Consuruction of naive Fab fragment phage library

Objective:To construct a naive human Fab fragment phage display library.Methods:Peripheral blood lymphocytes were isolated from 800 ml blood,which was obtained from four healthy blood donors.The heavy chain Fd and light chain cDNA synthesized from the total RNA of lymphocytes were PCR amplified and the amplification products were ligated into the phagemid vector pComb3,then the ligated sample was transformed into competent E.coli XL1-Blue.The transformed cells were infected with VCSM13 helper phage to yield recombinant phage antibody Fabs.The phagemids abstracted from amplified E.coli were cut with endonucleases such as SacⅠ,XbaⅠ,SpeⅠand XhoⅠto monitor the insertion of the light chain or heavy chain Fd genes.Results:By combination of light chain and heavy chain genes,an antibody library containing 5.4×10~6 clones was obtained,and the cutting of enzymes showed that there were light chain or heavy chain Fd genes of the phagemids.Conclusion:This study can construct a human naive Fab phage display library which is useful to immunotheropy.