Abstract Serological assays have been the mainstay of diagnosis of hepatitis B virus (HBV) infection and have had substantial impact on public health. However, not all interpretation of hepatitis B serology is straightforward. In addition, serology has proven inadequate in determining how new anti-HBV agents and therapeutic strategies can be assessed and compared. Some of these deficiencies can be overcome by the new HBV DNA quantification assays, using improved signal amplification or real-time PCR and which have been standardized to the WHO HBV reference standard. Molecular diagnostics have now also turned to the examination of liver tissue. Biopsy material, usually taken for histological examination, can also be assayed to quantify intrahepatic HBV DNA replicative intermediates, in particular HBV covalently closed circular (ccc) DNA and HBV RNA transcripts. Measurement of such replicative species may offer additional options for monitoring and predicting treatment efficacy, but further validation and standardization are required before they can become truly diagnostic assays. Similar obstacles will apply to newer technologies, such as tissue-based microarrays and gene arrays, which may lead to even more insights into HBV pathogenesis.