• Clinical and Experimental Reproductive Medicine
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• 제14권1호
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• pp.51-59
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• 1987

The present experiments have been bone to verify the effects of the warming rate and the degenerated blastomere(s) on further development of the frozen and thawed 4- and 8-cell mouse embryos. The embryos obtained from the mouse superovulated and mated were frozen in the solution of 15M DMSO in PBS containing 10% FCS at a slowly cooling rate($0.3^{\circ}C/min$). Two methods of warming slowly($8^{\circ}C/min$) and quickly ($450^{\circ}C/min$) were applied for thawing embryos. The thawed embryos were grouped according to the number of healthy blastomere(s) in the embryos. Some of the embryos were eliminated their degenerated blastomere(s) by means of a micromanipulation technique. The embryos were examined their developmental phases after 48 or 72 hrs incubation. The rates of blastocyst development from the frozen and thawed 4- and 8-cell embryos were 72.7% and 73.5%, respectively in case of thawing slowly, and were 78.9% and 80.0%, respectively in case of thawing quickly. The rate in case of thawing quickly was significantly higher than that in case of thawing slowly. The rates of blastocyst development from the frozen and thawed 4- and 8-cell embryos eliminated their degenerated blastomere(s) increased 5.9% and 24.4%, respectively compared with those of control groups not eliminated. The more number of degenerated blastomere(s) were eliminated from the embryos, the higher rate of blastocyst development was shown. It may be concluded from the results that the quickly thawing method is better for increasing survival rate than the slowly thawing one, and that the degenerated blastomere(s) in the frozen and thawed embryos affects as an interfering factor for further development of the embryos.

• 한국수정란이식학회지
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• 제24권4호
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• pp.275-279
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• 2009

The beneficial effect of glycerol as a cryoprotectant, especially for sperm cryopreservation, has been shown in many studies. However, glycerol is toxic to living cells, and boar sperm in particular show greater sensitivity to glycerol than sperm from other domestic animals. Amides have been studied as alternative cryoprotectants for freezing stallion sperm. Sperm frozen in methylformamide or dimethylformamide as cryoprotectants show similar motility when thawed compared with sperm frozen in glycerol. We evaluated the cryoprotective effects of dimethylformamide on boar sperm freezing. To test the effect of amides, the concentration of boar semen was adjusted to $10^9sperm/mL$, and seminal plasma was removed using Hulsen solution. After centrifugation, the pellet was diluted in modified-Modena B extender. Lactose-egg yolk (LEY) extender was used as the cooling extender. The freezing extender was madeed aaddition of the optimal amount of glycerol and amides to LEY-Glycerol-Orvus ES Paste extender, and this extender was used for the second dilution. Diluted sperm were frozen in liquid nitrogen using the 0.5 mL straw method. Sperm frozen in extender with glycerol as a cderol were compared with those frozen in extender including the different amides. Sperm were tested for motility, viability, the sperm chromatin structure assay, and normal apical ridge after thawing. The percent of motile sperm diluted in glycerol was as high as that in the stallion study (61%). Dimethylformamide showed positive effects on sperm quality and was better than glycerol. Methylformamide provided similar sperm quality as glycerol. Therefore, dimethylformamide is useful for reducing cryoinjury in boar sperm and is expected to be useful as an alternative cryoprotectant.

• Geomechanics and Engineering
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• 제29권3호
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• pp.281-290
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• 2022

The direct shear test is commonly used to evaluate the shear behavior of frozen soil-structure interfaces under normal stress. However, failure criteria, such as the Mohr-Coulomb failure criterion, are needed to obtain the unconfined shear strength. Hence, the punch shear test, which is usually used to estimate the shear strength of rocks without confinement, was examined in this study to directly determine the adfreezing strength. It is measured as the shear strength of the frozen soil-structure interface under unconfined conditions. Different soils of silica sand, field sand, and field clay were prepared inside the steel and concrete ring structures. Soil and ring structures were frozen at the target temperature for more than 24 h. A punch shear test was then conducted. The test results show that the adfreezing strength increased with a decrease in the target temperature and increase in the initial water content, owing to the increase in ice content. The adfreezing strength of field clay was the smallest when compared with the other soil specimens because of the large amount of unfrozen water content. The field sand with the larger normalized roughness showed greater adfreezing strength than the silica sand with a lower normalized roughness. From the experiment and analysis, the applicability of the punch shear test was examined to measure the adfreezing strength of the frozen soil-structure interface. To find a proper sample dimension, supplementary experiments or numerical analysis will be needed in further research.

• Carbon letters
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• 제13권4호
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• pp.248-253
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• 2012

Dielectrophoretic filtering and alignment of single-walled carbon nanotubes (SWCNTs) were tested using deuterium oxide as a solvent. A solution of deuterium oxide-SWCNTs was dropped on top of a silicon chip and an ac electric field was applied between pre-defined electrodes. Deuterium oxide was found to be a better solvent than hydrogen oxide for the dielectrophoresis process with higher efficiency of filtering. This was demonstrated by comparing Raman spectra measured on the initial solution with those measured on the filtered solution. We found that the aligned nanotubes along the electric field were not deposited on the substrate but suspended in solution, forming chain-like structures along the field lines. This so-called pearl chain formation of CNTs was verified by electrical measurements through the aligned tubes. The solution was frozen in liquid nitrogen prior to the electrical measurements to maintain the chain formation. The current-voltage characteristics for the sample demonstrate the existence of conduction channels in the solution, which are associated with the SWCNT chain structures.

• 한국수정란이식학회지
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• 제14권2호
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• pp.131-137
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• 1999

This experiment was carried out to investigate the effects of osmolarity of thawing diluents, seminal plasma added in thawing diluents on the sperm viability and the effects of thawing temperature, the temparature of the thawing diluents on the sperm viability and acrosomal morphology of boar spermatozoa by the straw method. The result obtained were summarized as follows: 1. The sperm viablilty after thawing of the frozen semen was shown greater in the high osmolarity(392~492mOsm) than low osmolarity(300mOsm) in thawing diluent. The added levels of seminal plasma in thawing diluent did not affect the viability of frozen-thawed boar semen. 2. In terms of thawing temperature, the sperm viability was shown higher in the frozen semen thawed at 5$0^{\circ}C$ for one min. (p<0.01) than those thawed at 2$0^{\circ}C$ or 37$^{\circ}C$ for one min. The sperm viability was not significant at the diluent temparature of 2$0^{\circ}C$or 37$^{\circ}C$ after thawing: but the sperm viability was higher in thawing diluent at 2$0^{\circ}C$ than in that at 37$^{\circ}C$. However, the effects of thawing temperature and diluent solution on normal acrosomal rate were not significant. 3. Cleavage rates of oocytes fertilized with frozen semen were 46.4% and 43.3%, respectively, which were thawed at 5$0^{\circ}C$ for one min. and then diluted in mBTS medium at 2$0^{\circ}C$or 37$^{\circ}C$. To sum up, the sperm viability was shown greater at the high of thawing diluents of frozen boar semen. In terms of thawing conditions, the sperm viability was shown greater, when semen was thawed at a high temperature for a short time and then diluted at the same temperature as that in the straw.

• 한국가축번식학회지
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• 제15권3호
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• pp.173-177
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• 1991

In vitro survival of the mouse morulae frozen by vitrification method(Kasai et al., 1990) was investigated in the present study. The embryos were plunged into LN2 directly after exposure to the vitrification solutions(EFS, GFS and DFS). The results were obtained as follows. The viability of morulae after freezing and thawing was high in EFS(96.7∼100.0%) and GFS vitrification solution(93.3∼96.7%), and the lowest in DFS vitrification solution(0.00∼0.03%).

• Bulletin of the Korean Chemical Society
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• 제11권2호
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• pp.115-118
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• 1990

Solution and frozen solution EPR spectra of $\alpha-1,2,3-[HPV(IV)V_2W_9O_{40}]^{6-}$ have been analyzed. The isotropic hyperfine coupling constants remain constant at 350-77 K, indicating that the unpaired electron is delocalized over three vanadium atoms probably even in the ground state.

• Journal of Applied Biological Chemistry
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• 제58권1호
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• pp.55-60
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• 2015

수출용 냉동딸기 제조 시 미생물학적 안전성 확보를 위한 수단으로 이산화염소수와 acetic acid 병합처리를 최적 비가열 전처리 기술로써 적용하여 -20, -70, $-196^{\circ}C$로 냉동한 후 미생물 수, 품질변화 및 관능평가를 조사하였다. 냉동방법 및 세척 처리에 따른 색도 차이는 나타나지 않았고, 비타민 C 함량은 $-70^{\circ}C$ 냉동에서 35.33 mg/100 g FW로 가장 대조구와 유사하였으며, drip loss도 $-70^{\circ}C$ 냉동이 14.39%로 가장 낮게 나타났다. 관능평가 역시 $-70^{\circ}C$ 냉동이 -20, $-196^{\circ}C$ 냉동보다 높은 점수를 받았으며, 세척처리는 비타민 C 함량, drip loss 및 관능평가에 큰 영향을 끼치지 않는 것으로 나타났다. 또한, 냉동딸기에 50 ppm 이산화염소수와 1% acetic acid를 병합 처리하여 냉동 후 미생물 수 변화를 측정한 결과, 냉동 전과 같이 병합처리 된 딸기 시료에서 미생물이 검출되지 않았다. 따라서 저장성이 높은 수출용 냉동딸기 생산을 위해서는 gas nitrogen convection chamber를 이용한 $-70^{\circ}C$에서의 급속냉동 처리가 보다 효과적인 냉동방법이며, 냉동 처리만으로는 미생물 제어가 어렵기 때문에 냉동 전 비가열 전처리를 통해서 냉동딸기의 미생물학적 안전성을 확보해야 한다고 판단된다.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2002년도 국제심포지엄
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• pp.78-78
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• 2002

The role of heat shock proteins in shielding organism from environmental stress is illustrated by the large-scale synthesis of these protein by the organism studied to date. However, recent evidence also suggests an important role for heat shock protein in fertilization and early development of mammalian embryos. Effects of elevated in vitro temperature on in vitro produced bovine embryos were analysed in order to determine its impact on the expression of heat shock protein 70 (HSP70) by control and frozen-thawed after in vitro fertilization (IVF) or nuclear transfer (NT). The objective of this study was to assess the developmental potential in vitro produced embryos with using of the various containers and examined expression and localization of heat shock protein 70 after it's frozen -thawed. For the vitrification, in vitro produced embryos at 2 cell, 8 cell and blastocysts stage after IVF and NT were exposed the ethylene glycol 5.5 M freezing solution (EG 5.5) for 30 sec, loaded on each containers such EM grid, straw and cryo-loop and then immediately plunged into liquid nitrogen. Thawed embryos were serially diluted in sucrose solution, each for 1 min, and cultured in CRI-aa medium. Survival rates of the vitrification production were assessed by re-expanded, hatched blastocysts. There were no differences in the survival rates of IVF using EM grid, cryo-loop. However, survival rates by straw were relatively lower than other containers. Only, nuclear transferred embryos survived by using cryo-loop. After IVF or NT, in vitro matured bovine embryos 2 cell, 8 cell and blastocysts subjected to control and thawed conditions were analysed by semiquantitive reverse transcription polymerase chain reaction methods for hsp 70 mRNA expression. Results revealed the expression of hsp 70 mRNA were higher thawed embryos than control embryos. Immunocytochemistry used to localization the hsp70 protein in embryos. Two, 8-cell embryos derived under control condition was evenly distributed in the cytoplasm but appeared as aggregates in some embryos exposed frozen-thawed. However, under control condition, blastocysts displayed aggregate signal while Hsp70 in frozen-thawed blastocysts appeared to be more uniform in distribution.

• 한국동물생명공학회지
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• 제35권1호
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• pp.58-64
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• 2020

This present study was conducted to investigate protective effect of discontinuous Percoll gradient containing alpha-linolenic acid (ALA) before freezing process on viability, acrosome damage, mitochondrial activity, and oxidative stress of frozen-thawed boar spermatozoa. The separation of spermatozoa by discontinuous Percoll gradient was performed by different concentration of Percoll solution (45/90%) containing ALA combined with bovine serum albumin (BSA), and collected sperm in each Percoll layer was cryopreserved. To evaluate viability, acrosome damage, mitochondrial activity, and reactive oxygen species (ROS) level of frozen-thawed sperm, flow cytometry was used. Morphological abnormalities were observed under light microscope. In results, viability of sperm from 90% Percoll layer was higher than control and 45% Percoll group (p < 0.05). Separated sperm in 90% Percoll layer had lower acrosome damage and morphological abnormalities than control as well as viability, whereas 45% Percoll group was higher (p < 0.05). Similar with acrosome damage and abnormalities, mitochondrial activity was slightly enhanced and the population of live sperm with high ROS level was decreased by 90% Percoll separation, however, there was no significant difference. Supplementation of 3 ng/mL ALA into Percoll solution increased sperm viability and decreased population of live sperm with high ROS compared to control (p < 0.05). In conclusion, discontinuous Percoll gradient before freezing process could improve efficiency of cryopreservation of boar sperm through selection of sperm with high freezing resistance, and supplement of ALA during Percoll gradient might contribute suppression of ROS generation via stabilizing of plasma membrane during cryopreservation.