With CCK-8 assay kits (Dojindo, Tokyo, Japan), a determination of cell viability was made. We seeded transfected BEAS-2B cells (3 × 103/well) into plates each having 96 wells and altogether having 100 μL of the medium. We added a total of 10 μL CCK-8 solutions (Dojindo, Tokyo, Japan) 24 h after diverse treatments, and then we incubated the cells in an atmosphere whose temperature remained 37 °C and had 5% of CO2 for 2 h. With the help of an enzyme-labelled instrument (Thermo, Waltham, MA, USA), in every well we measured the absorbance at 450 nm.
Enzyme labelled instrument
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Enzyme-labelled instrument is a piece of laboratory equipment designed for the detection and quantification of specific biomolecules. The core function of this instrument is to utilize enzyme-based labeling techniques to generate a measurable signal that corresponds to the presence and concentration of target analytes in a sample.
Automatically generated - may contain errors
Lab products found in correlation
Cck 8 solution, by Beyotime (2 mentions)
Cck 8 solution, by Dojindo Laboratories (1 mentions)
Cck 8 assay kit, by Dojindo Laboratories (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Merck Group (1 mentions)
Erastin, by Merck Group (1 mentions)
Deferoxamine dfo, by Merck Group (1 mentions)
Erastin, by Dojindo Laboratories (1 mentions)
Ferrostatin 1 ferr 1, by Merck Group (1 mentions)
9 protocols using enzyme labelled instrument
1
Cell Viability Assay with CCK-8
2
Cell Viability Assay Protocol
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NP cells were injected into 96-well plates (8 × 103cells/well) incubated for 24 h. Subsequently, 10 µL cell counting kit-8 (CCK-8) solution (Beyotime, Jiangsu, China) was added to each well and the cells were incubated for another 2 h. Then, cells were detected by enzyme-labelled instrument (Thermo Fisher Scientific, Waltham, MA, USA) with a wavelength at 450 nm. The cell survival rate was calculated as follows: cell survival rate = [(experimental OD - blank OD)/(control OD - blank OD)] × 100%.
3
Evaluating Cardiomyocyte Viability and LDH Levels
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Cell viability was measured by CCK8 assay, cardiomyocyte was incubated on a 96‐pore plate and pretreated (2000 cells/pore), and 10 µL CCK8 solution (Beyotime, Shanghai, China, C0037) was added to each pore. After incubation in cell incubator for 1 hour, the cardiomyocyte was detected by enzyme‐labelled instrument (Thermo, America) with a wavelength at 450 nm. The supernatant was collected after various treatments to measure the LDH levels using a commercial available kit (Beyotime Institute of Biotechnology, Shanghai, China; C0016) according to the manufacturer's instructions.
4
MTT Cell Proliferation Assay
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Cell proliferation assay was performed using MTT assay. The VSMCs were harvested into flat-bottomed 96-well plates at a density of 5 × 104/ml. After that, the culture medium was removed and substituted with serum-free DMEM. The cells were then rinsed twice with PBS, followed by incubation along with 10 μl of MTT (Sigma, USA) at 37°C for 4 h in the dark. Consequently, to dissolve MTT crystals, 150 ml of DMSO (Sigma, USA) was added. The absorbance was calculated using an enzyme-labelled instrument (Thermo Fisher Scientific) at 490 nm.
5
Quantification of Inflammatory Mediators
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The content of IL-1β, TNF-α and IL-8 in BALF was assessed using Rat IL-1β ELISA Kit (TW-reagent, Shanghai, China), Rat TNF-α ELISA Kit (TW-reagent) and Rat IL-8 ELISA Kit (TW-reagent). The assay was performed according to the manufacturer’s instructions. The optical density values of samples were detected at 450 nm wavelength using enzyme-labelled instrument (Thermo Fisher Scientific).
6
Cell Proliferation Assay Using CCK-8
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Cell proliferation was determined using the CCK-8 assay. PK15 cells were seeded at 6 × 104 cells per well in 24-well plates overnight and then untransfected or transfected with 200 ng of pcDNA3-Flag or pcDNA-Flag-UL13. The proliferative ability of the cells was evaluated 0, 12, 24, and 36 h post transfection. The cells were then digested and seeded into 96-well plates. CCK-8 reagent (10 μL) was added to the wells, and the plates were incubated at 37 °C for 2 h. The optical density of each sample was measured at a wavelength of 450 nm by using an enzyme-labelled instrument (Thermo Fisher Scientific).
7
Ferroptosis Inhibition in Osteoblasts
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Cell culture and treatment
The hFOB1. Erastin (Sigma-Aldrich) was used as positive control for cell ferroptosis. Deferoxamine (DFO, Sigma-Aldrich) and Ferrostatin-1 (Ferr-1, Sigma-Aldrich) were used to inhibit Fe or Erastin induced cell death. All samples were collected for relative assays.
Cell proliferation assay hFOB1.19 osteoblast cells were seeded (10 3 cells per well) in 96-well plates. After cells were treated with Erastin or Fe or/and Ferr-1 or/and DFO, a solution containing fresh medium (90 μl) and CCK-8 reactant (10 μl) (Dojindo) was added to each well, and cells were incubated at 37 °C for 1.5 h in the dark followed by addition of stop buffer. Absorbance at 450 nm was then measured using an enzyme-labelled instrument (Thermo). All operations were performed in accordance with the manufacturer's instructions.
The hFOB1. Erastin (Sigma-Aldrich) was used as positive control for cell ferroptosis. Deferoxamine (DFO, Sigma-Aldrich) and Ferrostatin-1 (Ferr-1, Sigma-Aldrich) were used to inhibit Fe or Erastin induced cell death. All samples were collected for relative assays.
Cell proliferation assay hFOB1.19 osteoblast cells were seeded (10 3 cells per well) in 96-well plates. After cells were treated with Erastin or Fe or/and Ferr-1 or/and DFO, a solution containing fresh medium (90 μl) and CCK-8 reactant (10 μl) (Dojindo) was added to each well, and cells were incubated at 37 °C for 1.5 h in the dark followed by addition of stop buffer. Absorbance at 450 nm was then measured using an enzyme-labelled instrument (Thermo). All operations were performed in accordance with the manufacturer's instructions.
8
CCK8 Assay for Cell Viability Analysis
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CCK8 assay was performed to explore the cell viability of CGM1 cells at 24 and 48 h after drug treatment. The cells were suspended in DMEM containing 10% FBS and the cell density was adjusted to 1 × 105/mL. The cell suspension was seeded into 96-well plates, with 100 μL cell suspension in each well. CCK8 reagent (10 μL) was added into each well, and the cells were cultured for 4 h in an incubator at 37 °C. The absorbance of samples was detected at 450 nm using enzyme-labelled instrument (Thermo Fisher Scientific, Waltham, MA).
9
SMCC-7721 Cell Proliferation Assay
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An MTT assay was performed to explore the proliferation of the SMCC-7721 cells at 24 and 48 hours. SMCC-7721 cells were incubated with the cell supernatant of the wild or modified TAMs. Subsequently, the SMCC-7721 cells were resuspended in RPMI-1640 medium containing 10% FBS and the cell density was adjusted to 10 5 /mL. The cell suspension was seeded into 96-well plates, with 100 µL cell suspension in each well, and incubated for 24 or 48 hours. Then, 10 µL MTT (5 mg/mL) was added to each well, and the cells were cultured for 4 hours in an incubator at 37°C. After that, supernatant of cells was removed and DMSO (100 µL) was added and mixed in each well. The absorbance of samples was detected at 490 nm wavelength using enzyme-labelled instrument (Thermo Fisher Scientific).
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