We report an accurate method, based on capillary on-column gas chromatography, for determining triglycerides in human serum. After serum extraction and chemical hydrolysis, glycerol is directly measured by gas chromatography (GC). With our extraction method no free glycerol is extracted from serum. The accuracy of this method was compared with that of a method based on the original procedure of Carlson (J. Atheroscler. Res. 3, 334-336 (1963)) and which is standardized by the Centers for Disease Control (CDC, Atlanta, U.S.A.). Orthogonal regression analysis of the GC method (y) and the CDC reference method (X) resulted in y = 0.996 X + 0.000 with a correlation coefficient of 0.999. The variances of analytical data, collected over a two month period showed that, for the GC method, the within-day coefficient of variation (CV) was less than 1.43%; the between-day CV less than 1.35%. The data for the CDC method were less than 3.36% (within-day) and less than 6.38% (between-days). The CDC method is linear up to 3.7 mmol/l and the GC method to 22 mmol/l.