Mito Stress Test: Seahorse Analysis

UM cell lines were seeded on poly-Lysine coated 96-well Seahorse plates (Agilent Technologies); 24 hours later the cells were treated with IACS-010759 at 1.5 nm (the lowest effective dose tested) for the next 24 hours. The level of oxygen consumption rate (OCR) was measured in a Seahorse Mito stress test using the Agilent Seahorse XF Cell Mito Stress Test Kit. OCR, as well as reserve capacity, were measured following the protocol described in the kit. For normalization purposes, after Seahorse testing, cells were stained with Hoechst and propidium iodide, and scanned in Operetta and cell numbers utilized for normalization.