| Solution Information | help | |
| Enzyme: | Tyrosine-protein phosphatase non-receptor type 7 | |
| inhibitor: | BDBM46235 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay materials: 1)HePTP protein was provided by Dr. Mustelin (Sanford-Burnham Medical Research Institute, San Diego, CA). 2) Assay Buffer: 50mM Bis-Tris pH 6.0, 1mM DTT, 0.1% BSA, 0.005% Tween-20 3) 3-O-Methylfluorescein phosphate (OMFP) cyclohexylammonium salt (Sigma-Aldrich, M2629) Dose response protocol: 1) Dispense 10 ul of 600 uM OMFP in assay buffer into all columns of black 384-well small-volume Greiner plates (786076) 2) Dispense 10 uL of assay buffer in columns 1-2. 3) Using Labcyte Echo550, dispense 200 nL of serially diluted compounds to columns 3-22 and DMSO to columns 1-2 and 23-24. Final concentration of DMSO was 1 % 4) Using Thermo Scientific MultiDrop Combi, dispense 10 uL of 3 nM HePTP working solution to wells in columns 3 through 24. 5) Incubate lidded plate for 30 min. at room temp. 6) Read plate on M5 plate reader in fluorescence mode (Ex: 480nm, Em: 520nm) | |
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