Identification of tumor associated single-chain Fv by panning and screening antibody phage library using tumor cells.
OBJECTIVE
To study the feasibility of panning and screening phage-displaying recombinant single-chain variable fragment (ScFv) of anti-tumor monoclonal antibodies for fixed whole cells as the carriers of mAb-binding antigens.


METHODS
The recombinant phage displaying libraries for anti-colorectal tumor mAb MC3Ab, MC5Ab and anti-gastric tumor mAb MGD1 was constructed. Panning and screening were carried out by means of modified fixation of colorectal and gastric tumor cells expressed the mAb-binding antigens. Concordance of binding specificity to tumor cells between phage clones and parent antibodies was analyzed. The phage of positive clones was identified with competitive ELISA, and infected by E.coli HB2151 to express soluble ScFv.


RESULTS
The ratio of positive clones to MC3-ScF-MC5-ScFv and MGD1-ScFv were 60 %, 24 % and 30 %. MC3-ScFv had M(r) 32 000 confirmed by Western blot. The specificity to antigen had no difference between 4 positive recombinant phage antibodies and MC3Ab.


CONCLUSIONS
The modified process of fixing whole tumor cells is efficient, convenient and feasible to pan and screen the phage-displaying ScFv of anti-tumor monoclonal antibodies.
Nie
Yong-Zhan
YZ
Institute of Digestive Diseases,Xijing Hospital, Fourth Military Medical University, Changle West Road, Xi'an 710032,Shannxi Province China.
He
Feng-Tian
FT
Li
Zhi-Kui
ZK
Wu
Kai-Chun
KC
Cao
Yun-Xin
YX
Chen
Bao-Jun
BJ
Fan
Dai-Ming
DM
D000911:Antibodies, Monoclonal / Q000235:genetics*; D000912:Antibodies, Neoplasm / Q000235:genetics*; D000918:Antibody Specificity; D000951:Antigens, Neoplasm; D015179:Colorectal Neoplasms / Q000276:immunology; D005404:Fixatives; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D019151:Peptide Library; D013274:Stomach Neoplasms / Q000276:immunology; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D005404:Fixatives; D007135:Immunoglobulin Variable Region; D019151:Peptide Library