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目的利用噬菌体展示随机十二肽库淘选出与胃癌BGC823细胞特异性结合的多肽,为寻找胃癌早期诊断和治疗的标志物打下基础。方法以胃癌细胞BGC823为靶细胞,胃黏膜永生化上皮细胞GES为吸附细胞,在常温下对噬菌体随机十二肽库进行三轮消减淘选,挑取单克隆、扩增纯化噬菌体,并经ELISA和免疫组化法初步鉴定噬菌体克隆亲和力;测序阳性克隆,合成荧光素FITC标记多肽,鉴定其与胃癌细胞和胃癌组织的亲和力及特异性。结果三轮淘选后,经ELISA法初步鉴定,随机挑选的20个单克隆中11号(GC-11)多肽对BGC823细胞亲和力最高。细胞、组织免疫荧光实验进一步证明,荧光素标记的多肽FITC-GC-11对BGC823细胞和胃癌组织具有高亲和力。结论利用噬菌体随机十二肽库成功淘选出与胃癌细胞BGC823和胃癌组织具有较高亲和力的多肽GC-11。
OBJECTIVE: To use phage display random peptide library to screen out the specific binding peptide of gastric cancer BGC823 cells, so as to lay a foundation for searching the early diagnosis and treatment of gastric cancer. Methods BGC823 gastric cancer cells as the target cells, gastric mucosa immortalized epithelial cells GES as adsorbed cells at room temperature phage randomized twelve peptide library three rounds of abatement panning, picked monoclonal, amplified purified phage, and by ELISA Immunohistochemistry was used to identify the phage cloning affinity. The positive clones were sequenced and the FITC-labeled polypeptides were synthesized. The affinity and specificity of FITC-tagged polypeptides were evaluated. Results After three rounds of panning, preliminary identification by ELISA showed that the 20 monoclonal GC-11 polypeptides randomly selected had the highest affinity to BGC823 cells. Cell and tissue immunofluorescence experiments further demonstrated that the fluorescein-labeled polypeptide FITC-GC-11 has high affinity for BGC823 cells and gastric cancer tissue. Conclusions The peptide GC-11 with high affinity to gastric cancer cells BGC823 and gastric cancer was successfully screened by phage display randomized 12-mer peptide library.