Conventional DCs, but not macrophages or B cells, are necessary for anti-capsid CD8⁺T-cell responses. (A) Experimental timeline of WT, CD11c-DTR, gadolinium chloride (GdCl₃)-treated, or μMT mice that were IM injected with AAV2-SIINFEKL (n = 4/group). CD11c-DTR mice received diphtheria toxin (DT) on days −1 and 0. Mice received GdCl₃ on days −1 and 0. (B) Quantification of capsid-specific CD8⁺ T-cell response in peripheral blood of APC-depleted mice as a function of time. (C) Time-course of anti-CD20 depletion experiment. WT mice received phosphate-buffered saline (PBS) or anti-CD20 in 2 doses 3 weeks apart. One day after the second injection, mice were IM injected with AAV2-SIINFEKL, and capsid-specific CD8⁺ T-cell responses were assessed 7 days later (n = 4/group). (D) Quantification of capsid-specific CD8⁺ T-cell response. The dotted line at 0.1% represents the limit of detection of capsid-specific CD8⁺ T cells using the tetramer. Data points are averages ± SEM. (E) Time-course of DC depletion with chlodronate liposomes, and (F) quantification of capsid-specific CD8⁺ T-cell response. (G) Time-course of AAV2 administration and DC depletion in CD11c-DTR mice using DT or WT BALB/c mice using chlodronate liposomes, followed by measurement of CD8⁺ T-cell frequency of response to native capsid epitopes. (H) Quantitation of IFN-γ ELISPOT assays (n = 4/group; data are averages ± SEM for epitope-stimulated minus mock-stimulated cultures). P values: (B) *P =.02 for WT vs CD11c-DTR; (F) **P =.005 for day 7 and *P =.02 for day 10; (H) *P =.03 for both experiments. ns, not significant.