Screening and identification of mimotope of gastric cancer associated antigen MGb1-Ag.
OBJECTIVE
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 of enrichment 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) %.


CONCLUSIONS
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.
Han
Zhe-Yi
ZY
Institute of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi Province, China.
Wu
Kai-Chun
KC
He
Feng-Tian
FT
Han
Quan-Li
QL
Nie
Yong-Zhan
YZ
Han
Ying
Y
Liu
Xiao-Nan
XN
Zheng
Jian-Yong
JY
Xu
Mei-Hong
MH
Lin
Tao
T
Fan
Dai-Ming
DM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000951:Antigens, Neoplasm / Q000276:immunology / Q000302:isolation & purification*; D036022:Capsid Proteins; D004268:DNA-Binding Proteins; D000939:Epitopes; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D051379:Mice; D018716:Molecular Mimicry; D019151:Peptide Library; D013274:Stomach Neoplasms / Q000276:immunology*; D014407:Tumor Cells, Cultured; D014760:Viral Fusion Proteins
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D036022:Capsid Proteins; D004268:DNA-Binding Proteins; D000939:Epitopes; D007074:Immunoglobulin G; D019151:Peptide Library; D014760:Viral Fusion Proteins