Thermal expansion of substrate may affect adhesion of Chinese hamster fibroblasts to surfaces during freezing

Abstract

Despite success in cryopreservation of cells in suspension, cryopreservation of cells in monolayers is still challenging. One of the major problems is detachment of the cells from the substrate which occurs during cryopreservation. We hypothesized that this detachment may be due to a mismatch in the coefficient of linear thermal expansion α_L between glass and the frozen cell layer which manifests as residual stress and stress relaxation. This mismatch results in a difference between the thermal expansion of ice and glass as they undergo temperature changes. Rinzl plastic coverslips were selected as a possible substitute for glass because Rinzl has an α_L (60 × 10^-6/K) similar to that of ice (51 × 10^-6/K) whereas glass has a much lower α_L (5 × 10^-6/K). V79-4 Chinese hamster fibroblasts were cultured on both glass and Rinzl coverslips until confluent and the area of coverage was measured before and after freezing at -9 °C. The glass coverslips showed significant loss of cells (coverage = 77.9 ± 8.0%) compared with Rinzl (coverage = 97.9 ± 1.4%). We concluded that Rinzl coverslips may improve cell attachment in future monolayer cryopreservation experiments.

Keywords: Cell adherence; Freezing; Monolayer; Thermal expansion mismatch.