Portal de Periódicos da CAPES

Dual AO/EB Staining to Detect Apoptosis in Osteosarcoma Cells Compared with Flow Cytometry

2015; International Scientific Information Inc.; Volume: 21; Linguagem: Inglês

10.12659/msmbr.893327

2325-4416

Kuan Liu, Pengcheng Liu, Liu R, Xing Wu,

Photodynamic Therapy Research Studies

Background:The aim of this study was to evaluate the ability of dual acridine orange/ethidium bromide (AO/EB) staining to detect tumor cell apoptosis.According to apoptosis-associated changes of cell membranes during the process of apoptosis, a clear distinction is made between normal cells, early and late apoptotic cells, and necrotic cells. Material/Method:We cultured human osteosarcoma cells with 30, 60, and 120 µg/ml kappa-selenocarrageenan.To assess the rates of cell proliferation and apoptosis, cells were fluorescently stained with acridine orange/ethidium bromide (AO/EB) or stained with propidium iodide (PI) and analyzed by flow cytometry.All experiments were repeated at least 3 times. Result:Normal tumor cells, early and late apoptotic cells, and necrotic cells were examined using fluorescent microscopy.Early-stage apoptotic cells were marked by crescent-shaped or granular yellow-green acridine orange nuclear staining.Late-stage apoptotic cells were marked with concentrated and asymmetrically localized orange nuclear ethidium bromide staining.Necrotic cells increased in volume and showed uneven orange-red fluorescence at their periphery.Cells appeared to be in the process of disintegrating.The percentage of apoptotic osteosarcoma cells detected by dual acridine orange/ethidium bromide (AO/EB) staining was not significantly different from that detected using flow cytometry (P>0.05). Conclusions:Our results suggest that dual acridine orange/ethidium bromide staining is an economic and convenient method to detect apoptosis in tumor cells and to test tumor chemosensitivity compared with flow cytometry.