Synthesis of hapten, production of monoclonal antibody, and development of immunoassay for ribavirin detection in chicken.

Ribavirin (RBV) is an effective antiviral drug, whose use is prohibited in animal husbandry worldwide. In this work, a novel immunizing hapten of RBV, named Hapten 4, was designed by comparing the conformational and electronic properties of RBV and haptens based on computational chemistry. Hapten 4 was synthesized and conjugated with carrier proteins to produce monoclonal antibody (mAb). The obtained mAb 4C3 for RBV exhibited an IC50 value of 6.24 ng/ml in an indirect competitive enzyme-linked immunosorbent assay (icELISA) and displayed no cross-reaction with five other antiviral drugs, including amantadine. The applicability of the developed icELISA was verified in chicken, with a calculated limit of detection of 4.23 µg/kg. The recoveries in spiked chicken were 79.2%-107.3% with a coefficient of variation less than 15.9%. The results indicated that the produced antibody from the new hapten was reliable and would be useful for RBV screening in chicken. PRACTICAL APPLICATION: RBV is a broad-spectrum antiviral drug, which is commonly used illegally in poultry farms. A high-affinity mAb 4C3 against RBV was produced and used to develop icELISA with acceptable sensitivity and accuracy. The constructed icELISA has excellent performance for detecting RBV residues in chicken.