Chapter 13
Use of In Situ Fourier Transform Infrared Spectroscopy
in Cryobiological Research
Willem F. Wolkers and Harrie ¨ tte Oldenhof
Abstract
In this chapter, we describe how Fourier transform infrared spectroscopy (FTIR) can be applied in
cryobiological research to study: structure and thermal properties of biomolecules in cells and tissues,
physical properties of cryopreservation and freeze-drying formulations, and permeation of molecules into
cells and tissues. An infrared spectrum gives information about characteristic molecular vibrations of specific
groups in molecules, whereas the temperature dependence of specific infrared bands may reveal information
about conformational and phase changes. Infrared spectroscopy is minimally invasive and does not require
labeling, whereas spectra can be recorded in any physical state of a sample. Data acquisition and spectral
processing procedures are described to study phase state changes of protective formulations, cell membrane
phase behavior during freezing and drying, protein denaturation during heating, and permeation of
protective molecules into tissues. The latter can be used to estimate incubation times needed to load tissues
with sufficient amounts of protective agents for cryopreservation or freeze-drying.
Key words Dehydration, Freeze-drying, Freezing, Diffusion, Fourier transform infrared spectroscopy (FTIR), Membrane phase behavior, Protein denaturation
1 Introduction
Fourier transform infrared spectroscopy (FTIR) is emerging as a
powerful technique for cryobiological research. In situ FTIR studies during freezing, heating, or dehydration provide information on
conformational and phase changes of endogenous biomolecules,
including membrane lipids and proteins [1, 2]. FTIR can also be
used to study physical properties of cryopreservation and freezedrying solutions including glass and water-to-ice phase transitions
[3–5]. Moreover, FTIR can be applied to study permeation of
protective agents into tissues or scaffolds, to estimate incubation
times needed for loading prior to freezing or drying them [6–8].
IR spectroscopy applied to cells or tissues relies on studying
characteristic molecular group vibrations of endogenous biomolecules. The vibrational frequency of a molecular group primarily
Willem F. Wolkers and Harrie ¨ tte Oldenhof (eds.), Cryopreservation and Freeze-Drying Protocols, Methods in Molecular Biology,
vol. 2180, https://doi.org/10.1007/978-1-0716-0783-1_13, © Springer Science+Business Media, LLC, part of Springer Nature 2021
331
Use of In Situ Fourier Transform Infrared Spectroscopy
in Cryobiological Research
Willem F. Wolkers and Harrie ¨ tte Oldenhof
Abstract
In this chapter, we describe how Fourier transform infrared spectroscopy (FTIR) can be applied in
cryobiological research to study: structure and thermal properties of biomolecules in cells and tissues,
physical properties of cryopreservation and freeze-drying formulations, and permeation of molecules into
cells and tissues. An infrared spectrum gives information about characteristic molecular vibrations of specific
groups in molecules, whereas the temperature dependence of specific infrared bands may reveal information
about conformational and phase changes. Infrared spectroscopy is minimally invasive and does not require
labeling, whereas spectra can be recorded in any physical state of a sample. Data acquisition and spectral
processing procedures are described to study phase state changes of protective formulations, cell membrane
phase behavior during freezing and drying, protein denaturation during heating, and permeation of
protective molecules into tissues. The latter can be used to estimate incubation times needed to load tissues
with sufficient amounts of protective agents for cryopreservation or freeze-drying.
Key words Dehydration, Freeze-drying, Freezing, Diffusion, Fourier transform infrared spectroscopy (FTIR), Membrane phase behavior, Protein denaturation
1 Introduction
Fourier transform infrared spectroscopy (FTIR) is emerging as a
powerful technique for cryobiological research. In situ FTIR studies during freezing, heating, or dehydration provide information on
conformational and phase changes of endogenous biomolecules,
including membrane lipids and proteins [1, 2]. FTIR can also be
used to study physical properties of cryopreservation and freezedrying solutions including glass and water-to-ice phase transitions
[3–5]. Moreover, FTIR can be applied to study permeation of
protective agents into tissues or scaffolds, to estimate incubation
times needed for loading prior to freezing or drying them [6–8].
IR spectroscopy applied to cells or tissues relies on studying
characteristic molecular group vibrations of endogenous biomolecules. The vibrational frequency of a molecular group primarily
Willem F. Wolkers and Harrie ¨ tte Oldenhof (eds.), Cryopreservation and Freeze-Drying Protocols, Methods in Molecular Biology,
vol. 2180, https://doi.org/10.1007/978-1-0716-0783-1_13, © Springer Science+Business Media, LLC, part of Springer Nature 2021
331