Subcellular localization of the antigenic sites of relaxin in the luteal cells of the pregnant rat using an improved immunocytochemical technique

Abstract

Using an improved immunocytochemical technique, the present work has demonstrated the antigenic sites of relaxin in the luteal cells of the pregnant rat with preservation of the ultrastructural details. Tissues collected were fixed with either 4% paraformaldehyde (PFA), 4% PFA containing 0.3% glutaraldehyde (GLA) or Zamboni solution followed or not by 1% osmium. Ultrathin sections of the tissues embedded in epoxy resin were incubated with rat relaxin antiserum and antibody binding sites were visualized with protein A-gold. Among these fixatives, the use of 4% PFA-0.3% GLA, followed by osmium, allowed excellent preservation of fine cellular structure with maintenance of antigenic activity and immobilization of the antigen. The majority of the gold particles indicative of relaxin immunoreactivity were localized on the dense core of the membrane-limited granules, while a minor fraction was localized on the Golgi apparatus. The results suggest that the subcellular localization of relaxin can be detected after post-fixation with osmium and that the antigenic sites of relaxin exist mostly in the dense granules with a small fraction in the Golgi apparatus.