Supplemental file (full blot)

Published: 17 December 2020| Version 1 | DOI: 10.17632/65n6hrsvtk.1
Contributor:
Asfar Azmi

Description

Full blots for western blots

Files

Steps to reproduce

50,000 BON-1 cells or 100,000 QGP-1 were grown in 100 mm petri dishes overnight. The following day, each cell line was treated with specified concentrations of KPT-9274, PF-3758308, Everolimus, FK866, and combinations for 72 hours. 50 μg of protein lysates from treated and untreated cells were separated in a 10-12% SDS-PAGE and transferred into a nitrocellulose membrane (GE Healthcare Life Sciences). Mouse monoclonal antibodies anti-PAK4, anti-NAMPT, anti-NAPRT, anti-β-tubulin from Santa Cruz Biotechnology (Dallas, Texas), and anti-pmTOR, anti-RICTOR, anti-RAPTOR, anti-Pp70S6K, anti-P70S6K, anti-mTOR, from cell signaling were used at a 1:1000 dilution in (3% non-fat milk or 5% FBS) PBS Tween-20.