摘要
采用可生物降解的合成高分子聚DL-乳酸-聚乙二醇共聚物(DL-polylactide–co–polyethylene glycol,PELA),包裹哈维氏弧菌(Vibrio harveyi,Vh)重组外膜蛋白OmpK后,制备成PELA-OmpK微球。微球粒径小于10μm,且94%粒径小于5μm,平均粒径为2.8μm。蛋白包裹率达79.4%。斜带石斑鱼(Epinephelus coioides)口灌四甲基异硫氰酸罗达明(TRITC)荧光素标记的OmpK-PELA微球后,第1天在其后肠观察到大量红色荧光微球,第3天荧光微球数量明显减少,第7天荧光微球基本消失。免疫组化研究结果表明,口服PELA-OmpK组在斜带石斑鱼后肠组织黏膜层可见较强的棕黄色阳性着色。口服PELA-OmpK组血清抗体效价(峰值210)显著高于口服OmpK蛋白溶液组(峰值24)与对照组(P<0.05),低于注射组(峰值212)(P<0.05);口服PELA-OmpK组的黏液抗体效价显著高于口服OmpK蛋白组(P<0.05)。初次免疫30d后用活菌攻击,口服PELA-OmpK组的相对保护率(62.5%)显著高于口服OmpK蛋白组(12.5%)(P<0.05);低于注射组(87.5%)(P<0.05)。结论为采用可生物降解材料PELA作为鱼类口服疫苗的投递载体是可行的。
To prepare oral biodegradable microsphere vaccine, the recombinant outer membrane protein （OmpK） as a common antigen of Vibrio harveyi was obtained from Escherichia coli DH5α containing the recombinant plasmid pBV220-OmpK, and the OmpK was encapsulated in the biodegradable delivery system comprised of DL-polylactide-co-polyethylene glycol （PELA）microspheres. The diameter of microsphere vaccine was smaller than 10 μm and 94% of them were smaller than 5 μm, with an average diameter of 2.8 μm. The encapsulation rate of the microsphere vaccine reached 79.4%. Marked with tetramethyl rhodamine isotheynate （TRITC）, the PELA-OmpK microspheres were orally administrated to the grouper, Epinephelus coioides and a large amount of red microspheres were found in the hindgut tissue at the first day after oral administration, then the amount reduced at the third day, and almost could not be found at the 7th day. The hindgut tissue slice was studied by immunohistochemistry method, and the result showed that antigen could be found in the mucous membrane layer of hindgut in the oral PELA-OmpK vaccine treatment group. After being orally immunized with the microsphere vaccine, the maximal antibody titer in serum of oral PELA-OmpK vaccine treatment group （2^10） was obviously higher than that of oral OmpK treatment group （2^4） （P〈0.05）, but lower than that of the injecting adjuvant OmpK treatment group （2^12） （P〈0.05）. The antibody titer in hindgut mucus of the oral PELA-OmpK vaccine treatment group was also obviously higher than that of the oral OmpK treatment group （P〈0.05）. The relative percent survival rate （PSR） of the oral PELA-OmpK vaccine treatment group （62.5%） was significantly higher than that of oral OmpK treatment group （12.5%） after 4 weeks （P〈0.05）, lower than that of injection group （87.5%） （P〈0.05）. These results show that the biodegradable micropartical can be a candidate as the vector system of oral microsphere vaccine in fish.
出处
《中国水产科学》
CAS
CSCD
北大核心
2008年第5期837-844,共8页
Journal of Fishery Sciences of China
基金
国家高技术研究发展计划(863计划)项目(2006AA100308)
农业部农业结构调整重大技术研究专项(05-06-02B).
