasaka Y, Kogure A, Kato H, Ito S, et al. Priming Phosphorylation of TANK-Binding Kinase 1 by IκB Kinase β Is Essential in Toll-Like Receptor 3/4 Signaling. Mol Cell Biol. 2020;40: pubmed publisher