Influenza PR8 HA-specific Fab fragments produced by phage display methods.
Anti-influenza hemagglutinin (HA) Fabs were isolated from a phage display library using purified HA of influenza virus A/Puerto Rico/8/34 (PR8; H1N1) as an antigen. Four Fab clones displaying a 25-50-fold higher binding signal to PR8 HA than the control were selected for further analysis and comparison with anti-PR8 monoclonal antibody (mAb). All four Fabs and mAb recognized the PR8 HA under non-reducing conditions but rarely bound to reduced PR8 HA. Inhibition of influenza virus infection on MDCK cells was observed with Fab1 and mAb in a dose-dependent manner while Fab3 and 4 exhibited only a partial inhibitory effect. Moreover, Fab1 clone and mAb exhibited cross-reactivity with the A/Peking/262/95 (A/Peking; H1N1) strain. The inhibitory effects of mAb on both influenza strains were more potent than Fab1, which is likely attributed to its higher affinity for the antigen. SPR analyses, in fact, revealed that Fab1 and mAb have K(D) of 1.5 x 10(-8) and 3.2 x 10(-9)M, respectively. These results strongly suggest that phage library-derived Fabs can be readily prepared and that such HA-specific Fabs with inhibitory action on influenza infection may be used to treat influenza patients.
Asanuma
Hideki
H
Department of Applied Biochemistry, School of Engineering, Tokai University, 1117 Kitakaname, Hiratsuka-shi, Kanagawa 259-1292, Japan; CREST, Japan Science and Technology Agency (JST), Japan.
Matsumoto-Takasaki
Ayano
A
Suzuki
Yujiro
Y
Tamura
Shin-Ichi
S
Sata
Tetsutaro
T
Kusada
Yu
Y
Matsushita
Misao
M
Fujita-Yamaguchi
Yoko
Y
D000818:Animals; D006389:Hemagglutinins, Viral / Q000276:immunology* / Q000302:isolation & purification*; D007140:Immunoglobulin Fab Fragments / Q000276:immunology* / Q000302:isolation & purification*; D053118:Influenza A Virus, H1N1 Subtype / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009975:Orthomyxoviridae / Q000276:immunology*; D019151:Peptide Library
D006389:Hemagglutinins, Viral; D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library