Agilent fluorescence spectrophotometer

Rhodamine 123 (Rh 123) staining was used to evaluate the mitochondrial membrane potential. As mitochondria transition from a polarized to a depolarized state during apoptosis, dye leakage occurs, leading to a decrease in the fluorescence intensity of Rh 123. MDA-MB-231 cells were seeded onto 24 well plates and treated with DOX, ADA, or combination of both for 24 hrs. Cells were stained for 15 minutes at 37 ℃ in the dark with 10 µM Rh 123 and washed thrice with 1x PBS and imaged using FLoid™ Cell Imaging Station. Also, following staining, cells were collected in PBS and fluorescence intensity was determined using Agilent Fluorescence Spectrophotometer (31 (link)). In some tests, cells were pretreated for 2 hrs with 5 mM N-acetyl cysteine (NAC) before exposure to the drugs.