Lipiradical green

Detection of intracellular Fe²⁺ was performed using FerroOrange (excitation: 561 nm, emission: 570–620 nm) (Dojindo). Cells were cultured in glass-bottomed dishes for 3 d and washed with HBSS (Thermo Fisher Scientific), after which 1 μM FerroOrange was added for 30 min at 37°C, followed by observation under a microscope (BZ-X800; Keyence). Lipid radicals were detected by LipiRADICAL Green (Funakoshi) (excitation: 470 nm, emission: 520–600 nm). Cells were cultured in glass-bottomed dishes for 3 d and washed with HBSS, after which 1 μM LipiRADICAL Green was added for 10 min at 37°C, followed by observation under a microscope (BZ-X800; Keyence).

Detection of intracellular Fe²⁺ was performed using FerroOrange (excitation, 561 nm; emission, 570–620 nm) (DOJINDO, Kumamoto, Japan). Cells were cultured in glass-bottomed dishes for 3 days and washed with HBSS (Thermo Fisher), after which 1 µM FerroOrange was added for 30 min at 37°C, followed by observation under a microscope (BZ-X800; KEYENCE). To detect Fe²⁺ in mitochondria, 5 µM Mito-FerroGreen working solution (excitation, 488 nm; emission, 500–550 nm) (DOJINDO, Kumamoto, Japan) was added to the cells for 30 min at 37°C, followed by observation under a microscope. Lipid radicals were detected using LipiRADICAL Green (Funakoshi, Tokyo, Japan) (excitation, 470 nm; emission, 520–600 nm) and Liperfluo (excitation, 524 nm; emission, 535 nm) (DOJINDO, Kumamoto, Japan). Cells and cardiomyocyte were cultured in glass-bottomed dishes for 3 days and washed with HBSS, after which 1 µM LipiRADICAL Green or 4 µM Liperfluo working solution was added for 10 min at 37°C, followed by observation under a microscope (BZ-X800; KEYENCE).