Application of a monoclonal antibody-based enzyme-linked immunosorbent assay for the determination of ractopamine in incurred samples from food animals.

A monoclonal antibody-based ractopamine immunoassay has been applied to incurred samples from sheep and cattle. Results obtained by immunoassay were compared with those from high-performance liquid chromatography (HPLC). Three sets of sample extracts containing primarily unmetabolized ractopamine were analyzed. Correlation of HPLC with enzyme-linked immunosorbent assay (ELISA) for beef liver samples gave an r(2) = 0.98 despite rather low ractopamine concentrations (range 1.1-13.4 ng/mL, n = 6). Ractopamine concentrations in cow urine samples treated by solid phase extraction, to remove ractopamine metabolites, also showed a high correlation between the HPLC and the ELISA results (r(2) = 0.95, range 1.0-275 ng/mL, n = 61). In contrast, HPLC and ELISA analyses of ractopamine in sheep urine were not well-correlated (r(2) = 0.58, range 0.85-51 ng/mL, n = 34). When ractopamine conjugates in urine samples were hydrolyzed with hydrolytic enzymes, ELISA and HPLC methods were highly correlated [r(2) = 0.94 for sheep (range 123-10 554 ppb, n = 60) and an r(2) = 0.98 for cattle (range 14-8159 ppb, n = 62)]. Tissues contained only minute amounts of ractopamine, and after 7-day withdrawal periods, less than 1 ppb of free ractopamine was detected. Ractopamine was rapidly metabolized in both cattle and sheep. The difference in ractopamine concentration of urine samples before and after hydrolysis indicated that only 1-5% of ractopamine was excreted unmetabolized. Results from this study indicate that the monoclonal antibody-based ELISA could be useful for a sensitive, quantitative, or qualitative ractopamine screening assay.