Anti adar1 mouse monoclonal antibody

Various antibodies were used, including anti-ADAR1 rabbit polyclonal antibody (Thermo Fisher), anti-Normal rabbit IgG (Sigma Aldrich), anti-GAPDH rabbit monoclonal antibody (Abcam), anti-KYNU rabbit polyclonal antibody (Thermo Fisher), anti-ADAR1 mouse monoclonal antibody (Thermo Fisher). Secondary antibodies were used, including goat anti-mouse (Abcam) and goat anti-rabbit IgG HRP (Thermo Fisher). For western blotting analysis, the primary antibodies dilutions were 1:1000 and GAPDH was used as the loading control. Secondary antibodies for western blotting analysis were diluted to 1:7000. Additional reagents used include pierce protein A/G agarose (Thermo Fisher), Protease inhibitor cocktail, EDTA-Free (100X) (Thermo Fisher), RIPA buffer, Pierce ip lysis buffer (Thermo Fisher), Sterile conical centrifuge tubes 15 ml (Thermo Fisher), cell culture flask 25 cm (Thermo Fisher).

Cell were seeded on coverslips until they reached between 80 to 90% confluency. The fixation step was accomplished by incubating the cell coated coverslips in 4% paraformaldehyde for 12 min at room temperature (RT). The permeabilization step was performed by incubating the cell coated coverslips in (0.2% Triton X-100) for 20 min at RT. Cells were then blocked for 45 min at RT with 2% goat serum. Cells were incubated overnight at 4 °C with a specific primary antibody (anti-ADAR1 mouse monoclonal antibody (Thermo Fisher). The primary antibody was diluted 1:200. The cells were then incubated for one hour at RT with the secondary antibody. Then, cells were incubated with the mounting media with the DAPI for few minutes. Cells were imaged using confocal microscopy (Zeiss LSM 700) fitted with a × 20 and × 40 objective.