目的·探究非小细胞肺癌 （non-small cell lung cancer，NSCLC）免疫相关特征，发掘V-J基因中的肿瘤潜在标志物，为建立T细胞受体（T-cell receptor，TCR）-抗原识别预测模型提供基础。方法·收集704例NSCLC样本，用于构建一套系统的TCR组库分析流程，涵盖从原始数据到质控、过滤、TCR序列的识别和提取等上游分析部分，以及组库克隆分布、克隆分型、V-J基因共享性、互补决定区3（CDR3）分布特征以及克隆追踪等下游分析流程。使用Shannon-Weiner指数、Chao1等指数分析样本克隆分布。根据克隆扩增数目进行克隆分型，以探索不同分型间的差异。选取2例甲状腺乳头状癌（papillary thyroid carcinoma，PTC）样本进行去克隆扩增权重V-J基因共享分析，以探究低频克隆型对V-J基因共享性的影响。最后，通过分析V基因及高频克隆型CDR3分布特征和克隆追踪分析，监测肿瘤免疫前后克隆型频率的变化，寻找潜在的肿瘤标志物。结果·① NSCLC不同组织间克隆分布和克隆分型均存在显著差异，且在不同年龄与性别间存在显著差异。②在V-J基因共享性分析中发现了特定高共享V-J基因，同时在CDR3分布特征分析中存在非正态分布的高克隆V基因与氨基酸高频克隆型。③高频克隆型克隆追踪分析中，观察到NSCLC中高表达或新表达的高频克隆型，提示这些克隆型可作为潜在的肿瘤相关抗原或新抗原结合CDR3参考序列。④通过去克隆扩增权重V-J基因共享分析，发现原本低表达的TRBJ2-5基因表达频率显著增加，提示该基因可能为肿瘤免疫反应的潜在低频关键基因。结论·TRAV21和TRBV6.5基因在NSCLC中高克隆扩增，可作为潜在肿瘤标志物。

Objective ·To explore the immune-related characteristics of non-small cell lung cancer (NSCLC), discover potential tumor markers in V-J genes, and lay the foundation for establishing a TCR-antigen recognition prediction model. Methods ·A total of 704 NSCLC samples were collected to establish a comprehensive T-cell receptor (TCR) repertoire analysis workflow. The upstream analysis included steps such as raw data processing, quality control, filtering, TCR sequence identification, and extraction. The downstream analysis included repertoire clone distribution, clone typing, V-J gene sharing, CDR3 distribution characteristics, and clone tracking. The sample clone distribution was analyzed by using indices such as Shannon-Weiner index and Chao1 index. Clone typing was performed based on the number of clone amplifications to explore differences among different types. The degree of V-J gene segment sharing was analyzed, and the sharing of low-frequency clone types was determined through clone amplification weight analysis of V-J genes by using two samples of papillary thyroid carcinoma. Finally, analysis of the distribution characteristics of V genes and high-frequency clone type CDR3, and clone tracking analysis were conducted to monitor changes in tumor immune clone frequencies before and after analysis, aiming to identify potential tumor markers. Results ·① Significant differences were observed in clone distribution and clone typing among different NSCLC tissues, as well as among different ages and genders. ② Specific highly-shared V-J genes were identified in the analysis of V-J gene sharing, and non-normal distribution of high-clone V genes and amino acid high-frequency clone types were found in the CDR3 distribution analysis. ③ In the analysis of high-frequency clone type clone tracking, highly expressed or newly expressed high-frequency clone types were observed in NSCLC, suggesting that these clone types could serve as potential tumor-associated antigens or bind with CDR3 reference sequences of new antigens. ④ It was found that the expression frequency of TRBJ2-5 gene, originally low-expressed, significantly increased, indicating its potential role as a key low-frequency gene in tumor immune response. Conclusion ·The TRAV21 and TRBV6.5 genes show high clone amplification in NSCLC and could serve as potential tumor biomarkers.