Computational protocol: Tetraarsenictetrasulfide and Arsenic Trioxide Exert Synergistic Effects on Induction of Apoptosis and Differentiation in Acute Promyelocytic Leukemia Cells

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Protocol publication

[…] The proportions of apoptotic cells were measured with the Annexin V-FITC and propidium iodide (PI) apoptosis detection kit (KeyGEN Biotech, China) using flow cytometry [,,]. NB4 and primary APL cells were treated with 2.0 μM As4S4, 2.0 μM As3+, or 1.0 μM As4S4 and 1.0 μM As3+ for 48 h. After treatment, the cells were washed twice with Ca2+-free phosphate buffer (PBS), stained with Annexin V-FITC and PI in the dark at room temperature for 15 min, and detected using a BD LSRL Fortessa flow cytometer. The percentages of apoptotic cells were analyzed using the BD FACSDiva software. [...] Cellular ROS levels were assayed using DCFH-DA. DCFH-DA can be hydrolyzed to DCFH by esterases and oxidized to 2’,7’-dichlorofluorescein (DCF) by cellular ROS []. Thus, the DCF fluorescence intensity is positively correlated with cellular ROS levels. After treatment with As4S4 (0.5, 1.0 and 2.0 μM), As3+ (2.0 μM), or a combination of 1.0 μM As4S4 and 1.0 μM As3+ for 24 h and 36 h, NB4 and primary APL cells were washed twice with PBS and then incubated in RPMI-1640 medium containing 10.0 μM DCFH-DA at 37°C for 30 min. Excess probe was washed out with PBS, and the percentages of DCF-positive cells were detected by flow cytometry and analyzed using FlowJo.7.6. […]

Pipeline specifications

Software tools BD FACSDiva, FlowJo
Application Flow cytometry
Organisms Homo sapiens
Diseases Leukemia, Promyelocytic, Acute, Drug-Related Side Effects and Adverse Reactions
Chemicals Oxygen, Tretinoin