Identification of a target antigen recognized by a mouse monoclonal antibody to the bile canalicular surface of rat hepatocytes with a random phage display library.
We developed a monoclonal antibody (MoAb) (clone 5E8) against an antigen on the bile canalicular membrane of rat hepatocyte. By immunoblotting, MoAb 5E8 detected a band of 110 kD. In this study, we used the phage display technique to identify the target antigen recognized by MoAb 5E8. We screened a random phage display library expressing 12-mer peptide sequences and identified a peptide sequence, FHFNPYTGHPLT, as an epitope. We compared this peptide sequence with those of dipeptidyl peptidase IV (DPP IV, E.C.3.4.14.5) and Cell-CAM105, which proteins were located by a database search based on the information of tissue localization and approximate molecular weight of the MoAb 5E8 antigen, and sequence similarity with a region in DPP IV (amino acids 225-233) but not with Cell-CAM105 was found. In addition, we immunohistochemically stained various tissues (liver, small intestine, and kidney) of Japanese Fischer 344 rats, known to be deficient for DPP IV, with MoAb 5E8 and showed that the expression of MoAb 5E8 antigen was negligible or weak. In contrast, tissues sampled from the same organs of Sprague-Dawley rats, known to express DPP IV, were positively stained. These findings suggest that the antigen recognized by MoAb 5E8 is DDPIV and its major epitope is located in amino acids at positions 225-233.
Department of Medicine and Medical Science, Okayama University Graduate School of Medicine and Dentistry, Japan.
Masanori
M
Ariyoshi
Motowo
M
Mizuno
Yoshiko
Y
Morisue
Morizou
M
Shimada
Shirou
S
Fujita
Junichirou
J
Nasu
Hiroyuki
H
Okada
Hiroyuki
H
Shimomura
Kazuhide
K
Yamamoto
Takao
T
Tsuji
D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000941:Antigens / Q000235:genetics / Q000276:immunology*; D001648:Bile Canaliculi; D022781:Hepatocytes / Q000276:immunology*; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D019151:Peptide Library; D051381:Rats; D011916:Rats, Inbred F344; D017207:Rats, Sprague-Dawley; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D000941:Antigens; D019151:Peptide Library