PDE5A1 Assay Kit1

The PDE5A1 Assay Kit is a fluorescence polarization (FP), homogeneous, 96-well assay kit designed for the screening and profiling of PDE5A1 (Phosphodiesterase 5A1) inhibitors. This assay takes advantage of a specific fluorescent phosphate-binding nanoparticle. The kit contains enough purified recombinant PDE5A1, fluorescent probe, PDE assay buffer, Binding Agent, and diluent for 100 reactions.Figure 1: Illustration of the PDE5A1 Assay Kit principle.The assay uses a fluorescein-labeled cyclic guanosine monophosphate (cGMP-FAM for PDE5A1), in which the phosphate group is engaged within the cyclic nucleotide. This is a very small molecule that rotates fast (low FP) . PDE5A1 catalyzes the hydrolysis of the phosphodiester bond in the cyclic nucleotide and frees the phosphate group. In a second step the free phosphate group is recognized by a specific phosphate-binding nanobead (Binding Agent) leading to the formation of a large complex, with restricted movement (high FP) . FP is proportional to PDE5A1 activity.This assay requires a fluorescent microplate reader capable of measuring fluorescence polarization (FP) and equipped with the required parts to read the FP signal. For more information FP technology, visit our Tech Note:FP, assay principles and applications.Note:As of July 2024, this protocol has been re-optimized for performance. Previous versions of this kit are available upon request.Need us to run inhibitor screens or profile your compounds against PDE5A1? Check out ourPhosphodiesterase Screening Services.This product has beencited 14 times.