| Assay Method Information | |
| | IL-17A Enzyme-Linked Immunosorbent Assay (ELISA) |
| Description: | The inhibition of receptor-ligand binding by human IL-17A inhibitors was quantified by competitive ELISA. 0.2 ug/ml of IL-17A (Sino Biological Inc. Cat #12047-H07B) was added in a 96-well pated at 100 ul per well for incubation for 30 min under 37C. The 96-well plate was washed with PBST (PBS, 0.05% Tween-20) at 200 ul per well for 4 times, and then added with 200 ul of 5% skim milk and placed on a shaker for incubation at 25C for 30 min. The compound to be tested was prepared at 100x concentration ranging from 0.003 uM to 30 uM. After washed with PBST (PBS, 0.05% Tween-20) 4 times, the plate was added with 89 ul of PBST and 1 ul of the compound to be tested at 100x concentration followed by uniformly mixing and pre-incubation at 25C for 10 min. Then the plate was added was 10ul of 16nM IL-17R, and incubated on a shaker at 25C for 30 min. After washed for 4 times, the plate was added with 100 ul of ant-Fc-tag horseradish peroxidase (HRP)-conjugated antibody, and incubated on a shaker at 25C for 30 min. After washed 4 times, the plate was added with 100 ul of 3,3', 5,5'-tetramethylbenzidine (TMB) substrate solution, and incubated at 25C in the dark. Then the plate wa added with 20% HCl, followed by detection of the light absorption value at a wavelenght of 450 nm by a microplate reader. |
| Affinity data for this assay | |
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