Development of latex agglutination test for detection of antibodies against capripoxvirus

Abstract

Currently the diagnostic tests in use for Capripoxvirus infection are immunofluorescent antibody test (IFAT) and virus neutralization test (VNT), which are slow because they are tissue culture dependant. IFAT is subjective while VNT is inconsistent. Earlier work using whole Capripoxvirus to sensitize latex beads proved to be effective in detection of Capripoxvirus antibodies. A major structural protein P32 which is the first to raise antibody response and is common to all the strains of Capripoxvirus has been identified and its gene cloned. The gene Q13L coding for the Capripoxvirus group specific structural protein P32 was expressed in Escherichia coli using plasmid pGEX-2T as a fusion protein with glutathione-s-transferase and then purified on glutathione sepharose affinity chromatography column. A latex agglutination Test (LAT) was developed using the purified P32 antigen and guinea pig detector antiserum raised against the P32 antigen. The LAT was used to screen one hundred livestock field sera for antibodies to Capripoxvirus. The serum samples from infected animals tested by LAT and VNT showed that the LAT assay was 23% more sensitive, much simpler and rapid as compared to the VNT. LAT picked antibodies earlier after inoculation as well as at higher levels than VNT. In addition LAT was found to be specific for Carpripoxvirus since it did not pick antibodies to Orthopoxvirus (vaccinia) and parapoxvirus (orf). The LAT test will reduce the reliance of diagnostic laboratories on tissue culture facilities.