【摘要】 目的:寻找被T细胞识别的食管癌细胞抗原肽,为食管癌免疫治疗奠定基础。方法:应用pH3.3的枸橼酸磷酸盐缓冲液间隔24h多次酸洗贴壁生长的Eca109细胞,获得人主要组织相容性复合体Ⅰ类分子(HLAⅠ)类分子结合的多肽,经超过滤,反相HPLC色谱层析得到不同组分多肽,DC递呈抗原肽刺激特异性细胞毒T淋巴细胞(CTL)反应,51Cr杀伤实验检测CTL对肿瘤细胞杀伤作用。结果:Mr小于3000的混合多肽经过反相HPLC可获得50多个组分,混合肽、P17、P29和P41组分体外诱导实验表明可以诱导出较强的CTL反应。结论:酸洗法能有效获得HLAⅠ类分子结合的多肽抗原,Mr小于3000的多肽成分中3个组分存在能引起特异性的抗肿瘤CTL免疫反应的肿瘤抗原,具有潜在的免疫治疗作用。更多还原

【Abstract】 Aim: To seek the antigenic peptides from the esophageal carcinoma cell, which could be identified by T lymphocytes and lie a foundation of immune theraphy for esophageal carcinoma.Methods: Eca-109 cell line was treated by citrate-phosphate buffer at pH 3.3, to denature HLA class Ⅰ molecules and release HLA class Ⅰ molecules-associated peptides into the supernatant. The acid solution containing peptides was desalted, vacuum concentrated, and centrifuged on Microcon YMGAAB23. Peptides were subsequently fractioned by reverse-phase high performance liquid chromatography (RP-HPLC). PBMCs were stimulated by peptide-pulsed dendritic cells (DCs). The specific cytotoxicity against the Eca-109 cell line was determined by 51Cr release assay.Results: RP-HPLC analysis showed that about fifty different fractions were obtained from M_r<3 000 HLA-Ⅰ molecules-associated peptides. P17, P29 and P41 fractions excited cytotoxic activity of CTLs in vitro.Conclusion: Mild acid wash method can efficiently elute the HLA-Ⅰ molecules-associated peptides from Eca-109 cell line. There are three fractions (P17, P29, P41) that can stimulate specific CTLs reaction against Eca-109 cell line.更多还原