Prediction Of The B Cell Epitopes Of TRIM34 By Bioinformatic Analysis And Preparation Of Polypeptide Antibody

Purpose:TRIM34 belongs to Tripartite motif(TRIM)family proteins.Human TRIM34 gene is located on chromosome 11p15,and there are homologous molecules TRIM5,TRIM6 and TRIM22 nearby.Although there are commercial TRIM34 antibodies,the specificity and titer of the antibodies are not ideal.In this study,the specific segments of TRIM34 compared with TRIM6,TRIM5 and TRIM22 were selected through Vector NTI software analysis.Then,through Lasergene software,according to the secondary protein structure,antigen index,hydrophilicity,plasticity and other indicators of TRIM34 amino acid sequence,the appropriate sequence was selected to synthesize peptide antigen(amino terminal and KLH coupling).The New Zealand white rabbits were immunized with synthetic polypeptide antigen to prepare trim34 polypeptide antibody and to detect the titer and specificity of the antibody.Therefore,the purpose of this study was to:1.Predict the epitope of TRIM34 protein by bioinformatics software;2.Preparation of rabbit-derived trim34 peptide antibody;3.To verify the specificity of trim34 polypeptide antibody;4.To provide an important experimental tool for the localization of TRIM34 and the mechanism of the occurrence and development of viruses,tumors,degenerative diseases and other diseases.Methods:This study aims to predict B cell epitopes of TRIM34 and prepares polypeptide antibodies specifically against TRIM34 protein.B cell epitopes are screened by Vector NTI 11.5 and Lasergene7.1 software via analying TRIM34 amino acid sequence.The synthesized peptides of TRIM34 are cross-linked with keyhole limpet hemocyanin(KLH)at N-terminal and used to immunize rabbits for preparing antibody.ELISA and Western blot are respectively used to detect the antibody titer and specificity for TRIM34 proteins.Results:ELISA shows that the antibody titer in rabbit serum aganinst TRIM34 reaches 1:8000.Western Blot shows that the antibody can specifically recognize exogenous and endogenous TRIM34 proteins.at the same time TRIM34 paralogous molecules such as TRIM6,TRIM5,TRIM22 can not be recognized by the TRIM34 polypeptide antibody.Moreover,the TRIM34 polypeptide antibody can be used to showed the subcellular location of TRIM34 protein by indirect immuno-fluorescent staining using.Conclusion:In this study,antibody titer was successfully prepared with high specificity and strong anti-trim34 peptide antibody.This antibody is expected to provide an important experimental tool for the localization of TRIM34 and the mechanism of the occurrence and development of viruses,tumors and other related diseases.