EOSINOPHILS ISOLATED WITH 2 DIFFERENT METHODS SHOW DIFFERENT CHARACTERISTICS OF ACTIVATION

Eosinophils can be isolated from a mixed suspension of granulocytes by different procedures. We compared functional responses of human eosin ophils purified according to two different principles: (1) an fMLP-ind uced difference in specific gravity between eosinophils and neutrophil s and (2) selective removal of neutrophils by means of immunomagnetic beads coated with CD16 mAb. The results showed that eosinophils isolat ed with the CD16 beads method have a higher capacity to synthesize pla telet activating factor (PAF) after stimulation with serum-treated zym osan (STZ) than eosinophils purified with the fMLP method. Binding of STZ and subsequent activation of the respiratory burst were also incre ased in CD16-isolated eosinophils. Furthermore, eosinophils isolated w ith the CD16 beads showed stronger chemotactic responses towards C5a a nd PAF. The difference in activity of these eosinophil preparations mi ght be explained by a loss of the more active cells during the isolati on with the fMLP method: only 30-60% of the eosinophils were recovered with this method, in contrast to a recovery of more than 95% with the CD16 beads method. Indeed, this 'lost' population of eosinophils, sub sequently purified with CD16-coated beads, had a higher respiratory bu rst activity. The alternative explanation i.e., an enhancement of eosi nophil function by the beads method, appeared not to be valid, because repurification of fMLP-isolated eosinophils in the presence of fresh neutrophils and CD16-coated beads did not change the reactivity of the eosinophils. We conclude that the fMLP method leads to selective puri fication of eosinophils with a resting (or 'unprimed') phenotype.