Reactivity of monoclonal antibodies EG1 and EG2 with eosinophils and theirgranule proteins

Use of the murine monoclonal antibodies EG1 and EG2 has been based on the a ssumption that EG2 recognizes activated eosinophils. We examined the reacti vity of EG1 and EG2 with eosinophil cationic protein (ECP), eosinophil-deri ved neurotoxin (EDN), and stimulated and nonstimulated eosinophils from nor mal donors, By radioimmunoassay, EG1 recognized only ECP, whereas EG2 recog nized both ECP and EDN, By Western blot, EG1 reacted with ECP, EG2 reacted with both ECP and EDN, but EG2 could not distinguish between lysates of sti mulated and nonstimulated eosinophils. By immunofluorescence, EG1 and EG2 a t 20 mu g/mL stained 95-100% of nonstimulated eosinophils, regardless of fi xative; EG1 and EG2 at 0.1 mu g/mL stained 61-90% of acetone- and paraforma ldehyde-fixed and only 5-21% of methanol-fixed nonstimulated eosinophils, T hus, the reactivity of EG1 and EG2 with eosinophils depends on the method o f fixation and antibody concentration; and EG2, in contrast to previous rep orts, cannot reliably discriminate between resting and activated eosinophil s.