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Alkaline phosphatase from calf intestine was conjugated to thyroxine using cyanuric chloride, glutaraldehyde and 1-cyclohexyl-3(−2 morphoinoethyl)-carbodiimide as coupling reagents. The conjugates produced were studied for incorporation of thyroxine, retention of enzyme activity and immunoreactivity in a thyroxine radioimmunoassay system against a range of thyroxine antisera. The use of these conjugates in thyroxine enzyme immunoassay was studied using second-antibody separation. Comparison of the results with radioimmunoassay data was satisfactory and indicates the potential of these conjugates for thyroxine immunoassay. The use of cyanuric chloride as a coupling reagent was unsatisfactory, but good results were obtained with glutaraldehyde. Although satisfactory immunoreactivity of carbodiimide conjugates was observed in a radioimmunoassay system, these results were not reflected in the enzyme immunoassay systems studied.

The Preparation and Characterization of Alkaline Phosphatase—Thyroxine Conjugates and Their Use in the Enzyme Immunoassay of Thyroxine