• Development and Reproduction
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• v.12 no.3
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• pp.221-229
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• 2008

For the clinical application, it is needed to keep characteristics of stem cells after storage for a long time. In the present study, we examined stem cell properties of human cord-derived stem cells (HUC) after cryopreservation. Cells were isolated from human umbilical cord and cultured in vitro. At passage 2 or 3, HUC were suspended at a concentration of $1.0{\times}10^6/m{\ell}$ in cryomedium consisting of DMSO and FBS. After freezing at $-80^{\circ}C$ overnight, HUC were cryopreserved at $-196^{\circ}C$ nitrogen gas. After 6 months, HUC were thawed and cultured in vitro. Assessment for the stem cell properties was made upon the morphology, population doubling time, and expression profiles of genes and various proteins. Cryopreserved HUC showed more than 70% viability and maintained fibroblast-like morphology similar to HUC before cryopreservation. Throughout the culture, they underwent average 42.8 doublings and produced $6.75{\times}{10^{18}}$ cells. RT-PCR analyses showed that cryopreserved HUC expressed Oct-4, nanog, SCF, NCAM, nestin, GATA-4, BMP4, and HLA-1 genes. They did not express Brachyury and HLA-DR genes. Immunocytochemical studies showed that cryopreserved HUC reacted with antibodies against SSEA-3, -4, Thy-1, vimentin, fibronectin, HCAM, ICAM, HLA-1 proteins. They did not react with antibody against HLA-DR protein. Theses genes and proteins expression patterns of cryopresserved HUC were similar to those of HUC before cryopreservation. These results suggest that cryopreserved HUC could retain proliferative potential and they expressed various genes and proteins similar to HUC before cryopreservation. Thus, cryopreservation might be useful for HUC for future research and clinical application.

• Journal of Embryo Transfer
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• v.19 no.3
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• pp.307-313
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• 2004

본 연구는 고양이의 신선 및 동결 정소상체 정액과 정소상체 정액 성상과 및 동결보존시의 생존성 및 난포란과 정소상체 정자의 체외수정 후 체외수정율과 분할율에 대해 조사하였다. 고양이 정소상체 정액의 정자농도는 3.25$\pm$0.75${\times}$$10^{6}$ cells/$m\ell$, 정자의 활력은 70.85$\pm$4.20%, 기형정자 수는 8.55$\pm$1.85%로서 대조군인 사출정액의 정자농도는 5.05$\pm$0.40${\times}$$10^{6}$ cells/$m\ell$, 정자의 활력은 90.24$\pm$455%, 기형정자 수는 4.20$\pm$0.50%와 비교할 때 정자농도와 활력은 낮았으며, 기형정자 수는 많았다. 고양이 정액과 tris-buffer로 희석한 정액을 20분간 배양했을 때 정자농도는 3.50$\pm$0.40${\times}$$10^{6}$ cells/$m\ell$, 정자활력 은 75.50$\pm$2.55%, 기 형정자 수는 6.75$\pm$0.58%로서 희석하지 않은 정소상체 정액의 성상에 비해 약간 높게 나타났다. Tris-buffer로 희석한 고양이 정소상체 정액을 동결 융해했을 때 생존율은 54.50$\pm$4.45, 활력은 47.50$\pm$6.40%로서 희석하지 .않은 대조군의 생존을 74.50$\pm$6.25%와 활력 78.50$\pm$5.20%에 비해 현저히 높게 나타났다. 고양이의 난포란과 신선 및 동결 정소상체 정자를 수정시켰을 때 체외수정율과 분할율은 68.30$\pm$5.35%, 57.25$\pm$4.35% 및 48.65$\pm$4.95%, 35.65 $\pm$4.75%로서 신선 정자에 비해 동결 정소상체 정자로 수정시킨 군의 분할율이 유의하게 낮았다.

• Korean Journal of Plant Resources
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• v.17 no.2
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• pp.75-81
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• 2004

This study was conducted to investigate effects of cryopreservation by vitrification on the seed germination rate and growth and physiological properties of seedlings of Maackia amurensis. Cryopreservation significantly decreased the germination rate of seeds of M. amurensis, but the reduction of germination rate was mitigated by the treatment of cryoprotectant (plant vitrification solution, PVS2) before plugging into liquid nitrogen and fast thawing rate after cryopreservation. Long-term PVS2 exposure decreased seed germination rate, whereas cryopreservation time didn't have influence on seed germination rate. In addition, growth and physiological properties of seedlings were not affected by PVS2 exposing time and cryopreservation time. Therefore cryopreservation could be widely used as a technique of long-term ex situ conservation without any damage and deterioration of cells or tissues of the forest seeds. However, in order to increase the effect of cryopreservation, we have to develope the lower toxic cryoprotectant and suitable techniques to the structural or chemical properties of a variety of seeds.

• Korean Journal of Food Science and Technology
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• v.54 no.2
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• pp.224-227
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• 2022

In this study, the protective effects of turanose on Saccharomyces cerevisiae (Baker's yeast) were examined during the freeze-drying process to evaluate the feasibility of utilizing turanose as a novel cryoprotectant. The survival rate of the Baker's yeast cells improved substantially with a cryoprotective medium containing turanose in a substitution-dependent manner. In accordance with these survival rates, the yeast cell surfaces became smoother as the turanose content increased. Turanose with skim milk maintained the viability of the Baker's yeast, which improved substantially upon storage at -20℃. Thus, it is thought that turanose will exhibit excellent preservation effects during the distribution of Baker's yeast. Finally, these results suggest that turanose has the potential to be used as a novel cryoprotectant against various microorganisms.

• Korean Journal of Animal Reproduction
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• v.24 no.3
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• pp.263-268
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• 2000

The development of single blastomeres isolated from 2-, 4- and 8-cell mouse embryos and the ability of such blastomeres to survive slow freezing were studied. Of 223, 60 and 188 single blastomeres isolated from 2-, 4- and 8-cell mouse embryos, respectively, 111 blastomeres (49.8%) from 2-cell embryos, 12 blastomeres (20.0%) from 4-cell embryos and blastomeres (16.5) from 8-cell embryos developed into blastocysts after culture for 96 hrs. The recovery rate was 54.2% (65/120), 46.4% (13/28) and 24.3% (17/70) of blastomeres derived from 2-, 4- and 8-cell embryos following freezing and thawing and the survival of frozen-thawed blastomeres was 27.1% (16/59), 36.4% (4/11) and 17.6% (3/17), and respectively. The apparently six normal fetuses were obtained from frozen-thawed blastomere from 2-cell embryos after transferring into the recipients. These results indicate that mouse btastomeres isolated from preimplatation stage embryos can survive storage in liquid nitrogen following slow freezing.

• Conservation Science in Museum
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• v.5
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• pp.37-41
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• 2004

The immersed ancient wood relics excavated from the low swampy area of historic site in Shinchangdong, Gwangju are extremely weak. Since they are quickly dried inducing shrinkage when exposed to air, special preservative treatment is required to exhibit and preserve such items. In the case of the heavily varnished lacquerware such as the lacquered sheaths, in particular, it is extremely difficult to preserve them because chemicals are not easily penetrated and the coating might swell or come off during the process of coating. The lacquer sheaths have been preserved by carrying out freeze-drying after impregnating them in aqueous solution with low concentration of PEG#4000 40%.

• Korean Journal of Poultry Science
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• v.41 no.1
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• pp.21-27
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• 2014

To preserve chicken genetic materials like cryopreserved spermatozoa, various kinds of freezing agents like glycerol, dimethylsuloxide, dimethylformamide or dimethylacetamide have been used for rooster semen preparation. Recently, the usage of N-methylacetamide (MA) for Ogye rooster semen preservation resulted in hatched chicken successfully. In this study, we investigated the effects of 7, 9 and 11% of MA on the viability, fertility and hatchability of frozen-thawed rooster semen using artificial insemination. The results of viability, fertility and hatchability in frozen semen with 7%, 9% or 11% MA were $35.16{\pm}6.12%$, $67.83{\pm}15.3%$ and $66.2{\pm}16.3%$ of motile sperm rate, 21.5%, 34.7% and 25% of fertility rate, and 100%, 89.5% and 87.5% of hatchability rate. The results of control group with frozen semen were 96.0% of fertility rate and 92.2% of hatchability rate. With these results, the concentration range of MA as a freezing agent of rooster semen could be 7~9% of media. The higher concentration of 9 % MA could decrease the fertility rate of thawed semen not the rate of hatchability rate. So the use of MA without affecting fertility rate would be a key point of freezing method of rooster semen for poultry genetic resource preservation.

• Korean Journal of Animal Reproduction
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• v.20 no.1
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• pp.35-42
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• 1996

This study was carried out to investigate the effect of straw loading method and thawing protocol on the in vitro development of in vitro produced mouse blastocysts cryopreserved by vitrification. Three loading types of straw I, Il and III on loading and sealing method were made for vitrification. The ability of the solution on straw loading methods to remain vitreous during warming was tested by exposed in air for 1 to 10 s sec. and then plunged the vitrified straws into water bath at 25°C. Embryos to be vitrified were equilibrated to the 20% EG for 5min. and exposed in EFS 40 for 1min. The plug ends of Straw I and Straw II were sealed with straw powder and Straw III was treated straw powder, followed by heat sealing and then plunged into LN$_2$. The results obtained in these experiments were summarized as follows; 1) Straw I embryo column mostly changed from transparent to opaque upon thawing without exposure in air for 3-6 sec. Straw II embryo column was I improved partially but was not remained completely vitreous during warming. However, when Straw lll loading method was used, the embryo column was remained vitreous completely. 2) High survival rates and development rates of each groups (middle blastocysts and hatching blastocysts) of vitrified embryos were obtained by using Straw III loading method than Straw I method (P<0.05). And the range of s standard error was low in Straw lll method. 3) When the embryos vitrified-frozen were placed in air for 3, 5 and l0sec. and then warmed rapidly in water bath at 25$^{\circ}C$, the survival rates after 24h of culture were 72.7-87.1% and the development rates to hatching stage after 48h of culture were 34.0-48.4%. There were no significantly differences according to exposure time in air during warming. In conclusion, the present results showed that highly survival and low standard error of vitrified-frozen mouse bIastocysts were obtained by using straw lll loading, double sealing and appropriate 2 step warming method.

• Journal of Embryo Transfer
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• v.29 no.3
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• pp.241-248
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• 2014

This study was carried out to evaluate the effects of embryonic stage, cryoprotectant, and freezing-thawing method on the rates of survival and development of the cryopreserved mouse early embryo and finally to establish the cryopreservation method of surplus embryos obtained during assisted reproductive technology (ART). Two to eight cell embryos were obtained from oviducts of mated $F_1$ hybrid female mice superovulated by PMSG and hCG. Two-step EG, DMSO and 4-step EG, DMSO were used as cryoprotectant and dehydration and rehydration method of embryos, and slow-cooling or rapid-cooling method was used as frozen program. The survival rates of embryos were measured after thawing and rehydration, and the developmental rates of embryos were compared and observed during culturing embryos for 24, 48, 72, 96 hrs. As for the survival and development rates of embryos according to embryonic stage, the survival rate of 2 cell stage in EG and DMSO was significantly higher than 4~8 cell (65.4% versus 61.2%, 81.1% versus 72.5%) (p<0.01, p<0.01), but the development rates of 4~8 cell embryos in EG and DMSO were significantly higher than 2 cell embryos for whole culture period (p<0.01) and the development rates of 4~8 cell embryos in EG were significantly higher than 2 cell embryos in DMSO (p<0.01). As for the survival and development rates of embryos according to cryoprotectant, the survival rate of 2 cell embryo in DMSO was significantly higher than that in EG(77.0% versus 64.4%) (p<0.01), whereas the development rate of embryos was not differ till 24 hrs. The development rate from morular to hatching blastocyst, however, was sinificantly higher in EG than in DMSO during 48 hr (p<0.01). The survival rate of 4~8 cell embryo was 62.5% in EG and 73.3% in DMSO. The development rates of embryo in EG were significantly higher for whole culture periods (p<0.01, 0.05). In respect to the effect of freezing and thawing program on the survival and development rates of embryos, method of slow cooling and rapid thawing was more effective than that of rapid cooling and rapid thawing. The survival rate of embryo in 2 cell stage was higher than in 4~8 cell stage, and EG appears more effective cryoprotectant than DMSO because EG showed better development rates of embryos in 2 and 4~8 cell stage. Moreover, slow cooling and rapid thawing method was considered as the best cryopreservation program.

• Journal of Plant Biotechnology
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• v.44 no.4
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• pp.478-483
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• 2017

The expression profiles of low-temperature-related genes were examined in in vitro potatoes exposed to a cold condition for 1-3 days. The expression levels of PPI1 and CI21B genes were lineraly elevated from day 1 to day 3, while the opposite trend was observed for CBF4 and CI21A. In addition, the expression of the genes CI21A and CI21B varied, along with specific tissues (leaf, stem, and tuber) and the treatment periods. Therefore, potato shoot tips were cryopreserved with LS and PVS2 containing different oncentrations of AFP. It can thus be inferred that the presence of AFP in LS and PVS2 was likely to elevate expression pattern of the genes. Furthermore, the concentration of AFP (1,500 ng/ml for LS and 500 ng/mg for PVS2) was the best for the cryopreservation of potatoes.