Cellular localization of pneumolysin in various types of pneumococci was studied by the immunoelectron microscopy. Cells of pneumococcal type 19F, type 8~10 and 8-CB, and non-capsulated Rx1 strain were fixed in paraformaldehyde and treated with rabbit anti-pneumolysin serum, followed by goat-anti-rabbit IgG labeled with colloidal gold. The cells were then reacted with chrome osmium fixative, dehydrated, embedded in epoxy resin sectioned and examined with an electropn microscope. The cellular localization of pneumolysin was similar in all 4 strains examined. No labeling was found on the exterior of the cell, either cell wall or capsule; indicating the absence of ply in these locations. In addition, totally intact cells had no bound intracellular gold particles, due to the inability of the antibodies to penetrate the dense cell wall matrix. Pneumolysin labeling was associated with a structure adherent to the cell membrane and extending into the cytoplasm. Gold particles were seen on the periphery of the adherent structures inside lysed cells. The amount of this material seems to correlate with the measured pneumolysin activity; Rx1 and type 8-10 strains that showed higher pneumolysin activity, demonstrated more adherent material and labeled antibodies than type 8-CB that showed a low pneumolysin activity.
