There is evidence that glutaraldehyde used routinely in the fixation process of bioprosthetic heart valves may be a major factor in their subsequent calcification. A further complication is introduced by the use of a formaldehyde treatment step, whether for sterilization or storage. The effects of this second aldehyde on calcification is also unknown. The aim of the present work is to determine the degree of glutaraldehyde and formaldehyde incorporation into commercial valve leaflets and to establish whether there is a relationship between the aldehyde treatment and tissue calcification. Both pre- and post-implantation concentrations of aldehydes were estimated by high performance liquid chromatography of acid hydrolyzates of commercial valve tissue. Control samples were taken from freshly prepared porcine aortic valves and bovine pericardium fixed in glutaraldehyde. The degree of calcification was investigated using the rat subcutaneous implant model. Samples were retrieved after 56 days and calcium estimated by atomic absorption spectroscopy. The results indicated that storage of both porcine and pericardial valves for periods of 5 years or longer reduced calcification. A greater reduction in calcification was noted when tissue was treated sequentially with glutaraldehyde and formaldehyde rather than glutaraldehyde alone. From this work it can be concluded that bioprosthetic valves calcify less in the rat model when they are stored for extended periods of time in glutaraldehyde solution. This effect is enhanced when formaldehyde storage is employed.