A general method for the production of 125I-labelled low-molecular-weight tracers for radioimmunoassay

A general method is described for the separation of low-molecular-weight 125I-labelled tracers from a chloramine-T labelling mixture, containing the starting material, the tracer, labelled by-products, free radioiodine and reagents used in the labelling procedure. Small molecules like tyrosine, thyronine and steroids as well as their tyrosine methyl ester conjugates are adsorbed on Sephadex LH-20 dextran gel. The introduction of iodine atoms increases the elution volume and the distribution coefficient in proportion to the number of iodine atoms per molecule. Organic solvents like ethanol are also adsorbed on dextran gel, which results in desorption of the starting material or its iodine-substituted derivatives. The elution order follows the number of iodine substituents per molecule, i.e. unsubstituted starting material, mono-, di-, tri- and tetra-substituted molecules. Using a water/ethanol binary eluent in which water can be considered as inert diluent, the elution is governed by the ethanol concentration. The finding that the logarithm of the distribution coefficient of the solutes is linearly correlated with the logarithm of the ethanol concentration permits an optimization of the separation. (author)