We have prepared and characterized semisynthetic [GlyA1-3H]insulin. The preparation was carried out at specific radioactivities ranging from 1Ci/mmol to 44Ci/mmol. The largest quantity prepared in any one synthesis was 3.5 mCi. Chemical degradation showed that the label was in its expected position in the molecule. The semisynthetic product behaved authentically on reversed-phase h.p.l.c. and radioimmunoassay. It gave the expected profiles of biological activity as regards depression of blood sugar concentration in rats and stimulation of conversion of glucose into lipid in isolated rat fat-cells. We discuss some applications for which this tracer would be particularly suited. An expanded version of this paper, containing full experimental details of the semisynthesis and characterization of [GlyA1-3H]insulin, has been deposited as Supplementary Publication SUP 50129 (30 pages) at the British Library (Lending Division), Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1985) 225, 5.

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