PurposeTo determine the feasibility of using a portable carbon dioxide (CO2) sensor to calibrate a pneumatic esthesiometer and then to calibrate the chemical stimuli.MethodsThe chemical stimuli in ocular surface experiments are combinations of medical air and added CO2 (%CO2). These stimuli were calibrated using a portable CO2 sensor (COZIR CM-0041) and data logger, delivered for 100 seconds by using the Waterloo Belmonte esthesiometer. The distances between the sensor and esthesiometer tip were 0 (to measure feasibility), 3, 5, and 10 mm. In experiment I, 100% CO2 was tested using four different flow rates (50, 100, 150, and 200 mL/min) at three working distances. In experiment II, flow rates of 20 to 100 mL/min and concentrations of 20% to 100% CO2 were tested in 20 steps at 3 working distances.ResultsThe CO2 sensor correctly reported the esthesiometer extremes of 0% and 100% CO2 when placed at the esthesiometer tip. There were progressive, systematic increases in concentrations reaching/reported by the sensor with increasing flow rates and nominal concentrations and progressive decreases in measurements with increases in working distance.ConclusionsCO2 concentrations in pneumatic esthesiometers can be calibrated and, as expected, vary with flow rate and distance, highlighting the importance of calibration and standardization of CO2 stimuli in these instruments.Translational RelevanceCalibrated CO2, a chemical sensory stimulus in humans, may be used in testing the surface of the eye as well as other membranes within which the CO2 can be dissolved (e.g., mucous) to produce an acidic stimulus.