Improved 1251 Radioimmunoassay for Cotinine by Selective Removal of Bridge Antibodies

We describe an 1251-based RIA for cotinine, the major metabollte of nicotine. The slope of the dose-response curve was quite shallow (6-8 % change in binding per doubling dose), resulting in between-assay CVs of 15 to 20%. This effect occurred because the radioligand formed by linking a cotinine derivative to tyramine manifested greater affinity for the anti-cotinine antibodies than did cotinine itself. We ab-sorbed the serum with a derivative of nicotine coupled to the carrier protein via a chemical bridge similar to that used to form the cotinine/carrier protein immunogen. An AlA in which we used such absorbed serum showed a significantly in-creased slope of the dose-response curve (11-13 % change in binding per doubling dose), and between-assay CVS were only 6 to 8%. We suggest that this improvement results because absorption removes anti-bridge antibodies directe