The immune response in the rat to Streptococcus pneumoniae type 3 and type 4 capsular polysaccharide. Detection by double immunocytochemical staining of antibody-containing cells in situ and ELISA.

Two different methods have been used to study immune responses in the rat to Streptococcus pneumoniae type 3 and type 4 capsular polysaccharides (PPS). First, for simultaneous detection of the specificity and isotype of anti-PPS antibody-containing cells (ACC) in cryostat sections of lymphoid tissue, a double immunocytochemical method was developed. This method is a combination of a three-step immunoperoxidase method to demonstrate specific anti-PPS ACC as bright red cells and a two-step immunophosphatase method to detect the isotype of ACC as blue cells. Double positive cells appear violet. Using this staining procedure, the detection of antigen was also possible. Second, to study the anti-PPS response in serum, an ELISA procedure was modified. In this ELISA, polyvinylchloride microtiter plates are coated directly with type-specific pneumococcal polysaccharide. After intraperitoneal (i.p.) immunization of rats with PPS-3 or PPS-4, both antigen (PPS) and specific ACC could be detected. Specific ACC were found in the spleen and mesenteric lymph nodes. In the spleen, the specific ACC were found in the red pulp, marginal zone, outer PALS, and follicles. Most of these ACC were IgM-positive and to a lesser extent IgG-positive and IgA-positive. However, specific ACC in mesenteric lymph nodes were predominantly of the IgA isotype, with only few IgM or IgG positive cells. The anti-PPS response in serum, as measured by the ELISA, consisted mainly of IgM antibodies with small amounts of IgG and IgA. Both methods were found to be valuable in studies of immune responses against bacterial polysaccharides.