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- W2355673107 abstract "[Objective] To construct a naive human Fab fragment phage display library,gp96 purified from the tissue of renal cell carcinoma to screen the anti-gp96 antibodies and provide a platform for human antibody preparation and a basis for new therapy of the malignant tumors.[Methods] With lymphoprep,peripheral blood lymphocytes were isolated from 50ml of blood,which was obtained from a healthy blood donor.The heavy chain Fd and light chain cDNA synthesized from the total RNA of lymphocytes were amplified by PCR with variable regions 5' and 3' primers of heavy and light chain,and the amplifica-tion products were ligated into the phagemid vector pComb3,then the ligated sample was transformed into competent E.coli XL1-Blue by Electroporation.The transformed cells were infected with VCSM13 helper phage to yield recombinant phage anti-body Fabs.The phagemids abstracted from amplified E.coli were cut with endonucleases such as SacⅠ,XbaⅠ,SpeⅠand Xho Ⅰ,and the phage antibody Fabs or phagemids abstracted from amplified E.coli were amplified by PCR to monitor the insertion of the genes of light chain or heavy chain Fd fragment.The gp96 purified from the renal cell carcinoma tissue by affinity chromatography on concanavalin-A sepharose and DEAE-sepharose ion exchange chromatography were utilized as antigens to pro-cess three rounds of panning to the original Fab antibody library.[Results] The quantity of total RNA and cDNA were quali-fied.By combination of light chain and heavy chain genes,an antibody library containing 1.2×106 clones was obtained,and both the cutting of enzymes and PCR showed that there were the genes of light chain or heavy chain Fd fragment in the phagemids.The gp96 protein was obtained from renal cell carcinoma tissue.After having been panned by gp96,the original an-tibody library gained enrichment in 71.5 times.[Conclusion] Utilizing the technology of phage surface display,special antibody can be gained from the human naive Fab phage display library,which can be used as a new therapy for tumors." @default.
- W2355673107 created "2016-06-24" @default.
- W2355673107 creator A5053096377 @default.
- W2355673107 date "2010-01-01" @default.
- W2355673107 modified "2023-09-23" @default.
- W2355673107 title "SCREENING OF THE FAB ANTIBODIES AGAINST GP96 FROM RENAL CELL CARCINOMA WITH HUMAN NAIVE PHAGE ANTIBODY LIBRARY" @default.
- W2355673107 hasPublicationYear "2010" @default.
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