Anti pho ikkβ

Cells were transfected with the appropriate vectors, as indicated in each figure. Western blotting and IP assays were performed as described previously (25 (link)26 (link)27 (link)28 (link)). HEK293T cells were transfected with a mock vector as a control vector and appropriate vectors. At 38 h post-transfection, the transfected cells were extracted and the cell lysates were subjected to immunoprecipitation with anti-Flag or anti-Myc antibodies followed by immunoblotting (IB) using anti-Flag, anti-Myc, anti-HA, anti-p62, anti-TRAF6, or anti-ECSIT antibodies. WT p62^+/+ MEF and p62^−/− MEF cells were stimulated without or with LPS for different times, lysed in lysis buffer, and the lysates were examined by Western blotting with anti-p62, anti-IKKαβ, anti-pho-IKKβ, anti-pho-TAK1, anti-TAK1, and anti-GAPDH antibodies (Cell Signaling Technology).

Specific anti-HA, anti-Flag anti-Myc, anti-TRAF6, anti-pho-TAK1, anti-TAK1, anti-pho-IKKβ, anti-IKKαβ, and anti-GAPDH antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA). The anti-p62 and anti-ECSIT antibodies were purchased from Abcam (Cambridge, MA, USA). Mouse TrueBlot ULTRA: anti-Mouse Ig HRP was purchased from Rockland Immunochemicals, Inc. (Limerick, PA, USA).