Macrophages were separately seeded into 96-well plates at a density of 5 × 104 cells/ml and cultured in a 10% FBS RPMI-1640 medium for 24 hours. Following another 24-hour treatment with 11.1 mM glucose and 33.3 mM glucose (above NG and HG concentrations were based on the results from Method 2.2), Ka (National Institutes for Food and Drug Control, ≥ 98%) at 4, 8, and 12 μM [18 (link)], and HSYA (National Institutes for Food and Drug Control, ≥ 99%) at 100, 200, and 300 μM [19 ] were added into HG. The supernatants were removed, and each well washed with PBS before the addition of 10% FBS RPMI-1640 medium and 10 μl CCK-8 reagent (Boster Biological Technology, Wuhan, China). Cell viability was determined by measuring the absorbance at 450 nm using a microporous plate reader (Model 550; Bio-Rad Laboratories, Inc., Hercules, CA, USA) after an incubation period of 2 hours at 37°C. The average optical density was determined by examining six wells per group.
Cck 8 reagent
Manufactured by Boster Bio
Sourced in China, United States
The CCK-8 reagent is a colorimetric assay kit used for the determination of cell viability and cytotoxicity. It measures the number of living cells in proliferation, cytotoxicity, or activation assays.
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Lab products found in correlation
16 protocols using cck 8 reagent
1
Macrophage Viability under Glucose Levels
2
Cell Viability Assay using CCK-8
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Cells were plated in 96-well plates (1 × 103 cells/well) with 100 μl of the medium. The absorbance at 450 nm was measured to estimate the relative number of viable cells after culturing with 10 μl of CCK-8 reagent, which was purchased from Boster Biological Technology Co., Ltd. (Wuhan, China). The analysis was performed in three replicate wells for each sample and repeated for three times.
3
Cell Proliferation, Migration, and Invasion Assays
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4.0×103 cells were seeded into 96-well-plate in quintuplicate. After 72 hours of incubation, the absorbance was measured at 450 nm by adding a CCK8 reagent (Boster Biological Technology) to detect cell proliferation. Cell migration was evaluated by scratch assays as described previously [25 (link)]. After 16 hours of incubation, the migrated area was photographed and calculated by ImageJ software. For invasion assay, 8 mm pore transwell inserts (Corning Life Sciences) were coated with 75μl Matrigel matrix (Corning, 200 μg/mL) as upper chambers. 700μl DMEM medium with 20% FBS was added into the 24-well-plate. Meanwhile, 5.0×104 cells were suspended in a serum-free medium and were added to the inserts which were placed in a 24-well-plate. After 48h, the cells were fixed with 10% formalin and stained with 0.1% crystal violet solution. The numbers of invaded cells were counted at 200x magnification.
4
Cell Proliferation, Migration, and Invasion Assays
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Cell proliferation was measured by CCK8 reagent (Boster, catalog no.AR1160) according to the manufacturer's instructions. Briefly, 2,000 cells were seeded in 96-well plates and incubated with 100 μL culture medium. An aliquot of 10 μL CCK8 was added and incubated for 2 hours. The absorbance at 450 nm was measured to calculate the numbers of viable cells. Migration and invasion assays were performed with transwell and Matrigel chamber plates (24-well format; 8-μm pore size; Corning Costar, catalog no. 3422) as described previously (32 ). ImageJ software was used to measure the cell area in the bottom chamber to quantify the cells that migrated across the filter. Each measurement was performed in triplicate, and the experiments were repeated three times.
5
Cell Viability Assay in AR42J Cells
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AR42J cells were seeded in 96-well plates at 8 × 103 cells/well density. The SAP model was initiated when the cell density reached 80% confluence. After the treatment, 10 μL of CCK-8 reagent (Boster, Wuhan, China) was added to each well according to the CCK-8 kit guidelines. The absorbance of each well was measured at 450 nm using a microplate reader after the plates were incubated at 37 °C for 1 h. The cellular activity was then assessed based on the absorbance values obtained.
6
Evaluating Cell Injury via CCK-8 and LDH
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Cell counting kit-8 (CCK-8) assay and lactic dehydrogenase (LDH) content were used to assess cell injury. For CCK-8 assay, HepG2 cells were inoculated into 96-well plates at a cell density of 5 × 103 cells/well, and H/R was performed when the cell density reached 80% confluency. After reoxygenation, 10 μL of CCK-8 reagent (Boster, Wuhan, China) was added to each well according to the instructions of the CCK-8 kit, and the absorbance of each well was detected at 450 nm with microplate reader after incubation for 1 h at 37 °C. The cell activity was calculated according to the absorbance value. For LDH assay, after reoxygenation, cell culture medium was collected and operated according to the instructions of the detection kit.
7
Cell Proliferation Assay Protocol
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Cells were seeded in a 96-well plate at a density of 2×103. After culture for 24, 48, 72, and 96 h, 10 μL of CCK-8 reagent (Boster, Wuhan, China) was injected, and the cells were further cultured for another 24 h. Finally, the optical density in each well was detected at a wavelength of 450 nm.
8
CCK-8 Assay for Cell Viability
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Cells were plated into 96-well plates (2 × 103 cells/well). After various times points (6, 24, 48, or 72 h), CCK-8 reagent (Boster, Wuhan, China) was added and samples were incubated for 2 h. Absorbance was read at 450 nm using a microplate reader.
9
Cell Viability Assay for PANC-1 and MIA PaCa-2
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PANC-1 and MIA PaCa-2 cells were seeded into 96-well plates at a density of 5×103 cells/well (6 replicates per treatment). After 24, 48, and 72 h, 100 μl of DMEM containing 10 μl CCK-8 reagent (Boster Bio, Wuhan, China) was added to each well. The optical density (OD) of each well was measured at 450 nm.
10
CCK-8 Assay for Cal's Effects on A549 Viability
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The CCK-8 assay was used to assess the effects of Cal on A549 cell viability. Cells were seeded onto 96-well plates at a density of 1×104 cells/well and then treated with various concentrations of Cal (1, 5, 10 and 20 µM) for 24 h. DMSO (0.1%) was added to the 0 µM group. After treatment, the cells were incubated with 10 μl of CCK-8 reagent for 1 h (Bosterbio, United States) and then the absorbance at 450 nm was measured using a Microplate Reader.
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