Edu imaging kit

Cells were seeded in 96-well plates with 1.0 × 10⁴ cells per well. EdU labeled cells according to the steps in the EdU imaging kit instructions (Beyotime, Shanghai, China) after incubation for 24 h. After cell fixation and infiltration promotion, EdU detection, DNA restaining, and other series of operations, the culture plates were observed under an inverted fluorescence microscope. Three fields were randomly selected to take photos. ImagePro Plus 6.0 professional image analysis software (Media Cybernetics, Silver Spring, MD, USA) was used to count the fluorescence cells to calculate the DNA synthesis rate.

Cells planted into 24-well plates (5 × 10⁴ cells per well) were cultured at 80% confluence by using EdU Imaging Kit (C0075S, Beyotime Institute of Biotechnology, Jiangsu, China). In a nutshell, cells in each chamber were marked with 10 μM EdU solution for 2 h. Next, cells were immobilized utilizing 4% paraformaldehyde for 10 min, then the nucleus was stained 5 min by DAPI in the dark. Photographs were obtained using a fluorescence microscope (Olympus, Tokyo, Japan). The Image J software (v.1.8.0) was utilized to assess the number of EdU-positive cells.