The present study aimed at investigating the influence of fixation methods on the ultrastructure and calcium content of five different aortic biovalves. Biovalves subcutaneously implanted in Wistar rats for 12 weeks demonstrated significant differences in their calcium contents. Using Scheffe tests, two different groups of biovalves could be distinguished: (i) valves with high calcium content, such as Toronto-SPV (165 +/- 42 mg/g dry tissue) and Intact (193 +/- 36 mg/g dry tissue), and (ii) valves with low calcium content, such as Mosaic (2.7 +/- 1.8 mg/g dry tissue), Freestyle (2.3 +/- 1.1 mg/g dry tissue) and Hancock-II (3.6 +/- 1.2 mg/g dry tissue) (P < 0.05). All biovalves with an ultrastructurally preserved endothelium exhibited a low calcification tendency. The data suggest that if the endothelium is lost as a result of the fixation procedure, then calcification can only be prevented by appropriate anticalcification methods.