CAJ | 학술논문

为了对壳聚糖分子作为佐剂的缓释效果进行免疫学评价,本研究合成了牛冠状病毒DB2毒株N蛋白基因的3'端第487位～1287位碱基,用大肠杆菌对其进行了高效表达,通过SDS-PAGE电泳分离、电洗脱纯化该牛冠状病毒N蛋白(BCV N).以戊二醛为交联剂,采用乳化交联的方法制备空白壳聚糖微球.利用吸附法制备载BCVN蛋白的壳聚糖微球,通过肌肉注射途径免疫BALB/c小鼠,用间接ELISA方法检测免疫鼠血清中的抗体水平,由此评价壳聚糖微球对BCVN的缓释作用和佐剂效应.结果表明,壳聚糖微球的形态较好、表面光滑、分散性较好,平均粒径为6.50±1.77μm,电势分布在36 mV,吸附BCV N蛋白的壳聚糖微球在体内所产生的免疫效果明显优于唯N蛋白组,说明载BCV N蛋白的壳聚糖微球在体内具有一定的缓释效果,使N蛋白在体内缓慢释放而长时间诱导抗体的产生,该研究结果为以壳聚糖微球作为疫苗佐剂积累了实验数据.
위료대각취당분자작위좌제적완석효과진행면역학평개,본연구합성료우관상병독DB2독주N단백기인적3'단제487위～1287위감기,용대장간균대기진행료고효표체,통과SDS-PAGE전영분리、전세탈순화해우관상병독N단백(BCV N).이무이철위교련제,채용유화교련적방법제비공백각취당미구.이용흡부법제비재BCVN단백적각취당미구,통과기육주사도경면역BALB/c소서,용간접ELISA방법검측면역서혈청중적항체수평,유차평개각취당미구대BCVN적완석작용화좌제효응.결과표명,각취당미구적형태교호、표면광활、분산성교호,평균립경위6.50±1.77μm,전세분포재36 mV,흡부BCV N단백적각취당미구재체내소산생적면역효과명현우우유N단백조,설명재BCV N단백적각취당미구재체내구유일정적완석효과,사N단백재체내완만석방이장시간유도항체적산생,해연구결과위이각취당미구작위역묘좌제적루료실험수거.
A DNA fragment corresponding to the 3' terminus 487-1 287 bases of the N protein gene of bovine coronavirus DB2 strain were artificially synthesized and expressed in E, coil, Blank chitosan microspheres were prepared using glutaraldehyde as crosslinkers followed by emulsification. The expressed BCV N protein was purified by electric elution and loaded on to chitosan microspheres by absorptive method. These microspheres were used to immunize BALB/c mice. The dynamic antibody level in serum was monitored by indirect ELISA to evaluate the slow-release and adjuvanted effect of the BCV N-loaded chitosan microspheres. The results showed that the microspheres had a good shape with smooth surface and an average size of 6.50 ± 1.77μm. The BCV N-loaded chitosan microspheres induced better antibody response than the N protein alone in mice, indicating that the BCV N-loaded chitosan microspheres had slow-release effect as adjuvant.