An assay for infection with one or more herpes simplex viruses that comprises a) dividing an antibody-containing serum sample into at least three serum subsamples, and b) separately incubating each with a physical matrix of cell antigens from i) uninfected cells, (ii) HSV-1-infected cells, or (iii) HSV-2-infected cells to form at least three serum subsample admixtures so that antibodies present in each admixture can immunoreact to form matrix-bound antibodies and at least three preadsorbed serum subsample portions. Each preadsorbed serum subsample portion is incubated with a mixture of matrix-bound antigens from cells uninfected by HSV-1 or HSV-2, infected by each of HSV-1 and by HSV-2 to permit antibodies to immunoreact with antigens present to form three matrix-bound immunoreactants. The amount of each immunoreaction is determined, and reaction amounts are prognostic for whether the subject whose serum was tested is infected by one, both, or neither of HSV-1 and HSV-2.