Human monoclonal antibodies to cytomegalovirus. Characterization and recombinant expression of a glycoprotein-B-specific antibody.
Human monoclonal antibodies (mAb) to human cytomegalovirus (HCMV) were established from spleen cells of a HCMV-positive donor. The antibodies (gamma 3, lambda) secreted from a stable heterohybridoma cell line were further characterized by immunoprecipitation and immune-fluorescence microscopy using HCMV infected cells and recombinant cell lines expressing HCMV glycoprotein B. The antibody reacted with the entire glycoprotein B or the extracellular domain expressed as glycoprotein-B--beta-galactosidase fusion protein in the native state, but the antibody was not neutralizing HCMV. Denatured and reduced forms of glycoprotein B were not recognized by this antibody, however, native glycoprotein B on the surface of infected cells was detected efficiently. The genes encoding the Fab part of the antibody were cloned and expressed in Escherichia coli. Recombinant Fab fragments specifically binding the extracellular domain of glycoprotein B could easily be isolated from the periplasmic space. Recombinant antibodies provide the opportunity to modify effector functions and to add tags to diagnostic antibodies for more efficient detection of CMV-infected cells.
Böldicke
T
T
Gesellschaft für Biotechnologische Forschung mbH, Department of Applied Genetics, Braunschweig, Germany.
Haase
B
B
Böcher
M
M
Lindenmaier
W
W
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000737:chemistry; D001483:Base Sequence; D002478:Cells, Cultured; D006224:Cricetinae; D003587:Cytomegalovirus / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis / Q000737:chemistry; D007135:Immunoglobulin Variable Region / Q000737:chemistry; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry; D014759:Viral Envelope Proteins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C040495:glycoprotein B, Simplexvirus