We describe here an efficient procedure for the precise quantitation of leukotriene E4 (LTE4) in a small volume of urine, which was achieved mainly by the use of an Empore extraction disk cartridge. After addition of [3H]LTE4 to 2 ml of urine, an Empore C18 cartridge was used for initial extraction of the urine, which resulted in the extraction of LTE4 in a small volume of solvent. The eluate could then be injected onto a high-performance liquid chromatography column without further concentration. After separation by high-performance liquid chromatography, LTE4 was extracted from the effluent using an Empore C18 cartridge. The concentration of LTE4 was subsequently quantified by enzyme immunoassay. LTE4 can be recovered from urine with sufficient efficiency (69.9+/-4.7%, mean+/-S.D., n=101). The coefficient of variation of the assay procedure was less than 10%. When urine was spiked with different amounts of LTE4, the recovery of LTE4 added to the urine specimen was less than 120%. The concentration of LTE4 in urine from normal healthy subjects was 48.0+/-15.3 pg/mg creatinine (n=15).