Multicycle cell cycle software

Cell cycle analysis in HMEC was performed as described previously [11 (link)]. Briefly, 10⁵ cells were fixed in 70% (v/v) ice-cold ethanol at 4 °C for 24 h. Thereafter, the fixed cells were stained with CyStain DNA 2 step kit (Partec GmbH, Münster, Germany) and filtered through a 50 μm filter. The samples were then analyzed in a Galaxy flow cytometer (Partec GmbH) using the MultiCycle cell cycle software (Phoenix Flow Systems Inc., San Diego, CA, USA).

The cell cycle analysis was performed as described previously [16 (link)]. Briefly, 9.3 × 10³ cells/cm² were seeded in culture plates (diameter 10 cm; Greiner Bio-one GmbH, Frickenhausen, Germany) overnight to allow attachment of the cells and adjustment to the culture conditions. Following incubation with 1 μM cisplatin, or 1 μM carboplatin, or 2nM epothilone B for 48 h, the cells were fixed in 70% (v/v) ice-cold ethanol at 4°C for 24 h. Thereafter, about 5 × 10⁵ fixed cells were stained with CyStain DNA 2 step kit (Partec GmbH, Münster, Germany) and filtered through a 50 μm filter. The samples were then analyzed in a Galaxy flow cytometer (Partec) using the MultiCycle cell cycle software (Phoenix Flow Systems Inc., San Diego, CA).