LNCaP and MDA PCa 2b cells overexpressing PIK3CD-L or PIK3CD-S were subjected to apoptosis assays for evaluating the apoptosis capacities upon different drug treatments. The PCa cells were seeded in 96-well plates at initial cell density of (5 × 103 cells/well) and were grown for 24 h, then were treated with four small-molecule inhibitors (Idelalisib, Seletalisib, Wortmannin, and Dactolisib) at concentrations of 0, 1, 10, and 25 μM. After drug treatment for 48 h, the PCa cells were harvested for the apoptosis assays using Apo-ONE Caspase-3/7 Assay Kit (Promega Corporation, Madison, WI, USA) according to the protocol described by the manufacturer. The apoptosis activities were measured by detecting fluorescence signals at wavelengths of 498 nm and 521 nm (for excitation and emission, respectively) using Biotek Synergy HT Microplate Reader (BioTek, Winooski, VT, USA).
Apo one caspase 3 7 assay kit
Manufactured by Promega
Sourced in United States
The Apo-ONE Caspase-3/7 Assay Kit is a fluorometric assay designed to detect and measure the activity of caspase-3 and caspase-7, two key enzymes involved in the execution phase of apoptosis. The kit provides a simple and sensitive method for quantifying caspase-3/7 activity in cell lysates or purified enzyme preparations.
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Lab products found in correlation
4 protocols using apo one caspase 3 7 assay kit
1
Apoptosis Evaluation of LNCaP and MDA PCa 2b Cells
2
Apoptosis Assay for miRNA Modulation in PCa
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For the measurement of apoptosis initiation by transfection of miRNA mimic or antagomir in PCa cells, the transfected cells 5000 cells/well were seeded onto 96-well cell culture plates (Corning, Corning, NY, USA) and then incubated overnight. The miRNA mimic/antagomir transfected cells were then treated with vehicle or 11 mM of Docetaxel [26 (link)]. 24 h after the vehicle or drug treatment, the apoptosis assays were performed using Apo-ONE Caspase-3/7 Assay Kit (Promega Corporation, Madison, WI, USA) according to the protocol described by the manufacturer. 100 µL of homogeneous Caspase-3/7 reagent was added to the sample plate and incubated at room temperature for 1 h. Fluorescence was detected for measuring the apoptosis state (Caspase 3/7 activity) using Biotek Synergy HT Microplate Reader (BioTek, Winooski, VT, USA) at wavelengths of 499 nm and 521 nm for excitation and emission, respectively.
3
Apoptosis Assay of Cancer Cell Lines
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HT-29, SW620, MDA MB 231, MCF7, A549, and H1299 were seeded at a density of 3 × 104 cells/well in 96-well plates. The cells were grown overnight and then followed by transfections. After 24 h, fresh media were applied to replace the transfection reagent-containing media with 11 mM of docetaxel or vehicle, then the cells were incubated for an additional 24 h. The Apo-ONE Caspase-3/7 Assay Kit (Promega Corporation, Madison, WI, USA) was used to measure apoptosis according to the protocol described by the manufacturer. Then, 100 µL of homogeneous Caspase-3/7 reagent was added to each well and the plate incubated at room temperature for 30min to 2h. Fluorescence was detected for measuring (at wavelengths of 499/521 nm excitation/emission) the apoptosis state (Caspase 3/7 activity) using Biotek Synergy HT Microplate Reader (BioTek, Winooski, VT, USA).
4
Quantifying Apoptosis by Annexin V and Caspase 3/7
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Cell apoptosis was detected by flow cytometry with an Annexin V/PI apoptosis kit (Cat. no. 559763, BD Biosciences, San Jose, CA, USA), according to the manufacturer's instructions. Cells were digested and centrifuged at 100 rcf at 4 °C for 5 min. Then, the cells were resuspended in 100 μL 1 × binding buffer and incubated with 5μL Annexin V-APC/PI in the dark at 25 °C for 15 min. The cells were suspended in 400 μL 1 × binding buffer to remove unbound moieties and subjected to flow cytometry (BD Biosciences, Oxford, UK). Bel7404 cells were incubated in a 96-well plate, and three blank wells were set up. Caspase 3/7 intracellular activity was detected by using the Apo-ONE® caspase 3/7 assay kit (Cat. no.G8091, Promega, Mannheim, Germany), and cell fluorescence intensity at 499 nm was measured by using Tecan Infinite plate reader. The experiments, including technical duplicates, were repeated at least three times.
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