摘要
AIM:To study the feasibility of panning and screeningphage-displaying recombinant single-chain variablefragment (ScFv) of anti-tumor monoclonal antibodiesfor fixed whole cells as the carriers of mAb-bindingantigens.ME,ODS: The recombinant phage displaying librariesfor anti-colorectal tumor mAb MC3Ab, MC5Ab andanti-gastric tumor mAb MGD1 was constructed.Panning and screening were carried outby means ofmodified fixation of colorectal and gastric tumor cellsexpressed the mAb-binding antigens. Concordance ofbinding specificity to tumor cells between phageclones and parent antibodies was analyzed. Thephage of positive clones was identified withcompetitive ELISA, and infected by E.coli HB2151 toexpress soluble ScFv.RESULTS:The ratio of positive clones to MC3-ScF-MC5-ScFv and MGD1-ScFv were 60 %, 24 % and 30 %. MC3-ScFv had Mr 32 000 confirmed by Western blot. Thespecificity to antigen had no difference between 4positive recombinant phage antibodies and MC3Ab.CONCLUSION:The modified process of fixing wholetumor cells is efficient, convenient and feasible to panand screen the phage-displaying ScFv of anti-tumormonoclonal antibodies.
AIM: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.coliHB2151 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. CONCLUSION: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.
基金
the National"863"Strategy for Science(102-10-01-06)
National Natural Science Foundation of China,No.39525020
