PDE3A Assay Kit
Catalog # 60330

Size: 96 reactions

DESCRIPTION:  Phosphodiesterases  (PDEs)  play  an  important  role  in  dynamic
regulation  of  cAMP  and  cGMP  signaling.  PDE3A,  also  known  as  cGMP-inhibited
phosphodiesterase,  has  been  implicated  in  cardiovascular  function  and  fertility.  The  PDE3A
Assay  Kit  is  designed  for  identification  of  PDE3A  inhibitors  using  fluorescence
polarization.  The  assay  is  based  on  the  binding  of  a  fluorescent  nucleotide
monophosphate generated by PDE3A to the binding agent. 

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.

The  PDE3A  inhibitor  screening  assay  kit  comes  in  a  convenient  96-well  format,  with
purified  PDE3A  enzyme,  fluorescently  labeled  PDE3  substrate  (cAMP),  binding  agent,
and  PDE  assay  buffer  for  100  enzyme  reactions.  The  key  to  the  PDE3A  Assay  Kit  is  the
specific  binding  agent.  Using  this  kit,  only  two  simple  steps  on  a  microtiter  plate  are
required  for  PDE3A  reactions.  First,  the  fluorescently  labeled  cAMP  is  incubated  with  a
sample  containing  PDE3A  for  1  hour.  Second,  binding  agent  is  added  to  the  reaction mix  to
produce  a  change  in  fluorescent  polarization  that  can  then  be  measured  using  a
fluorescence reader equipped for the measurement of fluorescence polarization.

COMPONENTS:

 
APPLICATIONS: Great  for  studying  enzyme  kinetics  and  screening  small molecular
inhibitors for drug discovery and HTS applications.

REFERENCE:  Maurice DH. Front. Biosci. 2005; 10:1221-8.

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