The rapid evolution of molecular biology techniques has a significant impact on laboratory medicine. Nowadays a large number of diagnostic tools are available to diagnose and to characterize the different phases of hepatitis B virus (HBV) infection. The advent of the assay for nucleic acid amplification and detection enables clinicians to initiate and monitor antiviral therapy whilst allowing basic scientists to carry out studies on HBV biology. This review will focus on the evolution of the diagnostic tools to detect and monitor HBV infection.