Enzyme immunofiltration technique for rapid diagnosis of herpes simplex virus eye infections in a rabbit model

Abstract

A rapid enzyme immunofiltration assay for herpes simplex virus (HSV) was developed which is sensitive enough to detect viral antigens in eye swabs from rabbits with primary herpes keratitis. This assay employs a specially designed filter manifold to immobilize whole cells and cell debris dissociated from the swabs. Viral antigens trapped on the filters are then detected in an indirect immunoassay utilizing staphylococcal protein A conjugated with horseradish peroxidase. The assay required only 2.5 h to perform and could be read visually. Reconstruction experiments indicated that antigen from as few as 49 HSV-infected cells could be detected. Calcium alginate swabs were shown to recover more viral antigen than dacron swabs. The enzyme immunofiltration assay detected HSV antigens on 95% of the eye swabs from which infectious virus was recovered. HSV antigen was also detected in several swabs from infected eyes which did not yield infectious virus, presumably because the virus was neutralized by native antibody present in the lacrimal fluid. This enzyme immunofiltration assay technique lends itself to the elution of native antibody bound to the viral antigens, and this may be especially applicable in the diagnosis of recurrent HSV keratitis, where antiviral antibody in the lacrimal fluid may interfere with virus isolation and fluorescent-antibody or other virus detection assays.