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Cck 8 agent

Manufactured by Dojindo Laboratories
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Sourced in Japan, United States

The CCK-8 agent is a cell counting kit used for the quantitative determination of cell viability and proliferation. The kit utilizes a water-soluble tetrazolium salt that is reduced by the dehydrogenase activity of viable cells, producing a colored formazan dye that can be measured with a spectrophotometer.

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Lab products found in correlation

Microplate reader, by Bio-Rad (2 mentions) Microplate reader, by Agilent Technologies (1 mentions) Perifosine, by Apexbio (1 mentions) Lapatinib, by Selleck Chemicals (1 mentions) 12 well plate, by Merck Group (1 mentions) Epoch microplate spectrophotometer, by Agilent Technologies (1 mentions) μ slide 8 well chamber slide, by Ibidi (1 mentions) 96 well plate, by Corning (1 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)

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CCK-8

13 protocols using cck 8 agent

1

Evaluating BMSC Proliferation on Composite Hydrogels

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BMSCs were cultured in α–minimum essential medium (Hyclone, Thermo Fisher Scientific, USA) containing 10% fetal bovine serum (Gibco) and 1% penicillin-streptomycin. BMSCs were seeded on composite hydrogels and cultured for 48 hours. Then, they were fixed with 4% paraformaldehyde for 40 min at room temperature. Afterward, the cells were washed with phosphate-buffered saline (PBS) and sealed and permeabilized with 3% bovine serum albumin solution containing 0.3% Triton X-100 for 12 hours. Then, the sample was stained with rhodamine phalloidin and 4′,6-diamidino-2-phenylindole (DAPI). Images were acquired with an inverted fluorescence microscope. For SEM scanning, BMSCs were seeded on composite hydrogels (5 × 104 cells per well) and incubated for 48 hours. Then, they were fixed with 4% paraformaldehyde for 40 min, dehydrated with gradient ethanol (10, 30, 50, 70, 85, 90, and 100%), and observed by SEM after critical point drying. Cell proliferation was evaluated by the CCK-8 assay. Briefly, BMSCs were seeded on composite hydrogels (1 × 103 cells per well). At the time points of 1, 3, and 5 days of incubation, cells were incubated with a diluted CCK-8 agent (Dojindo, Japan) for 2 hours (CCK-8 agent:culture medium = 1:10). Then, the absorbance was measured at 450 nm of wavelength.
Li J., Ma J., Sun H., Yu M., Wang H., Meng Q., Li Z., Liu D., Bai J., Liu G., Xing X., Han F, & Li B. (2023). Transformation of arginine into zero-dimensional nanomaterial endows the material with antibacterial and osteoinductive activity. Science Advances, 9(21), eadf8645.
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2

Cell Viability Assay Using CCK-8 Reagent

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The cells (1 × 105) were placed on a 96-well culture dish. After treatment in the corresponding groups, the cells were incubated with CCK-8 agent (DOJINDO, Laboratories, Kumamoto, Japan) for 1 h at 37°C. Next, cell viability was determined through measuring the absorbance at 450 nm [28 ]. For this procedure, the microplate reader (Bio-Tek, Winooski, USA) was used.
Li T., Quan H., Zhang H., Lin L., Ou Q, & Chen K. (2020). Silencing cyclophilin A improves insulin secretion, reduces cell apoptosis, and alleviates inflammation as well as oxidant stress in high glucose-induced pancreatic β-cells via MAPK/NF-kb signaling pathway. Bioengineered, 11(1), 1047-1057.
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3

Evaluating Cell Viability with CCK8 Assay

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Cell viability was determined using the CCK8 assay. First, the three groups of cells were seeded on 96-well plates at 40,000 cells/well. Following attachment, cells were incubated with different concentrations of lapatinib (S2111, Selleckchem, Houston, TX, USA) and/or perifosine (1.5–100 μM APExBIO, Houston, TX, USA) and cultured for 48 h. CCK8 agent (Dojindo Molecular Technologies, Inc., Japan) was added, and after 2 h, the absorbance at 450 nm was measured using a microplate reader (Bio-Rad Laboratories, Hercules, CA, USA).
Yang Z.Y., Yang L., Xu C.W., Wang X.J, & Lei L. (2020). An insertion mutation of ERBB2 enhances breast cancer cell growth and confers resistance to lapatinib through AKT signaling pathway. Biology Open, 9(1), bio047662.
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4

Oleandrin Cytotoxicity in hFOB1.19 Cells

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hFOB1.19 cells were seeded in a 96-well dish at a final density of 6 × 103 cells/well and incubated overnight. Then, cells were treated with various concentration of oleandrin (0, 25, 50, 75 and 100 nM) for 24 h. Then, CCK-8 agent (Dojindo Laboratories, Kumamoto, Japan) was added to each well and incubated for another 3 h. The absorption at 450 nm was determined using an automatic ELIASA microplate reader as reported above. Five replicate wells were used for each treatment.
Ma Y., Zhu B., Yong L., Song C., Liu X., Yu H., Wang P., Liu Z, & Liu X. (2016). Regulation of Intrinsic and Extrinsic Apoptotic Pathways in Osteosarcoma Cells Following Oleandrin Treatment. International Journal of Molecular Sciences, 17(11), 1950.
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5

Glioma Cell Proliferation Assays

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The proliferation of glioma cells was tested using CCK‐8 assay and colony formation assay according to manufacturer's instructions. CCK‐8 agent (Dojindo, Rockville) was administrated into the 96‐well plates at a density of 2 × 103 per well. The absorbance was recorded at 450 nm using microplate reader. Glioma cells were cultured in RPMI1640 containing 10% FBS and seeded into the 6‐well plates at 2 × 103 per well for 2 weeks. Then, the cells were washed and fixed with methanol and stained with 0.5% crystal violet. The colonies greater than 150 μm were counted under a microscope.
Li C., Liu H., Yang J., Yang J., Yang L., Wang Y., Yan Z., Sun Y., Sun X, & Jiao B. (2019). Long noncoding RNA LINC00511 induced by SP1 accelerates the glioma progression through targeting miR‐124‐3p/CCND2 axis. Journal of Cellular and Molecular Medicine, 23(6), 4386-4394.
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6

Astrocyte Viability Assessment under Oxygen-Glucose Deprivation

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Upon reaching 80% confluency, human astrocytes in 12-well plates (Merck Sigma) were subjected to OGD for 8 h. The media were subsequently replaced with 0% FBS-containing DMEM (HyClone). Various concentrations of KRGE were added, and the cells were incubated for 23 h. Thereafter, 30 μL of a cell-counting kit-8 (CCK-8) agent (Dojindo, Fukuoka, Japan) was added to each well, and cells were incubated at 37 °C for 1 h. We used a plate reader (Epoch Microplate Spectrophotometer; BioTek, Santa Clara, CA, USA) to determine the absorbance at a wavelength of 450 nm. The background wave was determined at 630 nm and subtracted from the value measured at 450 nm.
Kim M., Moon S., Jeon H.S., Kim S., Koh S.H., Chang M.S., Kim Y.M, & Choi Y.K. (2022). Dual Effects of Korean Red Ginseng on Astrocytes and Neural Stem Cells in Traumatic Brain Injury: The HO-1–Tom20 Axis as a Putative Target for Mitochondrial Function. Cells, 11(5), 892.
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7

CCK-8 Cell Proliferation Assay

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Cell proliferation was evaluated by conducting CCK-8 assays. 1000 cells were seeded per well in 96-well plates in triplicate. After incubation with 10uL of CCK8 agent (Dojindo, Kumamoto, Japan) for 3 hours in the dark in an incubator at 37°C, the absorbance value was measured at 450 nm every 12 hours.
Wang J., Xie G.F., He Y., Deng L., Long Y.K., Yang X.H., Ma J.J., Gong R., Cen W.J., Ye Z.L., Zeng Y.X., Wang H.Y, & Shao J.Y. (2019). Interfering Expression of Chimeric Transcript SEPT7P2-PSPH Promotes Cell Proliferation in Patients with Nasopharyngeal Carcinoma. Journal of Oncology, 2019, 1654724.
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8

CCK-8 Viability Assay for HFL-1 Cells

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Cell viability of HFL-1 cells was determined using the CCK-8 assay. The CCK-8 agent (Dojindo, Japan) was administrated to HFL-1 cells following different treatments. After further incubation at 37°C with 5% CO2 for 2 h, the absorbance at 450 nm was detected by a microplate reader.
Wu W., Zhang G., Qiu L., Liu X., Zhou S, & Wu J. (2022). Contribution of Adiponectin/Carnitine Palmityl Transferase 1A-Mediated Fatty Acid Metabolism during the Development of Idiopathic Pulmonary Fibrosis. Oxidative Medicine and Cellular Longevity, 2022, 5265616.
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9

Cytotoxicity Evaluation of DS001 and MMAE

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Cell viability was assayed as previously described.17 (link) In brief, 3000 to 5000 cells/well were seeded into 96-well plates and incubated overnight. The next day, cells were treated with different concentrations of DS001 or MMAE alone (3-fold serial dilutions starting at 300 nM) for 96 hrs. CCK8 agent (Dojindo, Kumamoto, Japan) was added according to the manufacturer’s instructions and incubated for 1–2 h. Growth inhibition was measured as percentage of growth relative to untreated cells. Dose-response curves were generated from the means of triplicate determinations, IC50 (half maximal inhibitory concentration) values and Imax (%) values (maximal inhibition rate) were calculated by nonlinear regression (four-parameter) using Prism GraphPad Software. Imax (%) values were calculated using the formula: [1-minimum viability (%)] ×100.
Liu X., Zhou T., Wang Y., Pei M., Wang G., Chu W., Wang Q., Du S., Wang H, & Wang C. (2022). TROP2 as Patient-Tailoring but Not Prognostic Biomarker for Breast Cancer. OncoTargets and therapy, 15, 509-520.
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10

In vitro Photodynamic Therapy Evaluation

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In vitro PDT was evaluated by CCK8 assay. PC3pip and PC3flu cells were seeded in 96-well plates at 1 × 104 cells/well. After 1 day of incubation, AuNPs-Pc158 conjugates were added at Pc158 concentrations of 0.0625, 0.125, 0.25, 0.5, and 1 μmol/L. After coincubation for 6 or 24 h, the medium was removed and cells were washed with PBS, then another 200 μL of medium was added to each well prior to light irradiation. The 96-well plate was irradiated (Appolo Horizon projector, Acco Brands) with radiant exposure at 1 J/cm2, after which the cells were incubated overnight. After incubation, CCK8 agent (DojinDo Laboratories) was added to each well (10 μL/well) and incubated for 3 h at 37 °C, and absorbance at 450 nm was measured for each well.
The intracellular ROS generation after light irradiation was evaluated with a DCFH-DA assay. PC3pip and PC3flu cells were cultured in μ-Slide 8-Well Chamber Slide (ibidi) and incubated with AuNPs-Pc158 conjugates for 6 and 24 h at a Pc158 dose of 1 μmol/L. Culture medium was removed, and cells were washed with PBS and incubated with 20 μM DCFH-DA HEPES buffer for 30 min. After incubation, the cells were washed again with PBS and irradiated with light at 1 J/cm2. The cells were counterstained with DAPI and fixed for fluorescence imaging.
Luo D., Wang X., Walker E., Wang J., Springer S., Lou J., Ramamurthy G., Burda C, & Basilion J.P. (2020). Nanoparticles Yield Increased Drug Uptake and Therapeutic Efficacy upon Sequential Near-Infrared Irradiation. ACS nano, 14(11), 15193-15203.
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