Anti hsp90 antibody

Whole cell extracts (WCEs) were prepared in RIPA buffer. Protease inhibitor cocktail (Roche) and phosphatase inhibitor cocktail (Sigma) were included in lysis buffer. Secreted proteins in culture media were precipitated by trichloroacetic acid. For immunoprecipitation, anti-RIPK3 (Prosci) or anti-Caspase 8 (Enzo Life Sciences) antibodies were used. Western blotting was performed with anti-RIPK1 (BD Biosciences), RIPK3 (Prosci), FADD (kindly provided by A. Winoto), Caspase 8 (Enzo Life Sciences), MyD88 (eBioscienes), IL-1β, cIAP1/2 (R&D systems), GFP (Roche), Caspase-1 (Santa Cruz), and IL-18 (BioVision) antibodies. Anti-HSP90 antibody (BD Biosciences) was used as a loading control.

Anti-NgBR rabbit polyclonal antibody was generated as described in our previous publication (Zhao et al., 2010 (link)). Anti-total ERK, Akt, phos-Akt, and phos-Erk antibodies were purchased from Cell Signaling Technology (Beverly, MA). Anti-Hsp90 antibody was purchased from BD Biosciences (San Jose, CA). Anti-CCM1, CCM2 and CCM3 antibodies were purchased from Acris (San Diego, CA). Non-silencing (NS) control siRNA and siRNAs targeting NgBR were purchased from QIAGEN (Frederick, MD). Tek-Cre (Tie2-Cre) (Stock number 008863), ROSA26Sor-lacZ (stock number: 003474), and ROSA26Sor-EYFP (stock number: 006148) mice were purchased from Jackson laboratory (Bar harbor, Maine), and β-actin Cre mouse (Lewandoski et al., 1997 (link)) was a generous gift from Dr. Katherine Shim (Children’s Research Institute, MCW). Cdh5(PAC)-CreERT2 mice were transferred from Dr. Ralf Adams (Cancer Research UK).