DESCRIPTION

• Notes: MATERIALS OR INSTRUMENTS REQUIRED BUT NOT SUPPLIED: Fluorescent microplate reader capable to measure fluorescence polarization
• Shiptemp: -80°C (dry ice)
• Synonyms: inhibitor screening, assay kit, PDE8A
• Warnings: Avoid freeze/thaw cycles.
• Category: Phosphodiesterase/Assay Kit
• Background: Phosphodiesterases (PDEs) play an important role in the dynamic regulation of cAMP and cGMP signaling. PDE8 is involved in many biological processes, including T-cell activation, testosterone production, adrenocortical hyperplasia, and thyroid function. Phosphodiesterases catalyze the hydrolysis of the phosphodiester bond in dye-labeled cyclic monophosphates. Beads selectively bind the phosphate group in the nucleotide product. This increases the size of the nucleotide relative to unreacted cyclic monophosphate. In the polarization assay, dye molecules with absorption transition vectors parallel to the linearly-polarized excitation light are selectively excited. Dyes attached to the rapidly-rotating cyclic monophosphates will obtain random orientations and emit light with low polarization. Dyes attached to the slowly-rotating nucleotide-bead complexes will not have time to reorient and therefore will emit highly polarized light.
• References: Wang, H., et al. Biochemistry 2008; 48: 12760-8.
• Description: The PDE8A Assay Kit is designed for identification of PDE8A inhibitors using fluorescence polarization. The assay is based on the binding of a fluorescent nucleotide monophosphate generated by PDE8A to the binding agent. The key to the PDE8A Assay Kit is the specific binding agent. Using this kit, only two simple steps on a microtiter plate are required for PDE8A reactions. First, the fluorescently labeled cAMP is incubated with a sample containing PDE8A for 1 hour. Second, a binding agent is added to the reaction mix to produce a change in fluorescent polarization. The FP signal is measured using a fluorescent microplate reader capable of measuring fluorescence polarization.
• Methodology: Fluorescence Polarization
• Supplied As: The PDE8A inhibitor screening assay kit comes in a convenient 96-well format,including purified PDE8A enzyme, fluorescently labeled PDE8A substrate (cAMP),binding agent, and PDE assay buffer for 100 enzyme reactions.
• Unspsc Code: 41121811
• Unspsc Name: Laboratory microchemistry kits
• Applications: Great for studying enzyme kinetics and screening small molecular inhibitors for drug discovery and HTS applications.
• Product Type: Assay Kit
• Biosafety Level: Not applicable (BSL-1)
• Contraindications: DMSO >1%, strong acids or bases, ionic detergents, high salt
• Storage Stability: At least 6 months from date of receipt, when stored as directed. Kit components require different storage conditions. Be sure to store each component at the proper temperature upon arrival.
• Scientific Category: Phosphodiesterase
• Instructions for Use: See assay kit datasheet for detailed protocol.