Three dimensional multi-cell spheroids assembly using thermoresponsive Gel probe

In this paper, we achieved three dimensional (3D) assemblies of multi-cell spheroids towards 3D in vitro cell structures. The 3D assembly of cells in vitro is one of the important techniques to reveal cell growth, differentiation and hystogenesis. The techniques can be used for applications in tissue engineering, drug screening, and cell characterization. The multi-cell spheroids were used as building blocks for 3D structure, and thermoresponsive gel was used for spheroids assembly. The mouse myoblast cells C2C12 and mouse fibroblast cells NIH 3T3 were cultured in an ultra-low atachment surface flask. The cells were naturally aggregated in the flask and cell spheroids were prepared in self-assembly process. The rat liver cell RLC-18 was also cultured to prepare spheroids. The thermoresponsive Gel probe (GeT probe) we proposed was employed for assembly of the prepared spheroids in 3D. The handled spheroids by GeT probe were cultured inside the thermoresponsive gel and the cell viability was confirmed. The cultured structure was handled by the GeT probe for further assembly. The results validate that the GeT probe can manipulate spheroids with keeping live condition of cells and achieve 3D cell assembly inside the thermoresponsive gel with arbitrary and precise patterning.