CO2 was measured in rat cortical bone samples with an accurate manometric technique. Bone CO2 content increased with the age of the animals. When heated to constant weight, bone samples lost 15.5% of their initial fresh CO2 content, indicating that CO2 exists in several chemical forms in the bone CO2 pool. Bone samples were exposed in vitro to atmospheres of increasing PCO2. Bone CO2 increased by 15% of its original content when exposed for 60 min. to pure CO2. CO2 uptake by bone in vitro was shown to be time- and PCO2-dependent, evoking a saturating mechanism and requiring the presence of water in bone. These findings lead to the conclusion that the bone CO2 compartment consists of 30% bicarbonate and 70% carbonate. The exact location of the bicarbonate pool and its availability as a buffer store for the whole organism are discussed.