Oestrogen receptor beta (ERbeta) is expressed in human skeletal muscle tissue. In the present study, we have developed an immunohistochemical method to reveal if ERbeta is located within the muscle fibres as well as within capillaries. Skeletal muscle biopsies were obtained from m. quadriceps femoris vastus lateralis in four healthy young subjects. Immunohistochemical triple staining was applied to transverse sections of paraffin-wax-embedded tissue. The basement membrane of muscle fibres and capillaries was identified by using an antibody to collagen IV, endothelial cells using an antibody to CD34 and ERbeta using a corresponding antibody. The ERbeta-positive (ERbeta+) nuclei were located within the muscle fibre defined by the localisation of collagen IV. ERbeta+ nuclei were also, for the first time, found in endothelial cells of capillaries in skeletal muscle tissue. Quantification was performed on transverse cryostat sections after performing a double staining (collagen IV and ERbeta). It was shown that 24% of the ERbeta+ nuclei were located within capillaries, and 76% were located within muscle fibres. In conclusion, ERbeta in human skeletal muscle tissue is expressed not only in the muscle fibres themselves, but also within the capillary endothelial cells. This observation might improve understanding of the physiological role of oestrogen and its receptor.