国家高技术研究发展计划(863计划)资助项目(102-09-03-04).
利用噬菌体随机肽库筛选抗TNF单抗表位的研究中,就抗体的选择、肽库富集的检测、筛选得到的表位多肽的验证及如何提高筛选的成功率等,进行了一些初步探索. 实验结果表明,由识别线性抗原位点的抗体较容易筛选到噬菌体呈现表位,具有较强中和活性的抗体,因多识别空间构型抗原位点而增加筛选难度. NC膜斑点印迹、ELISA及DNA测序均可作为筛选富集的检测方法. 用与天然抗原同源比较的方法及竞争性ELISA分析,可以帮助确定噬菌体呈现多肽是否是抗体表位.
Three monoclonal antibodies against tumuor necrosis factor(TNF-α) were selected to biopan a 6-mer phage-displayed random peptide library. After biopannings, the enrichment of binding phages were estimated using a dot blot, ELISA, DNA sequence methods.These results showed that three to four rounds of biopanning could be enriched enough and the results also revealed that the screening of epitope would be easier with an antibody recognized a conformation-independent antigen than conformation-dependent one. Competitive ELISA assay and homological comparasion between the original antigen and the motifs were helpful to identification of epitopes.
杨子义,王鑫,董家新,姚志建,沈倍奋.噬菌体显示技术用于抗体表位的筛选[J].生物化学与生物物理进展,1999,26(2):176-179
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