Confirmation of human cytomegalovirus by reverse passive hemagglutination with monoclonal antibodies reactive to the major glycosylated peptide (GP-66)

Abstract

Sheep erythrocytes coated with three monoclonal antibodies, each reactive to a different epitope of the 66-kilodalton cytomegalovirus (CMV) matrix protein, were used in a reverse passive hemagglutination test with CMV-infected cell lysate to identify and confirm the CMV. The test is specific only for CMV, since 5 laboratory strains of CMV (AD169, Davis, Espilat, C-87, and Towne) and 10 clinical isolates reacted well, but uninfected MRC-5 cell lysate, lysates of herpes simplex virus types 1 and 2, varicella-zoster virus, and adenoviruses did not react. The reactive CMV lysate was confirmed by the pretreatment of CMV lysate with the three monoclonal antibodies followed by the addition of antibody-coated erythrocytes. The reverse passive hemagglutination test and the confirmatory blocking test are performed at the same time, requiring 2 h to complete. Since V-bottom microtiter 96-well plates and a 25-microliter pipette can be used to perform the test, it is ideal for CMV confirmation, especially when the equipment to read the fluorescent-antibody test or enzyme-linked immunosorbent assay is not available.