Purification of mouse alpha-1-fetoprotein and preparation of specific peroxidase conjugates for its cellular localization
Wolf D. Kuhlmann
Immunocytochemistry SFB 136 and Institut für Nuklearmedizin, DKFZ Heidelberg, Germany
Mouse alpha-1-fetoprotein (AFP) was isolated from amniotic fluid by immunoadsorbent columns and preparative electrophoresis. Specific immune sera against AFP were obtained by hyperimmunization of rabbits from which pure antibody molecules were isolated by the use of immunoadsorbents. The isolated antibodies were subsequently conjugated with horseradish peroxidase (HRP). Cellular localization of AFP in fetal liver were done by direct and indirect immunoperoxidase staining methods. Several fixation schedules were tried. Both localization of AFP and tissue preservation were satisfactory when embryonic liver blocks were fixed with ethanol-acetic acid or with formaldehyde-picric acid-glutaraldehyde mixture. Inhibition of endogenous peroxidase activity was obtained by incubation of the sections in diluted hydrogen peroxide solutions.