transferred to liquid nitrogen without raising the sample temperature. When using Mr. Frosty or similar 1
C/min-freezing
containers, extracellular ice spontaneously forms at a random
temperature, which introduces variability into the freezing
outcome.
8. To maintain sterility, do not remove 0.5-cc straws from their
bag. Just open one end of the bag where cotton-plugged ends
of straws are located, and label only cotton-plugged ends of
straws. Compared to cryovials, 0.5-cc straws are easy to seed
extracellular ice and are preferable if small volumes
(0.3–0.5 mL for each straw) of cell suspensions are cryopreserved. Nevertheless, similar viability rates were obtained after
cryopreservation of ASCs in 0.5-cc straws and 1.5-cc
cryovials [19].
9. Use extreme caution when handling liquid nitrogen. Severe
frostbite and even suffocation may occur as a result of direct
liquid nitrogen exposure and displacement of air by evaporating liquid nitrogen, respectively.
10. In our previous study, we compared thawing in air (RT) and in
a water bath at 37
C after cryopreservation of ASCs in 0.5-cc
straws. We did not observe any significant difference in terms of
viability and plating efficiency (see Fig. 1) [19]. However, we
recommend thawing cryovials in a water bath at 37
C.
References
1. Gimble JM, Guilak F, Bunnell BA (2010) Clinical and preclinical translation of cell-based
therapies using adipose tissue-derived cells.
Stem Cell Res Ther 1:1–8
2. Boquest AC, Shahdadfar A, Brinchmann JE,
Collas P (2006) Isolation of stromal stem cells
from human adipose tissue. Methods Mol Biol
325:35–46
3. Kern S, Eichler H, Stoeve J, Kluter H, Bieback
K (2006) Comparative analysis of mesenchymal stem cells from bone marrow, umbilical
cord blood, or adipose tissue. Stem Cells
24:1294–1301
4. Yoshimura K, Shigeura T, Matsumoto D,
Sato T, Takaki Y, Aiba-Kojima E, Sato K,
Inoue K, Nagase T, Koshima I, Gonda K
(2006) Characterization of freshly isolated
and cultured cells derived from the fatty and
fluid portions of liposuction aspirates. J Cell
Physiol 208:64–76
5. Gimble JM, Katz AJ, Bunnell BA (2007)
Adipose-derived stem cells for regenerative
medicine. Circ Res 100:1249–1260
6. Safford KM, Hicok KC, Safford SD, Halvorsen
YDC, Wilkison WO, Gimble JM, Rice HE
(2002) Neurogenic differentiation of murine
and human adipose-derived stromal cells. Biochem Biophys Res Comm 294:371–379
7. Guilak F, Lott KE, Awad HA, Cao QF, Hicok
KC, Fermor B, Gimble JM (2006) Clonal analysis of the differentiation potential of human
adipose-derived adult stem cells. J Cell Physiol
206:229–237
8. Zuk PA, Zhu M, Mizuno H, Huang J, Futrell
JW, Katz AJ, Benhaim P, Lorenz HP, Hedrick
MH (2001) Multilineage cells from human
adipose tissue: Implications for cell-based
therapies. Tissue Eng 7:211–228
9. Ashjian PH, Elbarbary AS, Edmonds B,
DeUgarte D, Zhu M, Zuk PA, Lorenz HP,
Benhaim P, Hedrick MH (2003) In vitro differentiation of human processed lipoaspirate
cells into early neural progenitors. Plastic
Reconstr Surg 111:1922–1931
10. Yanez R, Lamana ML, Garcia-Castro J,
Colmenero I, Ramirez M, Bueren JA (2006)
Adipose tissue-derived mesenchymal stem cells
have in vivo immunosuppressive properties
applicable for the control of the graft-versushost disease. Stem Cells 24:2582–2591
566
Melany Lo ´ pez and Ali Eroglu
C/min-freezing
containers, extracellular ice spontaneously forms at a random
temperature, which introduces variability into the freezing
outcome.
8. To maintain sterility, do not remove 0.5-cc straws from their
bag. Just open one end of the bag where cotton-plugged ends
of straws are located, and label only cotton-plugged ends of
straws. Compared to cryovials, 0.5-cc straws are easy to seed
extracellular ice and are preferable if small volumes
(0.3–0.5 mL for each straw) of cell suspensions are cryopreserved. Nevertheless, similar viability rates were obtained after
cryopreservation of ASCs in 0.5-cc straws and 1.5-cc
cryovials [19].
9. Use extreme caution when handling liquid nitrogen. Severe
frostbite and even suffocation may occur as a result of direct
liquid nitrogen exposure and displacement of air by evaporating liquid nitrogen, respectively.
10. In our previous study, we compared thawing in air (RT) and in
a water bath at 37
C after cryopreservation of ASCs in 0.5-cc
straws. We did not observe any significant difference in terms of
viability and plating efficiency (see Fig. 1) [19]. However, we
recommend thawing cryovials in a water bath at 37
C.
References
1. Gimble JM, Guilak F, Bunnell BA (2010) Clinical and preclinical translation of cell-based
therapies using adipose tissue-derived cells.
Stem Cell Res Ther 1:1–8
2. Boquest AC, Shahdadfar A, Brinchmann JE,
Collas P (2006) Isolation of stromal stem cells
from human adipose tissue. Methods Mol Biol
325:35–46
3. Kern S, Eichler H, Stoeve J, Kluter H, Bieback
K (2006) Comparative analysis of mesenchymal stem cells from bone marrow, umbilical
cord blood, or adipose tissue. Stem Cells
24:1294–1301
4. Yoshimura K, Shigeura T, Matsumoto D,
Sato T, Takaki Y, Aiba-Kojima E, Sato K,
Inoue K, Nagase T, Koshima I, Gonda K
(2006) Characterization of freshly isolated
and cultured cells derived from the fatty and
fluid portions of liposuction aspirates. J Cell
Physiol 208:64–76
5. Gimble JM, Katz AJ, Bunnell BA (2007)
Adipose-derived stem cells for regenerative
medicine. Circ Res 100:1249–1260
6. Safford KM, Hicok KC, Safford SD, Halvorsen
YDC, Wilkison WO, Gimble JM, Rice HE
(2002) Neurogenic differentiation of murine
and human adipose-derived stromal cells. Biochem Biophys Res Comm 294:371–379
7. Guilak F, Lott KE, Awad HA, Cao QF, Hicok
KC, Fermor B, Gimble JM (2006) Clonal analysis of the differentiation potential of human
adipose-derived adult stem cells. J Cell Physiol
206:229–237
8. Zuk PA, Zhu M, Mizuno H, Huang J, Futrell
JW, Katz AJ, Benhaim P, Lorenz HP, Hedrick
MH (2001) Multilineage cells from human
adipose tissue: Implications for cell-based
therapies. Tissue Eng 7:211–228
9. Ashjian PH, Elbarbary AS, Edmonds B,
DeUgarte D, Zhu M, Zuk PA, Lorenz HP,
Benhaim P, Hedrick MH (2003) In vitro differentiation of human processed lipoaspirate
cells into early neural progenitors. Plastic
Reconstr Surg 111:1922–1931
10. Yanez R, Lamana ML, Garcia-Castro J,
Colmenero I, Ramirez M, Bueren JA (2006)
Adipose tissue-derived mesenchymal stem cells
have in vivo immunosuppressive properties
applicable for the control of the graft-versushost disease. Stem Cells 24:2582–2591
566
Melany Lo ´ pez and Ali Eroglu