Diffusion kinetics and perfusion times in tissue models obtained by bioorthogonal Raman m- spectroscopy

The penetration kinetics of small-molecule compounds like nutrients, drugs, and cryoprotective agents into artificial cell aggregates are of pivotal relevance in many applications, from stem cell differentiation and drug screening through to cryopreservation. Depending on compound and tissue properties as well as aggregate size and shape, the penetration behavior can differ vastly. Here, we introduce bioorthogonal Raman microspectroscopy as a contactless technique to investigate the penetration of various compounds into spheroids, organoids, and other tissue models in terms of diffusion coef fi- cients and perfusion times. We showcase the potential of the method by applying it to the radial perfusion of neural stem cell spheroids with the prevalent cryopreservation additive dimethyl sulfoxide. Employing a diffusion model for spherical bodies, the spectroscopic data were quantitatively analyzed. Perfusion times were obtained for spheroids in the sub-mm region, and interesting findings about the spheroid-size dependence of the diffusion coef ficient are reported.