EdU incorporation assay was performed using a BeyoClick™ EdU Cell Proliferation Kit with Alexa Fluor 488 in vitro Imaging Kits (Beyotime C0071s, Shanghai, China). Briefly, HCT116 cells were transiently transfected with pcDNA3.1, F-YY1, and F-K183R. The cells were re-plated on a 12-well plate 48 h later. After 24 h, 10 μM of EdU (5-ethynyl-2′-deoxyuridine) was added and incubated in a 37 °C incubator for 2 h. The cells were then fixed with 4% paraformaldehyde for 15 min and rinsed with PBS containing 0.5% Triton-X-100. The Hoechst33342 (GC10939, GLPBIO) was used to stain the nuclei. The cell proliferation rate was calculated according to the manufacturer’s recommendations (BeyoClick EdU-488 Kit, C0071s, Shanghai, China). Using a fluorescent microscope, pictures of three randomly selected regions of each group were captured.
Beyoclick edu cell proliferation kit with alexa fluor 488 in vitro imaging kits
Manufactured by Beyotime
The BeyoClick™ EdU Cell Proliferation Kit with Alexa Fluor 488 is an in vitro imaging kit used to detect and quantify cell proliferation. The kit utilizes the EdU (5-ethynyl-2'-deoxyuridine) labeling method and Alexa Fluor 488 dye to visualize and analyze proliferating cells.
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2 protocols using beyoclick edu cell proliferation kit with alexa fluor 488 in vitro imaging kits
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EdU Incorporation Assay for Cell Proliferation
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EdU Incorporation Assay for Proliferation Analysis
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EdU incorporation assays were performed using BeyoClick™ EdU Cell Proliferation Kit with Alexa Fluor 488 in vitro Imaging Kits (Beyotime). Briefly, VSMCs in the logarithmic growth were plated at 5 × 104 cells per well into 12-well plates. After 24 h of the indicated treatment, each well was incubated with 1 ml of 10 μM EdU medium for 2 h. The cells were fixed with 4% paraformaldehyde for 30 min. After washing with PBS in 3 times, the cells were washed with PBS containing 0.5% TritonX-100 for 10 min. Then, click additive solution and click reaction solution were prepared according to the manufacturer's instructions, then added into each well and incubated in the dark for 30 min at room temperature. The staining solution was discarded, and the cells were washed with PBS for 10 min. For the nuclear staining, 1 × Hoechest 33,342 was added for 30 min incubation at room temperature. After washing with PBS, the positive cells were observed by fluorescence microscopy.
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