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. 2017 Aug 4;7:349. doi: 10.3389/fcimb.2017.00349

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Copyright © 2017 Guo, Yang, Tang, Yin, Liu, Gong, Wei, Zhang, Xu and Liu.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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CWAE expression, purification, and antigen characteristics. (A) Protein expression and purification of CWAE. (Lane 1) protein marker, (lanes 3 and 5) the inclusion bodies of E. coli BL21(DE3)/pETCWAE, (lanes 2 and 4) the soluble proteins of E. coli BL21(DE3)/pETCWAE, (lane 6 and 7) the purified CWAE proteins. (B) Protein expression and purification of CTB-UE. (Lane 1) protein marker, (lanes 2, 3, and 5) the inclusion bodies of E. coli BL21(DE3)/pETCUE, (lane 4) the soluble proteins of E. coli BL21(DE3)/pETCUE, (lane 6, 7, and 8) the purified CTB-UE proteins. (C) Immunogenicity and immunoreactivity of CWAE analyzed by Western blot. (i) CWAE and CTB-UE reaction with Rabbit anti-H. pylori polyclonal antibody (Rabbit anti-Hp PcAb). (Lane M) protein marker, (lane 1) CWAE proteins, (lane 2) CTB-UE proteins. (ii) The H. pylori antigens (UreA, UreB, HpaA, Hsp60 and NAP) reaction with antiserum induced by CWAE vaccine. (Lane M) protein marker; (lane 1, 60 KD) Hsp60; (lane 2, 64 KD) UreB; (lane 3, 30 KD) UreA; (lane 4, 31 KD) HpaA; (lane 5, 14 KD) NAP. (D) The adjuvanticity of CTB component in CWAE and CTB-UE vaccine analyzed by GM1-ELSIA. In order to confirm the CTB component in CWAE or CTB-UE with the ability to bind GM1 gangliosides, GM1-ELISA was performed. ELISA plates were coated with 1 μg/well of GM1 ganglioside or BSA. The recombinant proteins CWAE, CTB-UE, CTB, and UreB with a concentration of 100 μg/ml were used to evaluate their capability of binding GM1. Data are mean ± SD. p < 0.05 was considered as statistically significant. ^**p < 0.01; ^***p < 0.001; ns, not significant.