Gfp sc 9996

Ammonium chloride (NH₄Cl), arsenic trioxide (As₂O₃) and oleic acid (OA) were purchased from Sigma-Aldrich (Milwaukee, WI, USA); Ambroxol hydrochloride (Amb) and cyclohexylamine were from Santa Cruz Biotechnology (Santa Cruz, CA, USA). An arsenite solution was prepared by dissolving As₂O₃ in minimal volumes of 1 N sodium hydroxide (NaOH). The arsenite solution (referred to in the manuscript as As III) was then diluted with phosphate buffered saline to a concentration of 10 mM as a stock solution.
For Amb and OA, the desired concentrations were obtained by dissolving Amb in DMSO and OA in ethanol. Other compound solutions were prepared as per the manufacturer's recommendations. The AR (N-20, used at a 1:1000 dilution) and GFP (sc-9996; also used at a 1:1000 dilution) antibodies were purchased from Santa Cruz Biotechnology. The β-catenin antibody (#4270; 1:1000) was purchased from Cell Signaling Technology (Danvers, MA, USA) and the anti-α-tubulin antibody (1:3000) from Sigma-Aldrich; the anti-BCL2 antibody (clone 124; 1:500) was from Dako (Santa Clara, CA, USA).

Western blots were performed as previously described (Ramos et al. 2019 (link)). Briefly, cellular extracts and purified protein samples were fractionated on NuPAGE Bis-Tris polyacrylamide gels (Thermo Scientific) followed by transfer to Immobilon FL PVDF membrane (Millipore) for immunoblotting. For analysis of skin fibroblasts, 1 × 10⁶ cells were harvested and proteins were extracted using radioisotope immunoprecipitation assay (RIPA) buffer (50 mM TrisHCl, pH 7.5, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, 150 mM NaCl, 2 mM EDTA). Antibodies were against the following proteins : 6xHis tag (MA1-21315, Thermo Fisher), GFP (sc-9996, Santa Cruz Biotechnology), Strep-tag II-tag (NC9261069, Thermo Fisher), ADAT3 (Abcam, ab192987), ADAT3 (H00113179-B01P, Abnova), ADAT2 (ab135429, Abcam), and actin (CST). Primary antibodies were detected using IRDye 800CW Goat anti-Mouse IgG (SA5-35521, Thermofisher) or Rabbit (SA5-35571, Thermofisher) or Rat (925-32219, LI-COR Biosciences), or IRDye 680RD Goat anti-Mouse IgG (926-68070, LI-COR Biosciences) or Rabbit (925-68071). Immunoblots were scanned using direct infrared fluorescence via the Odyssey System (LI-COR Biosciences).