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INFLAMMATORY BOWEL DISEASE |
1 Department of In Vitro Diagnostics, Otsuka Life Science Initiative, Otsuka Pharmaceutical Co. Ltd, Tokushima, Japan
2 Department of Internal Medicine, Division of Gastroenterology, Hyogo College of Medicine, Nishinomiya, Japan
3 Medical Chemistry Research Institute, Otsuka Pharmaceutical Co. Ltd, Tokushima, Japan
Correspondence to:
Correspondence to:
H Saito, Department of In Vitro Diagnostics, Otsuka Life Science Initiative, Otsuka Pharmaceutical Co. Ltd, 463-10 Kagasuno, Kawauchi-cho, Tokushima 771- 0192, Japan;
saitohh{at}assay.otsuka.co.jp
Background: Phage displayed random peptide technology has been utilised to identify binding epitopes of antibodies or receptor ligands.
Aim: To isolates peptides from a phage library which are specifically recognised by antibodies in serum from patients with Crohn’s disease (CD).
Methods: A phage displayed random peptide library composed of nine amino acids was established and sequentially screened using serum immunogloblin G obtained from CD patients.
Results: Five different CD specific peptides were isolated from the phage library. No homology in amino acid sequences was observed among four (CDP-1, -3 to -5) of the five peptides exhibiting different binding characteristics with each CD patient’s serum. In contrast, two peptides (CDP-1 and -2) had similar amino acid sequences and similar binding characteristics. Four multiple antigenic peptides (MAP, CDP-1, -3 to -5) were synthesised, and an enzyme linked immunosorbent assay (ELISA) using the four peptides was developed to detect serum antibodies against them. Fifty two of 92 CD patients (56.5%) were detected by ELISA, none of 20 ulcerative colitis (UC) patients, only one of 25 duodenal ulcer patients, and only three of 48 healthy subjects.
Conclusions: ELISA using the four peptides isolated in this study may be useful for the differential diagnosis of CD and UC.
Keywords: Crohn’s disease; phage displayed random peptide technology; inflammatory bowel disease; ulcerative colitis; serological markers
Abbreviations: CD, Crohn’s disease; UC, ulcerative colitis; DU, duodenal ulcer; IBD, inflammatory bowel disease; pANCA, perinuclear antineutrophil cytoplasmic antibodies; ASCA, anti-Saccharomyces cerevisiae antibodies; ELISA, enzyme linked immunosorbent assay; HRP, horseradish peroxidase; MAP, multiple antigenic peptide; IPTG, isopropyl-ß-D-thiogalactopyranoside; BSA, bovine serum albumin; D-PBS, Dulbecco’s phosphate buffered saline; LB, Luria-Bertani; TMB, 3,3',5,5'-tetramethylbenzidine; OD, optical density; ROC, receiver operating characteristic
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