*2.6. In Vivo SLP Distribution*

The Fe3O<sup>4</sup> nanoparticles in the nanocarrier allowed the determination of the SLP distribution in the different tissues using inductively coupled plasma optical emission spectrometry (ICP-OES) (ICPE-9000 SHIMADZU; Kyoto, Japan). For this purpose, tissues of 4 mice per treatment were collected at 20 days postinjection, fixed in formalin, weighed, and calcined at 450 ◦C for 12 h to remove all the organic matter. The obtained ashes were dispersed in concentrated chlorhydric acid (HCl 37%) overnight. Finally, samples were diluted to 10 mL with Milli-Q water and analyzed via ICP-OES.

Confocal microscopy imaging was used to visualize particles in lungs 3 and 8 h after intravenous administration. They were embedded in tissue-Tek® O.C.T (optimum cutting temperature) solution (Sakura, Japan), frozen, and cut into 15 µm sections with cryostat. Sections were then fixed in paraformaldehyde (4%) and stained with Hoechst (33258; Sigma; Germany).
