The U251 or A172 glioma cells were seeded into 96-well plates at 5 × 103 cells/well and incubated for 12, 24, 36, 48, 72, and 96 h. An amount of 10 μL of CCK8 solution (0.5 mg/mL; Sigma, Saint Louis, MO, USA) was added into each well and incubated at 37 °C for 2 h. Finally, the absorbance was measured at 570 nm to assess cell viability. EdU Apollo 567 Cell Tracking Kit (Rib-oBio, Guangzhou, China) was used for cell proliferation. Hochest 33342 (Sigma, Saint Louis, MO, USA) was used for nuclei staining. The percentage of EdU-positive cells in total cells was calculated based on counting 500 random cells in three independent experiments.
Edu apollo 567 cell tracking kit
Manufactured by RiboBio
Sourced in China
The EdU Apollo 567 Cell Tracking Kit is a laboratory tool used to detect and visualize cell proliferation. It incorporates the nucleoside analog 5-ethynyl-2'-deoxyuridine (EdU) to label dividing cells. The kit provides the necessary reagents and protocols to perform this analysis.
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6 protocols using edu apollo 567 cell tracking kit
1
Cell Viability and Proliferation Assay
2
Cell Proliferation Measurement Using EdU
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Cell proliferation was measured using the EdU Apollo 567 Cell Tracking Kit (Ribo-bio; Guangzhou, China). Cells (2 × 104) under different treatments were seeded onto 24-well plates, exposed to 200 μM of 5-ethynyl-20-deoxyuridine for 2 h at 37 °C, fixed with 4% paraformaldehyde for 20 min, and treated with 0.5% Triton X-100 for 10 min. Cells were rinsed with PBS three times, and incubated with 100 μL of Apollo reagent for 30 min. Nuclie were stained with Hochest33342. The percentages of EdU-positive cells were determined from 500 cells and three independent experiments were performed.
3
Cell Viability, Proliferation, and Colony Formation Assays
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For the CCK-8 assay, 5 × 103 cells were seeded into 96-well plates. And cell viability was examined daily for 3 days using CCK-8 (Dojindo Laboratories, Japan) according to the instruction provided.
For colony formation assay, 800 cells were seeded into 12-well plates and cultured for 4–5 days. Then, the medium was discarded and cells were fixed with 4% paraformaldehyde for 20 min, washed with PBS, and stained with 0.1% crystalline violet solution for analysis.
For EdU assays, 5 × 103 cells were seeded into 96-well plates and EdU Apollo 567 Cell Tracking Kit (RiboBio, China) was used according to the instruction provided. And cell nucleus was stained with DAPI.
For colony formation assay, 800 cells were seeded into 12-well plates and cultured for 4–5 days. Then, the medium was discarded and cells were fixed with 4% paraformaldehyde for 20 min, washed with PBS, and stained with 0.1% crystalline violet solution for analysis.
For EdU assays, 5 × 103 cells were seeded into 96-well plates and EdU Apollo 567 Cell Tracking Kit (RiboBio, China) was used according to the instruction provided. And cell nucleus was stained with DAPI.
4
EdU Proliferation Assay Protocol
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EdU assays were performed using the EdU Apollo 567 Cell Tracking Kit (RiboBio, China). Treated and control cells (5 × 10 [3 (link)]/well) were seeded onto 96-well plates and incubated with EdU (200 μM) for 2 h at 37 °C. The cells were fixed with 4% paraformaldehyde for 20 min, treated with 0.5% Triton X-100 for 10 min, rinsed with PBS three times, and incubated with 100 μl Apollo reagent for 30 min. Nuclei were labelled with Hoechst 33342. The percentage of EdU-positive cells was calculated based on counts in three independent experiments.
5
Measuring GBM Cell Proliferation
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GBM cells (2 × 104) were seeded into 24-well plates, allowed to attach, and treated. Following treatment, cell proliferation was measured using the EdU Apollo 567 Cell Tracking Kit (Ribo-bio, Guangzhou, China). Briefly, cells were incubated with EdU reagent (200 μM) for 2 h at room temperature, fixed with 4% paraformaldehyde for 20 min and permeabilized with 0.5% Triton X-100 for 10 min. Cells were rinsed with phosphate buffered saline (PBS) three times, incubated with 100 μl of Apollo reagent for 30 min and stained with 4′,6-diamidino-2-phenylindole (DAPI). Three fields of view were randomly selected for counting EdU-positive cells, and three independent experiments were performed.
6
EdU Assay for Cell Proliferation
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EdU assay was performed as described.13 (link) Subsequently, 5×103 cells were seeded into 96-well plates. EdU Apollo 567 Cell Tracking Kit (RiboBio, Guangzhou, China) was utilized for EdU determination. The percentage of EdU-positive cells was counted in three independent experiments.
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