The HDAC activity was analyzed using a colorimetric assay kit (Bio Vision, USA), according to the manufacturer’s protocol. Briefly, nuclear cell lysates were extracted using the Nuclear Extract Kit (Active Motif, Belgium) and were incubated with the HDAC colorimetric substrate, the density was read at 405 nm on a Multimode Reader (Promega, USA).
Multimode reader
Manufactured by Promega
Sourced in United States
The Multimode Reader is a versatile lab equipment that can perform various assays and measurements. It has the capability to detect and quantify different types of signals, including luminescence, fluorescence, and absorbance, across a wide range of microplates, cuvettes, and other sample types.
Automatically generated - may contain errors
2 protocols using multimode reader
1
Colorimetric HDAC Activity Assay
2
NF-κB Activation Assay Protocol
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The NF-κB-luciferase reporter assay was performed as previously described. [56] [57] Briefly, HEK293 cells were transfected with the NF-κB-luciferase reporter genes in a 10 cm plate with Lipofectamine 3000. The next day, cells were lifted with TrypLE and resuspended in phenol red-free DMEM. Transfected cells (7500 cells/well) were dispensed in 96-well white, solid-bottom plates and incubated with increasing concentrations of ABY TNFR1-1 (1 pM -10 μM) or PBS (negative control) in the presence (0.6 nM) and absence of TNF-α (ab9642, Abcam) for 24 hours at 37 °C. After incubation, 70 μL of Dual-Glo Luciferase Reagent (Promega, Madison, WI) was added and incubated at room temperature for 15 min, and firefly luminescence was measured using a Cytation 3 Cell Imaging Multi-Mode Reader luminometer. Next, 70 μL of Dual-Glo Stop & Glo Reagent was added and incubated at room temperature for 15 min, and luminescence was measured using a luminometer.
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