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目的:从天然的单链大容量噬菌体抗体库中筛选特异性的抗TNF-α的人单链抗体(scFv)。方法:以TNF-α为抗原,通过“吸附-洗脱-扩增”过程从大容量抗体库中筛选特异性抗体,对其抗原结合活性和序列进行分析鉴定。结果:经过4轮筛选,获得62个与TNF-α结合的阳性克隆,其中27个特异性结合的克隆,指纹分析有7种不同的可变区片段;其中5种获得可溶性表达;基因序列分析表明其轻重链分属VλⅠ和VHⅣ亚群。结论:利用单链大容量抗体库技术可以不经免疫制备获得特异性抗TNF-α的scFv,为今后的研究和应用提供依据。
OBJECTIVE: To screen specific anti-TNF-α human scFv from natural single-chain high-capacity phage antibody library. Methods: The specific antibody was screened from bulk antibody pool by “adsorption-eluting-amplification” process using TNF-α as antigen, and its antigen binding activity and sequence were analyzed and identified. RESULTS: After 4 rounds of screening, 62 positive clones that could bind to TNF-α were obtained. Of 27 specific binding clones, there were 7 different variable region fragments in fingerprinting analysis; 5 of them were soluble, and the sequence analysis Indicating that the light and heavy chains belong to Vλ Ⅰ and VH Ⅳ subgroups. Conclusion: It is possible to obtain specific anti-TNF-α scFv without immunization by using single-chain high-capacity antibody library technology, and provide the basis for future research and application.