| Solution Information | help | |
| Enzyme: | M1 family aminopeptidase | |
| inhibitor: | BDBM48805 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | M1AAP Assay Protocol for 1536-well HT Purified recombinant M1AAP enzyme was provided by Prof. John Dalton, Institute for the Biotechnology of Infectious Diseases (IBID) University of Technology Sydney. Compound Dosing/Plating: For the primary screen 15 nl of compounds in DMSO were dispensed into 1536-well black non-binding surface plates. This resulted in a concentration of 30 uM of compound in the assay. For the follow up dose response assays 8 concentrations of each compound ranging from 100-1 uM were used. Assay Setup: 2.5 uL of M1AAP reagent mix, which included the fluorogenic peptide substrate (H-Leu-NHMec) in assay buffer, was added to each well of the previously compound dosed 1536-well plates. The reaction was initiated with the addition of 2.5 uL of the M1AAP diluted in assay buffer. The final concentrations in the reaction were 0.1 mM H-Leu-NHMec and 50 ug/ml M1AAP diluted in assay buffer (50 mM Tris-HCl (pH 7.5), 0.01% Triton X-100, and 2% DMSO). The test plate was immedi | |
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