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High-performance liquid chromatographic method for the determination of insulin synthesis in biological systems
G Calvaruso1, G Tesoriere, R Vento
1Istituto di Chimica Biologica, Università di Palermo, Italy.
Abstract:
This paper reports a two-step high-performance liquid chromatographic procedure which permits the study of the incorporation of [3H]leucine into insulin in biological systems. The first step of the procedure was size exclusion chromatography, performed on a GPC-100 column, which was eluted with 0.1 M KH2PO4-methanol (9:1, v/v). By this step the bulk of both protein and radioactivity was separated from tritiated insulin. The second step, which employs reversed-phase chromatography on an octadecylsilyl column, permits the separation of insulin from other contaminants by means of a linear gradient of acetonitrile. This simple and reproducible method was employed to test insulin synthesis in cultured human retinoblastoma Y79 cells.
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