Hrp conjugated anti rabbit igg or anti mouse igg

A polyclonal antibody against the MIB2-134aa protein produced by circMIB2 was obtained by inoculating rabbits (GenScript). Cellular and tissue lysates were generated by using 1 × SDS-PAGE loading buffer. Proteins were extracted from cells and measured with the BCA Protein Assay kit (Vazyme), then subjected to SDS-PAGE (8%) gel and transferred to PVDF (Millipore) membranes by semidry blotting (Bio-Rad Trans Blot Turbo System). The antibody against TRAF6 was diluted at 1:500 (Abcam); The antibody against MIB2 was diluted at 1:500 (Abcam); The antibody against MIB2-134aa was diluted at 1:200 (GenScript); anti-Flag, anti-HA, anti-Myc, and anti-Tubulin monoclonal antibody were diluted at 1:2000 (Sigma); and HRP-conjugated anti-rabbit IgG or anti-mouse IgG (Abbkine) at 1:5000. The immunoreactive proteins were detected by using WesternBright^TM ECL (Advansta). The digital imaging was performed with a cold CCD camera.

A polyclonal antibody against the Ythdc2-170aa polypeptide produced by circYthdc2 was obtained by inoculating rabbits (GenScript). Cellular lysates were generated using 1 × SDS-PAGE loading buffer. Proteins were extracted from cells and measured with the BCA Protein Assay kit (Vazyme), then subjected to SDS-PAGE (8%) gel and transferred to PVDF (Millipore) membranes by semidry blotting (Bio-Rad Trans Blot Turbo System). The membranes were blocked with 5% BSA. Protein was blotted with different antibodies. The antibody against STING was diluted at 1: 500 (Abcam); The antibody against Ythdc2 was diluted at 1: 500 (Abcam); the antibody against Ythdc2-170aa was diluted at 1: 200 (GenScript); anti-Flag, anti-HA, anti-Myc, and anti-Tubulin monoclonal antibody were diluted at 1: 2,000 (Sigma); and HRP-conjugated anti-rabbit IgG or anti-mouse IgG (Abbkine) at 1: 5,000. The results were representative of three independent experiments. The immunoreactive proteins were detected using WesternBright™ ECL (Advansta). The digital imaging was performed with a cold CCD camera.

Cellular lysates were generated by using 1× SDS-PAGE loading buffer. The proteins were extracted from the cells, and the concentrations were measured with a BCA Protein Assay kit (Vazyme), then subjected to SDS-PAGE (10%) gel and transferred to PVDF (Millipore) membranes by semidry blotting (Bio-Rad Trans Blot Turbo System). The membranes were blocked with 5% BSA. The Proteins were blotted with different antibodies. Antibodies against RIPK2 was diluted at 1:400 (Abcam); anti-Flag and anti-β-actin monoclonal antibodies were diluted at 1:2,000 (Sigma, USA); HRP-conjugated anti-rabbit IgG or anti-mouse IgG (Abbkine) was diluted 1:5,000. The results are representative of three independent experiments. Immunoreactive proteins were detected using WesternBright™ ECL (Advansta). Digital imaging was performed by using cold charge-coupled device (CCD) camera. The Image J analysis was used to analyze the grayscale of the protein band and get the results of the gray value. The ratio of the gray value of target protein against the gray value of β-actin was the relative expression of protein.

Cellular lysates were generated by using 1×SDS-PAGE loading buffer. Proteins were extracted from cells and measured with the BCA Protein Assay kit (Vazyme), then subjected to SDS-PAGE (10%) gel and transferred to PVDF (Millipore) membranes by semidry blotting (Bio-Rad Trans Blot Turbo System). The membranes were blocked with 5% BSA. Protein was blotted with different antibodies. The antibody against TBK1 was diluted at 1: 400 (BOSTER, BA3138); anti-Flag, and anti-Tubulin monoclonal antibody were diluted at 1: 2,000 (Sigma); and HRP-conjugated anti-rabbit IgG or anti-mouse IgG (Abbkine) at 1: 5,000. The results were the representative of three independent experiments. The immunoreactive proteins were detected by using WesternBright™ ECL (Advansta). The digital imaging was performed with a cold CCD camera.

Cellular lysates were generated by using 1 × SDS-PAGE loading buffer. Proteins were extracted from cells and measured with the BCA Protein Assay kit (Vazyme), then subjected to SDS-PAGE (10%) gel and transferred to PVDF (Millipore) membranes by semidry blotting (Bio-Rad Trans Blot Turbo System). The membranes were blocked with 5% BSA. Protein was blotted with different antibodies. The antibody against MAVS was diluted at 1: 500 (Abcam); anti-Flag and anti-Tubulin monoclonal antibody were diluted at 1: 2,000 (Sigma); the anti-GFP monoclonal antibody was diluted at 1: 1,000 (Sigma); and HRP-conjugated anti-rabbit IgG or anti-mouse IgG (Abbkine) at 1: 5,000. The results were the representative of three independent experiments. The immunoreactive proteins were detected by using WesternBright^TM ECL (Advansta). The digital imaging was performed with a cold CCD camera.

Cellular lysates were generated by using 1×SDS-PAGE loading buffer. Proteins were extracted from cells and measured with the BCA Protein Assay kit (Vazyme), then subjected to SDS-PAGE (8%) gel and transferred to PVDF (Millipore) membranes by semidry blotting (Bio-Rad Trans Blot Turbo System). The membranes were blocked with 5% BSA. Protein was blotted with different antibodies. The antibody against TRIF was diluted at 1: 500 (Abcam); anti-Flag and anti-Tubulin monoclonal antibody were diluted at 1: 2,000 (Sigma); and HRP-conjugated anti-rabbit IgG or anti-mouse IgG (Abbkine) at 1: 5,000. The results were representative of three independent experiments. The immunoreactive proteins were detected by using WesternBright ECL (Advansta). The digital imaging was performed with a cold CCD camera.