Horseradish peroxidase hrp labeled goat anti rabbit igg

Total proteins were extracted from trigeminal ganglion and nasal mucosa tissues using RIPA lysate containing protein phosphatase inhibitor. Western blotting was performed as previously described.32 (link) After 3 washes in Tris-Buffered Saline Tween-20 (TBST), the PVDF membranes were incubated for 2h at room temperature using horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (Cell Signal Technology). After washing 3 times with TBST, ECL reagent was used for visualization and imaging was conducted by chemiluminescence imaging system.

Total protein was extracted from CAFs-derived exosomes or supernatant using radioimmunoprecipitation assay lysis buffer (Beyotime Institute of Biotechnology). Briefly, 40 mg protein were loaded on each lane, separated using 10% SDS-PAGE and transferred onto polyvinylidene fluoride membranes. Following blocking with 5% skim milk at room temperature for 30 min, the membranes were subsequently incubated with the primary antibodies (all 1:1,000) against CD9 antigen (cat. no. ab92726; Abcam), CD63 antigen (cat. no. 55051; Cell Signaling Technologies, Inc.), tumor susceptibility gene 101 protein (cat. no. ab125011; Abcam), heat shock protein 70 (HSP70; cat. no. 4873; Cell Signaling Technologies, Inc.), CD81 antigen (cat. no. 10037; Cell Signaling Technologies, Inc.) overnight at 4°C, and with horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (cat. no. G-21234; Invitrogen; Thermo Fisher Scientific, Inc.; 1:5,000) at room temperature for 2 h. Blots were visu-alized using chemiluminescence detection (GE Healthcare) and analyzed by ImageJ 1.8.0 software (National Institutes of Health). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH; cat. no. D190090; Shanghai Sangon Biological Engineering Technology & Services Co., Ltd.) was applied as an internal reference.