Purification of [125I]-vasoactive intestinal peptide by reverse-phase HPLC.
VIP was labeled with sodium 125iodide, and 125I-VIP was purified by reverse-phase high performance liquid chromatography. Optimal separations of 125I-VIP and unlabeled VIP were obtained using two C18-Novapak columns in series and a gradient of acetonitrile in triethylamine phosphate for elution. The specific activity of the 125I-VIP was 1.99 +/- 0.21 Ci/mumole, approaching the maximum specific activity of monoiodinated VIP (2.26 Ci/mumole). Radioimmunoassay and radioreceptorassay for VIP were more sensitive (2.6-fold, and 2.5-fold, respectively) using 125I-VIP purified by HPLC compared to 125I-VIP obtained from an open-end cellulose column. These results demonstrate the advantage of preparing purified 125I-VIP by HPLC for the accurate assay of VIP and VIP-receptors in tissues and biological fluids.
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D007457:Iodine Radioisotopes; D011863:Radioimmunoassay; D011956:Receptors, Cell Surface / Q000032:analysis; D018005:Receptors, Vasoactive Intestinal Peptide; D014660:Vasoactive Intestinal Peptide / Q000302:isolation & purification*
D007457:Iodine Radioisotopes; D011956:Receptors, Cell Surface; D018005:Receptors, Vasoactive Intestinal Peptide; D014660:Vasoactive Intestinal Peptide