Detecting RIPK3 and MLKL Oligomers

Western blotting experiment was described previously (Zhao et al., 2019 (link)), and the detection of RIPK3 or MLKL oligomers was performed according to the method described by Wang et al. (2019) (link). The following antibodies were used for the experiments: anti-FADD (1:1000, 610399) and anti-RIPK1 (1:3000, 610458) (BD Transduction Laboratories, San Jose, CA); anti-phospho-RIPK3 (1:1000, ab195117), anti-MLKL (1:1000, ab172868), anti-phospho-MLKL (1:1000, ab196436) (Abcam, Cambridge, MA, United States); anti-RIPK1 (1:3000, 3493), anti-RIPK3 (1:3000, 15828), anti-Myc tag (1:2000, 2276), anti-HA tag (1:2000, 3724), anti-Flag tag (1:2000, 14793), anti-TRAF2 (1:1000, 4724), anti-phospho-RIPK1 (1:1500, 31122) and anti-phospho-RIPK3 (1:1000, 57220) [Cell Signaling Technology (CST), Beverly, MA]; anti-TRADD (1:1000, ABP52634) and anti-GAPDH (1:1500, ABP52783) (Abbkine, Redlands, CA, United States); anti-β-actin (1:3000, A5441) (Sigma-Aldrich). All western blot assays were performed three times, and the representative results are shown.