Elephant endotheliotropic herpesvirus (EEHV), a persistent threat, has caused significant mortality among young Asian elephants (Elephas maximus), raising serious concerns for the conservation of this endangered species. Given the urgent need for protective measures, research into EEHV vaccine development has become increasingly critical. This study evaluated the immune response in mice following immunization with an EEHV1A-glycoprotein B (gB) subunit vaccine. The vaccine incorporated gBF1 and gBF2, corresponding to segments of the gB ectodomains I and IV, respectively, along with emulsion adjuvants Montanide™ ISA 206 VG and incomplete Freund's adjuvant (IFA). The findings revealed that both gBF1 and gBF2, when paired with these adjuvants, were capable of inducing strong humoral immune responses against EEHV-gB, as demonstrated by the ability of sera from immunized mice to detect EEHV-gB ex vivo. Additionally, in terms of cellular immunity, the vaccine formulations predominantly activated CD4+ T cells, including both Th1 (IFN-γ+) and Th2 (IL-4+) cells, with no significant activation of CD8+ T cells. If also applicable in elephants, gB-based vaccines would be a significant step forward in the fight against EEHV.