Introduction: The international ICALIC initiative aims at developing a new indirect calorimeter according to the needs of the clinicians and researchers in the field of clinical nutrition and metabolism. The project initially focuses on validating the calorimeter for use in mechanically ventilated acutely ill adult patient. However, standard methods to validate the accuracy of calorimeters have not yet been established. This paper describes the procedures for the in-vitro tests to validate the accuracy of the new indirect calorimeter, and defines the ranges for the parameters to be evaluated in each test to optimize the validation for clinical and research calorimetry measurements. Methods: Two in-vitro tests have been defined to validate the accuracy of the gas analyzers and the overall function of the new calorimeter. 1) Gas composition analysis allows validating the accuracy of O2 and CO2 analyzers. Reference gas of known O2 (or CO2) concentration is diluted by pure nitrogen gas to achieve predefined O2 (or CO2) concentration, to be measured by the indirect calorimeter. O2 and CO2 concentrations to be tested were determined according to their expected ranges of concentrations during calorimetry measurements. 2) Gas exchange simulator analysis validates O2 consumption (VO2) and CO2 production (VCO2) measurements. CO2 gas injection into artificial breath gas provided by the mechanical ventilator simulates VCO2. Resulting dilution of O2 concentration in the expiratory air is analyzed by the calorimeter as VO2. CO2 gas of identical concentration to the fraction of inspired O2 (FiO2) is used to simulate identical VO2 and VCO2. Indirect calorimetry results from publications were analyzed to determine the VO2 and VCO2 values to be tested for the validation. Results: O2 concentration in respiratory air is highest at inspiration, and can decrease to 15% during expiration. CO2 concentration can be as high as 5% in expired air. To validate analyzers for measurements of FiO2 up to 70%, ranges of O2 and CO2 concentrations to be tested were defined as 15e70% and 0.5e5.0%, respectively. The mean VO2 in 426 adult mechanically ventilated patients was 270 ml/min, with 2 standard deviation (SD) ranges of 150e391 ml/min. Thus, VO2 and VCO2 to be simulated for the validation were defined as 150, 250, and 400 ml/min. Conclusion: The procedures for the in-vitro tests of the new indirect calorimeter and the ranges for the parameters to be evaluated in each test have been defined to optimize the validation of accuracy for clinical and research indirect calorimetry measurements. The combined methods will be used to validate the accuracy of the new indirect calorimeter developed by the ICALIC initiative, and should become the standard method to validate the accuracy of any future indirect calorimeters.