Direct intramuscular injection of AAV vectors does not activate DCs. Eight- to 10-week-old C57BL/6 mice (n = 4 per cohorts) received intramuscular injection of 10¹¹ vg of AAV1-hFIX or 6 × 10¹⁰ vg of AAV2-hFIX. Control mice of similar age were administered PBS. After 24 hours, cells from the draining lymph nodes and spleen were collected and stained for the appropriate antibodies, and flow cytometry analyses were performed, as described. (A) No change in DC numbers in draining lymph nodes and spleen after intramuscular injection of AAV. The dot plots show the expression of CD11c on cells. Numbers indicate the percentage of CD11c⁺ cells among the total live cells in either the draining lymph nodes or spleen. (B) No activation of DCs in draining lymph nodes and spleen after intramuscular injection of AAV. The dot plots show the expression of either MHCII and CD80 or MHCII and CD86 on cells gated with CD11c. Numbers indicate the percentage of MHCII⁺CD80⁺ or MHCII⁺CD86⁺ among the CD11c⁺ population. Data shown are mean ± SEM. A representative of 2 independent experiments is shown.