Protein microarray epitope mapping of monoclonal antibody 5H2 raised by human immunization with the 4CMenB vaccine formulation containing the fusion antigen NHBA-NMB1030.

The aim of the study was to determine the epitope targeted by 5H2 human Fab directed against NHBA and the crossreactivity aginst a panel of nine different NHBA long and short peptide variants (p2, p3, p5, p10, p17, p20, p21, p24, p29). 5H2 were diluted at 1:2000 and incubated on a non-commercial Protein Microarray platform printed with NHBAp2 specific recombinant protein fragments and full length NHBA of different variants. A panel of recombinant fragments of different length spanning the entire sequence of NHBAp2 and NHBAp3 plus different nine different NHBA long and short peptide variants (p2, p3, p5, p10, p17, p20, p21, p24, p29) were printed in 8 replicates on a no-commercial Protein Microarray. All fragments were expressed in E. coli as either glutathione S-transferase-, His-tagged or TRX-fusions, purified from the cytoplasmic fraction as soluble forms. Human IgG (at 8 different concentration points) was distributed in each array copy as control. HuFab 5H2 was profiled for anti-Human IgG specific analysis of reactivity to the spotted proteins.