Irdye donkey anti rabbit 800 and goat anti mouse 680 secondary antibodies

For assessment of protein expression in Figure 1—figure supplement 2 , parental, TRE3G::BRAF^WT and TRE3G::BRAF^V600E cells were plated in 6-well plastic culture plates, and starved with 0.5% HS, DMEM/F12 containing P/S with 1 mM Na Pyruvate and 10 mM HEPES overnight before treatment with media or doxycycline (2 µg/ml) for 24 hr. Samples were lysed with RIPA buffer (CST) containing HALT protease and phosphatase inhibitors (Thermo), and reduced in Laemelli SDS buffer (BioRad) with BME (Sigma). Samples underwent electrophoresis on 4–15% gradient polyacrylamide gels (BioRad) and were immunoblotted with Rabbit anti-BRAF (CST) and mouse anti-HSC70 (Santa Cruz Biotechnology), and IRDye donkey anti-rabbit 800 and goat anti-mouse 680 secondary antibodies (LiCor) before imaging. For validation of ADAM17 CRISPR-KOs in Figure 3 , suspected clones were grown, lysed, and run on a gel as described above, before immunoblotting with Rabbit anti-ADAM17 (CST) and mouse anti-HSC70 (Santa Cruz Biotechnology) primary and IRDye donkey anti-rabbit 800 and goat anti-mouse 680 secondary antibodies (LiCor). All images were acquired on an Odyssey Infrared Scanner (LiCor).