A large variety of antiherpes compounds was compared for their inhibitory activity against laboratory strains and clinical isolates of herpes simplex virus (HSV) type 1 and type 2. From studies performed in primary rabbit kidney cell cultures, six, E-5-(2-bromovinyl)-2'-deoxyuridine, E-5-(2-iodovinyl)-2'-deoxyuridine, 5-vinyl-2'-deoxyuridine, 2'-fluoro-5-iodoaracytosine, acycloguanosine, and 5-iodo-2'-deoxycytidine, emerged as the most potent and selective antiherpes agents. For HSV type 1, the 50% inhibitory doses (ID50) were 0.008, 0.012, 0.018, 0.017, 0.04, and 0.06 micrograms/ml, respectively; those for HSV type 2 were 1, 2, 0.1, 0.05, 0.04, and 0.3 microgram/ml, respectively. These compounds did not inhibit host-cell metabolism or replication of vaccinia virus except at concentrations 100--10,000 times greater than the ID50 for any HSV. All were significantly less inhibitory for a thymidine kinase (TK)-deficient mutant of HSV type 1 than for normal strains, suggesting that phosphorylation by virus-induced TK was required to produce specific inhibition of HSV replication.