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加载HPV6E7抗原肽的HLA-A2单体及其四聚体的制备和鉴定 被引量：1

Preparation and Identification of The Soluble HLA-A2 Monomer and Tetramer Loaded with HPV6E7 Antigenic Peptide
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摘要 目的制备可生物素化的可溶性HLA-A2/HPV6E7单体,进一步组装成PE标记的可溶性HLA-A2/HPV6E7四聚体。方法以原核表达的sHLA-A2BSP为重链,β2m为轻链,与人工合成的HPV6E7(22-30)抗原肽(GLH-CYEQLV)利用稀释法进行共折叠复性得到单体。利用HLAI类分子单抗(W6/3)和抗Bzm抗体进行Westernblot和ELISA鉴定折叠产物的构象。以BirA酶对其进行生物素化,经过超滤离心后得到纯化的sHLA-A2/HPV6E7单体,再与Streptavidin-PE按4:1比例混合形成四聚体。结果该四聚体具有与HLA-A2阳性HPV6感染的CA患者的抗原特异性CTL结合活性。结论成功获得了天然构象的sHLA-A2/HPV6E7单体及PE标记的HLA-A2/HPV6E7四聚体。 Objective To acquire the soluble HLA-A2 monomer loaded HPV6E7 peptide and tetramer labeled by Streptavidin-PE. Methods The heavy chain of soluble HLA-A2 and β2m were expressed highly as insoluble aggregates in E. coli, and then the two subunits were refolded to form a sHLA-A0201-monomer by dilution method in the presence of Human Papillomavirus E7 peptide（GLHCYEQLV）. The refolded product was detected by ELISA and Western blot, Refolded A2-GLH monomer was biotinylated with a commercial BirA. The tetramer was then formed by mixing A2-GLH monomer with Streptavidin-PE in a ratio of 4 ： 1. Results Flow cytometry analysis indicated that this tetramer could bind to specific CTL from HLA-A2 ＋ Condyloma Acuminata （CA）, Conclusion The HLA-A2/HPV6E7 monomer and tetramer loaded HPV6E7 were successfully gained.
作者 张彩娥 邓云华 吴雄文 梁智辉 翁秀芳 卢小玲 夏芳珍 钟茂华 陆盛军
机构地区 武汉工业学院医学院免疫学教研究 华中科技大学同济医学院附属同济医院皮肤科 华中科技大学同济医学院免疫学系
出处 《中国皮肤性病学杂志》 CAS 北大核心 2006年第9期519-522,共4页 The Chinese Journal of Dermatovenereology
基金 国家自然科学基金资助项目(编号:30271201) 科技部973计划课题资助项目(编号:2001CB510008)
关键词 HLA-A2 sHLA-A2/HPV6E7四聚体 尖锐湿疣 细胞毒性T细胞 HLA-A2 sHLA-A2/HPV6E7 Tetramer Condyloma Acuminate CTL
分类号 R392.11 [医药卫生—免疫学]
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参考文献9

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二级参考文献8

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共引文献22

同被引文献6

  • 1张彩娥,费世江,吴雄文,梁智辉,翁秀芳,陆盛军,夏芳珍,钟茂华,龚非力.同种T细胞前体频率与MHC-抗原肽种类呈正相关[J].中华微生物学和免疫学杂志,2006,26(12):1028-1031. 被引量：1
  • 2Altman JD, Moss PA, Goulder PJ, et al. Phenotypic analysis of antigen-specific T lymphocytes. Science, 1996, 274(5284) : 94- 96.
  • 3Coleman N, Birley HD, Renton AM, et al. Immunological events in regressing genital warts. Am J Clin Pathol, 1994 , 102(6) : 768 -774.
  • 4Rener M, Claesson MH, Bregenholt S, et al. An improved method for detection of peptide-induced upregulation of HLA-A2 molecules on TAP-deficient T2 cells. Exp Clin Immunogenet, 1996, 13(1) : 30-35.
  • 5Youde SJ, Dunbar PR, Evans EM, et al. Use of fluorogenic histocompatibility leukocyte antigen-A * 0201/HPV 16 E7 peptide complexes to isolate rare human cytotoxic T-lymphocyte recognizing endogenous human papillomavirus antigens. Cancer Res, 2000, 60(2) : 365-371.
  • 6洪少林,王家璧,刘跃华,司静懿,许雪梅,郭秀婵,曾毅.北京地区尖锐湿疣患者皮损中人乳头瘤病毒的检测和分型[J].中国医学科学院学报,2002,24(4):397-400. 被引量：17

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中国皮肤性病学杂志
2006年 第9期

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