It has recently been shown that several members of the tetraspan superfamily, including CD9 and CD63, associate with each other and with β1 integrins. In this study, we examined the distribution of a recently identified tetraspan, PETA-3 (CD151), and of CD9, CD63, α5β1, and the integrin β1 chain in normal human tissues by the indirect immunoperoxidase and alkaline phosphatase-anti-alkaline phosphatase techniques. PETA-3 showed a broad distribution and was expressed by endothelium, epithelium, Schwann cells, and dendritic cells and by skeletal, smooth, and cardiac muscle. Expression in skin was mostly restricted to the basal cells of the epidermis and was downregulated on differentiation. In the small intestine, PETA-3 was expressed by crypt and villous enterocytes with a mostly basolateral distribution, but was not detectable on the brush border. CD9 was expressed on the plasma membrane of enterocytes in crypts and at the bases of the villi whereas CD63 demonstrated a unique granular appearance concentrated in the apical cytoplasm below the brush border. The findings of this study show co-localization of PETA-3 with CD9, CD63, α5β1, and β1 in particular tissues, demonstrating that tetraspan/integrin complexes may occur. However, the lack of co-localization of these antigens in other tissues also implies distinct roles for these molecules.