Localization of Antibodies by Electron Microscopy in Developing Antibody-Producing Cells

Abstract

The development of antibody-producing cells in thc early stagcs of the sccondary or hypcrimmune response has becn studied with the electron microscope in lymph nodes of adult chinchilla rabbits immunized with fcrritin or apofcrritin. The intraccllular distribution of antifcrritin antibodies was determined in the lymph node cells at 1 to 5 days after a booster injcction, employing the labelling technique prcviously used by the authors (12) to dcmonstratc the localization of antibodies in mature plasma cells. Antibodies were first detected at 48 hours in blasts; i.e., cells which have a poorly dcveloped endoplasmic reticulum and a cytoplasm filled with many ribosomes grouped in clusters. The label was subscquently found in forms intermediate bctwccn blasts and plasma cells (plasmoblasts, immature plasma cells), in which the cndoplasmic reticulum appeared progressivcly more dcvclopcd. Antifcrritin antibodies wcrc also found in cells in mitosis. In all the above ccll types, antigcnantibody precipitates were consistently found in the perinuclear space and in the cisternae of the granular endoplasmic rcticulum, from an early stage in the development of thc latter. Evidcncc was also obtained for thc presence of antibody in the Golgi area. The results arc discussed in relation to the possible cellular sites of antibody synthesis.In a previous paper (12) de Petris, Karlsbad, and Pernis described the subcellular distribution of antiferritin antibodies in the typical plasma cells present in lymph nodes of hyperimmunized rabbits on the 4th and 5th day after a booster injection of antigen. The antibodies (marked with ferritin) appeared to be localized in the perinuclear space and cisternae of the endoplasmic reticulum, and possibly also in the Golgi region.We have now extended this research to the early stages of the immune response, namely to the study of the cells present in the lymph nodes at 24 to 72 hours after restimulation. These early stages are characterized by active cellular division, accompanied by a series of cellular transformations entailing marked ultrastructural changes. In an effort to throw light on the cellular aspects of antibody synthesis, we have tried to correlate the beginning and further development of the antibody-synthesizing activity, as deduced from the initial and subsequent intracellular distribution of labelled antibodies, with the morphological changes which occur in the antibody-producing cells and eventually lead to their full differentiation into plasma cells.The technique employed for labelling the antibodies was essentially the same as that used in the previous investigation (12): it consisted in incubating ferritin with lymph node cells of animals immunized with either ferritin or apoferritin, after suitable treatment aimed at allowing the penetration of the marker (ferritin) into the ceils. The localization of antibodies was deduced from the distribution of ferritin-antiferritin microprecipitates.Preliminary communications on this subject 759

Antigens in Immunity

“…Since we first drew attention to the fact that antibody-forming cells need not contain detectable antigen (38), this finding has been confirmed several times (5)(6)(7)39). Again, in the present ultrastructural study no evidence for passage of labeled antigen into plasma ceils was obtained but this finding is not a very telling one as it would have taken some thousands of molecules of antigen per tell to have caused convincing plasma cell labeling.…”

Ultrastructure of the Primary Immune Response

“…The ultrastructure of developing and mature plasma cells has been studied by a number of investigators [38,11,12,16]. The organelles and ultrastructural features of the cells described in this study are quite similar to those previously reported, except for the micropinocytotic processes and the microtubules observed here in close associa tion with the nuclei of many of the cells.…”

Ultrastructure of Cells Present in Lymph Nodes During the Development of Contact Sensitivity

“…It is of interest that the immunoblasts induced by the appli cation of a chemical sensitizing agent appear on the basis of this and earlier studies (25) to give rise to small lymphocytes. These immunoblasts arc, however, indistinguishable in the electron microscope from the immunoblasts which are seen in the early stages of the production of a humoral antibody (9,19). In the pro duction of humoral antibodies these cells give rise to plasmoblasts (with increasing amounts of endoplasmic reticulum), which them selves develop into plasma cells.…”