To verify the immunizing potential of these vectors against HIV we next sought to determine if they were capable of eliciting an anti-HIV immune response in mice. Equal numbers (1610 10 VP) of the vectors were used to immunize BALB/c mice via an intramuscular route. The sera were collected from mice for ELISA assays at various days after priming and boosting. Purified MPER antigenic peptide was bound to ELISA plates. The plates were then incubated with the immunized mice sera. The binding was detected with HRP-conjugated secondary antibody. The amount of anti-MPER antibody in the sera was calculated based on a standard antibody curve dilution. The data demonstrates no binding of the sera from mice immunized with Ad5, AdCMVEnv, or AdCMVGag to the MPER on the plates at any time point. On the contrary, immunization with Ad that contains capsid-incorporated MPER antigen elicits an anti-MPER specific response and allows the formation of MPER-specific antibody in the serum ( Figure 6 ). This immune response is observed at the earliest time point that we observed, which was 14 days post-prime (d.p.p). The peak immune response was observed with the capsid incorporated MPER vectors at 54 d.p.p, which also corresponds to 14 days postboost (d.p.b.). At 54 d.p.p the determined concentration of gp41 antibody was ,7 mg/ml. A slight decline of the MPER antibody in the sera at 95 d.p.p was observed in the mice vaccinated with capsid-modified vectors, which corresponded to 55 d.p.b. At 55 d.p.b. MPER antibody levels in the sera was determined to be ,5 mg/ml. In summary, Ad5/HVR2-MPER-L15DE1 and Ad5/HVR2-MPER-L15(Gag) elicited an anti-gp41 response in vaccinated mice. This MPER-specific response was increased after boosting. It is important to note that the envelope expressed in AdCMVEnv was from the 89.6 HIV variant. The epitope inserted into the Ad5 hexon and the peptide were identical sequences with the 89.6 Env MPER region. The transgene expressed from the Ad5 vector was 89.6 Env (i.e. gp160), which is processed into gp120 and gp41. There were no point mutations within the transgene expressed Env.