Isolation, immunochemical detection, and observations of the instability of vitellogenin from four teleosts

Abstract

AbstractVitellogenin was purified from plasma of estradiol‐17β‐treated cod (Gadus morhua) rainbow trout (Oncorhynchus mykiss), turbot (Scophthalmus maximus), and wolffish (Anarhichas lupus) by precipitation with EDTA:Mg2+, distilled water, and high‐performance ion‐exchange chromatography. Vitellogenin of high purity was obtained by precipitation followed by chromatography, as evaluated by an homologous antiserum developed for each species. The instability of vitellogenin demanded consistent low temperature and the use of protease inhibitor before blood sampling. When the necessary precautions were taken, vitellogenin from rainbow trout, turbot, and wolffish eluted as one regular peak during chromatography. Cod vitellogenin eluted as two peaks and these demonstrated identical migration patterns on SDS‐PAGE. The observed differences in stability between the four species suggest that isolation procedures should be modified according to the requirements for each species. Electrophoresis of plasma from treated fish revealed the presence of several smaller proteins, with a molecular mass around 50 kDa, that were considered to be vitelline envelope proteins. Other minor plasma proteins were immunoreactive to antisera, directed against vitellogenin and therefore judged to be fragments of degraded vitellogenin. © 1993 Wiley‐Liss, Inc.