Abstract
Monoclonal antibodies were prepared against ceftiofur using its hydrolyzed form, desfuroylceftiofur, as the hapten. Desfuroylceftiofur was prepared in situ and conjugated directly to maleimide-activated carrier proteins bovine serum albumin and keyhole limpet hemocyanin (KLH). Balb/c mice were immunized with the desfuroylceftiofur−KLH conjugate (KLH-SMCC−desCef). Splenic lymphocytes from these mice were fused with SP2/0 myeloma cells to produce hybridomas. Two stable monoclonal antibodies, Cef-68 and Cef-116, of subclasses IgG2a and IgG1, respectively, were isolated. An indirect competitive inhibition enzyme-linked immunosorbent assay (ciELISA) was developed for the quantitative detection of ceftiofur in liquid samples using a heterologous assay system to provide a more sensitive ELISA. Detection limits for ceftiofur were 32 and 0.3 ppb for the two clones Cef-68 and Cef-116, respectively. The ciELISA described here provides a sensitive assay for the detection of ceftiofur. Cross-reactivity studies with a variety of related cephalosporins and penicillins indicated that only a limited number of the cephalosporins and none of the penicillins that were tested competed in the ciELISA. Keywords: Ceftiofur; immunoassay; monoclonal antibody; residue analysis
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