Development, characterisation and diagnostic application of monoclonal antibodies against Yersinia pestis fibrinolysin and coagulase

A library of monoclonal antibodies (MAbs) which recognised different epitop es of Yersinia pestis fibrinolysin (Fib) was developed. These MAbs were spe cies-specific and demonstrated no cross-reaction in indirect immunofluoresc ence tests (IIFT) with other gram-negative bacteria possessing plasminogen activator activity. All the MAbs provided equally high levels of immunofluo rescence with pPst(+) Y. pestis strains cultivated at 37 degrees C and at 2 8 degrees C, In all cases, the MAbs inhibited both fibrinolytic and coagula se (Coag) activities of Y pestis in Fib-activity inhibition and coagulase-a ctivity inhibition reactions, and reacted with 35- and 37-kDa proteins of Y , pestis in immunoblotting, demonstrating bifunctional activity possibly si milar to the properties of MAbs produced by hybrid hybridomas, On the basis of these and earlier studies, the immunochemical identity of Fib and Coag, two distinct subunits of a bifunctional fusion protein whose specific func tional activity depends upon the temperature factor, was established. A new rapid, cheap, strictly specific and safe dot-ELISA based on the use of MAb against Y. pestis Fib (MAb-Fib) for reliable identification of Y. pestis s trains was developed. This technique has great advantages over monoclonal d iagnostic kits based on the use of MAb against Y. pestis fraction I (FI) be cause it allows detection of plague bacilli grown at 37 degrees C as well a s at 28 degrees C, This dot-ELISA will be valuable as a clinical diagnostic tool and might be applicable to field studies and plague surveillance.