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| Solution Information | help | |
| Enzyme: | Complement C1s subcomponent | |
| inhibitor: | BDBM32669 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Protocol 1. Coat assay plates with chicken ovalbumin (OVA) (1mg/ ml in PBS, 50uL per well) at 4deg C overnight. 2. Wash assay plates three times with 300uL PBS-0.05% Tween-20 at room temperature. 3. Block assay plates by treating with 200uL of 1% BSA-PBS for 1 hour at room temperature. 4. Wash assay plates three times with 300uL PBS-0.05% Tween-20 at room temperature. 5. Add 50uL Anti-OVA (1:2000 dilutions in the blocking buffer) and incubate for 1 hour at room temperature. 6. Serial dilute single compounds at 20x concentration in DMSO (8 two-fold dilutions from 1 mM to 7.8125 uM) in a 96-well Greiner V-bottom plate. 7. Wash assay plates three times with 300uL PBS-0.05% Tween-20 at room temperature. 8. For columns 1 to 11 (positive control/compound), dilute human serum 1:10 in Mg2+ -EGTA GVB++, take 57uL diluted serum and 3uL compound from the compound plate, mix them in the mother plate. Transfer 50uL mix per well from the mother plate to the assay plate and incubate for 1 hour a | |
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