| Assay Method Information | |
| | Factor D Esterolytic Assay |
| Description: | An established esterolytic assay for the measurement of Factor D activity and inhibition of Factor D activity was used. For this assay Z-Lys-SBzl, 1.29 mM was used as the substrate for Factor D (104 mM). Hydrolysis of this compound by Factor D liberated a free sulfhydryl group which is then reacted with 5,5′-dithiobis(2nitrobenzoic acid) producing an intense yellow color. The assays were performed in 96 well microtiter plates and rates of hydrolysis were monitored at 405 nm on a Biotek Synergy H1 plate reader. Hydrolysis rates were reported as change in mOD/mmn. The assay was conducted in 100 mMv HEPES, 500 mMv NaCl, pH 7.5 containing 10% DM50 in a final volume of 50 μL per well. An IC50, a compound concentration which inhibits 50% of the enzymatic activity, was calculated. Compounds in the examples were tested a minimum of three times. |
| Affinity data for this assay | |
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