A Raman microimaging-based approach has been used in the current study to evaluate formation and progression of calcification in situ in human stenotic aortic valves obtained during surgical valve replacement. The capability of the method to visualize distribution of the calcified deposits resulted in structural characterization of deposits in the various phases of development. A high spatial resolution of the method along with the confocal depth profiling enabled to identify extremely small salt inclusions (of ca. 0.5 µm in diameter), formed probably at the very early stage of calcification. Structurally, these inclusions are built from an octacalcium phosphate-like compound that during grains' growth transforms into tricalcium phosphate, mixed with the salt containing the acidic phosphate groups (HPO42−) and, finally, into stable B-type hydroxyapatite that is the only salt present in large-area calcium salt deposits. Copyright © 2013 John Wiley & Sons, Ltd.