Anti-FVIII antibody response after AAV-FVIII delivery can be complex and multiphasic. The anti-FVIII IgG titers were measured by ELISA. IgG titers are indicated by the red curve and quantified on the right y axis. The corresponding FVIII activity levels are indicated by the black curve and quantified on the left y axis. Bethesda (inhibitor) titers (BU/mL) were measured for every sample with a detectable IgG titer and/or loss of detectable FVIII activity. BU/mL values are indicated by a heat map at the top of each graph. Samples below the limit of quantification for any antibody assay were given a working value of 0 in data analysis and graphical representation; however, the actual value could be somewhere between 0 and the limit of quantification. An X terminating the IgG titer curve denotes death of the animal at the designated time point. (A) 6E13vg/kg AAV-E06.TTR-ET3. Animal IDs: (i) 1471L, (ii) 1475N, (iii) 1477B, (iv) 1483N. (B) 2E13vg/kg AAV-E06.TTR-ET3. Animal IDs: (i) 1451L, (ii) 1453R, (iii) 1455R, (iv) 1457L. (C) 6.67E12vg/kg AAV-E06.TTR-ET3. Animal IDs: (i) 1453N, (ii) 1479N. (D) 2.22E12vg/kg AAV-E06.TTR-ET3, animal 1437N. (E) 6E13vg/kg AAV-HCB-ET3. Animal IDs: (i) 1437R, (ii) 1451R, (iii) 1453LL, (iv) 1479L. (F) 2E13vg/kg AAV-HCB-ET3. Animal IDs: (i) 1437L, (ii) 1455N. (G) 6E13vg/kg AAV-E06.TTR-HSQ. Animal IDs: (i) 1475B, (ii) 1481B, (iii) 1507R.