Some hepatitis B virus (HBV) carriers with chronic hepatitis delta virus (HDV) superinfection show progressive chronic hepatitis, whereas others show no apparent signs of liver disease. In the present study, we established a sensitive method for the quantitation of the level of HDV RNA in serum on the basis of real-time reverse-transcription polymerase chain reaction (RT-PCR), to clarify the role that the level of HDV RNA in serum plays in the diverse natural course of clinical manifestation. In 48 subjects who were positive for hepatitis B surface antigen and for anti-hepatitis delta antibody, the levels of HDV RNA in serum were quantitated by RT-PCR. The levels of HBV DNA in serum were determined by a transcription-mediated amplification assay. The levels of HDV RNA in serum of subjects with chronic hepatitis and of subjects with liver cirrhosis were significantly higher than those in asymptomatic carrier subjects. The levels of HBV DNA in serum did not differ significantly among these 3 groups. In conclusion, HDV RNA quantification by real-time RT-PCR is possibly a useful tool for understanding the pathophysiology of HDV infection.