In order to demonstrate that cultured mammalian cells can be frozen, dried, rehydrated and revivified, three cell lines were chosen for experiments which will determine (i) the optimized conditions for entry of the dry protectant trehalose into cells via electroporation in order to stabilize the membranes and proteins, (ii) the prefreeze optimization of the cells for freezing and drying, (iii) the optimized cooling rates, (iv) the optimized drying techniques, (v) the optimized rehydration techniques, and (vi) the feasibility of scale up to accommodate increased numbers of cell samples. The outcome variables used to monitor the parameters of optimization will be structured as determined by light and electron microscopy, in situ hybridization with cDNA probes, immunoelectron microscopy of selected cell components, estimation of membrane transition by infrared spectroscopy, and water state determinations by differential scanning calorimetry. The goal will be maintenance of cellular structure, normal biochemical functionality and reproduction. The medical applications of this technology lie in cell preservation for research and transplantation.
