Production of monoclonal antibody and development of enzyme-linked immunosorbent assay for kanamycin in biological matrices

Monoclonal antibodies (MAbs) against kanamycin were prepared by using a kan amycin-bovine gamma-globulin conjugate for the immunization of mice. Spleno cytes from BALB/c immunized mice were fused with P3X63Ag8U.1 myeloma cells. This resulted in two hybridoma cell lines. Fifty per cent inhibition conce ntrations (IC50) for the MAbs were 2 and 5 ng ml(-1). One MAb (IC50 = 2 ng ml(-1)) was named #22 and was used to develop quantitative assays for kanam ycin by means of an enzyme-linked immunosorbent assay (ELISA). The detectio n limit was 0.2 ng ml(-1) and the standard deviations were 0.2-4.4% for int ra-assay and 0.6-4.7% for inter-assay, respectively. The detection limits u sing peroxidase were 4 ppb in cattle milk, cattle plasma, cattle urine, swi ne plasma, swine urine and chicken plasma. Using the MAb #22 produced, a ra pid test kit based on an immunochromatographic method was developed. The de tection limits using the kit were 50 ppb in cattle milk, cattle plasma, cat tle urine and chicken plasma.