Screening Of Rheumatoid Arthritis Associated Peptide By Phage Display Random Library And Its Diagnostic Value

Background Rheumatoid arthritis(RA) is a kind of crippled autoimmune disease.The cure rate of RA may be improved if the patients get early diagnosed and treated, at least they are biologically reversible. The early and specific diagnostic value in RA of anti-keratin antibody(AKA),one of the multiple autoantibodies,has been determined. But it is difficult to extract antigen used for AKA test.Now, the antigen mostly comes from the stratum of Wister rat oesopnagus or human's callosities. The individuality expresses heterogeneous levels of keratin for the gene variation, so it is not easy to establish the standard test method.The purpose of this study is to obtain the mimotope by screening a phage random peptide library using polyclonal AKA, and evaluate its diagnostic value. My research maybe provide some database to synthesize the peptide for early diagnosis in RA, to explore the pathogenesis and immunol therapy of RA.Methods IgG was purified from AKA positive pooled serum of the RA patients by 33.3% (NH4 )2SO4(AKA was measured with indirect immunofluorescence technique) , then the antibody was used as the ligand for biopanning of a 12-mer phage random peptide library for 4 cycles.The were identified by ELISA, and the ssDNA of the positive clones were extracted and purified for sequencing.The serum of patients with RA,SLE,and normal people were detected with the positive clone of different DNA sequence .Results After 4 rounds of screening,9 positive clones were acquired ,they expressed 2 kinds of DNA sequence named mimotope 1and mimotope 2.The diagnostic specificity,positive predictive value, And Youden are 94.4%,91.7% and 149.4%.The peptide of mimotope 1 is associated with RA, Its amino acid sequence is QSESAGPTTSRR.The sequence is similar to 137～148 amino acid sequence of human filaggrin.Conclusion 1.It is feasible to find antigen epitope by polyclonal AKA in sera of patient with RA.2.The peptide associated with RA is acquired by screening a phage random peptide library using polyclonal AKA of patients with RA.It is specific and confirmed in diagnosing RA.