Macroprolactin is a complex of prolactin with immunoglobulins (IgG) that has limited or no biological activity in vivo. Immunoassays for prolactin have variable reactivity with macroprolactin. Therefore the presence of macroprolactin should be considered in the differential diagnosis of hyperprolactinemia. We compared a valid screening test for macroprolactin, polyethyleneglycol (PEG) precipitation, with the determination of the ratio of the results of two prolactin assays: Elecsys with high cross-reactivity with macroprolactin and Centaur with low cross-reactivity. In 59 negative samples subjected to the PEG test (precipitation 60%), 33 had, as expected, an increased ratio (over 1.45), 1 a normal ratio and 1 a decreased ratio (1.07). This decreased ratio could be due to a particular form of macroprolactin poorly recognised by the Elecsys assay. Among 5 samples in the grey zone (precipitation between 50 and 60%), the ratio was increased in 2, normal in 1 and decreased in 2. Apart from one false negative case (normal ratio with positive PEG test), the results of the Elecsys/Centaur ratio method were in good agreement with those of the PEG test. The ratio method could be helpful for samples with PEG test results in the grey zone, before undertaking a complete analysis of circulating molecular forms by gel filtration chromatography. Out of the 5 five samples in the grey zone, the ratio was 4 times out of the reference range: 2 increased, 2 decreased. Our results also underline the necessity of reevaluating the Centaur prolactin reference range from samples without macroprolactin.