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. 2018 Jan 30;8:1888. doi: 10.1038/s41598-018-20205-7

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Biological production and characterization of antigen coated PHB beads. (a) Schematic representation of the hybrid genes mediating formation of antigen displaying PHB beads or production of the respective soluble antigen. (b) PHB inclusion production and beads size evaluation by Transmission Electron Microscopy (TEM) using whole cells of recombinant E. coli (Clearcoli^TM) and the respective purified PHB beads; (c) SDS-PAGE analysis of the protein profile of isolated PHB beads as well as the purified His-tagged proteins (selected antigens). Lane 1, MW (molecular weight) standard, (GangNam-Stain, iNtRON BIOTECHNOLOGY); Lane 2, non-antigen displaying PHB beads (PhaC ~64 kDa); Lane 3, NadA-His6 protein ~50 kDa; Lane 4, NadA-PhaC fusion protein (~113 kDa) on PHB beads; Lane 5, MW standard, (GangNam-Stain, iNtRON BIOTECHNOLOGY); Lane 6, non-antigen displaying PHB beads (PhaC ~64 kDa); Lane 7, GNA2091-fHbp-G1-His6 protein (~47 kDa); Lane 8, GNA2091-fHbp-G1-PhaC fusion protein (~111 kDa) on PHB beads; Lane 9, fHbp-G1-G1-PhaC fusion protein (~120 kDa) on PHB beads. (d) Immunoblotting using commercial monoclonal anti-fHbp antibodies (JAR4, NIBCS, UK) and monospecific polyclonal anti-PhaC antibodies were performed to further confirm the identity of the respective fusion proteins. Fusion proteins with the anticipated MW containing fHbp were detected. Lane 1, PhaC protein, (non-antigen displaying PHB beads); Lane 2, GNA2091-fHbp-G1-His6 protein; Lane 3, GNA2091-fHbp-G1-PhaC fusion protein on PHB bead; Lane 4, fHbp-G1-G1-PhaC fusion protein on PHB beads.