IMMUNOLABELINGOF ANTIBIOTIC-TREATED GRAMNEGATIVE BACTERIA

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Abstract

The use of cross-reactive antibodies to the core polysaccharide (CPS) of E. coli J5 to protect against the toxic effects of Gram-negative bacterial sepsis has been proposed. However, it has been shown that conserved lipopolysaccharide (LPS) epitopes on viable cells are not necessarily available for binding by cross-reactive antibody. To test the hypothesis that these conserved epitopes become available for binding after exposure to bacteriolytic antibiotics, Immunoelectron microscopy of control and antibiotic-treated bacterial cells was performed using a murine monoclonal antibody. M1B1 is an IgG directed to the CPS of LPS and binds purified LPS from E. coli K1:07 strain C94, but not viable cells, as demonstrated by a radiolabelled binding assay. Binding of M1B1 to control and antibiotic-treated C94 cells was detected with a gold bead-labelled goat anti-mouse antibody. M1B1 did not bind to intact C94 cells, but, after exposure to moxalactam, M1B1 bound to disrupted areas on the cell surface and to intracellular contents extruding from lysed cells. Specificity of M1B1 binding to LPS was confirmed using an irrelevant monoclonal antibody. We conclude that sequestered LPS epitopes within bacteria become exposed after antibiotic treatment. Exposure of these common epitopes may facilitate therapeutic intervention using cross-reactive antibodies.

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