The molecular characterization of the epitope repertoire on herpes simplex virus (HSV) antigens would greatly expand our knowledge of HSV immunity and improve immune interventions against herpesvirus infections. HSV glycoprotein D (gD) is an immunodominant viral coat protein and is considered an excellent vaccine candidate antigen. By using the TEPITOPE prediction algorithm, we have identified and characterized a total of 12 regions within the HSV type 1 (HSV-1) gD bearing potential CD4 ؉ T-cell epitopes, each 27 to 34 amino acids in length. Immunogenicity studies of the corresponding medium-sized peptides confirmed all previously known gD epitopes and additionally revealed four new immunodominant regions (gD 49-82 , gD 146-179 , gD 228-257 , and gD 332-358 ), each containing naturally processed epitopes. These epitopes elicited potent T-cell responses in mice of diverse major histocompatibility complex backgrounds. Each of the four new immunodominant peptide epitopes generated strong CD4 ؉ Th1 T cells that were biologically active against HSV-1-infected bone marrow-derived dendritic cells. Importantly, immunization of H-2 d mice with the four newly identified CD4 ؉ Th1 peptide epitopes but not with four CD4 ؉ Th2 peptide epitopes induced a robust protective immunity against lethal ocular HSV-1 challenge. These peptide epitopes may prove to be important components of an effective immunoprophylactic strategy against herpes.Genital, dermal, and ocular herpes simplex virus (HSV) infections cause prevalent, lifelong recurrent infections, with a spectrum of clinical manifestations, including cold sores, genital lesions, corneal blindness, and encephalitis (41,58,60,64,81). Despite the availability of many interventional strategies, there has been a constant increase of HSV prevalence during the last 3 decades (27,41,47). Several challenges face the development of an effective herpes vaccine that could help control this epidemic, including the uncertainty about the exact immune correlates of protection, the identification of immunogenic epitopes, and the development of an effective and safe immunization strategy (9,13,33,35,58,60).Despite previous emphasis on antibody (Ab) and CD8 ϩ T-cell responses (34, 42), there is growing evidence to support a pivotal role for the T-helper type 1 (Th1) subset of CD4 ϩ T cells in antiherpesvirus immunity (29,37,43,46,54,63,71). CD4 ϩ T cells are required for the protection of mice from HSV challenge (32,55,66). In humans, CD4 ϩ T cells are stimulated in vivo following an HSV infection and the integrated CD4 memory response to HSV type 1 (HSV-1) appears to occur in up to 0.2% of circulating CD4 ϩ T cells (2,45,67,70). Severe herpetic infections are often seen in immunocompromised individuals with impaired T-cell immunity, such as AIDS and transplant patients, where the immune defect is predominantly displayed in CD4 ϩ T cells (16). While it is believed that CD4 ϩ T-cell responses are important for protection in general, the importance of Th1-versus Th2-type immune responses for prot...