The Rat PDE4B Assay Kit is a fluorescence polarization (FP), homogeneous 96-well assay designed for the screening and profiling of rat PDE4B inhibitors. The kit contains enough purified recombinant rat PDE4B, fluorescent probe (cAMP), PDE Assay Buffer, Binding Agent, and diluent for 100 reactions. This assay requires a fluorescent microplate reader capable of measuring fluorescence polarization (FP) to read the FP signal. For more information on the principles of FP, visit fp_assays.pdf ().

Figure 1: Illustration of the PDE4B assay principle.
The assay uses a fluorescein-labeled cyclic adenosine monophosphate (cAMP-FAM), in which the phosphate group is engaged within the cyclic nucleotide. This is a very small molecule that can rotate fast (low FP). PDE4B 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 large complex, with restricted movement (high FP). FP values are proportional to PDE activity.