Targeting Ag to DC using anti-mClec9A-OVA elicits both CD4 and CD8 T cell proliferative responses. OVA-specific transgenic CD8 (OT-I) or CD4 (OT-II) T cells (10⁶) were adoptively transferred into naive C57/BL6 Ly5.1 mice. One day later mice were injected intravenously with 2.5 μg of anti–mClec9A-OVA (n = 3) or nontargeted isotype control-OVA mAb-2 (GL117; n = 3), or left unimmunized (n = 2). Three days after mAb injection, mice were killed and spleens harvested. Cells were stained with mAb against Ly5.2 (S.450-15.2-PE) and CD4 (GK1.5-APC) or CD8 (YTS169-APC) and proliferating CFSE-labeled transgenic T cells, (A) OT-I (Ly5.2⁺CD8⁺) or (B) OT-II (Ly5.2⁺CD4⁺), enumerated by flow cytometry. The proliferative response of OVA-specific T cells was seen as a loss of CFSE fluorescence by flow cytometry. The total number of OT-I cells and OT-II cells proliferating per spleen was enumerated as described in “Purification of transgenic T cells and in vivo proliferation assays.” One experiment with the 2.5-μg dose is presented as the mean response plus or minus SEM. The experiment was performed twice with 2.5 μg and once with 5 μg of OVA-conjugated mAb, with similar results.