Radiochlmica Actaculty. Our results, which can be observed in the Fig. 3 and 4, have some interesting consequences. In the zone of the maximal yield -(I _ /Thyr ratio approximately equal to 2) -the iodination degree is 1.7. In these conditions, a 50% of the radioactivity of the hydrolysed product correspond to diiodinated tyrosine; thus, considering that thyrotrophin ha» 10 tyrosine residues, it can be calculated that 4.25 % of those residues bear two iodine atoms. The same type of calculation shows that for a (I~/Thyr) ratio of 15.8 (the maximal ratio in our experiments), this value increases to 23.4%. On the other hand, bearing in mind that such a relatively high proportion of diiodinated tyrosine residues may eventually alter the behaviour of the labeled hormone, it seems advisable to make a compromise between the yield of the labeling process and the proportion of diiodinated tyrosine residues. In this sense, it can be calculated that, for a fairly high yield such as 70%, which has been obtained with a (I~/Thyr) ratio of 0.34, the iodination degree is 0.24 and only 0.2% of the tyrosine residues bear two iodine atoms. Thus, we think that these last conditions allow the obtention of a labeled hormone which is better suited for its subsequent utilization.From the foregoing discussion, it can be deduced also that the optimum ratio between the number of the iodide atoms which are present and the number of molecules of the polypeptide is independent of the relative proportion of Na 131 I in the total quantity of iodide. Thus, the conclusions of the present work are also valid for labeling with higher specific activities. Even though we studied only two molecules, the size and complexity of which are quite different, it should be mentioned that in some preliminary experiments we observed that in the labeling of human seroalbumin (molecular weight approximately equal to 69,000), it is possible to find a similar effect as that demonstrated in the case of thyrotrophin. These facts induce us to think that these phenomena may occur generally in any la-beling process of a polypeptide or a protein. For this reason we believe that similar studies as those reported here should be carried out for every particular polypeptide. This study should include basically the following points: 1. The determination of the optimum ratio between the number of iodide atoms and the number of polypeptidic molecules which are present in the reaction.2. The determination of the relative proportion of MIT and DIT residues with respect to the total number of polypeptidic molecules. This value is interesting because, as has been shown in the case of angiotensin [2], the presence of two iodine atoms in the same tyrosine residue, determines a significant decrease of the biological activity of the molecule. It seems possible that similar effects may be observed in other polypeptides having a given biological activity. Our results demonstrate also that it should be born in mind that the sole obtention of a high labeling yield is not a sufficient guara...

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