Alongside with SEM, confirmation of the presence of embedded NPs can be obtained by their labeling with a fluorescent dye followed by observation by confocal laser scanning microscopy (CLSM). For instance, fluorescent liposomes appeared as uniformly distributed dots in HA, ALG, CS, (Figure ) or GelMA matrices without any sign of aggregation and silicate HAP/WKT NPs were uniformly detected in HA patches. CLSM also revealed a non-homogeneous distribution of heparin/CS NPs in a hydrogel made of acrylate-graftedcarboxymethyl starch and methacrylic acid (CMS-g-AA/PMAA), likely due to the random diffusion of the NPs in the matrix that featured an irregular macroporous structure (Figure ). As an alternative to fluorescence, analysis of the elemental chemical composition of the sample surface using SEM coupled with energy-dispersive spectroscopy (EDS) can provide evidence of the presence of nano-objects enclosed in a 3D matrix. In this manner, by monitoring the phosphorous signal, the regular dispersion of liposomes (Figure ) and the incorporation HAP NPs could be easily visualized (Figure ). Elemental mapping also enables detection of the uniform distribution of Ag, Ti, Cu, Au, Si, and Fe. permission. Copyright 2018 American Chemical Society. (c) Distribution of phosphorus element within an ALG hydrogel loaded with liposomes. Adapted with permission. Copyright 2022 Elsevier. (d) Elemental mapping of a HAP NPs within a CS hydrogel. Adapted with permission. Copyright 2023 Elsevier. (e) Distribution of Ca and P elements in a hydrogel made of DA-modified Gel/oxidized Dex (ODex) in which are dispersed PDAfunctionalized HAP NPs. Adapted with permission. Copyright 2023 Elsevier.