Preparation and Identification of Specific and High-Affinity Monoclonal Antibodies against Morphine

A C6-hemisuccinate derivative of morphine was prepared and conjugated to bovine serum albumin and ovalbumin. High titer antibody producing spleen cells were removed and fused with myeloma cells of SP2/0 origin. Using a conventional immunization protocol generated twenty-six stable murine monoclonal antibodies (MAbs) producing cell lines to morphine. The donor mouse produced antiserum with a high titer of 1/640,000. Twelve MAbs were selected for further characterization since they showed high sensitivities (53 pg/well to inhibit 50% of the tracer) in improved group-selective immunoassay (IGSI). The assay, which maintains high sensitivity, high precision, and a wide range of optical density (OD) values, was developed using the conjugate M-6-S-OVA to screen and characterize the anti-morphine MAbs. After four successive limiting dilutions, antibodies produced by 12 clones had high affinities ranging from 109 to 1010 M-1. These clones were found to be of IgG class and IgM class with κ and λ light chain. Subcla...