[1]孔倩倩a,唐振华,相芬芬a,等.抗HPV18 E6多肽单克隆抗体的制备及鉴定[J].现代检验医学杂志,2016,31(03):30-32.[doi:10.3969/j.issn.1671-7414.2016.03.009]
　KONG Qian-qiana,TANG Zhen-hua,XIANG Fen-fena,et al.Preparation and Identification of A Peptide Monoclonal Antibody Against Human Papillomavirus 18E6[J].Journal of Modern Laboratory Medicine,2016,31(03):30-32.[doi:10.3969/j.issn.1671-7414.2016.03.009]
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抗HPV18 E6多肽单克隆抗体的制备及鉴定()
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《现代检验医学杂志》[ISSN:/CN:]

卷:
第31卷
期数:
2016年03期
页码:
30-32
栏目:
论著
出版日期:
2016-06-15

文章信息/Info

Title:
Preparation and Identification of A Peptide Monoclonal Antibody Against Human Papillomavirus 18E6
文章编号:
1671-7414(2016)03-030-04
作者:
孔倩倩1a; 唐振华2; 相芬芬1a; 詹月萍1b; 许 健1b; 吴 蓉1a; 康向东1a
1.上海中医药大学附属普陀医院 a.检验科;
b.实验中心,上海 200062;
2.上海交通大学医学院附属国际和平妇幼保健院检验科,上海 200030
Author(s):
KONG Qian-qian1a; TANG Zhen-hua2; XIANG Fen-fen1a; ZHAN Yue-ping1b; XU Jian1b;  WU Rong1a; KANG Xiang-dong1a(1a
Department of Laboratory Medicine;
1b.Central Lab,Putuo Hospital Affiliated to Shanghai University ofTraditional Chinese Medicine,Shanghai 200062,China;
2.Department of Laboratory Medicine, the International Peace Maternity and Child Hea
关键词:
人乳头瘤病毒;  18 E6;  多肽;  单克隆抗体
分类号:
R392-33
DOI:
10.3969/j.issn.1671-7414.2016.03.009
文献标志码:
A
摘要:
目的 制备特异性抗人乳头瘤病毒(human papillomavirus,HPV)18 E6多肽抗体。方法 通过在线分析软件ABCpred和Bcepred预测HPV 18E6特异性多肽序列,将多肽序列与牛血清蛋白(bovine albumin,BSA)偶联,以偶联后多肽为抗原免疫BALB/c小鼠,采用杂交瘤技术制备抗HPV18 E6多肽单克隆抗体; 以HPV18阳性分泌物标本DNA为模板,聚合酶链式反应(polymerase chain reaction,PCR)获得HPV 18 E6基因,将该基因插入表达质粒pET-28a,转化大肠埃希菌BL21(DE3),异丙基硫代-β-D-半乳糖苷诱导目的蛋白表达; 采用Western Blot方法鉴定抗HPV18 E6多肽抗体与目的蛋白的反应性。结果 建立的杂交瘤细胞株能稳定产生HPV18 E6多肽单克隆抗体; 重组表达质粒pET-28a-HPV18 E6测序证明构建正确,成功表达HPV18 E6蛋白; 纯化后的抗HPV 18 E6多肽抗体可特异性识别原核表达HPV 18 E6蛋白。结论成功制备抗HPV18 E6多肽序列,该抗体具有较好的特异性。
Abstract:
Objective To prepare a peptide monoclonal antibody(McAb)against human papillomavirus 18E6 separately,and identify its specificity and pathogenicity.Metheds The advantage epitope peptide was designed and synthesized by ABCpred and Bcepred,and then used to immunize BALB/c mice after coupling with bovine serum albumin(BSA).And the McAb was prepared by hybridoma technique.HPV18E6 gene was amplified from cervical swab specimen containing HPV18 and inserted into expression vector pET-28a.The constructed recombinant plasmid was transformed to E.coli BL21(DE3)for expression under induction of isopropyl thio-β-D-galactoside.The expressed protein was used to identified the McAb had been prepared.Results The hybridoma cell lines could constantly produce MAbs against HPV18E6 peptides.Sequencing proved that recombinant plasmid pET-28a-HPV18E6 was constructed correctly.Western blotting showed that the anti-HPV18E6 peptides antibodycould specifically recognize HPV18E6.Conclusion A monoclonal antibody against the advantage epitope peptide of human papillomavirus 18E6prepared could specifically recognize HPV18E6 specifically.

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备注/Memo

备注/Memo:
基金项目:上海市科委引导项目(124119b1602,134119b1900),普陀区科委重大课题(2012PTKW001)。 作者简介:孔倩倩(1987-),女,硕士,初级检验技师,主要从事医学微生物研究,Tel:15921044663,E-mail:zhenxi628qian@163.com。 通讯作者:唐向东,男,博士,教授,主要从事免疫性疾病以及肿瘤的发病机理、诊断、治疗研究。
更新日期/Last Update: 2016-06-25