• Maxillofacial Plastic and Reconstructive Surgery
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• v.15 no.4
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• pp.303-316
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• 1993

The auricular cartilage grafts have been widely used in replacement of the temporomandibular joint disk. Cartilage grafts itself have a low metabolism and high survival rate after grafting. In processing the grafting materials, it was important to preserve the properties of chondrocyte proper. We used 15% glycerol and 10% DMSO (Dimethyl Sulfoxide) solutions for cartilage fixation before deep freezing. We have performed the allogenic auricular cartilage graft in the temporomandibular joint of 20 rabbits which 10 specimen was treated with 15% glycerol and the other 10 specimen was treated with 10% DMSO respectively and examined in 1, 2, 4, 6 and 8 weeks after operation histopathologically. The result were : 1. Inflammatory cell infiltration around the grafted material appeared more glycerol groups than DMSO groups at 1 week, but each group has no differences after 2 weeks. 2. Degenerative changes of grafted auricular chondrocytes were more deveolped in glycerol group than DMSO groups till 4 weeks, but there were no differences between two groups after 6 weeks. 3. Fibrous union between grafted fragment and mandibular condyle was prominent in DMSO group. 4. Vascular proliferation of the grafted auricualr cartilage was more developed in DMSO groups than glycerol group in early stage. 5. Amount of the additional growth of grafted auricular cartilage was more existed in DMSO groups than glycerol group. 6. General survival rate after grafting was more prominent in DMSO group. In summary, allogenic auricular cartilage grafts treated with 15% glycerol and 10% DMSO solution have supported to survivalbility as a cryopreservative agents, especially DMSO groups have little inflammatory cell infiltration in early stages and degenerative changes and additional growth are more prominent than glycerol groups.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.1
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• pp.75-81
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• 2004

Objective: The purpose of this study was to evaluate the efficacy and effect of various cryopreservation method on the survival and the cytoskeletal stability of metaphase II mouse oocyte. Methods: Mouse ovulated oocytes were collected and cryopreserved by a modified slow-freezing method with 1.5 M 1, 2-propanediol (PrOH)+0.1 M sucrose or by vitrification using cryo loop and EM grid with 40% ethylene glycol+0.6 M sucrose. Four hours after thawing, intact oocytes were fixed and stained with fluorescein isothiocyanate (FITC)-conjugated monoclonal anti-$\beta$-tubulin antibody to visualize spindle and propidium iodide (PI) to visualize chromosome. Spindle morphology was classified as follows: normal (barrel-shaped), slightly and absolute abnormal (multipolar or absent). Results: Survival rate of the frozen-thawed oocytes in vitrification group was significantly higher than that of slow-freezing group (62.7% vs. 24.4%, p<0.01). Vitrification with cryo loop showed significantly higher survival rate than that with EM grid (67.7% vs. 53.5%, p<0.05). On the other hand, proportion of normal spindle and chromosome configurations of the frozen-thawed oocytes between two vitrification group was not significantly different. Conclusion: For mouse ovulated oocytes, vitrification with cryo loop may be a preferable procedure compared to slow-freezing method. Further study should be needed to investigate developmental competency of frozen-thawed mouse oocytes.

• Development and Reproduction
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• v.15 no.4
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• pp.301-308
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• 2011

The tolerance evaluation for abalone Haliotis discus hannai embryos was performed using different concentrations of cryoprotective agents (CPAs): dimethyl sulfoxide, ethylene glycol and propylene glycol added to 0.2 M sucrose, respectively. 4-cell, trochophore and veliger were exposed in each CPA with different concentration for 10, 20 and 30 minutes of immersion time. Developmental rates were increased with decreased concentration of every CPA and decreased immersion time, and differed from types of CPA. Developmental rates of veliger in all the CPAs were higher than those of 4-cell and trochophore. The developmental rates and larval activity indices in ethylene glycol were comparatively higher than those in other CPAs and the effective CPA and its concentration for the cryopreservation of the abalone embryos was suggested as 2.0 M ethylene glycol with equilibration time of 30 minutes.

• Journal of Veterinary Clinics
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• v.28 no.4
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• pp.394-398
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• 2011

Mesenchymal stem cells (MSC) are pluripotent cells that can be found in umbilical cord blood from new borne babies as well as placenta, bone marrow, adipose tissue, amniotic fluid, muscle, et al. MSC are capable of renewing themselves without differentiation in long-term culture, also can be differentiated into various tissues under specific condition. Formulating a cryopreservation protocol for the MSC is required because these cells cannot survive for long periods under in vitro culture conditions and a new formulation of harmless cryoprotectant is needed for the direct injection of MSC into patients. The undifferentiated MSC were frozen with a vitrification solution of 40% ethylene glycol, 20% Ficoll-70 and 0.3M sucrose. The survival rate after thawing and their proliferation rate were examined and compared with slow rate cooling methods using dimethylsulfoxide (DMSO). The vitrification method showed high survival rate after thawing and proliferation capacity comparable to DMSO. It can be suggested that ultra-rapid cooling method by vitrification is reliable methods for long term preservation of MSC and the vitrification solution with ethylene glycol, Ficoll-70 and sucrose will be more beneficially used for direct transplantation of MSC into patients than DMSO solution.

• Current Research on Agriculture and Life Sciences
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• v.3
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• pp.79-84
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• 1985

This research was attempted in order to develop a long-term storage method for sprouting foods such as potatoes, onions, garlic and chestnuts using Co-60 gamma irradiation combined with a natural low temperature. The sprouting of the irradiated groups, 150 Gy in potatoes, 50 Gy in onions and garlic, and 250 Gy in chestnuts was almost completely inhibited until 8 to 10 months of storage. The rotting rate of loss of weight influenced a little by irradiation with a sprout inhibiting dose, and the weight of loss of the optimum dose irradiated groups was reduced by about 6 to 24% as against that of the nonirradiated in the four stored foods. The chemical components relating to the quality of sprouting foods were better retained in the irradiated groups than in the nonirradiated until the latter period of storage. Therefore, it was shown that the long term storage of sprouting foods is possible using gamma irradiation of 50 to 150 Gy for potatoes, onions, and garlic and 250Gy for chestnuts followed by storage at a natural low temperature($10{\pm}5^{\circ}C$, R.H. 75-85%).

• Korean Journal of Fisheries and Aquatic Sciences
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• v.47 no.1
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• pp.39-44
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• 2014

This study aimed to find an optimal diluent and cryoprotective agent (CPA) during cryopreservation of common Korean bitterling Acheilognathus signifier sperm. The bitterling is an endangered species in Korea, and this study will enable conservation and further technical development of artificial seed production. We tested the effects of cryopreservation and toxicity by the type of diluent and CPA. The optimal combination of diluent and CPA for cryopreservation was 300 mM glucose+10% methanol, resulting in a survival rate and sperm activity index (SAI) of $96.3{\pm}1.5%$, and $3.0{\pm}0.0$ respectively, and no significant difference compared to fresh sperm. The survival rate and SAI of post-thawed sperm was $9.7{\pm}1.5%$, $0.8{\pm}0.3$ respectively, which was significantly higher than had been achieved with other diluents and CPAs.

• Clinical and Experimental Reproductive Medicine
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• v.20 no.2
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• pp.165-176
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• 1993

These studies were carried out to investigate the optimal freezing protocol for 2 cell mouse embryos and to find the probability of quality control with 2-cell embryos frozen. The embryos showed the best survival by the protocol composed of a freezing solution with the cryoprotectants(1.5M propanediol + 0.1M sucrose), and a 2-steop thawing method(room temperature, 20 sec-37$^{\circ}C$, 20 sec). The developmental ability of frozen-thaw 2-cell embryos did not differ from that of fresh 2-cell embryos in m-KRB medium with 0.4% bovine serum albumin. But development of frozen-thaw embryos was depended on the supplements of the medium. In the albumin-free medium, the developmental rate(rate of blastocysts) was significantly reduced, compared with that in the medium with 0.4% BSA. Also, when frozen-thaw embryos were cultured in the meduim with human fetal cord serum(HCS), the developmental rate of frozen-thaw embryos was sligtly reduced, compared with that of fresh 2-cell embryos. Finally, frozen-thaw 2-cell mouse embryos were more sensitive to the toxic agent of disposable-plastic syringe. Therefore, toxicity of medium could be effectively detected by frozen-thaw 2-cell mouse embryos.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.2
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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.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.32 no.2
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• pp.238-241
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• 1999

Experiments were performed to find out the physico-chemical properties of milt and the sperm motilities in various conditions using the grey mullet, Mugil cephalus. The average concentration of sperm in the milt was $1,11 \pm0.36\times10^{10}/ml$. Spermatocrit was 96.7$\pm$2.6. pH and osmolality of seminal fluid were 7.8$\pm$0.1, 370$\pm$6 mOsm/kg, respectively, Total protein concentration of sperm was higher than that of seminal fluid, but total lipid concentration of seminal fluid was higher than that of sperm. The sperm motility was high in the diluent of milt : artificial seawater (1:10, by volume) and in 822 mOsm/kg and 983 mOsm/kg similar to seawater osmolality, but it decreased after 20 minutes. But activity of sperm was highly maintained in 482 mOsm/kg which was a little higher than osmolality of seminal fluid, and was high in pH 7$\~$9.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.2
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• pp.251-256
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• 1999

The present study was conducted to evaluate whether vitrification could be used for ovarian tissue preservation. The important issue here is that the vitrification is very simple, easy, and economical compared to the conventional cryopreserving method that using automatic freezing instrument. Human ovarian cortical tissues were cryopreserved by vitrification with 5.5 M ethylene glycol and 1.0 M sucrose as cryoprotectant. Three points of temperature ($4^{\circ}C$, room temperature, and $37^{\circ}C$) and two points of duration (5 or 10 minutes) for cryoprotectant treatment were examined to determine the best condition for vitrification of the human ovarian cortical tissues. After thawing, viability of the isolated primordial follicles was examined by dye-exclusion method. Histological appearance of tissues before and after the cryopreservation was evaluated. There was no toxic effect of the 5.5 M ethylene glycol on the primordial follicles. However, when the tissues were treated with cryoprotectant at $37^{\circ}C$ for 10 minutes and exposed to liquid nitrogen, it seems likely that there is certain deleterious effects on the viability of the primordial follicles. The highest viability of the primordial follicles was obtained with the treatment of cryoprotectant at room temperature for 10 minutes. Follicles and oocytes survived after freezing and thawing had the similar normal shapes as was seen in the specimens before cryopreservation. It would be useful to apply vitrification in establishing ovarian tissue banking for clinical purposes.