Western Blot Analysis of Cyclophilin A Protein

Western blot analysis was performed as previously described (T. Liu et al., 2013 (link)). Briefly, 50 µg of protein was loaded per sample. Total protein was isolated and analyzed by blotting using appropriate antibodies: anti‐CypA antibody, which was a rabbit polyclonal antibody generated for CypA, cloned from M. bicoloratus hemocytes and expressed in Escherichia coli using the pET‐32a‐CypA plasmid (1:2000; Bioworld Technology, Inc.), anti‐rabbit AIF polyclonal antibody (1:2000; cat. no. bs‐0037R; Bioss Technology, Inc.), mouse anti‐GAPDH antibody (1:2000; cat. no. T0004; Affinity, USA), mouse anti‐VDAC1 monoclonal antibody (1:2000; cat. no. AF1027; Beyotime), anti‐mouse V5 Tag monoclonal antibody (1:2000; cat. no. R960‐25; Invitrogen), anti‐α tubulin monoclonal antibody (1:2000; Cat. no. K006154P; Solarbio), anti‐mouse GST tag monoclonal antibody (1:2000; Cat. no. AG768; Beyotime), and a goat anti‐mouse horseradish peroxidase‐conjugated secondary antibody (1:2000; Cat. no. 0216; Beyotime). ImageJ (National Institutes of Health) was used to measure protein band density.