Confocal Laser Scanning Microscopy Examination of Cell Distribution in Macroporous Microcarriers

Macroporous microcarriers are often used to cultivate animal cells. The pores in the interior of the beads provide surface and space for cell growth. It is not clear how anchorage-dependent and suspension cells populated these microcarriers during cultivation. Confocal laser scanning microscopy was employed to perform time lapse observation of the cells in the interior. The structure of the bead was stained with fluorescein isothiocyanate for visualization, while the cells were stained with dialkyl indocarbocyanines for tracking over time. It was observed that mouse fibroblastic cells CRE BaG2 did not move extensively after initial attachment. Some cell divisions were observed during the course of the experiments, and essentially all cells remained viable throughout. Few hybridoma cells were deposited into the pores in the interior of the microcarriers. The results suggest that the occupancy of the internal volume by cells after prolonged cultivation is largely due to the growth of cells that are deposited in the interior as opposed to the migration of cells from the external surface into the interior. This method of observing cell behavior in a three-dimensional structure may find applications in other three-dimensional cell culture systems. The animation of time lapse sections is available on the worldwide web at http:/ www.cems.umn.edu/ approximately wshu_grp/acre/microcarrier.html.