Beyoclick edu 594 cell proliferation kit

Cell viability assays, colony formation assays, chemotaxis assays, invasion assays were performed as described previously [17 (link)]. According to the manufacturer’s instructions, cell proliferation assay was performed using the BeyoClick™ EdU-594 Cell Proliferation Kit (Beyotime Biotech. Inc.).

Curdione was purchased from Solarbio Science&Technology Co. Ltd. (Beijing, China). Modified Eagle’s medium (MEM), Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS), Non-Essential Amino Acids (NEAA), Pymvate Sodium (NaP), and penicillin–streptomycin (PS) were purchased from Gibco (Waltham, MA, USA). The IDO1 inhibitor epacadostat was purchased from Selleck (S7910, Texas, USA) and autophagy inhibitor 3- Methyladenine was purchased from Selleck (S2767, Texas, USA), CCk8 was purchased from Dojindo (CK04, Kumamoto, Japan), Beyo Click™ Edu-594 Cell Proliferation Kit was purchased from Beyotime (C0078S, Shanghai, China), and Annexin V-fluorescein isothiocyanate (FITC) cell apoptosis kit was purchased from Invitrogen (V13241, New York, California, USA).

EdU staining was used to measure the proliferation of embryos in the present study. Following the protocols of BeyoClick™ EdU-594 Cell Proliferation Kit (Beyotime Biotech. Inc.), 10 μM EdU was added to KSOM medium and embryos were incubated at 37 °C for 2 h. After fixation and permeabilization, embryos were incubated with the Click Reaction mixture for 2 min and stained with DAPI. BX51 microscope (Olympus) and Image J software (Rawak Software Inc.) were respectively used to photograph and quantitate EdU intensity in each group.

For MTT assay, the transfected cells (3×10³ / 200 µL /well) were seeded in 96-well plates. Cell viability was examined over the next 4 days. After incubation, 20 μL MTT (5 mg/mL in PBS; Sigma) was added to each well and incubated for 3-4 hours, and the formed formazan crystals were dissolved in 150 µL DMSO (Sigma). The absorbance was measured at 570 nm using a micro-plate auto-reader (Bio-Rad). All results are presented as the mean ± SD of triplicate independent experiments.
For colony formation assay, the transfected cells were seeded into 6-well plates at a density of 1000 cells ⁄ well. After approximately 15 days, the colonies had reached an appropriate size and were stained with crystal violet solution. The number of colonies was counted, and the size of the colonies was recorded.
The EdU assay was detected by the BeyoClick™ EdU-594 Cell Proliferation Kit (Beyotime Biotech. Inc.) according to the manufacturer's protocol. Briefly, after transfection for 48 h, cells were incubated with 25 μM EdU for 12 h before fixation, permeabilization, and EdU staining. The percentage of EdU-positive cells was examined by fluorescence microscopy. The data are presented as the means ± SDs of triplicate dishes in the same experiment.

A cell proliferation assay (BeyoClick™ EdU-594 Cell Proliferation Kit, Beyotime, China) was used for the analysis according to the manufacturer’ s instructions. Using a Zeiss laser scanning microscope, EdU cells were imaged.