Screening and evaluating the mimic peptides as a useful serum biomarker of ankylosing spondylitis using a phage display technique

Abstract

To screen specific serum biomarker for ankylosing spondylitis (AS) using a phage random peptide library. A phage random peptide library of random peptide 12-mers was immunoscreened with purified immunoglobulin (Ig) G from sera of AS patients. Positive clones obtained after three rounds of biopanning were detected with ELISA and sequenced. Reaction of the screened positive clones with sera from AS patients, systemic lupus erythematosus (SLE) patients, rheumatoid arthritis (RA) patients, osteoarthritis (OA) patients and healthy controls was detected using phage ELISA. Correlation among erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and the absorbance value of the positive clone in phage ELISA was examined in AS patients. Seventeen out of twenty randomly selected phage clones exhibited specific reaction with purified sera IgG from AS patients, among them seven coming from the same clone whose inserted peptide sequence was LALPPLAPNHHH (named "AS1"). Phage ELISA results showed that the positive reaction rate of the AS1 clone was 92.0% with AS patients, significantly different (P<0.01) from those with SLE patients (56.7%), RA patients (50.0%), OA patients (13.3%), and healthy controls (14.0%). Absorbance value of the AS1 clone in phage ELISA was significantly higher than those in the other groups (P<0.05). In addition, the absorbance value of the AS1 clone showed no statistically significant correlation with ESR and CRP in AS patients, suggesting that AS1 detects AS patients through a unique mechanism other than inflammation. The short peptide AS1 obtained through screening of a phage random peptide library with purified serum IgG from AS patients can specifically react with the sera of AS patients, and thereby may be a candidate of AS-specific serum biomarkers.