We performed a statistical analysis to elucidate effects of standardized measurement of hemoglobin A1c (HbA1c) on analysis of factors that affect HbA1c values. Subjects were participants in the Japan Public Health Center-based Prospective Study on Cancer and Cardiovascular Diseases, and a total of 1789 men and 3150 women in three distinct areas who did not have overt diabetes (HbA1c> or =6.1% or prior diagnosis) were analyzed. A different method of HbA1c assay was used in each area: high-performance liquid chromatography in one area and a different immunochemical method in each of the other two areas. Then, calibration of HbA1c was performed using two HbA1c standards (5.5 and 10.5%) provided by the Japan Diabetes Society. Analysis of co-variance was performed separately in men and women. When raw HbA1c data were used as the outcome, 'area', which represents differences in assay systems, lifestyles, etc. had a significant effect on HbA1c levels. When calibrated HbA1c data were used, however, 'area' was no longer a significant factor. In the latter analysis, age and BMI were the principal contributors to HbA1c, and parental history of diabetes had a weak effect in women. Thus, standardization of HbA1c reduced the difference between assay systems, and uncovered two common factors to determine HbA1c levels.