Muse cell analyzer software

The cell-cycle analysis was measured using a Muse Cell Cycle Assay Kit (MCH100106, Merck, Millipore) according to the manufacturer’s instructions. In brief, cells were dissociated and washed once with 1X PBS and fixed with 70% ethanol overnight. Cells were then centrifuged and washed once with 1X PBS. Cells were then re-suspended in 200 μl of Muse Cell Cycle Assay Kit and analyzed using Muse Cell Analyzer software (Merck, Millipore).

Cell cycle analysis was performed as described previously (24 (link)), using the Muse Cell Cycle Kit (Merck-Millipore, Burlington, MA, USA) according to the manufacturer's instructions. Briefly, 5 × 10⁴ cells were seeded in 60-mm dishes and after 48 h incubated with different concentrations of calcitriol in medium supplemented with 2.5% FBS for 24 h. Cells were then detached with phosphate buffered saline (PBS) 1X/ethylenediaminetetraacetic acid (EDTA) (5 mM), centrifuged (1,500 rpm, 5 min) and fixed with pre-cooled ethanol. The cells were then treated with Muse Cell Cycle Reagent for 30 min and analyzed with Muse Cell Analyzer Software (Merck-Millipore, Burlington, MA, USA).

This assay was carried out using a Muse cell cycle kit (Merck Millipore). Briefly, MDA-MB-231 cells (5 × 10⁵ cells/well) were incubated in 6-well plate for 24 h at 37°C. The cells were then treated with 30 and 60 μg/ml of F1 fraction. The cells were fixed in ice-cold 70% ethanol for 3 h and then washed with PBS and mixed with 200 μl of Muse cell cycle reagent. After 30 min of incubation at 25°C, the percentage of cells in Go/G1, S and G2/M was analyzed using Muse cell analyzer software (Merck Millipore).