Secondary lymphoid tissue organogenesis requires tumor necrosis factor (TNF) and lymphotoxin α (LTα). The role of TNF in B cell positioning and formation of follicular structure was studied by comparing the location of newly produced naive recirculating and antigen-stimulated B cells in TNF−/− and TNF/LTα−/− mice. By creating radiation bone marrow chimeras from wild-type and TNF−/− mice, formation of normal splenic B cell follicles was shown to depend on TNF production by radiation-sensitive cells of hemopoietic origin. Reciprocal adoptive transfers of mature B cells between wild-type and knockout mice indicated that normal follicular tropism of recirculating naive B cells occurs independently of TNF derived from the recipient spleen. Moreover, soluble TNF receptor-IgG fusion protein administered in vivo failed to prevent B cell localization to the follicle or the germinal center reaction. Normal T zone tropism was observed when antigen-stimulated B cells were transferred into TNF−/− recipients, but not into TNF/LTα−/− recipients. This result appeared to account for the defect in isotype switching observed in intact TNF/LTα−/− mice because TNF/LTα−/− B cells, when stimulated in vitro, switched isotypes normally. Thus, TNF is necessary for creating the permissive environment for B cell movement and function, but is not itself responsible for these processes.