Cadmium (Cd) is an extremely toxic metal commonly found as an environmental contaminant from industrial and agricultural sources, posing severe risks to human health. In this study, the binding mechanism of Cd(II)-human serum albumin (HSA) complex and the effect of Cd(II) on the conformational stability and structural state of HSA were comprehensively investigated through a series of efficient and appropriate methods. X-ray photoelectron spectroscopy accurately described the microenvironmental changes around protein C, N, and O atoms in the presence of Cd(II). Fluorescence results indicated that the probable mechanism of Cd(II)-HSA interaction is a static quenching process. Fourier transform infrared spectroscopy and dynamic light scattering showed Cd(II) complexation altered HSA conformation and the microenvironments of Trp and Tyr residues, accompanied by the size increases of HSA aggregates. This research will be helpful for understanding the toxic effects of Cd(II) on protein function in vivo.