AN IMPROVED ASSAY FOR URINARY LTE(4)

An improved method for the measurement of urinary LTE(4) is described, based on the combined use of solid phase extraction (SPE)-HPLC-enzyme -immunoassay (EIA). This allows the use of homologous radioactive trac er in easily measurable amount for HPLC retention time evaluation and recovery estimation. Recovery linearly correlated with the total amoun t of LTE(4) present in the extracted sample, indicating the existence of a carrier effect. Identification of immunoreactivity in HPLC fracti ons as LTE(4) was based on parallel dilution assay and confirmed by th e observable isotopic separation between tritium labeled LTE(4) and im munoreactive LTE(4). Critical selection of urine sample size, on the b asis of creatinine content, together with efficient purification by SP E, resulted in total absence of aspecific immunoreactivity in fraction s surrounding those associated with LTE(4). Urinary LTE(4) was measure d in normal subjects and in cirrhotic patients, where an increased LTE (4) excretion has been reported. The method described fulfils the crit eria of specificity, sensitivity and accuracy necessary for a potentia l successful use in the study of sulfido-peptide leukotrienes formatio n in normal and pathological conditions.