| Solution Information | help | |
| Enzyme: | Mannose-6-phosphate isomerase | |
| inhibitor: | BDBM34811 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | PMI assay materials: 1) Human PMI protein was provided by Dr. Hudson Freeze (Sanford-Burnham Medical Research Institute, San Diego, CA). 2) Substrate working solution: 50 mM HEPES, pH 7.4, 0.4 mM Mannose-6-phosphate, 1.6 U/ml Diaphorase, 0.2 mM Resazurin. 3) Enzyme working solution: 50 mM HEPES, pH 7.4, 0.44 mM NADP+, 9.048 mM MgCl2, 0.01% Tween 20, 4.6 ug/ml phosphoglucose isomerase, 30 ng/ml PMI, 1.8 ug/ml G6PDH. PMI HTS protocol: 1) 2 uL of Substrate working solution was added to columns 3-48 of a Costar 1536-well black plate (cat #3724) using a Thermo Multidrop Combi dispenser 2) 2 ul of Substrate working solution without mannose-6-p was added to columns 1 and 2 (positive control) of a Costar 1536-well black plate (cat #3724) using a Thermo Multidrop Combi dispenser 3) 40 nL of 100% DMSO was added to columns 1-4 using a HighRes biosolutions pintool and V&P Scientific pins 3) 40 nL of 2 mM compounds in 100% DMSO were dispensed in columns 5-48 using a HighRes biosolutions pin | |
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