Screening and biological characteristics identification of human anti-CD147 single chain antibody fragments

LIU Yan; CHEN Xiuxiu; WANG Yuxiao; DING Li; FU Kaifei; ZHOU Lijun

doi:10.13431/j.cnki.immunol.j.20180009

Abstract：In recent years,the target therapy for tumors has becoming more and more important,therefore people are actively looking for more suitable targets for the treatment of tumors.CD147,widely expressed in various malignant tumor cells or inflammatory cell but not normal cell,is a potential research target,therefore the study was designed to screen the specific human anti-CD147 single chain antibody fragments(scFvs) through phage-displayed library technology and to identify their biological characteristics.At the beginning,the eukaryotic expression product of CD147 protein was used as antigen to screen for human anti-CD147 scfvs by phage-displayed library technology.After four rounds of enrichment screening,the binding activity of clones selected randomly were determined by ELISA.Then the binding specificity of phage antibodies and soluble scFvs obtained were identified by competitive ELISA and Western blot respectively.Finally,the DNA sequences of obtained clones were analyzed by gene sequencing and their relative affinity was determined by thiocyanate elution method.There were 4 specific positive clones against CD147 were obtained from 220 random clones,3 of which were achieved soluble expression,and their specificities were proved by competitive inhibition and Western blotting.At the same time,DNA sequence analysis confirmed that the 3 positive clones were human origin with different genotypes.And the relative affinity indexes of the 3 human anti-CD147 scFvs were measured by thiocyanate elution:0.20 mol/L,0.35 mol/L and 0.6 mol/L respectively.Taken together,specific human anti-CD147 scFvs have obtained from phage antibody library,which could benefit further study on the function and application of the antibody.