trehalose and albumin is stable for months even under accelerated
aging conditions [54]. Sugars also stabilize DNA in freeze-dried
somatic cells during storage [79]. Rehydration can be done by
directly adding water or by gradually rehydrating the sample
under high relative humidity conditions.
7.3 Formulations
for Freeze-Drying
of Tissues
In order to freeze-dry tissues that are used as biological scaffolds, it
is important to preserve the tissue histoarchitecture and biomechanical characteristics. In the absence of protective agents, ice
crystal formation during freezing is evident as pores in the tissue
visible after rehydration. Pores are still visible if lyoprotective sugars
are used at ~5% (w/v) concentrations; however, pore formation can
be avoided by osmotic dehydration of the tissue prior to freezedrying. Therewith, similarly as is done in “cold” vitrification procedures, first tissue is loaded with sugars by immersing in a 5%
(w/v) sugar solution whereafter tissues are exposed to a highly
concentrated sugar solution (>40% w/v) to reduce the water content [105, 106]. Freeze-dried acellular tissue matrices can be used
to replace diseased or malfunctioning tissue. Examples are acellular
skin tissue matrix [107], decellularized heart valve conduits [105],
and pericardium patches [108]. Both sucrose and trehalose can be
used as protective agents; however, this process requires large
amounts of sugars, and therefore use of sucrose may be preferred
above trehalose. Macromolecules are typically not used for freezedrying of tissue scaffolds, because their permeation rates are very
slow [109] and tissue proteins themselves function as bulking
agent.
References
1. Crowe JH, Carpenter JF, Crowe LM (1998)
The role of vitrification in anhydrobiosis.
Annu Rev Physiol 60:73–103
2. Crowe JH, Hoekstra FA, Crowe LM (1992)
Anhydrobiosis.
Annu
Rev
Physiol
54:579–599
3. Alpert P (2000) The discovery, scope, and
puzzle of desiccation tolerance in plants.
Plant Ecol 151:5–17
4. Boothby TC, Pielak GJ (2017) Intrinsically
disordered proteins and desiccation tolerance:
elucidating functional and mechanistic underpinnings
of anhydrobiosis.
BioEssays
39:1700119
5. Bartels D, Schneider K, Terstappen G,
Piatkowski D, Salamini F (1990) Molecular
cloning of abscisic acid-modulated genes
which are induced during desiccation of the
resurrection plant Craterostigma plantagineum. Planta 181:27–34
6. Amuti KS, Pollard CJ (1977) Soluble carbohydrates of dry and developing seeds. Phytochemistry 16:529–532
7. Buitink J, Walters-Vertucci C, Hoekstra FA,
Leprince O (1996) Calorimetric properties of
dehydrating pollen: analysis of a desiccation
tolerant and an intolerant species. Plant
Physiol 111:235–242
8. Crowe JH, Crowe LM, Oliver AE,
Tsvetkova N, Wolkers W, Tablin F (2001)
The trehalose myth revisited: introduction to
a symposium on stabilization of cells in the
dry state. Cryobiology 43:89–105
9. Leprince O, Pellizzaro A, Berriri S, Buitink J
(2017) Late seed maturation: drying without
dying. J Exp Bot 68:827–841
10. Tanaka S, Tanaka J, Miwa Y, Horikawa DD,
Katayama T, Arakawa K, Toyoda A, Kubo T,
Kunieda T (2015) Novel mitochondriatargeted heat-soluble proteins identified in
20
Willem F. Wolkers and Harrie ¨ tte Oldenhof
aging conditions [54]. Sugars also stabilize DNA in freeze-dried
somatic cells during storage [79]. Rehydration can be done by
directly adding water or by gradually rehydrating the sample
under high relative humidity conditions.
7.3 Formulations
for Freeze-Drying
of Tissues
In order to freeze-dry tissues that are used as biological scaffolds, it
is important to preserve the tissue histoarchitecture and biomechanical characteristics. In the absence of protective agents, ice
crystal formation during freezing is evident as pores in the tissue
visible after rehydration. Pores are still visible if lyoprotective sugars
are used at ~5% (w/v) concentrations; however, pore formation can
be avoided by osmotic dehydration of the tissue prior to freezedrying. Therewith, similarly as is done in “cold” vitrification procedures, first tissue is loaded with sugars by immersing in a 5%
(w/v) sugar solution whereafter tissues are exposed to a highly
concentrated sugar solution (>40% w/v) to reduce the water content [105, 106]. Freeze-dried acellular tissue matrices can be used
to replace diseased or malfunctioning tissue. Examples are acellular
skin tissue matrix [107], decellularized heart valve conduits [105],
and pericardium patches [108]. Both sucrose and trehalose can be
used as protective agents; however, this process requires large
amounts of sugars, and therefore use of sucrose may be preferred
above trehalose. Macromolecules are typically not used for freezedrying of tissue scaffolds, because their permeation rates are very
slow [109] and tissue proteins themselves function as bulking
agent.
References
1. Crowe JH, Carpenter JF, Crowe LM (1998)
The role of vitrification in anhydrobiosis.
Annu Rev Physiol 60:73–103
2. Crowe JH, Hoekstra FA, Crowe LM (1992)
Anhydrobiosis.
Annu
Rev
Physiol
54:579–599
3. Alpert P (2000) The discovery, scope, and
puzzle of desiccation tolerance in plants.
Plant Ecol 151:5–17
4. Boothby TC, Pielak GJ (2017) Intrinsically
disordered proteins and desiccation tolerance:
elucidating functional and mechanistic underpinnings
of anhydrobiosis.
BioEssays
39:1700119
5. Bartels D, Schneider K, Terstappen G,
Piatkowski D, Salamini F (1990) Molecular
cloning of abscisic acid-modulated genes
which are induced during desiccation of the
resurrection plant Craterostigma plantagineum. Planta 181:27–34
6. Amuti KS, Pollard CJ (1977) Soluble carbohydrates of dry and developing seeds. Phytochemistry 16:529–532
7. Buitink J, Walters-Vertucci C, Hoekstra FA,
Leprince O (1996) Calorimetric properties of
dehydrating pollen: analysis of a desiccation
tolerant and an intolerant species. Plant
Physiol 111:235–242
8. Crowe JH, Crowe LM, Oliver AE,
Tsvetkova N, Wolkers W, Tablin F (2001)
The trehalose myth revisited: introduction to
a symposium on stabilization of cells in the
dry state. Cryobiology 43:89–105
9. Leprince O, Pellizzaro A, Berriri S, Buitink J
(2017) Late seed maturation: drying without
dying. J Exp Bot 68:827–841
10. Tanaka S, Tanaka J, Miwa Y, Horikawa DD,
Katayama T, Arakawa K, Toyoda A, Kubo T,
Kunieda T (2015) Novel mitochondriatargeted heat-soluble proteins identified in
20
Willem F. Wolkers and Harrie ¨ tte Oldenhof