We used a recently devised HPLC method to quantify eight modified nucleosides, an emerging group of tumor markers, in human serum and then calculated their reference intervals in a healthy population from Italy and the United States. We used the statistical procedure of element analysis, which reveals the effects of chosen variables (in this case, nationality, sex, and age) on an analyte (here, modified nucleosides). Using element analysis, we calculated the exact weight of each variable on the reference values. We found that nationality has the greatest effect on the serum concentrations of all the modified nucleosides apart from pseudouridine, whereas sex significantly influences only the concentrations of 4-pyridone-3-carboxamide-N1-ribofuranoside, 1-methylinosine and N2,N2-dimethylguanosine; age affects only N2,N2-dimethylguanosine. Thus, the reference intervals of all the nucleosides except pseudouridine were calculated separately for Italians and Americans, and the reference values for 4-pyridone-3-carboxamide-N1-ribofuranoside, 1-methylinosine, and N2,N2-dimethylguanosine were calculated separately for men and women. Our data form the baseline for study of variations in serum concentrations of modified nucleosides in various pathophysiological conditions.