The objective of this study was to compare total carbon dioxide (TCO2) levels measured by three different methods. Two hundred jugular venous blood samples from dogs admitted to the Veterinary Teaching Hospital, Ontario Veterinary College with various clinical disorders were analyzed by the Radiometer blood gas analyzer (BGA) and the Coulter DACOS analyzer. In 70 of these samples, TCO2was also measured by the Kodak Ektachem DTE chemistry analyzer. Comparison of the agreement between methods revealed intraclass correlation coefficients (ICC) of 0.63, 0.79, and 0.82 for the DACOS‐Ektachem, DACOS‐BGA, and Ektachem‐BGA comparisons, respectively. Evaluation of the effect of storage time on TCO2content of stored serum samples measured on the DACOS analyzer revealed a decrease of almost 2 mmol/L and a decrease in the intraclass correlation coefficient values between the DACOS analyzer and other methods after 7 hours storage time. The results of this observational study revealed lower agreement between the three methods in this study than previously reported between other methods of TCO2measurement. Possible reasons for the lower than expected agreement in this study included changes in DACOS values because of storage and differences in methodologies between the me