Reactivity of monoclonal antibody E5 with endotoxin. I. Binding to lipid A and rough lipopolysaccharides.
The murine IgM monoclonal antibody (mAb) E5 was produced by a hybridoma derived from spleen cells of a mouse immunized with the J5 rough mutant of Escherichia coli O111:B4. In a multicenter randomized placebo-controlled clinical trial, E5 has been shown to reduce significantly the mortality and morbidity of patients with Gram-negative sepsis. The characteristics of E5 binding to endotoxin were studied in vitro. We report here the results of binding to an extensive panel of rough lipopolysaccharide (LPS) and lipid A preparations. Using standard immunologic techniques, including enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay (RIA), as well as an antibody capture assay using immobilized antibody and a chromogenic Limulus amebocyte lysate (LAL) detection system, E5 was shown to bind to all rough LPS (chemotypes Ra through Re from Salmonella minnesota and E. coli J5) and lipid A preparations tested. E5 displayed a Kd for Ra LPS of approximately 6.5 nM. These results confirm and extend those reported previously and provide evidence that E5 binds specifically to lipid A and to the lipid A moiety of rough LPS.
Wood
D M
DM
Department of Preclinical Science, XOMA Corporation, Berkeley, California.
Parent
J B
JB
Gazzano-Santoro
H
H
Lim
E
E
Pruyne
P T
PT
Watkins
J M
JM
Spoor
E S
ES
Reardan
D T
DT
Trown
P W
PW
Conlon
P J
PJ
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D000918:Antibody Specificity; D000942:Antigens, Bacterial / Q000276:immunology; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D004731:Endotoxins / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology*; D006825:Hybridomas / Q000276:immunology; D007075:Immunoglobulin M / Q000378:metabolism; D008050:Lipid A / Q000276:immunology*; D008070:Lipopolysaccharides / Q000276:immunology*; D051379:Mice; D009154:Mutation; D011863:Radioimmunoassay
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D004731:Endotoxins; D007075:Immunoglobulin M; D008050:Lipid A; D008070:Lipopolysaccharides