Dna prep epics workstation

The distribution of cell-cycle phases (G₀/G₁, S, and G₂/M) was determined using flow cytometry by measuring the DNA content of nuclei labeled with propidium iodide as described previously [15 (link)]. Briefly, human colorectal (SW1116 and SW837) and breast (HTB 26 and HTB132) cancer cell lines were plated (5 × 10⁵ cells/ml) in 24-well plates and incubated at 37°C in a non-CO₂ incubator for 18 h. The cells were then treated with Nar (3 mM) for 24 h. Untreated and treated human cancer cells were collected by trypsinization, washed with cold phosphate-buffered saline (PBS) and counted. Cells were processed using a DNA-prep kit (Beckman & Coulter) and a DNA-Prep EPICS workstation (Beckman & Coulter). During this process, the cells were treated with a cell-membrane permeabilizing agent (non-ionic detergent) followed by propidium iodide (PI) and RNase. The samples were incubated at room temperature for 15 min before analysis by flow cytometry (FC500, Beckman & Coulter). The percentages of cells in different cell cycle phases were calculated using the Phoenix statistical software package, advanced DNA cell-cycle software (Phoenix Flow System, San Diego, CA).

Flow cytometry was used to monitor the disruption in cell cycle phases (Go/G1, S and G2/M) by measuring the DNA content of the nuclei labeled with propidium iodide (PI), as previously described [20 (link)]. Briefly, SW1116 and SW837 cells were plated (2.5 × 10⁵ cells/ml) into 24-well plates and incubated at 37 °C in a non-CO₂ incubator. Cells were treated with MF (1.5 mM) for 24 h starting 18 h after seeding the cells in culture. Untreated and MF-treated cells were collected by trypsinization, washed with cold PBS and counted. Cells were processed using a DNA-prep kit (Beckman & Coulter, FL, USA) and a DNA-Prep EPICS workstation (Beckman & Coulter). During this process, cells were treated with a cell-membrane permeabilizing agent followed by a treatment with PI and RNAase followed by incubation at room temperature for 15 min before analysis by flow cytometry (FC500, Beckman & Coulter). The percentage of cells in different cell cycle phases was calculated using the Phoenix statistical software package (Phoenix Flow System, San Diego, CA).