6. Freeze-drying microscopy permits one to observe the gross
structural changes associated with collapse and can be directly
used to determine the collapse temperature. DSC has complementary advantages for formulation development optimization
of cooling protocols.
7. PVS1 contains 22% (w/v) glycerol, 15% (w/v) ethylene glycol,
15% (w/v) propylene glycol, 7% (w/v) dimethyl sulfoxide, and
0.5 M sorbitol in MS medium [5]. PVS2 contains 30% (w/v)
glycerol, 15% (w/v) ethylene glycol, 15% (w/v) dimethyl sulfoxide, and 0.4 M sucrose in MS medium [6]. VSL contains
20% (w/v) glycerol, 30% (w/v) ethylene glycol, 10% (w/v)
dimethyl sulfoxide, and 5% (w/v) sucrose in 10 mM CaCl 2 .
VSL+ contains 20% (w/v) glycerol, 30% (w/v) ethylene glycol,
10% (w/v) dimethyl sulfoxide, and 15% (w/v) sucrose in
10 mM CaCl 2 .
8. Meristems were pre-treated by placing on 0.75 M sucrose for
7 days, encapsulated in alginate and dehydrated in 0.57 M
liquid sucrose medium for 22 h. Samples were cooled at a
rate of 10
C per min from 25
C to À100
C and then at
5
C per min to À150
C. Sample was held isothermally for
5 min before ramping up to À100
C at a rate of 10
C/min
and at 5
C/min to 25
C.
References
1. Sun WQ (1999) State and phase transition
behaviors of Quercus rubra seed axes and cotyledonary tissues: relevance to the desiccation
sensitivity and cryopreservation of recalcitrant
seeds. Cryobiology 38:372–385
2. Sun WQ, Wagner CT, Connor J (2004) The
glass transition behaviors of hydroxyethyl
starch solutions. Cell Preserv Technol 2:55–65
3. Sun WQ (1997) Temperature and viscosity for
structural collapse and crystallization of amorphous carbohydrate solutions. CryoLetters
18:99–106
4. Rall WF, Fahy GM (1985) Ice-free cryopreservation of mouse embryos at À196 degrees C by
vitrification. Nature 313:573–575
5. Uragami A, Sakai A, Nagai M, Takahashi T
(1989) Survival of cultured cells and somatic
embryos of Asparagus officinalis cryopreserved
by vitrification. Plant Cell Rep 8:418–421
6. Sakai A, Kobayashi S, Oiyama I (1990) Cryopreservation of nucellar cells of navel orange
(Citrus sinensis Obs. var. brasiliensis Tanaka)
by vitrification. Plant Cell Rep 9:30–33
7. Suzuki M, Tandon P, Ishikawa M, Toyomasu T
(2008) Development of a new vitrification
solution, VSL, and its application to the cryopreservation of gentian axillary buds. Plant
Biotechnol Rep 2:123–131
8. Sherlock G, Block W, Benson EE (2005) Thermal analysis of the plant encapsulationdehydration cryopreservation protocol using
silica gel as the desiccant. CryoLetters
26:45–54
9. Sun WQ, Wagner CT, Liversey SA, Connor J
(2003) Instability of frozen human erythrocytes at elevated temperatures. Cell Preserv
Technol 1:255–267
10. Sun WQ, Davidson P (1998) Protein inactivation in amorphous sucrose and trehalose matrices: effects of phase separation and
crystallization.
Biochim
Biophys
Acta
1425:235–244
302
Wendell Q. Sun
structural changes associated with collapse and can be directly
used to determine the collapse temperature. DSC has complementary advantages for formulation development optimization
of cooling protocols.
7. PVS1 contains 22% (w/v) glycerol, 15% (w/v) ethylene glycol,
15% (w/v) propylene glycol, 7% (w/v) dimethyl sulfoxide, and
0.5 M sorbitol in MS medium [5]. PVS2 contains 30% (w/v)
glycerol, 15% (w/v) ethylene glycol, 15% (w/v) dimethyl sulfoxide, and 0.4 M sucrose in MS medium [6]. VSL contains
20% (w/v) glycerol, 30% (w/v) ethylene glycol, 10% (w/v)
dimethyl sulfoxide, and 5% (w/v) sucrose in 10 mM CaCl 2 .
VSL+ contains 20% (w/v) glycerol, 30% (w/v) ethylene glycol,
10% (w/v) dimethyl sulfoxide, and 15% (w/v) sucrose in
10 mM CaCl 2 .
8. Meristems were pre-treated by placing on 0.75 M sucrose for
7 days, encapsulated in alginate and dehydrated in 0.57 M
liquid sucrose medium for 22 h. Samples were cooled at a
rate of 10
C per min from 25
C to À100
C and then at
5
C per min to À150
C. Sample was held isothermally for
5 min before ramping up to À100
C at a rate of 10
C/min
and at 5
C/min to 25
C.
References
1. Sun WQ (1999) State and phase transition
behaviors of Quercus rubra seed axes and cotyledonary tissues: relevance to the desiccation
sensitivity and cryopreservation of recalcitrant
seeds. Cryobiology 38:372–385
2. Sun WQ, Wagner CT, Connor J (2004) The
glass transition behaviors of hydroxyethyl
starch solutions. Cell Preserv Technol 2:55–65
3. Sun WQ (1997) Temperature and viscosity for
structural collapse and crystallization of amorphous carbohydrate solutions. CryoLetters
18:99–106
4. Rall WF, Fahy GM (1985) Ice-free cryopreservation of mouse embryos at À196 degrees C by
vitrification. Nature 313:573–575
5. Uragami A, Sakai A, Nagai M, Takahashi T
(1989) Survival of cultured cells and somatic
embryos of Asparagus officinalis cryopreserved
by vitrification. Plant Cell Rep 8:418–421
6. Sakai A, Kobayashi S, Oiyama I (1990) Cryopreservation of nucellar cells of navel orange
(Citrus sinensis Obs. var. brasiliensis Tanaka)
by vitrification. Plant Cell Rep 9:30–33
7. Suzuki M, Tandon P, Ishikawa M, Toyomasu T
(2008) Development of a new vitrification
solution, VSL, and its application to the cryopreservation of gentian axillary buds. Plant
Biotechnol Rep 2:123–131
8. Sherlock G, Block W, Benson EE (2005) Thermal analysis of the plant encapsulationdehydration cryopreservation protocol using
silica gel as the desiccant. CryoLetters
26:45–54
9. Sun WQ, Wagner CT, Liversey SA, Connor J
(2003) Instability of frozen human erythrocytes at elevated temperatures. Cell Preserv
Technol 1:255–267
10. Sun WQ, Davidson P (1998) Protein inactivation in amorphous sucrose and trehalose matrices: effects of phase separation and
crystallization.
Biochim
Biophys
Acta
1425:235–244
302
Wendell Q. Sun