The raising and characterisation of antibodies to salicylate

Polarisation fluorescence was used to assess the antibody response in sheep immunised with 4-aminosalicylic acid coupled to keyhole limpet haemocyanin and to develop a polarisation fluoroimmunoassay for subsequent clinical use. The fluorescent label was prepared by coupling 4-aminosalicylic acid to fluorescein isothiocyanate. All immunised sheep produced antibodies and a simple assay was developed using salicylic acid as the standard. The simple fluorescence polarisation approach enabled the rapid and direct study of the immuno-reactivity of a large number of pure chemical analogues of salicylic acid. Each was assessed for its ability to compete with the fluorophore-labelled analyte for antibody-binding sites, relative to salicylate, thereby enabling investigation of the steric and chemical characteristics important in the recognition and binding of a hapten to antibodies.