Abstract
Application of special in vitro culture techniques for the cells, derived from different animal and human organs, makes possible the obtaining of multicellular spheroids (MSs), being the natural 3-D environment for cells unlike the standard culture with the cells in monolayer. Previously we have shown that MSs formed in the newborn piglet adrenal cell culture are capable to produce the neuroblast-like cells, expressing the neuronal marker β-III-tubulin. In the present work we have assessed the regimens for MSs cryopreservation using 5, 7 and 10% dimethyl sulfoxide (DMSO) and 25% fetal bovine serum (FBS). Analysis of some characteristics of cryopreserved MSs (surface adhesion, capability to produce the neuroblast-like cells and monolayer formation by fibroblast-like cells) allowed to choose the cryopreservation regimen with 1°C/min cooling rate in the presence of 10% DMSO as the most optimal one. The FBS supplement to the medium did not significantly affect the cryopreservation outcome, although there was found a tendency to increase the capability of cryopreserved MSs to produce the neuroblast- and fibroblast-like cells. Probl Cryobiol Cryomed 2017; 27(4): 322-333
Highlights
Application of special in vitro culture techniques for the cells, derived from different animal and human organs, makes possible the obtaining of multicellular spheroids (MSs), being the natural 3-D environment for cells unlike the standard culture with the cells in monolayer
Half of the MSs was transferred into the cryovials (Thermo Fisher Scientific, USA) for further cryopreservation, and the other half was placed under the standard culture conditions
The MSs acquired the shape of dense conglomerates with sharp contours consisting of morphologically homogeneous cells
Summary
Application of special in vitro culture techniques for the cells, derived from different animal and human organs, makes possible the obtaining of multicellular spheroids (MSs), being the natural 3-D environment for cells unlike the standard culture with the cells in monolayer. Analysis of some characteristics of cryopreserved MSs (surface adhesion, capability to produce the neuroblast-like cells and monolayer formation by fibroblast-like cells) allowed to choose the cryopreservation regimen with 1 deg/min cooling rate in the presence of 10% DMSO as the most optimal one. The FBS supplement to the medium did not significantly affect the cryopreservation outcome, there was found a tendency to increase the capability of cryopreserved MSs to produce the neuroblast- and fibroblast-like cells. The derivation of multicellular spheroids (MSs) is one of the promising approaches in culturing the cells of different tissues This approach is based on a natural property of cells to function among similar ones, in contrast to the classical monolayer culture, where the cells contact in one plane only.
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