• Clinical and Experimental Reproductive Medicine
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• 제26권2호
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• pp.127-135
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• 1999

Cryopreservation is able to store the surplus pre-embryos for freezing and furthermore thawing and transfer in a subsequent cycle. Cryopreserving cells which are maintaining their viability are the very complex process. This study has been carried out in order to find the effects of cryopreservation steps, freezing media and embryonic stages on the rates of viability and development of cryopreserved mouse embryos. Female ICR mice ($6{\sim}8$ weeks old) were induced to superovulate by sequential intraperitoneal injection of 5 IU PMSG and 5 IU hCG 48h apart. Mouse embryos were collected according to its developmental stage after the injection of hCG. Embryos were cryopreserved not only by cryoprotectant step (1 step${\sim}$4 step) but also in a variety of media (HTF, IVF medium, D-PBS) and cell stage. The results were as follows: There is no clear advantage in these freezing media of rapid method, but 4 cell and 8 cell of slow method (2, 3, 4 step) have advantage in D-PBS. The development of embryos according to cell stage become greater in 8 cell stage. In the treatment steps of cryopreservation, the development of embryo to blastocyst was similar among rapid method, but the development of 4 cell and 8 cell embryos to blastocyst according to slow method was better than rapid method.

• 한국수정란이식학회지
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• 제22권4호
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• pp.251-255
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• 2007

This study was carried out to examine on developmental competence of Hanwoo embryos cultured in vitro according to culture conditions and freezing methods. The in vitro developmental competence to blastocyst stage at Day 8 of culture in SOF was significantly (p<0.05) higher than that in CR1aa (30.3% vs. 18.4%). The in vitro developmental rate of morula and blastocysts cultured in group culture was significantly (p<0.05) higher than that in individual culture (41.4% and 36.0% vs. 21.1% and 10.5%, respectively). The cell number of Day 8 blastocysts in group culture was significantly (p<0.05) higher than that in the individual culture ($120.1{\pm}12.8\;vs.\;94.1{\pm}12.1$, respectively). The survival rates of frozen-thawed balstocysts that were exposed in 1.5 M ethylene glycol or 1.5 M ethylene glycol containing 0.1 M sucrose were 77.5% and 78.7%, respectively. The survival rates of blastocysts cultured for 48 h in slow freezing and vitrification was not significantly different (73.3 and 74.0%). In conclusion, in vitro developmental competence of bovine embryos was influenced on the culture medium (SOF) and culture method (Group culture). Survival rate of frozen-thawed of bovine embryos was not influenced on freezing solutions and freezing methods.

• 한국축산식품학회지
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• 제41권6호
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• pp.1012-1021
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• 2021

This study evaluated the effect of freezing rate on the quality characteristics of pork loin to establish an objective standard for rapid freezing. To generate various freezing rates, three air flow rates (0, 1.5, and 3.0 m/s) were applied under three freezing temperatures (-20℃, -30℃, and -40℃). Based on the results, freezing rates ranged from 0.26-1.42 cm/h and were graded by three categories, i.e, slow (category I, >0.4 cm/h), intermediate (category II, 0.6-0.7 cm/h) and rapid freezing (category III, >0.96 cm/h). Both temperature and the air flow rate influenced the freezing rate, and the freezing rate affected the ice crystal size and shear force in pork loin. However, the air flow rate did not affect thawing loss, drip loss or the color of pork loins. In the comparison of freezing rates, pork belonging to category II did not show a clear difference in quality parameters from pork in category I. Furthermore, pork in category III showed fresh meat-like qualities, and the quality characteristics were clearly distinct from those of category I. Although the current standard for rapid freezing rate is 0.5 cm/h, this study suggested that 0.96 cm/h is the lowest freezing rate for achieving meat quality distinguishable from that achieved with conventional freezing, and further increasing the freezing rate did not provide advantages from an energy consumption perspective.

• Reproductive and Developmental Biology
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• 제32권2호
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• pp.111-115
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• 2008

The aim of this study was to examine more effective cryoprotectant for the cryopreservation of mouse preantral follicles. Enzymetically isolated preantral follicles from 12-day-old mice were cryopreserved by a slow freezing protocol with 1.5 M propanediol (PROH), dimethyl sulphoxide (DMSO) or glycerol (GLY) and then grown and matured in vitro for 11 days after thawing. The survival of preantral follicles immediately after freezing and thawing was not different among the PROH (68.2%), DMSO (72.4%) and GLY (72.1%). After grown and matured in vitro, the rates of survival and metaphase II oocytes were 54.9% and 36.6% for PROH which was significantly higher rates (p<0.05) compared with the rates obtained from DMSO (16.9% and 9.0%) and GLY (16.3% and 7.5%). The diameter of metaphase II oocytes from pre antral follicles frozen in PROH ($67.4{\pm}1.8\;{\mu}m$) was significantly (p<0.05) smaller than that of the fresh preantral follicles ($69.1{\pm}2.3\;{\mu}m$). The results from the present study revealed that PROH is more suitable cryoprotectant for the cryopreservation of mouse preantral follicles.

• Journal of Animal Science and Technology
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• 제48권6호
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• pp.797-804
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• 2006

본 연구에서는 동결 보호제 EG l에 sucrose 첨가 농도에 따른 생존성의 실험의 결과를 요약하면 다음과 같다. 1.5 M EG와 1.8 M EG 만을 이용하여 동결융해 후 생존서의 조사한 결과 71.1%와 70.2%로 각각 나타났다. 총세포수에 있어서도 127±1.3개와 124±1.6개로 생존율과 총세포수에 있어서도 두 그룹간에는 유의적인 차이를 보이지 않았다. 1.5 M EG와 1.8 M EG에 0.1 M sucrose를 각각 첨가한 후 동결 보존하여 융해 하였을 때 생존율과 총세포수 조사 결과는 1.5 M EG에 0.1 M sucrose 처리구가 73.6% 그리고 1.8 M EG 에 0.1 M sucrose 첨가군은 76.9%의 결과를 보였으며 총세포수 에 있어서도 118±1.2 와 112±1.2 개의 결과를 보여 생존성과 총세포수에 있어서도 두 처리군 모두 유의적인 차이를 나타내지 않았으나 1.5 M EG 처리구에서 총세포수는 다소 높은 경향을 보였다. 1.5 M EG 와 1.8 M EG에 0.3 M sucrose를 첨가하여 각각 생존성과 총세포수 조사 결과는 70.8%와 88.7%의 생존율을 나타내어 1.8 M EG 에 0.3 M sucrose 처리구가 유의적으로(P<0.05) 높은 결과를 보였다. 따라서 소 체외수정란을 conventional slow-freezing 방법으로 동결 보존할 경우는 1.8 M EG 동결보호제에 0.3 M sucrose를 병행하여 사용하는 방법이 수정란을 최상의 상태로 유지할 수가 있어 수정란이식에 적용할 경우 효과적일 것으로 사료된다.

• 대한수의학회지
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• 제33권1호
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• pp.171-178
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• 1993

The present study was carried out to investigate the developmental potency to blastocyst after freezing and thawing of nuclear transplanted 2-cell embryos. The nuclei from 2-, 4- and 8-cell mouse embryos were transferred into enucleated 2-cell embryos, and the reconstituted embryos were submitted to direct current(DC) pulse at output voltage of 2.0 kV/cm for $100{\mu}$ sec to induce cell fusion. The recovery rate and developmental potency to blastocyst after freezing and thawing of nuclear transplanted 2-cell embryos was investigated. 1. The recovery rate of nuclear transplanted 2-cell embryos in normal morphology after freezing and thawing was significantly higher in rapid freezing(DMSO 4.5M) than in slow cooling(p<0.01). 2. When the recovered embryos in normal morphology were cultured in vitro, there were no significant differences in the developmental potency to blastocyst between the freezing methods and the concentrations of cryoprotectant. In summary, these experiments have proved that rapid freezing method(DMSO 4.5M) is effective in nuclear transplanted 2-cell mouse embryos. If improved micromanipulation techniques and freezing are combined, nuclear transplantation technique will contribute to the improvement of productivity in livestock animals.

• 대한생식의학회:학술대회논문집
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• 대한불임학회 2005년도 제48차 춘계 학술대회
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• pp.126-126
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• 2005

• 한국가축번식학회지
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• 제23권1호
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• pp.75-84
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• 1999

본 연구는 토끼 전핵배의 효율적인 생산을 위한 동결방법과 조건 등을 찾고자 유리화 동결 및 완만동결법으로 동결ㆍ융해 후 체외배양하여 생존율 및 발달률을 조사하였다. 과배란시킨 토끼의 난관으로부터 채란된 전핵배를 동결에 공시하였다. 유리화 동결은 동결보호제로 EFS 와 EPG-I을 완만동결시는 EPG-II를 사용하였다. 동결ㆍ융해 후 5%, 39$^{\circ}C$ $CO_2$incubator 에서 소 난관상피세포와 공배양하였다. 본 실험의 결과는 다음과 같다. 동결보존을 위하여 동결보호제에 적절한 평형시간과 독성 여부를 판단하기 위하여 전핵배를 EFS 용액에 0~5분간 평형시킨 후 부화배반포로의 발달률은 1 분 군에서 72.0%로 동결보호제에 노출시키지 않은 대조구 (84.1%)에 비하여 무해한 결과를 얻었으나, 그 이상에서는 유해한 결과를 보여 주었다. EFS 노출 후 희석제로 sucrose와 D-PBS를, sucrose 사용 없이 D-PBS 만으로 희석하였을 때 유의적인 (P＜0.05) 차이를 보이지 않았다. 동결보호제에 있어서는 독성검사 및 동결ㆍ융해 후 발달률을 보아 EFS, EPG-. EPG-II는 동결보존에 있어서 동결보호제로서의 가능성을 보여주었으며 서로간의 유의적인 (P＜0.05) 차이는 찾아볼 수 없었다. 유리화동결에 의한 전핵배의 부화배반포로 발달률은 6.1%를 나타내었고, 완만동결에 의한 부화배반포 발달률은 11.5%로서 동결방법간에는 유의적인 (P＜0.05) 차이가 없었다. 완만동결시 동결속도가 전핵배의 투명대 파열에 미치는 영향을 규명하기 위하여 동결속도 및 침지온도를 달리하여 조사하였을 때 －35$^{\circ}C$ (25%) 보다는 －85$^{\circ}C$ 0.9%) 에서 액체질소에 침지하였을 때가 투명대 파열률에 있어 유의적인 (P＜0.05) 차이를 보였다. 이상의 결과로부터 전핵배는 현 배양상태에서 유리화동결 및 완만동결에 의하여 동결보존이 가능하다고 사료되나 전핵배의 배반포로의 발달률은 다소 저조하였다. 유리화동결 및 완만동결에 의한 전핵배는 후기 단계의 수정란보다 물리적, 화학적 손상에 더욱 민감하여 생존율 및 발달률에 영향을 미친다고 사료된다.

• Journal of Plant Biotechnology
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• 제7권3호
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• pp.183-186
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• 2005

Cryopreservation has been recognized as a practical and efficient tool for the long-term storage of vegetatively propagated plants. This study was conducted to investigate the effects of slow-freezing techniques on the cryopreservation of potato. In vitro plantlets of the potato genotypes of 'Atlantic', 'Superior’, 'Namseo', 'J138', and 'CTO5-5' were cold acclimated, and the excised axillary buds were precultured, osmoprotected, exposed to plant vitrification solution, frozen slowly to $-40^{\circ}C$ and then rapidly plunged into liquid nitrogen, thawed and finally plated on the regeneration medium. It was found that the higher the sucrose concentrations in the subculture medium of donor plantlets, the higher the survival rates of shoot tips after cryopreservation, and the highest survival (20%) was observed in the medium added with 0.25 M sucrose. As for the effect of cooling, $0.3^{\circ}C/min$ cooling speed showed the highest survival (25%). Different varieties showed different responses over different cryopreservation treatments. Survival rate was increased by slow-freezing technique method as compared with that of the basic cryopreservation method of vitrification alone in the diverse potato genotypes. Leaf and tuber morphologies of potatoes regenerated after cryopreservation using slow freezing technique were similar to those derived from the in vitro stock plantlets.

• 한국식생활문화학회지
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• 제18권2호
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• pp.105-110
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• 2003

김치를 $10^{\circ}C$에서 8일간 숙성시켜 산도 $0.6{\sim}0.7%$로 숙성된 김치를 $-70^{\circ}C$와 $-20^{\circ}C$로 냉동하여 $-20^{\circ}C$에서 저장하면서 배추 조직의 elasticity, hardness, 세포 조직의 변화, 드립양을 실험한 결과는 다음과 같다. $-70^{\circ}C$에서 급속 동결한 것과 $-20^{\circ}C$에서 완만동결한 냉동 김치중 배추조직의 elasticity는 냉동저장 15일까지 감소하다 일정하게 유지되었고 hardness는 거의 변화가 없었으며 냉동 방법에 따른 변화도 거의 나타나지 않았다. 투과 전자현미경으로 관찰한 결과 control의 경우 세포벽이 매우 두꺼우며 세포의 모양들이 잘 보존되어 있는 것을 볼 수 있었고 $-20^{\circ}C$로 냉동 처리하여 해동시킨 세포벽들은 많이 손상되어 있음을 볼 수 있었으며 $-70^{\circ}C$로 급속 냉동 시료의 경우 세포벽의 손상 정도가 $-20^{\circ}C$로 냉동 처리한 시료보다 덜 파괴되어 있음을 볼 수 있었다. 한편 냉동 저장기간 동안 드립의 손실량의 변화는 $-70^{\circ}C$로 냉동 처리한 시료의 드립양은 $3{\sim}4%$정도로 $-20^{\circ}C$로 냉동 처리한 시료의 $5{\sim}6%$에 비해 적은 것을 알 수 있었다.