Production of neutralizing monoclonal antibodies to Escherichia coli heat-stable enterotoxin.
In an effort to develop new approaches to the study and control of infectious diarrhea, we prepared murine monoclonal antibodies to the Escherichia coli heat-stable enterotoxin (STa). The toxin was purified from E. coli culture media and conjugated to bovine serum albumin. The STa-bovine serum albumin conjugate was used to immunize BALB/c mice, and the immune spleen cells from these mice were fused with SP2/0 myeloma cells. Resultant hybridomas were screened in an enzyme-linked immunosorbent assay protocol against 500 ng of STa-bovine serum albumin bound to microtiter wells as the solid-phase antigen. Five stable clones were selected and grown further in ascites fluid, which demonstrated anti-STa activity at dilutions of up to 1:500,000 in the enzyme-linked immunosorbent assay for heat-stable enterotoxin. In a competitive enzyme-linked immunosorbent assay format, the antibodies recognized several human and porcine strains of STa to various extents, but did not recognize E. coli heat-labile toxin, cholera toxin, or staphylococcal enterotoxin B. The antibodies were all able to bind lactoperoxidase-labeled [125I]STa, and antibody 20B3 was also able to dissociate [125I]STa bound to toxin receptors on rat jejunal villous cells. Preincubation of STa with antibodies 20B3 or 20F5 led to a concentration-dependent neutralization of toxin activity in a suckling mouse intestinal secretion assay. These antibodies are likely to provide new tools for the continued study of STa structure-function relationships and may lead to improved diagnosis and treatment of E. coli-induced infectious diarrhea.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000936:Antigen-Antibody Complex; D001427:Bacterial Toxins / Q000276:immunology*; D003967:Diarrhea / Q000276:immunology / Q000382:microbiology*; D004768:Enterotoxins / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000276:immunology*; D004927:Escherichia coli Infections / Q000276:immunology*; D029968:Escherichia coli Proteins; D006801:Humans; D007583:Jejunum / Q000382:microbiology; D007700:Kinetics; D051379:Mice; D008807:Mice, Inbred BALB C; D051381:Rats
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; C034151:heat stable toxin (E coli)