| Chitosan(CS)is a positively charged natural polymer material with good biodegradability,biocompatibility and adhesion.Therefore,CS has been used widely in the field of vaccine delivery.To improve the safety of the vaccine,prolong the action time of the antigen in vivo,and enhance the ability that the antigen to be taken up and absorbed by the negatively charged antigen-presenting cells,CS-based micro-level and nano-level composite vaccine delivery vehicles were prepared in this paper.And ovalbumin(OVA)was regarded as the vaccine model and was delivered into body by oral or subcutaneous injection,and then the immune response ability of the CS-based composite vaccine delivery vehicle to encapsulate OVA was explored in mice.1.Polylactic acid/Chitosan/Ovalbumin microspheres(CS/PLA/OVA MPs)with a core-shell double-layer structure were prepared by water-in-oil-in-water(W/O/W)emulsion-evaporation technique.And polyethylene oxide(PEO),a compatibility agent,was added to the emulsion in order to improve the compatibility between PLA and CS.The as-constructed MPs,with a particle size range of 1-7 ?m,were spherical and smooth.Besides,by confocal laser scanning microscopy(CLSM),it can be seen that MPs possessed a core-shell double-layer structure,which can prevent the structure of OVA in MPs from damage.It can be confirmed by polyacrylamide gel electrophoresis(PAGE)analysis that the construction of OVA remains intact in CS/PLA MPs.The conditions of preparing CS/PLA/OVA microspheres were optimized,and the outcomes were as follows:PLA concentration was 10 mg/m L,OVA concentration was 30 mg/m L,and CS concentration was 4 mg/m L.Under these conditions,the resultant encapsulation efficiency(EE)and loading capacity(LC)of MPs can reach 76.4% ± 0.1% and 272.0 ± 15.6 ?g/mg,respectively.According to the release behavior studies of MPs in vitro,OVA was released from MPs in the way of exponential decline and was completely released within 8 h.Finally,the cytotoxicity of MPs was studied by MTT method,and the outcomes indicated that CS/PLA MPs are low-toxic and biocompatible.In short,CS/PLA MPs have huge application prospects in the field of vaccine delivery.2.(?)-Polyglutamic acid/Chitosan/Ovalbumin nanoparticles((?)-PGA/CS/OVA NPs)were prepared by ion cross-linking method and polyelectrolyte composite method.The as-constructed (?)-PGA/CS/OVA NPs,with a positive charge on the surface,were uniformly round.And the particle size range of the as-constructed (?)-PGA/CS/OVA NPs was 200-400 nm.The analysis of polyacrylamide gel electrophoresis(PAGE)showed that OVA in (?)-PGA/CS/OVA NPs has high structural integrity.The conditions of preparing(?)-PGA/CS/OVA NPs were optimized,and the outcomes were as follows: OVA concentration was 16.0 mg/m L,TPP concentration was 0.5 mg/m L,CS concentration was 2.0mg/m L,and (?)-PGA concentration was 1.0 mg/m L.Under these conditions,the resultant encapsulation efficiency(EE)and loading capacity(LC)of NPs achieved 40.9% ± 2.2%and 207.7 ± 11.1 ?g/mg,respectively.MTT assay showed that (?)-PGA/CS NPs have low toxicity and biocompatibility.The results of immune experiments in mice showed that compared with OVA not wrapped with (?)-PGA/CS composite carrier,(?)-PGA/CS/OVA NPs can induce a stronger immune response through humoral immunity in vivo.Therefore,(?)-PGA/CS NPs can be regarded as a good vaccine delivery system to enhance the immune response in vivo. |
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