Contribution of Methodological Artifacts to the Measurement of T3Concentration in Serum

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Abstract
Sources of artifactual error have been studied during the measurement of serum T8 using a commonly employed paper chromatographic method. Significant and variable artifactual elevation of the serum T3 concentration can result from solvents used in the method, from material eluted from the initial Dowex AG 50 resin or from Whatman chromatographic paper. The mechanism of this “blank” effect is not clear, but it is a problem common to all protein binding hormone assays. In addition, spillover of T4 into the T3 area of the paper chromatogram may occur, particularly if the serum extract is applied to wet paper. Since T4 displaces 2.5 times more labeled T3 from TBG than does an equivalent weight of T3, the effect of this T4 spillover will be magnified 2–3 times. The most important source of error is artifactual deiodination of T4 and the major radioactive product of the deiodination of 131I-T4 is iodide. However, a significant monodeiodination of T4 to T3 also occurs; over a several-month period, the radioactivity recovered as T3 after chromatography of purified 131I-T4 averaged about 0.7%. Such deiodination can be minimized by using wet paper, but the separation of T4 and T3 is less adequate under these circumstances. Presently available evidence would suggest that minimizing or correcting for these sources of artifact reduces the mean serum T3 concentration in euthyroid subjects to the range of 100–125 ng/100 ml.

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