Comparative study for ultrastructural localization of intracellular immunoglobulins using peroxidase conjugates
Wolf D. Kuhlmann, S. Avrameas, T. Ternynck
Medizinische Universitätsklinik Heidelberg and Institut für Nuklearmedizin
Deutsches Krebsforschungszentrum Heidelberg, Germany
Unité d’Immunocytochimie, Département de Biologie Moléculaire, Institut Pasteur Paris, France
Various experimental conditions were tried to stain intracellular IgG globulins within popliteal lymph node cells by the use of specific antibodies and their Fab fragments labelled with peroxidase following two different procedures. Fixation with 4% formaldehyde/24 hours, 1% formaldehyde and 1% glutaraldehyde/1-1.5 hours and 1-1.5% glutaraldehyde/1-1.5 hours provided satisfactory ultrastructural conservation of the tissues. None of these fixatives appeared to denature IgG globulins. Best results were obtained by using frozen sections prepared from well-fixed specimens. Apparently, the most important points are tissue fixation and sampling, above all other experimental steps. Conjugates with molar ratios of antibody or Fab fragment to peroxidase being one, seem more effective, although no significant differences were noted between peroxidase conjugates of whole antibody molecules and Fab fragments.