Construction And Screening Of Fab Fragment Phage Display Library From Lymphocytes Of A Rhesus Immuned With SMMC-7721

Aim: 1.To construct an immuned Fab fragment phage display library from lymphocytes of a rhesus immuned with SMMC-7721;2.To express the extracellular domain of CD147 in E.coli;3.To screen the antibodies angainst extracellular domain of CD147.Methods:Total RNA was extracted routinely from lymphocytes in blood of a rhesus immuned with HCC SMMC—7721,and reverse transcriptase PCR was employed to amplify cDNA of the heavy chain Fd fragment and light chain k fragment. The products from PCR that use the cDNA talked above as the template were inserted into vector pComb3 respectively to construct Fab fragment library. Clone the gene of extracellular domain of CD147 into vector pET-22b(+) and express it in E.coli BL21. Protein of extracellular domain of CD147 was used as the target to screen the Fab library.Conclusion: an immuned Fab fragment phage display library including 2.7x10 members was constructed with the recombination rate of 60 % of light chain and of 100% of heavy chain. The immuned Fab fragment phage display library was enriched 126 times after 3 cycles of panning with extracellular domain of CD147, and one special high affinity phage antibody was screened out.