Preparation of 125I-(Tyr 3)- and 125I-(Tyr 11)-neurotensin for radioimmunoassay

Abstract

The chloramine-T method for radioiodination of neurotensin for radioimmunoassay was studied. As conventional procedures produced heterogeneous preparations, labelling was performed with a low amount of chloramine-T (1.8 nmol) in the presence of excess of peptide (6 nmol). Purification and complete separation of labelled from unlabelled peptide was obtained by ion-exchange chromatography on SP Sephadex C-25. Four labelled components were identified by isoelectric focusing, enzymatic cleavage and studies of immunoreactivity. The two components representing monoiodinated preparations labelled at Tyr 3 or Tyr 11 could be isolated. Depending on the binding site of the particular antiserum the appropriate tracer could be selected for use in the radioimmunoassay. The specific radioactivities were high (2303 (2137–2407) μCi/nmol and 1927 (1608–2307) μCi/nmol (median and range)) and the stability of the label and the reproducibility of the procedure was good.