DETECTION OF HALOFUGINONE RESIDUES IN CHICKEN SERUM BY A MONOCLONAL-BASED IMMUNOASSAY AND HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

Halofuginone (Hal) is a coccidiostat used worldwide as a feed additive at a level of 3 ppm in the feed of broiler chickens. The analytical m ethods described in the literature to determine the levels of Hal in l iver are very lengthy and time consuming. This study evaluated the use fulness of determining Hal in chicken serum with a competitive ELISA ( cELISA). Ifa 4-day withdrawal time could be determined by serum Hal le vels, the method would greatly improve on the high-performance liquid chromatography (HPLC) methods currently used for Hal detection in live r tissue. A modification of a previously developed HPLC method was use d to validate the cELISA analysis of Hal in chicken serum. A serum mat rix effect that afforded a higher sensitivity by the cELISA for the de tection of Hal in chicken serum than in assay buffer or in highly dilu ted serum was observed. The sensitivity of the cELISA method improved when used in more concentrated serum. The chicken serum samples were e valuated by cELISA using a standard curve obtained in control chicken serum diluted two-fold with assay buffer. Incurred levels of Hal in br oiler chickens fed Hal-HBr-treated feed were detected in serum at with drawal times of 2 and 6 h. At and after 24 h, the residues were not de tected by immunoassay with a detection limit of 0.52 ppb or by HPLC wi th detection limit of 0.86 ppb. The instability of Hal in acidified se rum and its potential for methanolysis in the HPLC method were overcom e by using the cELISA methodology. Although the determination of Hal i n chicken serum by immunoassay is fast, requiring no clean-up steps, c hicken serum cannot be used to determine the required 4-day withdrawal time in broiler chickens because of the lack of residues in the serum at and after 24 h.