Construction of human phage display antibody ScFv library and identification of antibody ScFv against lung adenocarcinoma / 基础医学与临床

Objective To construct a human phage single chain-antibody library, and to sieve out the antibody ScFv against lung cancer from the library. Methods Total RNA was abstracted from lymph node tissue of the lung cancer, and was used to amplify V_H and V_L gene by RT-PCR. V_H and V_L were joined by a DNA linker by SOE-PCR to form the single chain variable fragment ( ScFv) gene. ScFv gene was coloned into the phage vector pCANT-AB5E. Panning against lung cancer cell line A549 was performed and positive clones were chosen for soluble expression. Results A recombination phage single chain-antibody library was constructed. After 4 rounds panning, the number of eluted phages increased by 115 times. Positive reactions to A549 were detected in 7 of 10 random clones. The human ScFvs against lung cancer were produced and confirmed by SDS-PAGE and ELISA analysis. Conclusion ScFvs against lung cancer were acquired by the construction of phage single chain-antibody library. The soluble ScFvs has specificall avidity to human lung cancer cells.

Construction of human phage display antibody ScFv library and identification of antibody ScFv against lung adenocarcinoma / 基础医学与临床

Objective To construct a human phage single chain-antibody library,and to sieve out the antibody ScFv against lung cancer from the library.Methods Total RNA was abstracted from lymph node tissue of the lung cancer,and was used to amplify VH and VL gene by RT-PCR. VH and VL were joined by a DNA linker by SOE-PCR to form the single chain variable fragment (ScFv) gene. ScFv gene was coloned into the phage vector pCANTAB5E. Panning against lung cancer cell line A549 was performed and positive clones were chosen for soluble expression.Results A recombination phage single chain-antibody library was constructed. After 4 rounds panning,the number of eluted phages increased by 115 times. Positive reactions to A549 were detected in 7 of 10 random clones. The human ScFvs against lung cancer were produced and confirmed by SDS-PAGE and ELISA analysis.Conclusion ScFvs against lung cancer were acquired by the construction of phage single chain-antibody library. The soluble ScFvs has specificall avidity to human lung cancer cells.