A scintillation proximity assay for human interleukin-5 (hlL-5) high-affinity binding in insect cells coexpressing hlL-5 receptor alpha and beta subunits

The high-affinity receptor for human interleukin-5 (hIL-5) is composed of a lpha and beta subunits. A baculovirus expression system was established in Sf9 cells capable of expressing hIL-5 receptor alpha and beta subunits simu ltaneously. By using wheat germ agglutinin (WGA)-coated scintillation proxi mity assay (SPA) beads to capture I-125-labeled hIL-5-bound Sf9 cells, a SP A was developed and used to measure hIL-5 high-affinity binding. The hIL-5 receptors expressed in the Sf9 cells represented a single class of high-aff inity binding sites with a dissociation constant (K-d) of 0.24 nM and a den sity of 2.95 x 10(5) sites/cell. This is the first study in which the high- affinity K-d value similar to that for hIL-5 binding to human eosinophils w as achieved using a recombinant expression system. The SPA compared favorab ly with the filter binding assay with regard to various binding parameters. We also found that several lectins, when coated on SPA beads, were even mo re effective than WGA-coated SPA beads for capturing the insect cells. We c onclude that the baculovirus expression system was highly efficient in prod ucing the high-affinity hIL-5 receptors and that the SPA was a simple and s ensitive assay that could be readily adapted into a high-throughput screeni ng format. The SPA described here could be a prototype for binding assays f or other multimeric receptors. (C) 1999 Academic Press.