We validated a time-resolved fluoroimmunoassay (TR-FIA) using the lanthanide element europium as a non-radiological tracer for measuring luteinizing hormone (LH) in porcine peripheral blood. The dose-response curve of the reference standard ranged from 0.2048 to 50 ng/ml. Good parallelism was noted between the LH standard and plasma sample. The profile of LH throughout the estrous cycle, assessed by daily blood sampling, was consistent with the previous findings obtained by radioimmunoassays (RIAs). Moreover, the secretory patterns of pulsatile LH in the follicular phase and the preovulatory LH surge were also similar to those obtained in previous RIAs. We conclude that TR-FIA can be used to measure LH levels in porcine blood, with practical and convenient applications.