Arrangement of regions and expressions of the tHIVconsvX immunogens. (a) Serial organization (scramble) of the six regions used to minimize induction T-cell responses by potentially newly formed junctional epitopes. These are irrelevant for protection, because they are not present in natural HIV-1 proteins. The delivery vectors, into which the individual immunogens were inserted for presentation to the immune system, are also indicated. Note that mosaic 1 and mosaic 2 are always used together in the same vaccine dose. (b) Expression of individual tHIVconsvX proteins from the plasmid DNA, ChAdOx1, and MVA vaccine vectors. HeLa cells were either transfected with 2.5 µg of pSG2.tHIVconsv1 and pSG2.tHIVconsv2 for 2 days, infected with MVA.tHIVconsv3 and MVA.tHIVconsv4 MOI of 5 for 1 day, or infected with ChAdOx1.tHIVconsv5 and ChAdOx1.tHIVconsv6 at MOI of 10 for 1 day. Recombinant proteins were detected using primary mAb ID (91-5) followed by FITC-conjugated goat-antihuman IgG secondary antibody (green). DAPI stain shows the positions of cell nuclei (blue).