A rapid method for the measurement of prostaglandin receptors

Abstract A method is presented which provides for a simple and rapid determination of prostaglandin receptors in large numbers of tissue samples. Binding of prostaglandin F 2 α, to preparations of microsomal membranes obtained from the ovaries of PMSG/hCG-treated rats served as the model system for this study. After incubating the membranes with [ 3 H]-PGF 2 α ± graded concentrations of PGF 2 α for 90 min at 22°C, dextran-coated charcoal was added to adsorb the free PGF 2 α during a 10 minute period. Centrifugation of the mixture for 3 minutes atop a 2.2M sucrose cushion resulted in the charcoal pelleting to the bottom while the [ 3 H]PGF 2 α-membrane complex remained at the interface. There was no further significant diffusion of PGF 2 α after centrifugation, enabling large numbers of samples to be handled within a short period of time. The binding data revealed both high and low affinity receptors: K d =5.8 nM and 77 nM with binding capacities of 360 and 4000 fmole/mg protein. Both PGE 1 and PGF 1 α partially displaced [ 3 H]PGF 2α from the binding sites whereas PGA 2 was without effect. Binding equilibrium was reached within 90–140 min. The results are compatible with those of investigators using other techniques.