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Colloidally stable cerium oxide nanoparticles (CeNPs), known for mimicking multiple antioxidant enzymes, are promising nanozymes for therapeutic applications targeting oxidative stress and inflammation. Although fluorescent dye labeling facilitates nanoparticle imaging and tracking, its influence on physicochemical properties and biological interactions remains insufficiently understood. In this study, poly(acrylic acid)‐coated CeNPs and their DiI‐encapsulated counterparts are synthesized to evaluate the effects of dye functionalization on catalytic performance, cellular uptake, and intracellular fate. While overall redox cycling is retained, DiI@PAA‐CeNPs show an 30% decrease in superoxide dismutase‐like activity, whereas catalase‐, peroxidase‐, and oxidase‐like activities remain largely preserved. Both formulations exhibit no cytotoxicity toward human osteoblasts. However, DiI labeling modifies surface chemistry and delays cellular uptake, particularly in the presence of serum proteins, as revealed by fluorescence microscopy. Multiscale imaging combining fluorescence microscopy, transmission electron microscopy, and focused ion beam‐assisted lamella preparation reveals differences in nanoparticle internalization but similar intracellular fate. Focusing on poly(acrylic acid)‐coated cerium oxide nanoparticles, DiI encapsulation can markedly alter CeNPs properties. This finding highlights the need to critically assess labeling effects in bio‐nano studies. Integrating label‐free techniques is crucial for accurate characterization and the rational design of nanozymes for diagnostic and therapeutic applications.