The present invention uses the principle that phosphomolybdate binds to
hydrophobic surfaces to isolate the phosphomolybdate complex from other
phosphate-containing molecules and further uses the SPA concept to bring a
radiolabeled phosphomolybdate complex in close contact with a scintillant
for measurement by scintillation counting. Generally, the present
invention provides an assay for detecting and measuring the amount of
orthophosphate (Pi) in an aqueous reaction mixture, wherein the amount of
Pi released is separated from the reaction mixture by: adding a solution
of molybdate to the reaction mixture to form a phosphomolybdate complex;
and contacting the phosphomolybdate complex with a hydrophobic surface,
wherein the surface is capable of being separated from the aqueous
reaction mixture to allow measurement of the Pi. Particularly, this
invention provides an assay for measuring the ATPase activity of enzymes,
more particularly, the HPV E1 helicase.
