A non-cannabinoid immunogen used to elicit antibodies with broad cross-reactivity to cannabinoid metabolites

A benzpyran derivative was linked to the lysines of bovine thyroglobulin (BTG) where 69% of the available lysines were modified. This derivative was designed to elicit antibodies that were directed towards the conserved epitopes of cannabinoid metabolites that appear in urine. Polyclonal antibodies from sheep and goats and murine monoclonal antibodies were generated using this immunogen. The cross-reactivity of the antibodies was compared with antibodies generated from the more traditional phenolic-linked or 9-linked immunogens. An ELISA assay was developed using delta 9-11-nor-9-carboxy-tetrahydrocannabinol (COOH-THC) to obtain a standard curve. The antibodies generated using the benzpyran immunogen showed an average of two to three times higher cross-reactivity towards 11-OH-delta 9-THC, 8 beta-OH-delta 9-THC, 8 alpha-OH-delta 9-THC, 11-OH-delta 8-THC, and 8 beta,11-di-OH-delta 9-THC than antibodies that were generated by traditional cannabinoid immunogens. The selectivity of the benzpyran-elicited antibodies was also compared with antibodies derived from traditional immunogens using clinical urine samples that were confirmed for cannabinoids by GC/MS. The total cross-reactive cannabinoid values obtained with the benzpyran-elicited antibodies were 49% higher than the values obtained using the traditional immunogen structures. The benzpyran immunogen-induced antibodies exhibited the same low cross-reactivity for non-structurally related compounds as antibodies derived from traditional immunogens. The novel benzpyran immunogen used in this study is the first non-cannabinoid immunogen used to generate cannabinoid-selective antibodies and demonstrates the usefulness of such a structure in developing broadly cross-reactive cannabinoid antibodies.