DNA plasmids are potent inducers of long‐lasting antigen‐specific CTL responses. Little is known about the distribution of antigen‐specific CD8+ T cells in the lymphoid tissue and the non‐lymphoid tissue after DNA immunization. HBsAg‐specific CD8+ T cells in peripheral blood mononuclear cells, spleen, lymph nodes, and the liver of Balb/c mice have been quantified after injection with a DNA plasmid expressing the major S protein of hepatitis B virus (HBV). The kinetics of CD8+ T‐cell responses in the circulation were measured after priming and boosting, showing that antigen‐specific CD8+ T cells undergo first expansion and then decline to a sustainable level in the circulation, although the frequencies of HBsAg‐specific CD8+ T cells in the circulation were lower than for the spleen. The greater frequencies of HBsAg‐specific CD8+ T cells were found in the liver, whereas the largest numbers of antigen‐specific CD8+ T cells were found in the spleen. By day 100 after priming, HBsAg‐specific CD8+ T cells were still detected in the circulation, the spleen and the liver. After boosting with the same plasmid DNA immunogen, HBsAg‐specific CD8+ T cells proliferated quickly and vigorously. By 150 days after boosting, HBsAg‐specific memory CD8+ T cells were sustained at higher levels than those recorded after the first, primary injection, both in the spleen and the liver: anti‐HBs antibody‐secreting plasma cells persisted in the bone marrow and in the spleen, consistent with the detection of anti‐HBs antibodies detected in the blood. These findings indicate that DNA immunization has considerable potential for inducing specific T cell responses in the liver and offers a strategy for the development of post‐exposure immunotherapy against persistent hepatitis B infections. J. Med. Virol. 80:225–232, 2008. © 2007 Wiley‐Liss, Inc.

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