5-Ethynyl-2′-deoxyuridine (EdU) is an analogue of thymidine, which is incorporated into proliferating cells during DNA synthesis. Therefore, a higher positive rate of EdU incorporation indicate a stronger cell proliferation ability. The EdU assay was performed by using an EdU detection kit (Beyotime Biotechnology, Shanghai, China) according to the manufacturer’s instruction.
Edu detection kit
Manufactured by Beyotime
Sourced in China
The EdU detection kit is a laboratory tool used to detect and analyze cell proliferation. It utilizes the incorporation of the synthetic nucleoside EdU (5-ethynyl-2'-deoxyuridine) into the DNA of actively dividing cells, allowing for the visualization and quantification of cell proliferation through fluorescent labeling.
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Lab products found in correlation
Hoechst 33342, by Merck Group (1 mentions)
6 well insert device, by Corning (1 mentions)
Pi staining kit, by Merck Group (1 mentions)
Biocoat matrigel, by BD (1 mentions)
Cck 8 assay kit, by Beyotime (1 mentions)
Zoetm fluorescent cell imager, by Bio-Rad (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Cck 8 solution, by Dojindo Laboratories (1 mentions)
Anti fluorescence quenching agent, by Beyotime (1 mentions)
12 protocols using edu detection kit
1
Evaluating Cell Proliferation Using EdU
2
Cell Proliferation Assay with EdU
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EdU assay was conducted to examine the cell proliferation using the EdU detection kit (Beyotime) following the ‘manufacturer’s protocols. Briefly, cells were incubated in DMEM containing 20 μM EdU for 2 h. After fixing and permeabilizing as described above, cells were stained with Apollo and Hochest reagents in the dark. Images were taken by fluorescence microscopy and analyzed for EdU-positive cells using ImageJ.
3
EdU Proliferation Assay with Wnt3a
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Cells were seeded into the wells of a 24‐well plate and treated with either BK alone or a combination of BK plus Wnt3a as indicated. An Edu Detection Kit was purchased from Beyotime Biotechnology (Cat. No. C0078S). After 48 h, 10 μM 5‐ethynyl‐2′‐deoxyuridine (EdU) was added to each well and the cells were incubated for another 2 h; after which, they were fixed with 4% formaldehyde for 10 min at room temperature. After permeabilization, the cells were incubated with Click Additive Solution included in the EdU Detection Kit. The cell nucleus was stained with 4′,6‐diamidino‐2‐phenylindole (DAPI).
4
Cell Proliferation Analysis using EdU Assay
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Cell proliferation was examined by EdU assay using EdU detection kit (Beyotime) according to the manufacturer's protocols. Briefly, after incubation for 2 h in DMEM containing 20 μM EdU, cells were fixed using 4% paraformaldehyde, permeabilized using 1% Triton X‐100 and stained with Apollo and Hochest in dark. Then, cells were imaged by fluorescence microscopy and counted for EdU‐positive cells by Image J. All experiments are performed in triplicates.
5
Colon Cancer Cell Proliferation Assay
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The proliferation ability of colon cancer cells was detected by EDU assay. Colon cancer cells were seeded into 24-well plates with 48 h culture. Then, the EDU detection kit (Beyotime, Shanghai, China) was used to treat cells for 2 h at 37°C. Subsequently, cells were fixed with formaldehyde for 30 min and then washed with PBS. Apollo reagent was added and incubated in the dark for 30 min, and then the cells were stained with Hoechst 33342 (Sigma-Aldrich) for 30 min in the dark. Cells labeled and unlabeled by EDU were counted under a microscope, and pictures were taken.
6
Cell Proliferation, Cycle, and Invasion Assays
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Cell proliferation was analyzed using a commercial CCK-8 assay kit (#C0038, Beyotime) and EdU detection kit (#C0075S, Beyotime). Fluorescence-activated cell sorting (FACS) was used to assess the cell cycle with a PI staining kit (#R40432, Sigma). Cell invasion was assessed by the transwell assay with a 6-well insert device (8 μm pore size; Corning Life Sciences, Bedford, MA) and Biocoat Matrigel (BD Biosciences) according to the manufacturer’s instructions.
7
EdU Proliferation Assay for GBC-SD Cells
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An EdU detection kit (Beyotime Institute of Biotechnology, Haimen, China) was used for assessing cell proliferation. Briefly, GBC-SD cells were incubated with EdU for 2 h, and then fixed in 4% paraformaldehyde for 15 min. Next, cells were incubated with Click solution and then stained with Hoechst 33342 solution (diluting with PBS at a volume ratio of 1:1000) in darkness for 30 min at room temperature. Finally, EdU-positive cells were captured using an Fi3 Nikon fluorescence microscope (objective: 20x). This assay was repeated in triplicate.
8
EdU Proliferation Assay Protocol
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The EdU detection kit (Beyotime, China) was used to evaluate cell proliferation. According to the manufacturer’s protocol, cells were treated with 20 μM EdU for 2 h at 37 °C and fixed with 4% paraformaldehyde at room temperature for 15 min. Following washing with wash buffer three times, cells were treated with 0.5% Triton X-100 for 15 min and stained with Click reaction cocktail for 30 min at room temperature. Following washing with wash buffer, 1× Hoechst 33342 dye was used to incubate cells at room temperature for 10 min. Images were captured using ZOETM Fluorescent Cell Imager (Bio-Rad). Following the merging of the images, the average OD of cells was calculated using ImageJ software (version 1.8.0; National Institutes of Health, USA). The assay was repeated in triplicate.
9
HSCs Transduction and Proliferation
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HSCs were transduced with Ad‐shHOTTIP for 48 hours. Then the cells were incubated with 5‐Ethyny‐2′‐deoxyuridine (EdU) for 12 hours. Cell proliferation was examined using an EdU detection kit (Beyotime Biotechnology, Jiangsu, China) in keeping with the manufacturer's instructions.
10
Proliferation of RA-FLSs Evaluation
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The proliferation of RA-FLSs was evaluated by an EdU detection kit (Beyotime, Shanghai, China). Shortly, EdU medium was utilized to incubate RA-FLSs for 3 h, Next, RA-FLSs were then fixed, permeabilized and stained with DAPI (Beyotime). Finally, a fluorescence microscope (Leica, Wetzlar, Germany) was utilized to estimate EdU-positive cells.
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