Although macrocytosis might occur with frequency in patients with thyroid disorders, there is controversial information on the metabolic relationship between thyroid stimulating hormone (TSH), folic acid and B12 in the general population. We performed a retrospective analysis to retrieve results of serum folic acid, B12, and TSH performed on consecutive outpatients referred by general practitioners for routine blood testing over the last 2 years. A positive, significant trend towards increased values of folic acid, but not of B12, could be observed across the spectrum of TSH values suggestive for hypo- and hyperthyroidism. However, the prevalence of subjects with folic acid or B12 deficiency did not differ significantly among the subgroups of subjects. In multivariable linear regression analysis folic acid, but not B12, was associated with TSH levels. These results do not support the routine screening for either B12 or folic acid deficiency in subjects with subclinical disturbances of thyroid function, though we can not rule out that it might still be useful in patients with overt thyroid dysfunction.