FIG. 2.

AAV-221-IV vector delivers factor IX to C57BL/6-CD4 knockout mice. Mice were injected intramuscularly with CsCl-purified AAV1, AAV2, or AAV-221-IV (1 × 10¹¹ VG/mouse) carrying an hFIX construct under the control of a CMV promoter/enhancer construct. (A) Human factor IX (hFIX) expression profile after vector administration. hFIX expression was measured by ELISA and average values and standard deviation are shown for each group (n = 3). (B) Immunofluorescence staining for human factor IX in AAV-injected mouse muscle. Shown are samples obtained from mice killed at week 14 after vector delivery. The signal for human factor IX is shown in red. “Untransduced” is from a CD4 mouse that did not receive vector. (C) Southern blot analysis for vector genomes after vector administration. Total cellular DNA was extracted 14 weeks postinjection. After EcoRV digestion, 20 μg of DNA was electrophoresed on a 0.8% gel. The resulting membrane was probed for human factor IX. The viral genome shown here is the 1.7-kb fragment. The reference standards were prepared by adding 0.35, 3.5, and 35 copies of plasmid per murine diploid genome before EcoRV digestion.