Predominance of Theta-Positive Lymphocytes in Gut-Associated and Peripheral Lymphoid Tissues of Newborn Mice

Morphologic and isotope-labeling studies on newborn mice have not only suggested a substantial thymocyte production (1) and peripheralization (2), but have also provided direct evidence for the thymic origin of a majority of lymphocytes in mesenteric lymph nodes and Peyer's patches (3, 4). The presence of θ-alloantigen on thymocytes of mice (5) has been utilized to evaluate the relative proportions of thymus-derived cells in lymphoid tissues (6). The values obtained by the use of cytotoxic anti-θ serum (7) are, however, significantly lower than those suggested by morphologic and isotope-labeling studies (3, 4). We have evaluated the relative numbers of θ-positive cells in the lymphoid tissues of newborn mice by indirect immunofluorescent staining (8). The results indicate that most of the lymphocytes in lymph nodes and Peyer's patches of mice are θ-positive. Materials and Methods. Anti-θ and control anti-θ reagents (anti-θ, control reagent, respectively) were prepared by immunizing AKR mice with C3H mouse thymocytes. Approximately 108 thymocytes in incomplete Freund's adjuvant were injected intraperitoneally at 2-week intervals. One week after the fifth injection, an intraperitoneal injectionof 0.25 ml of complete Freund's adjuvant was given. A week later the ascites was tapped and the pooled fluid inactivated at 56°C and passed through 3.0-μ Millipore filter.Anti-θ was obtained by absorbing nonspecific anti- bodies according to the following procedure:one-half of the fluid was incubated at room temperature with θC3H-negative AKR thymocytes by using cells from one thymus/ml of fluid for each of the three incubations. Finally the fluid was incubated with θC3H-negative AKR lymphocytes by using cells from one mesentericlymph node/ml of fluid. Control reagent was obtained from the remaining half of the fluid by absorptions against θC3H-positive, Charles River albino/ICR mouse thymocytes (three times), and θC3H-negative, AKR lymph node cells (one time). Anti-θ and control reagents were stored at -20°C.