骆驼体液免疫系统是目前发现的唯一缺失轻链天然存在重链抗体的哺乳动物。尽管由于缺少重链和轻链的组合降低了其抗体库的多样性，但重链抗体可变区（也称纳米抗体）尤其是CDR3的特殊结构弥补了其多样性的不足，而且还扩大了重链抗体识别特殊抗原表位的能力。为了全面了解骆驼抗体独特的体液免疫系统的形成机制和多样性组成，特别是了解重链抗体在抗体库中的组成和分布，同时建立高效的基于序列深度分析的纳米抗体大规模发现技术，本项目以当前广泛采用的高通量测序（HTS）技术为基础，开展以下研究：(1)HTS分析骆驼抗体库中VDJ胚系基因的分布、IgG抗体亚型的比例、VH(H)序列和CDR3的长度分布、BCR谱系分布以及体细胞高突变和亲和力成熟等；(2)建立针对骆驼抗体HTS数据的分析方法；(3)建立基于HTS和VHH序列丰度和数据挖掘的纳米抗体发现平台；(4)实验测定候选抗体的抗原结合特性和亲和力，评价该技术的可行性。

The humoral immune response of the Camelidae is unique as these animals are the only known mammals that possess functional homodimeric heavy-chain antibodies (HCAbs) devoid of light chain naturally. Although the contribution of combination between heavy and light chain to the diversity of antibody repertoire was missing，the overall diversity was not lowered owing to the diverse germline VHH and specific mechanisms combined with unique structure of VHH CDR3. In contrast, it enlarged the antigen binding sites of HCAbs, especially, the sites which classical heteromeric antibodies can not easily accessible. For comprehensively understanding the mechanisms of antibody repertoire development in camelids，especially the composition and distribution of HCAbs in camelids humoral immune system，the newly developed high throughput sequencing (HTS) platform would be employed to deep analysis and characterizing of camel antibody repertoires. Meanwhile，we would develop novel large scale nanobody discovery platform based on HTS produced VHH sequences and data mining. The application of this project consisted following contents: (1) HTS analysis camel antibody repertoire, including VDJ segment germline analysis, the content ratio of IgG isotypes, VH(H) identification, frequency distribution of abundant CDRH3, BCR linage，somatic hypermutation and affinity maturation etc；(2) Development of bioinformatics analysis pipeline for handling vast camel antibody gene repertoire HTS data；(3) Development of nanobody discovery platform based on sequence frequency and ranking combined with data mining; (4) Evaluation the feasibility of HTS antibody discovery platform by analysis of the antigen binding specificity and affinity of HTS discovery interested nanobodies experimentally.