Intracellular cytosolic Ca2+ dynamic was measured using the FLIPR Calcium 6 Assay Kit (Molecular Devices) according to the manufacturer’s instructions. In brief, HeLa cells were plated at a density of 10000 cells per well in black wall/clear bottom 96-multiwell plates from Costar (Tewksbury, MA, United States) and cultured for 24 h before treatment. After that, calcium 6 reagent was added directly to cells, and cells were incubated for an additional 2 h at 37°C and 5% CO2. 5 and 10 μM of LP-4 were then added to the wells and immediately subjected to data acquisition on the FLIPR Tetra High-Throughput Cellular Screening System (Molecular Devices) at room temperature using a 1-s reading interval throughout the experiments.
Black wall clear bottom 96 multiwell plates
Manufactured by Corning
Sourced in United States
The Black wall/clear bottom 96-multiwell plates are a type of lab equipment designed for various experimental procedures. These plates feature black walls and a clear bottom, which allows for optimal optical clarity and contrast during imaging and analysis. The plates provide a standardized 96-well format, ensuring compatibility with a wide range of laboratory equipment and protocols.
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5 protocols using black wall clear bottom 96 multiwell plates
1
FLIPR Calcium 6 Assay in HeLa Cells
2
Intracellular Ca2+ Dynamics Measurement
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Intracellular cytosolic Ca2+ dynamics was determined using the FLIPR Calcium 6 Assay Kit (Molecular Devices) according to the manufacturer's instructions. In brief, RASFs were plated in black wall/clear bottom 96‐multiwell plates (Costar, Tewksbury, MA, USA) at a density of 10,000 cells per well and incubated overnight before treatment. Next day, the RASFs were treated with calcium 6 reagent for 2 hr at 37°C and 5% CO2. Indicated concentrations of celastrol or thapsigargin were then added to the wells and immediately subjected to data acquisition on the SpectraMax Paradigm Multi‐Mode Microplate Reader (Molecular Devices) at room temperature using a 1‐s reading interval throughout five independent experiments.
3
Measurement of Intracellular Ca2+ Dynamics
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Intracellular cytosolic Ca2+ dynamic was measured using the FLIPR Calcium 6 Assay Kit (Molecular Devices, USA), which contains a proprietary Ca2+-sensitive fluorophore, according to the manufacturer's instructions. In brief, 10000 RASFs per well were seeded in black wall/clear bottom 96-multiwell plates from Costar (Tewksbury, MA, USA) and cultured for 24 h before treatment. After that, calcium 6 reagent was added directly to cells, and cells were incubated for an additional 2 h at 37°C and 5% CO2. One micromolar of celastrol (China Chengdu MUST, A000106) was then added to the wells and immediately subjected to data acquisition on the FLIPR Tetra High-Throughput Cellular Screening System (Molecular Devices, USA) at room temperature using a 1-s reading interval throughout the experiments.
4
Intracellular Calcium Dynamics Assay
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Intracellular cytosolic Ca2+ dynamic was measured using the FLIPR Calcium 6 Assay Kit (Molecular Devices) according to the manufacturer’s instructions. In brief, A2780s and A2780cp cells were plated at a density of 10,000 cells per well in black wall/clear bottom 96-multiwell plates from Costar (Tewksbury, MA, USA) and cultured for 24 h before treatment. After that, calcium 6 reagent was added directly to cells, and cells were incubated for an additional 2 h at 37 °C and 5% CO2. Ssd and/or CDDP were then added to the wells and immediately subjected to data acquisition on the FLIPR Tetra High-Throughput Cellular Screening System (Molecular Devices) at RT using a 1 s reading interval throughout the experiments.
5
Intracellular Cytosolic Calcium Dynamics
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Intracellular cytosolic Ca2+ dynamic was measured using the FLIPR Calcium 6 Assay Kit (Molecular Devices) according to the manufacturer’s instructions. In brief, HeLa cells were plated at a density of 10 000 cells per well in black wall/clear bottom 96-multiwell plates from Costar (Tewksbury, MA, USA) and cultured for 24 h before treatment. After that, calcium 6 reagent was added directly to cells, and cells were incubated for an additional 2 h in the absence of external Ca2+ in HBSS buffer at 37 °C and 5% CO2. 10 and 20 μM of neferine were then added to the wells and immediately subjected to data acquisition on the FLIPR Tetra High-Throughput Cellular Screening System (Molecular Devices) at room temperature using a 1- s reading interval throughout the experiments.
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