Human sex hormone-binding globulin (hSHBG) is a plasma glycoprotein composed of two identical subunits. The protein, which has high affinity for testosterone and estradiol has been purified to homogeneity. In this study we have investigated, on neuraminidase-treated serum samples, the presence of genetic variations of hSHBG by polyacrylamide gel isoelectric focusing (IEF). Based on IEF analyses of 110 serum samples from adult Mexican individuals we have identified two distinct IEF-patterns. The most frequent phenotype (95.45%) was characterized by two IEF-bands with pIs of 6.50 and 6.63, respectively. In five serum samples, a different 4-band pattern with pIs of 6.50, 6.63, 6.70 and 6.76 was identified. Family studies showed that this pattern was genetically determined. The frequency of this variant was 4.55%, and the observed phenotypes were consistent with the expression of an autosomal genetic system. The estimated gene frequencies for both alleles were shown to be in genetic equilibrium. Affinity constants, binding kinetics and serum concentrations of hSHBG from individuals having a 4-band pattern were similar to those obtained in individuals with a 2-band pattern, thus suggesting that the mechanism responsible for the generation of polymorphic variants of hSHBG reported herein did not involve the steroid binding site of the molecule. These findings may be of broad interest, as other serum binding proteins express genetic variants, which may permit their further structural and functional subclassification.