Hepatic steatosis is a frequent histological alteration in chronic hepatitis C virus (HCV) infection that sensitizes the liver to cell injury, inflammation, and fibrosis via unclear mechanisms. Although apoptosis has been implicated in various liver diseases, its importance in HCV-associated steatosis is largely unknown. In this study, we investigated the role of caspases, the key regulators of apoptosis, and employed two novel caspase assays, an immunological and a luminometric enzyme test, to detect hepatic caspase activation in sera from HCV patients with different grades of steatosis. Our data show that increased caspase activation can be found not only in liver biopsies, but also in sera from HCV patients with liver steatosis. Patients with steatosis exhibited significantly higher serum levels of caspase activity compared with normal healthy individuals. Moreover, the extent of steatosis closely correlated with serum caspase activity, whereas in particular in cases of low or moderate steatosis, no correlation was found with aminotransferase levels. In conclusion, apoptotic caspase activation is considerably elevated in HCV-associated steatosis. More importantly, our data imply that measurement of caspase activation might be a sensitive serum biomarker to detect liver steatosis in patients with chronic HCV infection and other liver diseases.