犬五联病毒活疫苗制备用犬肾细胞系的建库筛选与检定  

Establishment of Master Cell Stock and Working Cell Bank of MDCK Lines and Selection and Evaluation of the Lines as Candidate Viral Substrates for Approval Production of Combinational Canine Attenuated-live Virus Vaccines

作　　者：张德礼[1] 李六金[2] 夏耕田[3] 何旭玉[3] 高步先[3] 白晓鸿[3] 黄高升[4] 刘尚高[5] 阎隆飞[5] 方福德[6] 

机构地区：[1]北京大学医学部基础医学院免疫学系,北京100083 [2]第四军医大学动物保健品研制中心 [3]北京军区联勤部卫生防疫队,北京100071 [4]第四军医大学基础部病理学教研室,西安710032 [5]中国农业大学动物医学院和生物学院,北京100094 [6]中国医学科学院基础医学研究所/中国协和医科大学基础医学院,医学分子生物学国家重点实验室,北京100005

出　　处：《中国农业科学》2002年第6期709-713,共5页Scientia Agricultura Sinica

基　　金：国家火炬计划重大项目

摘　　要：以人类子宫颈癌细胞系Hela为阳性对照 ,以连传 3代的犬、猫肾原代细胞 (CKC、FKC)作为阴性对照 ,对来自国内外不同单位收集的另外 4株MDCK传代细胞系培养 2 5～ 6 7代的完整活细胞进行裸鼠致癌 /致瘤实验观察 ,筛选出致癌性极低、符合细胞遗传学要求、无传染因子污染的 2株MDCK细胞系用于制苗 ,并建立了相应的细胞种子库和工作库 ,供科研和生产使用 ,4年运转很好。不同代次MDCK细胞系染色体众数所占比率的相差率一般不超过 5 %～ 15 % ,结构畸变率一般为 0～ 3%。研究表明 ,MDCK细胞染色体遗传特征决定致瘤性质 ,并具有种属特异性 ,MD CK细胞不论核型如何 ,始终具有致癌性 ,但其致癌 /致瘤性差 (2 3/ 5 9) ,且一般致上皮源性恶性肿瘤 ,多为高中分化腺癌。降低制苗毒液中细胞系基因含量 ,完全可以将MDCK细胞系 (WB和H株 )用于犬五联苗生产。MDCK细胞超二倍体YB株和亚二倍体与超二倍体KA株不能用作病毒活疫苗培养基质。找出了MDCK细胞系的染色体变异率、软琼脂中克隆形成率、对植物凝集素的凝集性和在裸鼠体内形成癌肿的潜力之间的可能相关性 ,发现细胞系染色体数目增加、克隆形成率增高、凝集性增强 ,则致癌Under the prerequisite that the incidence of cancer or tumor in negative control nude mice inoculated subcutaneously with primary feline or canine kidney cell cultures purified in vitro at passage 3 was 0 (0/22) and 0 (0/10), respectively. The incidence of the progressively growing malignant tumor(MT) in positive control nude mice inoculated subcutaneously with Hela cell cultures of KB, X, or NM20/X strain was 10/10, 25/25 and 5/51, respectively. The results showed that the incidence of tumor in nude mice with di and hyperploid YB strain of MDCK cell during 17-23 passages, with hyper and hypoploid KA strain of MDCK cell during 6-8 passages, with hypoploid WB strain of MDCK cell on passage 6, with hyper and hypoploid H strain of MDCK cell during 8-24 passages was 2/24, 6/10, 5/10 and 10/15, respectively. The chromosomal analysis results showed that the ratio of difference in the rate of modal chromosome number between high(mcs+n) and lowest (mcs)passages was not more than 5%-15% and the structure aberrations was generally 0-3%. These results proved that the genetic characteristics of chromosomal number of cell lines determines their tumorigenicity, but it is species specific. MDCK line has tumorigenicity no matter what its chromosome karyotype is, at least it has very low tumorigenicity even when its modal chromosome number is hypoploid. It is thus evident that MDCK cell of WB or H strain can be approved as substrate for the preparation of attenuated viral vaccines, but MDCK cell of YB or KA strain can not be approved as substrate for the preparation of attenuated viral vaccines.

关 键 词：犬 五联病毒活疫苗 制备 犬肾细胞系 建库筛选 检定 

分 类 号：S859.797[农业科学—临床兽医学]