Rapid purification of radioiodinated peptides with Sep‐Pak® reversed phase cartridges and HPLC

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Abstract
A simple, rapid method is described for the purification of radioiodinated peptides for use in radioimmuno‐ and in radioreceptor assays. Iodinated reaction mixtures are applied directly onto Sep‐Pak® disposable, reversed phase cartridges equilibrated with phosphate buffer. Unreacted 125‐iodide and other non‐peptide reaction components are eluted with buffer. The peptide fraction is then eluted with 70% buffer:30% acetonitrile. The peptide fraction is further purified by reversed phase high pressure liquid chromatography to separate the native peptide and the mono‐ and diiodo‐derivatives. In this study the method is used to prepare 125‐iodide‐labeled monoiodo‐leucine enkephalin and monoiodo‐angiotensin II, which are free of the parent peptides and diiodo‐derivatives and are of maximum obtainable specific radioactivity. The usefulness of these labeled peptides in radioimmuno‐ and radioreceptor assays is demonstrated by their binding to specific antibodies and receptors, respectively.