Abstract: This study utilized scanning electron microscopy combined with energy-dispersive X-ray spectroscopy (SEM-EDS) to systematically compare the micromorphology and composition of dental calculus and renal calculi, aiming to elucidate their distinct biomineralization mechanisms and pathological microenvironments, while also demonstrating the applicability of this combined technique in biomedical sample analysis. The results revealed that dental calculus consists primarily of calcium phosphate, exhibiting diverse morphological features such as nanoneedles, flakes, and block-like structures, indicating a formation mechanism involving plaque-template-induced heterogeneous nucleation and phase transformation in the salivary environment. In contrast, renal calculi are mainly composed of calcium oxalate and calcium phosphate, with crystal morphologies including tetragonal bipyramids, flower-like clusters, and porous spherical architectures, reflecting rapid crystallization and organic-mediated regulation in highly supersaturated urine. Comprehensive analysis confirmed that SEM-EDS effectively correlates microscopic morphology with elemental composition in pathological specimens, enabling clear differentiation of the formation pathways of different types of calculi. This work not only clarifies the fundamental differences in composition, morphology, and formation mechanisms between the two types of calculi, but also provides a representative case for the integration of advanced instrumental analysis into interdisciplinary experimental teaching, thereby fostering students’ ability to utilize comprehensive characterization techniques in solving practical biomedical problems.