| Assay Method Information | |
| | CD73 Enzyme Inhibition Assay |
| Description: | To evaluate the inhibitory effect of selected compounds on CD73, Malachite Green Phosphate Detection Kit (R&D, Cat #DY996) was used. Briefly, compounds were dissolved and diluted to the desired concentration using phosphate-free buffer (Tris-HCl, pH 7.3). 25 μL of the compound solution was added to an equal volume of CD73 protein solution (2× concentration, 0.5 μg/mL, Novoprotein, Cat #C446), followed by a 5-minute incubation at room temperature. 10 μL of Malachite Green Reagent A was added to each well, mixed thoroughly and incubated for 10 minutes at room temperature. 10 μL of Malachite Green Reagent B was then added to each well, mixed thoroughly and incubated for 20 minutes at room temperature. Finally, the optical density of each well was determined using a microplate reader set to 620 nm. |
| Affinity data for this assay | |
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