Injection of Ag into the anterior chamber (AC) of the eye induces deviant immune responses. It has been proposed that Ag internalized by ocular APCs is presented in a tolerogenic fashion in the spleen. However, the nature and distribution of the Ag-bearing cells in the lymphoid organs remain unclear. Fluorescent-labeled Ag (dextran, BSA) injected into the AC of Lewis rats was detected in the subcapsular sinus of the right submandibular lymph nodes (LNs) and cervical LNs, the marginal zone of the spleen, and the medulla of the mesenteric LNs. In the spleen, Ag-bearing cells were CD1(+), CD11b(+), ED1(+), ED2(low), ED3(+), CD86(low), OX6(+), CD11c(-), ED5(-) and in the LNs were CD4(+), CD8(+), CD80(+), and OX41(+) suggesting these were lymphoid organ resident macrophages. These Ag-bearing macrophages were located adjacent to CD4(+) cells, CD8(+) cells, and NK cells in the LNs and spleen and to marginal zone B cells in the spleen. No interaction with gamma delta T cells was observed. The data demonstrates that Ag derived from the AC of the eye is mainly internalized by resident macrophages in the LNs and spleen which are ideally placed to interact with cells involved in the induction of deviant ocular immune responses. The extensive distribution of Ag in LNs draining the upper airway and gastrointestinal tracts, together with the phenotype of Ag-bearing cells in the lymphoid organs, suggests that Ag leaves the eye predominantly in a soluble form and implies other mechanisms of tolerance may contribute to ocular-specific immune responses.