The sacrotuberous ligament (STL) and the hamstrings are important structures that are mutually connected and influenced by the pelvis. However, the anatomical connectivity and histological characteristics of these structures remain unclear. The present study aimed to comprehensively investigate the relationship between the STL and the proximal hamstrings through histological analysis. Sixteen specimens were obtained from eight fresh cadavers (mean age at death, 73.4 years). Verhoeff Van Gieson, Masson's trichrome, and immunohistochemical staining were used to analyze the connectivity between the STL and the hamstrings and to verify the ratios of collagen and elastic fibers. Dense connective tissue that overlapped tightly between the STL and hamstrings was observed. The relative ratios of collagen and elastic fibers between the STL and hamstrings characteristically identified regional differences. The ratio of elastic fibers to collagen in the biceps femoris (BF) was ~38.6 ± 4.7%, and the lowest ratio was 5.9 ± 2.6% observed in the semimembranosus (SM). In the case of the BF, contractibility is well‐regulated due to a high content of elastic fibers; however, the muscular structure of the BF is relatively fragile due to the low content of collagen. In the SM, collagen content is higher than that in the STL. This ratio of elastic fibers in the collagen analysis could provide crucial information for understanding the differences in hamstring contractility and maintaining the morphology of these structures.