中华微生物学和免疫学杂志
中華微生物學和免疫學雜誌
중화미생물학화면역학잡지
CHINESE JOURNAL OF MICROBIOLOGY AND IMMUNOLOGY
2012年
4期
346-350
,共5页
袁玉兰%郭京蓉%周继唯%高华
袁玉蘭%郭京蓉%週繼唯%高華
원옥란%곽경용%주계유%고화
流感嗜血杆菌%血清型特异基因%荚膜型基因%荚膜多糖%免疫原性
流感嗜血桿菌%血清型特異基因%莢膜型基因%莢膜多糖%免疫原性
류감기혈간균%혈청형특이기인%협막형기인%협막다당%면역원성
Haemophilus influenzae%Sentype-specific gene%Capsular type gene%Capsular polysaccharide%Immunogenicity
目的 从分子水平检定b型流感嗜血杆菌,研究不同菌株荚膜多糖结合物的免疫原性.方法 提取基因组,通过型特异和荚膜型基因特异引物,利用PCR检定b型流感嗜血杆菌；不同纯化多糖分别与破伤风类毒素(TT)进行耦联结合,结合物原液经稀释免疫小鼠,通过两针免疫,采血进行免疫效力测定.结果 5株b型流感嗜血杆菌通过PCR法均能获得预期大小的型特异(482 bp)和荚膜型(343 bp)基因片段,BLAST分析显示,各菌之间型特异和荚膜型序列比对,其同源性均为100％,各菌型特异和荚膜型序列分别与GenBank X78559.1和M19995.1序列比对,同源性分别为99％和100％；ELISA检测显示,4株不同菌株来源的荚膜多糖结合物(PRP-TT)的小鼠免疫效力差异无统计学意义.结论 通过PCR法从分子水平检定b型流感嗜血杆菌,纯化的不同荚膜多糖结合物小鼠免疫原性基本一致.可供不同Hib多糖结合物免疫原性的研究参考数据.
目的 從分子水平檢定b型流感嗜血桿菌,研究不同菌株莢膜多糖結閤物的免疫原性.方法 提取基因組,通過型特異和莢膜型基因特異引物,利用PCR檢定b型流感嗜血桿菌；不同純化多糖分彆與破傷風類毒素(TT)進行耦聯結閤,結閤物原液經稀釋免疫小鼠,通過兩針免疫,採血進行免疫效力測定.結果 5株b型流感嗜血桿菌通過PCR法均能穫得預期大小的型特異(482 bp)和莢膜型(343 bp)基因片段,BLAST分析顯示,各菌之間型特異和莢膜型序列比對,其同源性均為100％,各菌型特異和莢膜型序列分彆與GenBank X78559.1和M19995.1序列比對,同源性分彆為99％和100％；ELISA檢測顯示,4株不同菌株來源的莢膜多糖結閤物(PRP-TT)的小鼠免疫效力差異無統計學意義.結論 通過PCR法從分子水平檢定b型流感嗜血桿菌,純化的不同莢膜多糖結閤物小鼠免疫原性基本一緻.可供不同Hib多糖結閤物免疫原性的研究參攷數據.
목적 종분자수평검정b형류감기혈간균,연구불동균주협막다당결합물적면역원성.방법 제취기인조,통과형특이화협막형기인특이인물,이용PCR검정b형류감기혈간균；불동순화다당분별여파상풍류독소(TT)진행우련결합,결합물원액경희석면역소서,통과량침면역,채혈진행면역효력측정.결과 5주b형류감기혈간균통과PCR법균능획득예기대소적형특이(482 bp)화협막형(343 bp)기인편단,BLAST분석현시,각균지간형특이화협막형서렬비대,기동원성균위100％,각균형특이화협막형서렬분별여GenBank X78559.1화M19995.1서렬비대,동원성분별위99％화100％；ELISA검측현시,4주불동균주래원적협막다당결합물(PRP-TT)적소서면역효력차이무통계학의의.결론 통과PCR법종분자수평검정b형류감기혈간균,순화적불동협막다당결합물소서면역원성기본일치.가공불동Hib다당결합물면역원성적연구삼고수거.
Objective To determine Haemophilus influenzae type b strains in molecular level using PCR,and to study the immunogenicity of capsular polysaccharide conjugates in mice.Methods Extracting genomes using bacterial DNA extract kit from Hoemophilus influenzae type b strains,and PCR for determining the strains through serotyping-specific and capsular genotyping primers respectively.Various capsular polysaccharides conjugated TT respectively,and the conjugates were administered subcutaneously to mice through dilution.After vaccination with two doses,blood samples were collected for the detection of antibody levels to polyribosylribitol phosphate ( PRP),the capsular polysaccharide of Hib.Results All five Haemophilus influenzae type b strains contain type-specific(482 bp) and capsular type (343 bp)DNA fragment through PCR detecting.The DNA fragments were sequenced.BLAST show that these sequences are 100％ homology comparing the above strains respectively,and are 99％ and 100％ homology comparing the GenBank X78559.1 and M19995.1 respectively.The immunogenicity of mice from various capsular polysaccharide conjugates (PRP-TT) was not significantly different by ELISA detecting.Conclusion Through PCR,Haemophilus influenzae type b strain can be determined in molecular level.The immunogenicity of mice from purified capsular polysaccharide conjugates was not different.The study provides a detection means for the features and heredity stability of Haemophilus influenzae type b strain and reference data for the immunogenicity of different polysaccharide conjugates in vaccine research and development and production.
