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Screening and identification of mimotope of gastric cancer associated antigen MGb1-Ag
Authors:Zhe-Yi Han  Kal-Chun Wu  Feng-Tian He  Quan-Li Han  Yong-Zhan Nie  Ying HAN  Xiao-Nan Liu  Jian-Yong Zheng  Mei-Hong Xu  Tao Lin  Dai-Ming Fan
Affiliation:1. Institute of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi'an 710033, Shaanxi Province, China
2. Department of Biochemistry, Third Military Medical University, Chongqing, 400038, China
3. Department of Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an 710033, Shaanxi Province, China
4. Department of Gerontology, 323 Hospital of PLA,Xi'an 710054, Shaanxi Province, China
5. Department of Gastroenterology, 451 Hospital of PLA, Xi'an 710054, Shaanxi Province, China
Abstract:AIM: Using a monoclonal antibody against gastric cancer antigen named MGb1 to screen a phage-displayed random peptide library fused with coat protein pⅢ in order to get some information on mimotopes.METHODS: Through affinity enrichment and ELISA screening,positive clones of phages were amplified. 10 phage clones were selected after three rounds of biopanning and the ability of specific binding of the positive phage clones to MGb1-Ab were detected by ELISA assay (DNA sequencing was performed and the amino acid sequences were deduced)By blocking test, specificity of the mimic phage epitopes was identified.RESULTS: There were approximately 200 times ofenrichment about the titer of bound phages after three rounds of biopanning procedures. DNA of 10 phage clones after the third biopanning was assayed and the result showed that the positive clones had a specific binding activity to MGb1-Ab and a weak ability of binding to control mAb or to mouse IgG. DNA sequencing of 10 phage clones was performed and the amino acid sequences were deduced.According to the homology of the amino acid sequences of the displayed peptides, most of the phage clones had motifs of H(x)Q or L(x)S. And these 10 phage clones could also partly inhibit the binding of MGb1-Ab to gastric cancer cell KATO-Ⅲ. The percentage of blocking was from (21.0±1.6)%to (39.0±2.7)%.CONCLUSION: Motifs of H(x)Q and L(x)S selected and identified show a high homology in the mimic epitopes of gastric cancer associated antigen. There may be one or more clones which can act as candidates of tumor vaccines.
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