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United States Department of Agriculture

Agricultural Research Service

Title: Development of a Blocking Enzyme-Linked Immunosorbent Assay (Belisa) for Detection of Serum Antibodies to O157 Antigen of Escherichia Coli

item Laegreid, William
item Hoffman, Mark
item Keen, James
item Elder, Robert
item Kwang, Hwei-Sing - NTL UNIV OF SINGAPORE

Submitted to: Clinical and Diagnostic Laboratory Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 24, 1997
Publication Date: N/A

Interpretive Summary: Identification of cattle infected with E. coli O157:H7 has been difficult due to low levels of organisms in feces, intermittent shedding of organisms and limited duration of shedding. These problems have significantly affected interpretation of epidemiologic studies of this important food-borne pathogen. Serologic identification of infected cattle has been difficult due to cross-reactivity with related bacteria. Using a highly specific monoclonal antibody to the O157 antigen, we have developed a serologic test with very high specificity and which does not react with antibodies to other bacteria such as Brucella abortus. The test has been validated using cattle experimentally infected with E. coli O157:H7 and shown to be both sensitive and specific.

Technical Abstract: The O157 antigen of Escherichia coli shares structural elements with lipopolysaccharide (LPS) antigens of other bacterial species, notably Brucella abortus and Yersinia enterocolitica O9, which confounds interpretation of assays for anti-O157 antibodies. To address this problem, a blocking ELISA (bELISA) was designed using E. coli O157:H7 LPS as antigen and a monoclonal antibody specific for E. coli O157, designated 13B3, as the competing antibody. The bELISA had equivalent sensitivity and significantly higher specificity than the indirect ELISA (iELISA) detecting anti-O157 antibodies in sera from cattle experimentally inoculated with O157:H7. Only 13% of sera from naive heifers vaccinated for or experimentally infected with B. abortus had increased anti-O157 bELISA titers while 61% of anti-O157 iELISA titers increased. The bELISA is a sensitive and specific method for the detection of serum antibodies resulting from exposure to E. coli O157.

Last Modified: 4/20/2015
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