Cryopreservation of cells: FT-IR monitoring of lipid membrane at freeze-thaw cycles

- FTIR spectroscopy was used to monitor the freeze-thaw cycle of two cellular lines suspended in three different media.
- We evidenced the main transition of lipid membrane as induced by ice formation.
- Adding DMSO led to a more rigid membrane and increased the amount of non-freezable extracellular water.
- A more rigid gel state of the membrane correlates with a larger extent of intact membrane recovery after thawing.

In the present study, FTIR spectroscopy was used to monitor the freeze-thaw cycle of two cellular lines (HuDe and Jurkat) suspended in three different media: phosphate buffer solution (PBS); dimethylsulfoxide (DMSO)/PBS solution at 0.1 DMSO molar fraction; and CryoSure (0.1 DMSO molar fraction PBS solution + dextran 5% w/v) solution. The Trypan Blue test was also applied before freezing and after thawing each cell sample to estimate the recovery of membrane integrity after thermal treatment, and correlate this datum with spectroscopic results. By following the temperature evolution of two different spectral components (the libration and bending combination mode νc(H2O) at 2000-2500 cm− 1, and the methylene symmetric stretching vibration νsym(CH2) at about 2850 cm− 1) in the − 120 ÷ 28 °C range, we evidenced the main transition of lipid membrane in connection with cell dehydration, as induced by ice formation in the extracellular medium. In particular, in DMSO/PBS and CryoSure samples we observed a transition to a more rigid state of the lipid membrane together with an increased amount of non-freezable water in the extracellular medium; these results are connected to the role of DMSO as a cryoprotective agent irrespective of the nature of cell type.