Cardiolipin oxidation analysis

Cardiolipin oxidation in mitochondria was determined with 10-N-Nonyl-Acridine Orange (NAO). Before and at different time (0, 1, 2 and 3 hours) after SDT treatment, macrophages were incubated with 10 μg/ml NAO for 15 minutes at 37°C in the dark and monitored by the fluorescence microscope. Then the cells were washed carefully with PBS twice. A total of 1×10⁶ cells were collected and measured by the fluorospectrophotometer at 480 nm excitation and 525 nm emission wavelengths.