A1celaphine herpesvirus 1 (AHV-1) is a causative agent of malignant catarrhal fever which is a fatal and a lymphoproliferative syndrome. AHV-1 is a gamma herpesvirus, which induces frequent latent infection and often difficult to detect its antigens or specific nucleic acids because of its low viral copies in the infected tissues. A new method, in situ PCR, is developed for the detection of AHV-1 nucleic acid in this study. Target sequences of AHV-1 open reading frame 50 gene were detected within AHV-1 infected MDBK cells. As compare with other molecular biological methods for the detection of AHV-1, in situ PCR was found to be more sensitive than in situ hybridization and to be less sensitive than nested PCR. However, nested PCR cannot afford to observe and differentiate AHV-1 infected cells. In situ PCR amplifies a target sequence within cells that can be visualized microscopically with increased sensitivity compared to detection by in situ hybridization. In situ PCR has wide applications for sensitive localization of low copy AHV-1 viral sequences within cells to investigate the role of viruses in a variety of clinical conditions and also provide the rapid, sensitive, and specific detection of AHV-1 infection.