Facile in vitro method for screening inhibitors of IgE binding to mast cells.
A method for rapidly testing large numbers of chemical structures as potential modulators of the interaction between immunoglobulin E (IgE) and its specific receptors on rat peritoneal mast cells is described. IgE, isolated from the ascitic fluid of a transplantable rat IgE immunocytoma, is labeled with iodine-125 under mild conditions employing the Bolton--Hunter reagent. The antibody is incubated with mixed periotoneal cells at 37 degrees, and the cell-bound IgE is separated from unbound label by sedimentation through an 8% sucrose--polymer solution in microsediment tubes. Optimal conditions for the interaction of 3 nM IgE with 3 X 10(5) mast cells in 150 mul are: incubation time, 2 hr; pH, 6.5--7.0; and ionic strength, equivalent to 150 mM NaCl. Mixed peritoneal cells bind IgE with an affinity equal to that of purified mast cells. Human IgE pentapeptide III and several antiallergic agents do not compete with rat IgE in this assay.
D000818:Animals; D001202:Ascitic Fluid / Q000166:cytology; D001665:Binding Sites; D006863:Hydrogen-Ion Concentration; D006967:Hypersensitivity / Q000188:drug therapy; D007073:Immunoglobulin E / Q000378:metabolism* / Q000494:pharmacology; D007128:Immunoglobulin Fragments / Q000494:pharmacology; D066298:In Vitro Techniques; D008407:Mast Cells / Q000187:drug effects / Q000276:immunology*; D009194:Myeloma Proteins / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D051381:Rats
D007128:Immunoglobulin Fragments; D009194:Myeloma Proteins; D007073:Immunoglobulin E