• 한국수정란이식학회지
• /
• 제29권1호
• /
• pp.51-57
• /
• 2014

본 연구는 돼지 정자의 동결 건조 시 동결 보호제인 trehalose의 효과와 동결 건조 시간과 동결 건조 후 저장 기간에 따라 정자의 생존성과 체외 성숙 난자 내 동결 건조 정자를 직접 주입한 후 전핵 형성율, 난할율 그리고 배발달 성적을 조사하였다. 동결 건조 후 정자의 생존율은 trehalose 무첨가구에 비해 trehalose를 첨가한 처리구에서 높은 생존율을 보였으며, 75 mM의 trehalose를 첨가하여 동결 건조한 정자들의 생존율이 가장 높은 것으로 나타났다. 또한, 동결 건조 후 저장 기간이 길어질수록 생존율이 낮아지는 경향이었다. 체외 성숙 난자 내 동결 건조 정자를 직접 주입 후 전핵 형성율은 trehalose 첨가구에서 유의적으로 높았으며(p<0.05), 난할율과 배발달 성적도 trehalose 첨가구에서 유의적으로 높았고(p<0.05), 정자의 동결 건조 시간이 짧을수록 높은 난할율과 배발달율을 보였다.

• 대한수의학회지
• /
• 제15권2호
• /
• pp.207-213
• /
• 1975

Skimmed goat milk heated at $92^{\circ}C$ for 10 minutes was used as a basal extender for bull semen. The extenders for liquid semen were prepared by adding simultaneously at various ratio of 5% dextrose solution and egg yolk to skimmed goat milk. After bull seven was diluted with the extenders at the rate of 20 million spermatozoa per ml of the extenders. The extenders were stored at $5^{\circ}C$ and the survival rates of spermatozoa were examined at 4 and 24 hours, and 3, 5 and 7 days after dilution. The extenders for frozen semen were prepared by adding various ratlo of glycerol to skimmed goat milk containing 20 parts of 5% dextrose solution and 3 parts of egg folk to 77 parts of skimmed goat milk. After bull semen was diluted with the extenders at the rate of 40 million spermatozoa per ml of the extenders, the extenders were frozen in liquid nitrogen tank. The frozen extenders were thawed at $40^{\circ}C$ for 2 minutes, and the revival rates of the spermatozoa in the extenders were examined. These thawed extenders were stored at $5^{\circ}C$ and the survival rates of the spermatozoa were examined at 10 minutes and 24 hours and 3 and 5 days after thawing. The results obtained were as follows: 1. Among the extenders stored at $5^{\circ}C$, the survival rate of the sperm was the highest in the extender including 20 parts of 5% dextrose solution and 3 parts of egg yolk to 77 parts of skimmed goat milk, and the survival rate was significantly higher that of the spermatozoa in egg folk-2.9% sodium citrate (1 : 4) extender. (P<0.05) 2.Among the extenders frozen in liquid nitrogen tank, the revival rate of the spermatozoa was the highest in the extender containing 7ml of glycerol per 100ml of the extender with consisted of 77 parts of skimmed goat milk, 20hparts of 5% dextrose solution and 3 parts of egg yolk, and the revival rate was significantly higher than that of the spermatozoa in egg yolk-2.9% sodium citrate (1 : 4) extender containing 8ml of glycerol per 100ml of the extender (p<0.01). 3. Among the extenders stored at $5^{\circ}C$ after thawing, the survival rate of the spermatozoa was the highest in the extender containing 7ml of glycerol per 100ml of extender which consisted of 77 parts of skimmed goat milk, 20 parts of 5% dextrose solution and 3 parts of egg yolk, and the survival rate was significantly higher than that of the spermatozoa in egg yolk -2.9% sodium citrate (1 : 4) extender containing 8ml of glycerol per 100ml of the extender (p<0.01).

• 한국가축번식학회지
• /
• 제14권3호
• /
• pp.199-204
• /
• 1990

Equine semen was analysed for its survival after storage under different conditions. Total 12 ejaculates from 2 Thoroughbred were analysed for general characterisitcs and preservation at low temperature. The sperm fraction, concentration, nd the rate of motile spermatozoa were 57.91ml per ejaculate, 2.18$\times$108/ml and 74.1%, respectively. The survival rate of spermatozoa was highest when diluted semen with E-Z Mixin was stored at 7~8$^{\circ}C$. The optimum survival rate(>54%) can be obtained upto 24h at 7~8$^{\circ}C$. However only 10% spermatozoa survived after 5h storage at 7~8$^{\circ}C$ without use of E-Z Mixin. Other ranges of temperature(15$^{\circ}C$ and room temperature) gave less survival rates(<25%). These results indicate that the extender could be used as a basic solution for the preservation of equine spermatozoa at low temperature. It also provides a practical method for short-term storage of collected equine semen in a simple manner.

• 한국수정란이식학회지
• /
• 제25권3호
• /
• pp.195-199
• /
• 2010

This studies were conducted to investigate the survival rate of frozen-thawed spermatozoa of Jindo Dog by monosaccharide and freezing rates. Experimental animals were prepared 12 males within 1~8 year's old and collected once in a couple of weeks by digital manuplation methods. Collected semen was diluted 1:1 with Tris-egg yolk extender and added 4, 6 or 8% of glycerol and none, 4 mM glucose or 4 mM fructose as cryoprotectant and was equilibrated for 2 hrs in $4^{\circ}C$. In monosaccharide groups, the freezing rate was 5 cm-5 min. above $LN_2$. The survival rates without monosaccharide were $50.7{\pm}19.0%$, $58.6{\pm}18.0%$, $40.0{\pm}10.0%$ in 4, 6 or 8% glycerol, respectively. In addition of glucose, the survival rates were $43.1{\pm}14.7%$, $38.1{\pm}16.5%$, $33.3{\pm}4.0%$ in 4, 6 or 8% glycerol, respectively and in fructose, were $47.9{\pm}21.1%$, $61.3{\pm}6.2%$, $34.3{\pm}12.6%$ in 4, 6 or 8% glycerol, respectively. There showed significantly different between glycerol groups and monosaccharides groups (p<0.05). The survival rates of freezing rate in 5 cm-5 min. group was $64.5{\pm}15.8%$, $51.9{\pm}27.6%$, $29.7{\pm}24.8%$ and in 10 cm-10 min. group was $62.5{\pm}20.3%$, $64.9{\pm}23.6%$, $34.5{\pm}27.4%$ in 4, 6 or 8% glycerol, respectively. There were significantly different between freezing rates (p<0.05). These results suggest that the addition of fructose with 6%-glycerol and slow freezing improve the survival of frozen-thawed sperm in Jindo Dog.

• Clinical and Experimental Reproductive Medicine
• /
• 제24권2호
• /
• pp.187-192
• /
• 1997

The survival rate and motility recovered after cryopreservation of testicular spermatozoa in testicular sperm extraction (TESE)-ICSI program is low. The purpose of this study was to assess the availability and efficiency of mouse empty zona pellucida in cryopreserving human TESE spermatozoa. Mouse empty zonae pellucidae were obtained by extraction of cytoplasm with or without cytochalasin B treatment. Motile sperm from proven-fertile donor and two azoospermic patients after TESE were individually inserted into empty zona pellucida and cryopreserved. Two to five days after cyropreservation, the frozen sperm were thawed and the rates of recovery and motility were observed. The ooplasmic extraction rates of control (N=80) and cytochalasin B treated oocytes (N=80) were 94.0% and 96.2%, respectively (p>0.05). The post-thaw recovery rates of spermatozoa and rates of motility recovery of ejaculate (N=70) and testicular (N=70) sperm were 97.1%, 97.1% and 95.7%, 94.3%, respectively (p>0.05). The results of this study showed that the mouse zone pellucida is useful for cryostorage of single testicular spermatozoa.

• 한국패류학회지
• /
• 제26권4호
• /
• pp.255-260
• /
• 2010

피조개, Scapharca broughtonii (Schrenck) 정자의 동결보존 위해 동결높이에 따른 예비동결 (0, 2, 4, 6 cm), 희석제 (150 mM NaCl, 300 mM glucose, Ringer's solution)와 동해방지제(glycerol, dimethylsulfoxide (DMSO), dimethylacetamide (DMA), methaol) 에 따른 효과를 정자의 운동성과 생존율로 비교하였다. 피조개 정자 동결보존에 있어 예비동결은 액체질소표면으로부터 2 cm가 가장 효과적이었으며, 희석제로는 Ringer's solution이 동해방지제로는 DMSO 10% 농도가 가장 적합하였다.

• Clinical and Experimental Reproductive Medicine
• /
• 제25권2호
• /
• pp.171-177
• /
• 1998

The combination of testicular sperm extraction (TESE) with ICSI can achieve normal fertilization and pregnancy rate and is effective method in obstructive and non-obstructive azoospermic patients. But, when pregnancy was not occurred, repeated testicular biopsies are not evitable. Therefore, in this study, we observed the survival rate of testicular spemratozoa and spermatozoa extracted from the seminiferous tubules after cryopreserved-thawed used for next IVF cycle with ICSI. In a total of 23 cases, obstructive azoospermia was 17 cases and non-obstructive azoospermia was 6 cases. In obstructive azoospermia, after thawing, motile spermatozua was observed in 13 cases (76.5%). The fertilization rate with 2PN was 67.6% and 5 pregnancies (29.4%) were achieved. In non-obstructive azoospermia, motile spermatozoa was observed in 2 case (33.3%) after thawing. The fertilization rates with 2PN was 53.7% and 3 pregnancies (50.0%) were achieved. A comparison of the results of motile spermatozoa after thawed testicular spermatozoa and spermatozoa extracted from the thawed seminiferous tubule section were 3 cases (60.0%) and 12 cases (66.6%), respectively. The fertilization and pregnancy rates of thawed testicular spermatozoa and spermatozoa extracted from the thawed seminiferous tubule section were 69.4% and 20.0%, 62.5% and 38.8%, respectively. Conclusively, thawed testicular spermatozoa and spermatozoa extracted from the thawed seminiferous tubule section can achieve normal fertilization and pregnancy and cryopreservation of testicular spermatozoa and seminiferous tubule may avoid repetition of testicular biopsies in azoospermic patients in whom the only source of spermatozoa is the testis.

• Reproductive and Developmental Biology
• /
• 제34권3호
• /
• pp.271-274
• /
• 2010

This study was conducted to investigate the survival rate of frozen-thawed spermatozoa in equine by glycerol concentration and freezing speed Two stallions (1 Thoroughbred-13 year old and 1 Arab-7 year old) bred in Korea Racing authority was examined for 1 times in a couple of weeks. Semen was collected by condom method standing heated mare and were centrifuged 650 g for 15 min. and isolated the seminal plasma. Thick fraction of semen was diluted EDTA-Lactose-egg yolk diluents to 1:1 and contained in 0.5 ml straw as $6{\sim}14{\times}10^7\;cells/ml$. Final concentrations of glycerol were 3, 5 and 7% in cryopreseved diluents and added 4 times for 2 hours equilibration. For the freezing, equilibrated straws were located 3 or 5 em above $LN_2$ gas for 5 or 10 min. Survival rates of pre-frozen sperm were $65.0{\pm}13.2%$, $68.3{\pm}10.4%$, $66.7{\pm}11.5%$ and post-frozen were $53.3{\pm}23.1%$, $45.0{\pm}15.0%$, $50.0{\pm}18.0%$ in 3, 5, 7% glycerol concentration, respectively. There was no difference between glycerol concentrations. Survival rates of frozen-thawed sperm on freezing speed were $36.7{\pm}10.4%$, $40.0{\pm}7.1%$, $30.0{\pm}13.2%$ at 3 cm-5 min and $33.3{\pm}11.5%$, $31.7{\pm}2.9%$, $21.7{\pm}10.4%$ at 3 cm-10 min in 3, 5, 7% glycerol concentration, respectively. Survival rates of frozen-thawed sperm on freezing speed were $43.3{\pm}15.3%$, $32.0{\pm}17.9%$, $22.3{\pm}15.7%$ at 5cm-5 min and were $47.5{\pm}15.0%$, $43.3{\pm}12.6%$, $48.3{\pm}15.3%$ at 5cm-10 min in 3, 5, 7% glycerol concentration, respectively. There were significantly different between groups (p<0.05). These results suggest that glycerol concentration did not affect cryopreservation of stallion semen within 3~7% but freezing speed affects. In our experiment, the best cryopreservation condition was at 5 cm above $LN_2$ gas for 10 min for pre-freezing and 7% of glycerol concentration. These results lead to commercial AI with frozen-thawed stallion semen.

• 한국어류학회지
• /
• 제23권1호
• /
• pp.75-79
• /
• 2011

양태, Platycephalus indicus 정자의 동결보존을 위해 적정 동해방지제와 동결높이에 따른 예비동결(0, 2, 4, 6cm), 그리고 해동온도에 따른 정자의 운동성과 생존율로 비교하였다. 적정 동해방지제로 dimethylsulphoxide(DMSO), glycerol, methanol에 대해 실험하였으며, DMSO와 glycerol에서 methanol에 비하여 높은 생존율과 운동성을 나타내었다. 또한, 동결보존에 있어 예비동결은 액체질소표면으로부터 2 cm 높이에서 높은 생존율과 운동성을 보였고, 해동온도로는 $20^{\circ}C$가 가장 적합하였다.

• 한국수정란이식학회지
• /
• 제16권2호
• /
• pp.133-138
• /
• 2001

개의 인공수정에 사용할 정자의 보존방법을 확립하기 위하여, 동결속도와 응해 온도를 설정하여 적절한 동결방법을 정립하고자 본 실험을 실시하여 다음과 같은 결과를 얻었다. 동결의 방법에 있어서는 액체질소의 표면으로부터 17 cm 높이에서 동결하는 -3$^{\circ}C$/min의 동결속도로 실시하여 37$^{\circ}C$에서 2분간 응해하는 방법이 가장 좋은 결과를 보였다. 생존성과 운동성에 있어서의 차이는 없지만 첨체의 intact한 비율은 약간 낮은 결과를 보였으며, 이의 보완을 위해, 액체질소 표면으로부터 10cm와 17cm의 높이를 세분화하여 동결속도를 설정한 후보다 나은 정액동결방범을 찾는다면 동결정액을 이용한 인공수정의 수태율은 더욱 향상될 수 있을 것으로 사료된다.