This study is intended to reveal immunohistochemical localizations of α-SMA and calcium binding wbS-100, S-100α and S-100β proteins in intraorbital and extraorbital lacrimal gland of adult male rats. After extraorbital and intraorbital sections of the lacrimal gland were extracted from the rats under anesthesia, Streptavidin-biotin-peroxidase immunohistochemical technique to determine wbS-100, S-100α, S-100β and α-SMA proteins were applied to the 4-μm sections taken from the tissue samples blocked following the routine histological procedure. Immunoreaction of wbS-100, S-100α and S-100β was determined both in cytoplasm and nucleus of the acinus epithelium, duct epithelium, myoepithelial and endothelial cells of extraorbital lacrimal gland. While S-100α immunoreaction in all structural components of the intraorbital gland was most densely observed in the nucleus, S-100β immunoreaction was negative in myoepithelial cells. Moreover, it is detected that wbS-100 immunreaction was positive in the lateral membranes of acinus epithelial cells of extraorbital gland and wbS100 and S-100β immunoreaction was positive in the lateral membranes of acinus and duct epithelial cells of intraorbital gland. α-SMA immunoreaction was detected in the myoepithelial cells and the smooth muscle cells in the wall of blood vessels of both glands. In conclusion, the differences detected between wbS-100, S-100α and S-100β proteins in all structural components of intraorbital and extraorbital lacrimal glands in terms of immunohistochemical staining density suggest that there is a functional difference between both lacrimal glands with respect to the secretory activity.