Human monoclonal anti-HCMV neutralizing antibody from phage display libraries.
Human cytomegalovirus (HCMV) infection in immunocompromised patients causes considerable morbidity and mortality. Although ganciclovir prophylaxis reduces the incidence of HCMV disease, severe side effects raise serious problems. Thus, the development of new strategies for prophylaxis are clearly needed, and human monoclonal antibodies offer a potential alternative. We describe the cloning, using the phage display system, of a recombinant human Fab fragment against HCMV. A phage display library with 4 x 10(6) clones was panned three times against lysates of HCMV-infected cells, and screened by ELISA. Of six antigen-binding clones, one monoclonal antibody reacted strongly to HCMV. In immunostaining analysis, this Fab was able to stain HCMV-infected cells from 24 h post-infection (pi) through to 96 h pi, but not at 6 h pi. In the presence of cytosine arabinoside, HCMV-infected cells were not stained, even at 24 h pi. These results indicate that an HCMV protein that was recognized by the Fab was synthesized in the late phase of infection. In addition, this Fab exhibited neutralizing activity: at 1 microg/ml it reduced HCMV plaque formation by 50%. The Fab was able to neutralize three HCMV strains, but it did not neutralize HSV-1 or -2 infection.
Department of Molecular Life Science, Tokai University School of Medicine, Isehara, Kanagawa, Japan.
M
M
Takekoshi
F
F
Maeda
H
H
Tachibana
H
H
Inoko
S
S
Kato
I
I
Takakura
T
T
Kenjyo
S
S
Hiraga
Y
Y
Ogawa
T
T
Horiki
S
S
Ihara
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000276:immunology*; D001435:Bacteriophages; D002460:Cell Line; D003001:Cloning, Molecular; D003587:Cytomegalovirus / Q000235:genetics / Q000276:immunology*; D018985:Epitopes, B-Lymphocyte / Q000276:immunology; D005455:Fluorescent Antibody Technique; D005814:Genes, Viral; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D019151:Peptide Library; D017421:Sequence Analysis
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D018985:Epitopes, B-Lymphocyte; D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library