Egg envelope organisation in the icefish Chionodraco hamatus

Abstract

Antarctic teleosts perform their physiological activities at constant subzero temperatures. We previously described the gross morphology and the biochemical composition of the Antarctic teleost Chionodraco hamatus eggshell. In this work, we investigate thoroughly the chorion ultrastructure, showing a previously unknown external layer, and describe the preparation and use of monoclonal antibodies against eggshell proteins of this species. The main chorion polypeptide at 46 kDa was purified by preparative electrophoresis and used as immunogen in mice. After spleen-myeloma fusion, hybdridomas were screened by immunoblotting against eggshell homogenates, and two of the most interesting hybridomas were cloned by limiting dilution, and established in culture: CHE-1 (IgG) and CHE-5 (IgM). They stained intensely the eggshell in indirect immunofluorescence and in immunoelectron microscopy. The CHE-5 localisation on thin sections by immunogold staining was peculiar for the various vitelline envelope layers. By western blot analysis of eggshell, CHE-1 recognised two polypeptides at 46 kDa and 92 kDa, whereas CHE-5 recognised a single polypeptide at 92 kDa.