Characterization of monoclonal antibodies to the 44-kilodalton major outer membrane protein of the human granulocytic ehrlichiosis agent.
The major outer membrane proteins (OMPs) of the human granulocytic ehrlichiosis (HGE) agent, with molecular sizes of 44 to 47 kDa, are immunodominant antigens in human infection. Monoclonal antibodies (MAbs) to the OMPs were made by immunizing BALB/c mice with the purified HGE agent and then by fusing spleen cells with myeloma cells. The immunologic specificities of three MAbs (3E65, 5C11, and 5D13) were examined with five human HGE agent isolates and one tick isolate. By Western blot analysis, all three MAbs recognized the HGE agent but not Ehrlichia chaffeensis, Ehrlichia sennetsu, Ehrlichia canis, or their host cells. MAb 3E65 reacted with a 44-kDa protein in the homologous human isolate but not in the remaining five isolates. The two remaining MAbs recognized proteins with molecular sizes of 44 to 47 kDa in all six isolates. Western blot results with the OMP fraction of the six isolates were consistent with results with the whole HGE agent. Immunofluorescent-antibody staining and immunogold labeling with these MAbs showed that these antigens were primarily present on the membrane of the HGE agent. MAbs 5C11 and 5D13 recognized the recombinant 44-kDa protein by Western immunoblot analysis, but MAb 3E65 did not. Passive immunization with MAb 3E65 was more effective in protecting mice from HGE agent infection than with MAbs 5C11 and 5D13. These MAbs would be useful for analyzing the role of the major OMP antigens in HGE agent infection and for serodiagnosis.
Kim
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Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University, Columbus, Ohio 43210-1093, USA.
Rikihisa
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Y
D000818:Animals; D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D000942:Antigens, Bacterial / Q000737:chemistry; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000276:immunology*; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004536:Ehrlichia / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D016873:Ehrlichiosis / Q000276:immunology / Q000382:microbiology* / Q000517:prevention & control; D005260:Female; D006098:Granulocytes / Q000382:microbiology; D018922:HL-60 Cells; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007116:Immunization, Passive; D051379:Mice; D008809:Mice, Inbred C3H; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D017931:DNA Primers; D011994:Recombinant Proteins