Different concentrations of farletuzumab-FuGi, farletuzumab-cFuGi (25 to 250 ng), or furin assay buffer (negative control) were preincubated with 25 ng of furin (50 μL, 0.5 ng/μL in furin assay buffer) in a total reaction volume of 100 μL for 10 min at room temperature. The reaction was started by adding 40 μL furin protease substrate (final concentration of the furin protease substrate in a 100 μL reaction was 2 μM). The fluorescence intensity of the reaction mixture was monitored at excitation wavelength of 380 nm and detection of emission at wavelength 460 nm using 96-well microplate reader (BioTek Instruments Inc., USA). For inhibitor control, 10 μL of chloromethylketone (0.5 μM) was added to 25 ng of furin and for positive control (only furin), no antibody and inhibitor were added. The protease activity of positive control well was considering as 100% activity and the decrease in furin activity was compared to that. For kinetic study, fluorescence was measured immediately after adding the protease substrate and data was recorded every 1 min for 15 min at Ex/Em = 380 nm/460 nm.
96 well microplate reader
Manufactured by Agilent Technologies
Sourced in United States
The 96-well microplate reader is a compact and versatile instrument designed for high-throughput optical measurements. It can measure absorbance, fluorescence, and luminescence in a 96-well microplate format, making it suitable for a wide range of applications such as enzyme activity assays, cell-based assays, and protein quantification.
Automatically generated - may contain errors
Lab products found in correlation
Bca protein assay kit, by Thermo Fisher Scientific (6 mentions)
Fatal plus, by Vortech Pharmaceuticals (3 mentions)
Bca assay, by Thermo Fisher Scientific (2 mentions)
Wst 1 cell viability assay kit, by Daeil Lab Service (2 mentions)
Ez cytox, by Daeil Lab Service (2 mentions)
Celltiter 96 aqueous one solution cell proliferation assay, by Promega (2 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Mts cell proliferation assay, by Promega (1 mentions)
Calcium colorimetric assay kit, by Beyotime (1 mentions)
Cell counting kit 8 cck 8, by Selleck Chemicals (1 mentions)
Bevacizumab, by Merck Group (1 mentions)
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt assay kit, by Merck Group (1 mentions)
Bevacizumab avastin, by Roche (1 mentions)
Mtt assay, by Merck Group (1 mentions)
Elisa kit, by Bioswamp (1 mentions)
Protease inhibitor, by Merck Group (1 mentions)
Sigmastat 3, by Grafiti LLC (1 mentions)
Aconitase activity kit, by Merck Group (1 mentions)
Bicinchoninic acid (bca), by Beyotime (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Merck Group (1 mentions)
45 protocols using 96 well microplate reader
1
Furin Activity Inhibition Assay
2
Hippocampal and Cortical Protein Analysis
3
Curcumin's Impact on Cell Viability
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Cells were treated with 20 or 40 µM curcumin for 24 or 48 h. The effects of curcumin on cell viability were determined using the Cell Counting kit-8 (CCK-8; Dojindo Molecular Technologies, Inc., Kumamoto, Japan) assay. Briefly, following incubation with the 20 or 40 µM curcumin for 24 or 48 h, the cells were incubated with 5 g/l CCK-8 solution for 2 h. Subsequently, the cells were placed in a 96-well microplate reader (BioTek, Winooski, VT, USA) for analysis and the optical density (OD) was detected at 450 nm. Cell viability was evaluated using the following formula: Cell viability (%) = [1 − (OD of the samples/OD of the control)] × 100%.
4
Evaluating Cell Viability with MTS Assay
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CP70 and SKOV3 cell viabilities were detected using the MTS cell proliferation assay (Promega). Briefly, CP70 and SKOV3 (1000 cells each) were seeded in 96-well plates containing 100 μL media/well overnight. Then, they were exposed to nano-NI or niclosamide for 72 h. The MTS solution was prepared from the CellTiter 96R aqueous MTS reagent powder and kept at −20°C for long-term storage, and at 4°C away from light before use. Next, 20 μL of the MTS solution was added to each well, where the viable cells generated soluble formazan. After an incubation period of 45 to 60 min, the fluorescence absorbance at 490 nm was measured using a 96-well microplate reader (BioTek). Each reaction was assayed at least thrice. The results are expressed as the ratio of the absorbance of each sample to that of the untreated samples with media alone. The proliferation assays were performed in triplicate. All in vitro studies were conducted in triplicate in two independent experiments in the different cell lines.
5
Enzyme Activity Assays in Microplates
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Enzyme activities were determined in a 96-well microplate reader (BioTek Instruments) from protein extracts. GS activity was measured monitoring the formation of γ-glutamilhydroxamate (γ-GHM) in a semibiosynthetic assay (Setién et al., 2013 (link)). GDH, PEPC, ICDH, and NAD-ME activities were measured monitoring the evolution of NAD(P)H monitored at 340 nm. GDH activity was determined in aminating sense (Setién et al., 2013 (link)). For ryegrass, wheat, and clover, novel data of carbon enzymes activities (PEPC, ICDH, and NAD-ME) were determined as previously described by Vega-Mas et al. (2015) (link) or Sarasketa et al. (2016) (link).
6
Assessing Bladder Cancer Cell Viability
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Cell viability was assessed by using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. 3000 cells in 100 µl of medium per well were seeded in 96-well plates. Cells were cultured for 24 h, 48 h, 72 h, 96 h, and then incubated with 0.5 mg/ml of MTT at 37°C for 4 h. Medium was replaced with 150 µl DMSO per well to dissolve the precipitates. Colorimetric analysis using a 96-well micro-plate reader (Bio Tek) was performed at wavelength of 490 nm. An effect curve was drawn and the IC50 value of gemcitabine to bladder cancer cells was calculated depending on cell proliferation and drug concentration.
7
Cell Viability Determination using EZ-CyTox
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Cell viability was determined using the EZ-CyTox (tetrazolium salt, WST-1) cell viability assay kit (Daeil Lab Service, Seoul, Korea), as previously described [33 (link)]. Absorbance at 570 nm was detected using a 96-well microplate reader (BioTek Instruments, Winooski, VT, USA). Cell viability is expressed as the fraction of the surviving cells relative to the vehicle-treated cells.
8
Determination of Lipoxygenase Activity
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The lipoxygenase activity was determined according to the previous method [47 ]. Briefly, 200 µL mixture contained 160 µL sodium phosphate buffer (100 mM, pH 8.0), 10 µL sample extracts (25 to 100 µg in 100 mM Tris buffer pH 7.4) and 20 µL 5-lipoxygenase enzyme. The contents were preincubated for 10 min at 25 °C. The reaction was started by the adding of 10 µL linoleic acid solution as a substrate. After 6 minutes, the absorbance was noted at 234 nm. All reactions were achieved in triplicates in 96-well microplate reader (BioTek, Seoul, South Korea). The positive and negative controls were included in the experiment.
9
MTT Assay for Cell Viability
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Cell viability was determined with the MTT assay. Briefly, HeLa cells were seeded at 1×104 cells/well in 96-well flat-bottom microtiter plates. After 24 hours, the medium was replaced with fresh DMEM containing 3 or 5 µg/mL dicerandrol B or dimethyl sulfoxide (DMSO; untreated control), and cells were incubated for 24, 48, or 96 hours. Subsequently, 20 µL of 5 mg/mL MTT was added to each well, and cells were incubated for 4 hours. Formazan was solubilized in 150 µL DMSO, and the OD at 490 nm was detected with a 96-well microplate reader (BioTek, Winooski, VT, USA). Cell viability was evaluated according to the formula: cell viability (%) = [1− (OD of the samples/OD of the control)] ×100%.
10
Hippocampal and Cortical Protein Analysis
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A total of 48 rats (n=6 per group at each time point) received an overdose of Fatal-plus (100mg/kg sodium pentobarbital) and the brains rapidly dissected at 1, 3, 7 or 14 days post-injury. After removal of the brain, the ipsilateral hippocampus and cortex were rapidly dissected on a chilled ice plate, immediately snap frozen with liquid nitrogen and tissues stored at −80°C. Samples were homogenized using a lysis buffer (0.1 M NaCl, 0.01 M Tris-Cl (pH 7.6), 0.001 M EDTA) with protease inhibitor cocktail (Pierce, Rockford, IL, Cat. No. 1861281). The homogenized whole cell lysates were centrifuged at 12,000xg at 4°C for 30 minutes and the supernatants collected. The protein concentration for each lysate was determined using a BCA assay (Thermo Scientific, Pittsburgh, PA) and a 96 well microplate reader (Biotek, Winooski, VT).
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