摘要
目的检测疫苗生产用人二倍体细胞2BS和MRC-5是否存在外源因子猪圆环病毒1型(porcine circovirus type 1,PCVl)和猪圆环病毒2型(PCV2)污染。方法对已建立的PCR方法进行灵敏度验证后,检测人二倍体细胞2BS和MRC-5中PCV1和PCV2污染情况。结果两种细胞上清中最低均可检出1 pg的PCV1和PCV2 DNA,灵敏度较高;两种细胞中PCV检测结果均为阴性。结论在生产用人二倍体细胞2BS和MRC5中,均未检出PCV1和PCV2污染。
Objective To determine the presence or absence of contamination with porcine circovirus type 1(PCV l)and porcine circovirus type 2(PCV 2) in human diploid 2 BS and MRC-5 cells used for vaccine production. Methods The developed PCR method was verified for sensitivity and used for determination of PCV1 and PCV2 in 2 BS and MRC-5 cells. Results Both the minimum detection limits of PCV1 and PCV2 in the supernatants of 2 BS and MRC5 cells were 1 pg,indicating a high sensitivity of developed PCR method. Both the determination results of PCV in two cell lines were negative. Conclusion No contamination with PCV1 or PCV2 was detected in 2 BS and MRC5 cells used for vaccine production.
引文
[1]TISCHER I,GELDERBLOM H,VETTERMANN W,et al.Avery small porcine virus with circular single-stranded DNA[J].Nature,1982,295(5844):64-66.
[2]FENAUX M,HALBUR P G,GILL M,et al.Genetic characterization of type 2 porcine circovirus(PCV-2)from pigs with postweaning multisystemic wasting syndrome in different geographic regions of North America and development of a differential PCR-restriction fragment length polymorphism assay to[J].JClin Microbiol,2000,38(7):2494.
[3]BEACH N M,C仵RDOBA L,KENNEY S P,et al.Productive infection of human hepatocellular carcinoma cells by porcine circovirus type 1[J].Vaccine,2011,29(43):7303-7306.
[4]YAO L,LIN C M,JENG C R,et al.The pathogenic role of torque teno sus virus 1 and 2 and their correlations with various viral pathogens and host immunocytes in wasting pigs[J].Vet Microbiol,2015,180(3-4):186-195.
[5]RICHARD D,LEFEBVRE D J,DIPONGKOR S,et al.Outcome of experimental porcine circovirus type 1 infections in midgestational porcine fetuses[J].Bmc,2011,7(1):64.
[6]GRASLAND B,BIGAULT L,LE C M,et al.Naked PCV-2cloned genomic DNA is infectious by mucosal(intratracheal or oro-nasal)inoculation[J].Virus Res,2012,165(1):90-94.
[7]黄敏,吕冰凌,袁良玉,等.乙型脑炎减毒活疫苗中猪圆环病毒PCR检测方法的建立及验证[J].中国生物制品学杂志,2016,29(12):1346-1348.
[8]VICTORIA J G,WANG C,JONES M S,et al.Viral nucleic acids in live-attenuated vaccines:detection of minority variants and an adventitious virus[J].J Virol,2010,84(12):6033-6040.
[9]CHEN Z H,XU J J.Development of freeze-dried live varicella vaccine[J].J Microbiol,1999,19(3):63.(in Chinese)陈志慧,徐建军.冻干水痘活疫苗的研制[J].微生物学杂志,1999,19(3):63.
[10]LIU X L,ZHANG A N,CHEN X M,et al.Preparation of a live attenuated measles,mumps,rubella and varicella combined vaccine[J].Int J Biologicals,2016,39(5):209-213.(in Chinese)刘晓琳,张安宁,陈晓梅,等.麻疹、腮腺炎、风疹和水痘联合减毒活疫苗的研制[J].国际生物制品学杂志,2016,39(5):209-213.
