Inhibition of respiration has been reported as a short-term response of tree roots to elevated measurement CO2 concentration ([CO2]), calling into question the validity of root respiration rates determined at CO2 concentrations that differ from the soil [CO2] in the rooting zone. Our objectives were to validate previous observations of a direct CO2 effect on root respiration in sugar maple (Acer saccharum Marsh.) and to determine if high [CO2] also inhibited root respiration in other tree species. Root respiration rates for nine common North American tree species were measured in the field at ambient soil temperature at both 350 and 1000 ul CO2/l. No evidence of direct inhibition of root respiration by elevated measurements [CO2] was found for any of the species tested. The ratio of respiration rates at 1000 and 350 ul CO2/l ranged from 0.97 to 1.07, and the 95% confidence intervals for this ratio included unity for all species tested. Tests of a respiration cuvette used in earlier experiments suggested that gas leakage from the cuvette/IRGA system created an apparent direct CO2 effect on respiration of sugar maple roots when none actually existed. Small sample masses used in those experiments exacerbated the error. Careful attention to the possibility of gas leaks and the avoidance of small sample masses should produce data that will allow researchers to accurately assess whether direct effects of measurement [CO2] exist. Our findings of no direct CO2 effect on respiration of roots of a wide variety of species suggest that such effects may be less common than previously thought for tree roots.