Human sex hormone binding globulin (SHBG) is encoded by a normal and a variant allele. The resulting SHBG phenotypes (the homozygous normal SHBG, the heterozygous SHBG and the homozygous variant SHBG phenotype) can be distinguished by their electrophoretic patterns. We developed a novel detection system allowing us to distinguish between the different electrophoretic patterns in small amounts of plasma or serum (10 μ1). Small aliquots of Blue Sepharose were added to diluted sera or plasma samples for removal of albumin and the supernatants were subsequently applied to Western blotting. This method of detection was used to determine the distribution of SHBG phenotypes in healthy controls of both sexes and in five different pathological conditions characterized by changes in the SHBG level or endocrine disturbances (malignant and benign ovarian neoplasms, hirsutism, liver cirrhosis and alcoholism). The distribution of SHBG phenotypes was independent of such conditions and in agreement with the expected phenotype distribution of a bi-allelic gene in both healthy controls and patients (Hardy Weinberg law). An allele frequency of 0.13 was found for the SHBG variant allele based on the experimental values. Differences in SHBG phenotypes do not appear to have any clinical significance and no sex difference was found in the SHBG phenotype distribution.