Adenovirus–MVA heterologous prime–boost using a PfMSP1 antigen insert is a leading viral vectored regime for antibody and T cell induction against this blood-stage P. falciparum antigen [3] and [5]. As a protein-adjuvant comparator, we used a Pichia pastoris-expressed recombinant PfMSP119 [33], adjuvanted by Montanide ISA720 (Seppic, France). Montanide

ISA720 is a squalene-based water-in-oil emulsion which has been shown to be a potent adjuvant in both animal and human studies [34], [35], [36] and [37]. Here we describe and compare in detail the immunogenicity of PfMSP1 PD0325901 price vaccines using a novel combination of three subunit vaccine platforms: simian adenovirus AdCh63 [5] and [38]; MVA; and recombinant protein in Montanide ISA720. We report that, when combined, these technologies can achieve simultaneous antibody and T cell responses

which Staurosporine purchase equal, or in some cases surpass, the best immune responses achieved with either technology alone. We describe in detail the responses induced, with data on antibody isotypes and avidity, splenic antibody secreting cell counts, T cell quality, and response longevity. All procedures were performed in accordance with the terms of the UK Animals (Scientific Procedures) Act Project Licence and were approved by the University of Oxford Animal Care and Ethical Review Committee. 5–6 weeks old female BALB/c (H-2d) and C57BL/6 (H-2b) mice (Harlan Laboratories, these Oxfordshire, UK) were anesthetized before immunization with medetomidine (Domitor, Pfizer) and ketamine (Ketaset, Fort Dodge) and revived subsequently with Antisedan reversal agent (Pfizer). All immunizations were administered intramuscularly (i.m.) unless otherwise specified, with vaccine divided equally into each musculus tibialis. The creation of simian adenovirus 63 (AdCh63) and modified vaccinia virus Ankara (MVA) vectors encoding the PfM128 antigen is described elsewhere [5]. Briefly,

this antigen is a bi-allelic fusion incorporating the MSP142 antigen from the K1/Wellcome and 3D7/MAD20 P. falciparum strains fused in tandem alongside four blocks of conserved sequence from the remainder of the 3D7 strain MSP1 molecule (blocks 1, 3, 5 and 12). The MVA used in the current study differs from the previously published vector [3] in that it lacked the green fluorescent protein (GFP) marker. To generate the markerless MVA expressing PfM128, the antigen was cloned into a transient-dominant shuttle vector plasmid such that PfM128 was expressed from the vaccinia P7.5 promoter, and inserted into the TK locus of MVA. The plasmid also expresses a GFP marker [39]. This plasmid was transfected into chicken embryo fibroblast cells (CEFs) infected with MVA expressing red fluorescent protein (RFP), as previously described [3]. Recombinant MVAs were generated by homologous recombination between regions of homology at the TK locus of MVA and in the plasmid shuttle vector.