| Assay Method Information | |
| | Inhibitory Activity of NLRP3 |
| Description: | ELISA plate, and incubated at 4° C. overnight. Discard the liquid in the ELISA plate and wash it 4 times with eluent. Add 300 μL/well reagent diluent and incubate the ELISA plate at room temperature for 1.5 hours. Wash the plate with eluent for four times. Then 100 μL/well samples were added to ELISA plate coated with IL1β and incubated at room temperature for 2 h. Wash the plate with eluent for four times. Each well was added with 100 μL detection antibody and incubated at 37° C. for 2 h. Wash the plate with eluent for four times. Add 100 μL of HRP labeled secondary antibody to each well. Incubate at 37° C. for 1 hour. Wash the plate with eluent for four times. Add 100 μL of A+B substrate per well. Incubate at 37° C. for 30 minutes. Add 100 μL STOP solution per well. Gently shake the plate for a few seconds. Read the absorption value at 450 nm on the EnVision multifunction reader and calculate the Inhibitory rate according to the following formula: Inhibition %=(Ave_H-Sample)/(Ave_H-Ave_L). Among them, Ave_H represents the average value of DMSO well, Sample represents the average value of compound well, and Ave_L represents the average value of 10 μM positive control well. The log value of concentration was taken as the X-axis and the percentage inhibitory rate as the Y-axis. The dose-effect curve was fitted with log (inhibitor) vs. response-Variable slope from software GraphPad Prism 5 to obtain the IC50 value of each compound. |
| Affinity data for this assay | |
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