Development of a novel enzyme-linked immunosorbent assay for blood and urinary eosinophil-derived neurotoxin: A preliminary study in patients with bronchial asthma

Background: Eosinophil-derived neurotoxin (EDN), also called eosinophil pro tein X (EPX), has been suggested to be a useful marker of eosinophilic infl ammation. However, no commercial enzyme-linked immunosorbent assay (ELISA) kit for EDN is available yet. Methods: EDN was purified from pooled urine f rom healthy male volunteers. Polyclonal and monoclonal anti-EDN antibodies were subsequently raised, and a sandwich ELISA for EDN was established. EDN levels in serum, plasma and urine from asymptomatic patients with bronchia l asthma were measured by the ELISA method. Some of the blood samples were also measured by a commercial radioimmunoassay (RIA) kit. Results: The ELIS A method detected human EDN with a minimum detection limit of less than 0.6 2 ng/ml and did not cross-react with other eosinophil granule cationic prot eins including eosinophil cationic protein. The intra- and interassay coeff icients of variation of the ELISA method ranged from 2.6 to 3.6% and from 6 .5 to 9.4%, respectively. Good linearity was observed with serially diluted different samples, and the recoveries of the purified EDN added to serum s amples ranged from 85 to 110%. Median EDN concentrations in serum (36.9 vs. 19.1 ng/ml), plasma (23.0 vs. 14.5 ng/ml) and urine (118.2 vs. 72.1 mu g/m mol Cr) were significantly raised in asymptomatic asthmatic patients compar ed with healthy control subjects. EDN levels in serum, plasma and urine fro m the patients significantly correlated with the number of peripheral blood eosinophils, but not total serum IgE levels. A significant relationship be tween EDN values measured by the EPX-RIA kit and the EDN-ELISA method was o bserved. Conclusions: We have developed a novel efficient ELISA method to s pecifically measure blood and urinary EDN, which may be useful to study the role of eosinophils in allergic diseases including bronchial asthma. Copyright (C) 2000 S. Karger AG, Basel.