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1.
This study aimed to evaluate various concentrations of egg yolk (5, 10, or 20%) in combination with different concentrations of glycerol (3% or 6%) added to a Tris‐based extender on the post‐thaw characteristics of sperm obtained from Tayassu tajacu. For this purpose, semen from 10 sexually male mature collared peccaries was collected by electroejaculation and evaluated for sperm motility, vigour, viability, morphology and functional membrane integrity. The ejaculates were initially extended in Tris‐fructose plus egg yolk (5%, 10% or 20%). After cooling, the semen was added to Tris‐egg yolk plus glycerol (6% or 12%), resulting in a final concentration of 3% or 6% glycerol of the extender. Straws were frozen using liquid nitrogen and thawed in a water bath at 37°C for 30 s. The frozen–thawed semen was evaluated as reported for fresh semen. After thawing, a significant decrease was verified for sperm motility and vigour, for all the samples in comparison with fresh semen. However, no differences were evidenced among treatments for any sperm characteristics evaluated (p > 0.05), except for the combination between 10% egg yolk and 6% glycerol, which provided the worst preservation of functional membrane integrity (p < 0.05). The interactions between higher concentrations of egg yolk (20%) and glycerol (6%) and also between lower concentrations of the same substances (5% egg yolk and 3% glycerol) added to the Tris‐based extender negatively affected the preservation of the normal sperm morphology after thawing (p < 0.05). In conclusion, the use of Tris‐based extender added to 10% or 20% egg yolk plus 3% glycerol is recommended for effective sperm cryopreservation in collared peccaries.  相似文献   

2.
The present study aimed to compare cat sperm quality after thawing using two different temperatures (37 and 70°C) and to investigate the effects of post‐thaw dilution on the sperm quality and longevity of ejaculated cat spermatozoa. Six ejaculates of each of six male cats were collected using an electroejaculator (total 36 ejaculates). The semen was frozen in 0.25‐ml straws using a Tris egg yolk extender containing Equex STM paste. Four straws prepared from each ejaculate were thawed at four different occasions; (i) at 37°C for 15 s, (ii) at 37°C for 15 s and diluted 1 : 2 with Tris buffer (v/v), (iii) at 70°C for 6 s, (iv) at 70°C for 6 s and diluted 1 : 2 with Tris buffer (v/v). The percentages of motile spermatozoa, the scores of progressive motility, the percentages of spermatozoa with intact plasma membrane (using SYBR‐14/EthD‐1 stains) and intact acrosome (using fluorescein isothiocyanate conjugated peanut agglutinin/propidium iodide stains) were evaluated in fresh semen at 0, 2, 4 and 6 h after thawing. The thawing temperature had no effect on any sperm parameters throughout the incubation period (p > 0.05). The dilution after thawing improved sperm motility, progressive motility and acrosome integrity (p < 0.05). The thawing of cat spermatozoa and subsequently diluting with Tris buffer resulted in an immediate (at 0 h) overall (combined over temperature) percentage of motile sperm of 64.8 ± 10.7 (mean ± SD), a score of progressive motility of 4.0 ± 0.5, a percentage of spermatozoa with intact plasma membrane of 64.4 ± 12.1 and intact acrosome of 44.8 ± 20.2. In conclusion, frozen cat semen can be thawed either at 37 or 70°C and post‐thaw dilution is recommended to reduce the toxic effect of some ingredients in the extender during post‐thaw incubation.  相似文献   

3.
To evaluate and compare the efficacy of various extenders for the cryopreservation of epididymal cat spermatozoa, two experiments were planned. Bovine and equine commercial extenders in the experiment 1 and TRIS–egg yolk–based extenders in experiment 2 were separately studied since the number of sperm collected per cat is reduced. Epididymal sperm samples were packaged into 0.25‐ml straws and frozen. Vigour, motility, morphology, acrosome status, sperm viability and functional membrane integrity were assessed at collection, after cooling and after thawing, while DNA integrity was evaluated at 0‐ and 6‐h post‐thaw. Experiment 1 compared the effect of three non‐feline commercial extenders – based on TRIS–egg yolk (Triladyl), egg‐yolk‐free medium (AndroMed) and skimmed milk‐egg yolk (Gent) – on the quality of frozen‐thawed epididymal cat sperm. Values for sperm motility and functional membrane integrity in cooled sperm diluted in Triladyl were higher (p < 0.001) than those recorded for Andromed and Gent. Except sperm morphology, the other assessed characteristics showed significant higher values in frozen‐thawed sperm diluted in Triladyl than in Andromed and Gent extenders. Experiment 2 analysed the effects of three TRIS–egg yolk–based extenders, one non‐feline commercial (Triladyl) and the other two prepared using different monosaccharides (glucose and fructose), on freezing‐thawed sperm. Results showed that specifically prepared extenders for cryopreservation of feline spermatozoa performed better than the commercial extender Triladyl, although sperm quality during the freezing‐thawing process did not significantly differ associated with the type of monosaccharide (glucose vs fructose) added to the mentioned extenders. Although TRIS–egg yolk–based extenders prepared in experiment 2 improved sperm cryoprotection, Triladyl remains a good option for practitioners who, for ease of use and availability, prefer to work with commercial extenders.  相似文献   

4.
本试验皆在研究添加不同浓度大豆卵磷脂（SL）冷冻保存东佛里生奶绵羊精液的效果。我们在Tris基础稀释液中,添加18%蛋黄为对照组,添加0.5%、1%、1.5%、2%、2.5%SL设为试验组,检测冷冻精液解冻后的精子活率和顶体完整率。结果显示,添加0.5%、2.5% SL冷冻稀释液稀释的精液,解冻后精子活率和顶体完整率与其他组之间存在显著差异（P<0.05）;添加18%蛋黄和1%～2% SL冷冻稀释液稀释的精液,冷冻解冻后精子活率和顶体完整率之间无显著差异（P>0.05）;添加18%蛋黄和1.0%～1.5% SL冷冻稀释液稀释后的精液,进行人工授精后母羊的妊娠率与对照组无显著差异（P>0.05）。因此,大豆卵磷脂可以作为冷冻保护剂用于东佛里生奶绵羊精液的冷冻保存,其最佳添加浓度为1～2%（g／L）。  相似文献   

5.
【目的】 探究在冷冻稀释液中添加大豆卵磷脂代替10%卵黄对梅花鹿精液冷冻保存效果的影响,为梅花鹿人工授精体系的完善提供参考。【方法】 采用电刺激法采集梅花鹿精液,以精液冷冻稀释液中分别添加1%、2%、3%、4%和5%大豆卵磷脂代替10%卵黄作为试验组,添加20%卵黄作为对照组,分别进行各组精液冷冻保存。5 d后,进行精液解冻,检测解冻后各组精子的活力、质膜完整率、顶体完整率、线粒体活性、存活时间,筛选合适浓度的大豆卵磷脂。选取4~5岁健康雌性梅花鹿,肌肉注射300 IU孕马血清促性腺激素（PMSG）和0.4 mg氯前列醇钠进行同期发情处理,发情后第20 h用20%卵黄组与筛选出的大豆卵磷脂组冻精进行人工输精,输精后30 d使用B超检测仪检测妊娠情况,统计妊娠率。【结果】 与对照组相比,1%大豆卵磷脂组冻融后的精子活力、向前活动力、快速前进活力、活率、质膜完整率、顶体完整率及线粒体活性均显著提高（P<0.05）;随着稀释液中大豆卵磷脂浓度的增加,其冻融后精子活力、向前活动力、快速前进活力、活率、质膜完整率、顶体完整率以及线粒体活性呈下降趋势,精子存活时间也随浓度的增加而减少。1%大豆卵磷脂组冻融精子人工授精梅花鹿的妊娠率为61.11%,高于对照组、2%和3%大豆卵磷脂组,但差异均不显著（P>0.05）。【结论】 在梅花鹿精子冷冻稀释液中添加1%大豆卵磷脂替代10%卵黄,能有效提高梅花鹿冻融精子的质量,为进一步筛选新型梅花鹿精液冷冻稀释液提供理论基础。  相似文献   

6.
The objective was to assess the effect of cooling to different subzero temperatures around ice formation (?5°C) on dog sperm cryosurvival and plasma membrane fluidity. Semen was centrifuged, and sperm were resuspended in a Tris‐egg yolk medium (3% glycerol). Diluted sperm were cooled from 22 to 5°C, and then, a Tris‐egg yolk medium containing 7% glycerol was added (final concentration of 5% glycerol and 200 × 106 cells/ml). Sperm were packaged in 0.5‐ml plastic straws, and equilibration was done 16 hr at 5°C before freezing. I. Straws (n = 47) at 5°C were exposed to nitrogen vapours to determine the freezing point. II. Other straws (from different ejaculates) processed as mentioned, were further cooled to ?3, ?5 or ?7°C and immediately rewarmed in a water bath at 37°C. Motility, plasma membrane functionality and acrosome integrity were assessed. III. Other straws (from different ejaculates) processed as mentioned were further cooled to ?3 or ?5°C, frozen over nitrogen vapours and stored in liquid nitrogen for one month. Straws were thawed in a water bath at 38°C for 30 s. Motility, plasma membrane functionality, plasma membrane integrity, acrosome integrity, capacitation status and plasma membrane fluidity were assessed. Ice nucleation temperature was ?14.3 ± 2.05°C (mean ± SD); cooling to +5, ?3, ?5 and ?7°C, without freezing, produces no differences on sperm quality between target temperatures; cooling to +5, ?3, and ?5°C produced no differences on sperm survival and plasma membrane fluidity after freeze–thawing. In conclusion, cooling of dog spermatozoa to different subzero temperatures did not improve sperm cryosurvival and had no effect on plasma membrane fluidity after thawing.  相似文献   

7.
为了提高猪冷冻精液品质和精子抵抗低温打击的能力,本研究以5%、10%、15%、20%和25%等不同浓度的鸵鸟卵黄作为冷冻保护剂,以20%的鸡蛋卵黄和20%的鸽蛋卵黄为对照,将冷冻-解冻后的精子活率、质膜完整率和顶体完整率作为评价指标,分析鸵鸟卵黄对猪精子的抗冷冻保护作用。结果表明：稀释液中添加20%鸽蛋卵黄时,精子活率、顶体完整率和质膜完整性分别为52.11%、55.62%和54.94%,显著高于其他组（P〈0.05）。虽然稀释液中添加15%鸵鸟卵黄时,冷冻-解冻后精子活率、顶体完整率和质膜完整率显著高于5%、10%、20%和25%鸵鸟卵黄组,但仍然显著低于稀释液中添加20%鸽蛋卵黄处理组。本研究表明,鸵鸟卵黄在冷冻过程中对猪精子具有一定的保护作用,但相对于鸽子蛋和鸡蛋卵黄效果并不理想。  相似文献   

8.
During the cryopreservation process, the level of polyunsaturated fatty acids, especially docosahexaenoic acid (DHA), in the sperm plasma membrane decreases significantly because of lipid peroxidation, which may contribute to sperm loss quality (i.e. fertility) of frozen–thawed semen. The aim of this study was to investigate the effect of supplementation of DHA (fish oil) in freezing extender II on frozen–thawed semen quality. Semen from 20 boars of proven motility and morphology, were used in this study. Boar semen was split into four groups, in which the lactose–egg yolk (LEY) extender used to resuspend the centrifuged sperm pellet was supplemented with various levels of fish oil to reach DHA level of 1X (group I, control, no added fish oil), 6X (group II), 12X (group III) and 18X (group IV). Semen solutions were frozen by using a controlled rate freezer. After cryopreservation, frozen semen was thawed and evaluated for progressive motility, viability by using SYBR‐14/Ethidiumhomodimer‐1 (EthD‐1) staining and acrosome integrity by using FITC‐PNA/EthD‐1 staining. There was a significantly higher (p < 0.001) percentage of progressive motility, viability and acrosome integrity in DHA (fish oil) supplemented groups than control group. Generally, there seemed to be a dose‐dependent effect of DHA, with the highest percentage of progressive motility, viability and acrosome integrity in group‐III. In conclusion, supplementation of the LEY extender with DHA by adding fish oil was effective for freezing boar semen as it resulted in higher post‐thaw plasma membrane integrity and progressive motility.  相似文献   

9.
Egg yolk is usually included in extenders used for preservation of dog semen. Lecithin is an interesting animal‐protein free alternative to egg yolk for semen preservation. The aim of our study was to evaluate soya bean lecithin for cryopreservation of dog semen. Five ejaculate replicates were divided in three equal parts, centrifuged and each pellet diluted with one of the three Tris‐based extenders containing 20% egg yolk, 1% soya bean lecithin or 2% soya bean lecithin. Extended semen was loaded in 0.5‐ml straws, cooled and diluted a second time and frozen in liquid nitrogen vapours. Sperm motility parameters (CASA), acrosome integrity (FITC‐PNA/PI) and sperm membrane integrity (C‐FDA) were evaluated 5 min post‐thaw and after 2 and 4 h of incubation. Total motility was significantly better in the egg yolk extender than in any of the lecithin‐based extender and was better in the 1% lecithin extender than in the 2% lecithin extender. Sperm membrane integrity was significantly better in the egg yolk extender than in any of the lecithin‐based extenders but did not differ significantly between the 1% and 2% lecithin extenders. Acrosome integrity was significantly better in the egg yolk extender than in the 2% lecithin extender but did not differ between the egg yolk extender and the 1% lecithin extender or between the two lecithin extenders. In conclusion, egg yolk was superior to lecithin in our study. The extender with 1% lecithin preserved sperm motility better than the extender with 2% lecithin.  相似文献   

10.
The aim of this study was to investigate the influence of boar breed on the optimal concentration of gamma‐oryzanol on the qualities of cryopreserved boar semen. Semen was collected from 20 boars (10 Duroc, 5 Large white and 5 Landrace boars). The semen sample was divided into five groups (A–E) according to the concentration of gamma‐oryzanol in extender II, that is 0, 0.08, 0.16, 0.24 and 0.32 mm , respectively. The semen was cryopreserved by nitrogen vapour and storage in nitrogen tank (?196°C). After storage for a week, samples were thawed at 50°C for 12 s and evaluated for progressive motility, sperm viability and acrosome integrity. The results demonstrated that gamma‐oryzanol significantly improved progressive motility, viability and acrosome integrity of frozen–thawed boar semen. Considering the influence of breeds on the optimal concentration of gamma‐oryzanol, for Duroc boar, gamma‐oryzanol at 0.16 mm (group C) yielded the highest percentage of progressive motility, sperm viability and acrosome integrity. For Large white and Landrace boars, gamma‐oryzanol at 0.24 mm (group D) showed a significantly higher percentage of progressive motility, viability (not significant in Landrace) and acrosome integrity than other concentrations. In conclusion, the optimal concentration of gamma‐oryzanol needed for boar semen cryopreservation in lactose–egg yolk (LEY) freezing extender is not only depended on individual boar but also breed of boar, that is 0.16 mm for Duroc and 0.24 mm  for Large white and Landrace.  相似文献   

11.
This study investigates the effects of iodixanol supplementation in varied concentrations to Tris egg yolk (TEY) extender on the quality and fertilization ability of frozen–thawed sperm of Thai native bulls. Each ejaculate was divided into four different groups, as follows: sperm were treated with TEY extender (control group) and TEY extender supplemented with three different concentrations of iodixanol (1.25%, 2.50% and 5.00%). Semen straws were frozen in liquid nitrogen vapor. After thawing, sperm motility characteristics, viability, plasma membrane integrity and acrosome integrity were determined. Also, frozen–thawed spermatozoa from all groups were used for in vitro fertilization and artificial insemination (AI) in natural estrus Thai native cows. The results showed that the post‐thaw quality of the 2.50% iodixanol group was superior to the other iodixanol groups (P < 0.05). However, iodixanol had no beneficial effect on post‐thaw sperm in vitro fertilization ability and pregnancy rate after AI (P > 0.05). It can be concluded that the supplementation of 2.50% iodixanol extender significantly improves the progressive motility, viability, plasma membrane integrity and acrosome integrity of cryopreserved semen from Thai native bulls, but it has no beneficial effect on in vitro fertilization ability and pregnancy rate after AI.  相似文献   

12.
This study assessed the influence of three different anaesthetic protocols on semen quality obtained from the epididymis. Sixty male dogs undergoing to routine sterilization were assigned to three anaesthetic protocols: thiopental group (TG, n = 20), propofol group (PG, n = 20) and ketamine–dexmedetomidine group (KDG, n = 20). Immediately after orchidectomy, the cauda epididymides and vas deferent ducts were isolated and then a retrograde flushing was performed to collect spermatozoa. In experiment 1, after the initial evaluation of the semen (sperm concentration, sperm motility and the percentages of live spermatozoa, abnormal spermatozoa and acrosome membrane integrity), semen samples were diluted in Tris‐glucose‐egg yolk extender and chilled for 48 hr, and the sperm motility was assessed at 6, 24 and 48 hr. In experiment 2, semen samples were diluted in Tris‐glucose‐egg yolk extender and chilled for 24 hr, and then samples were frozen in two extenders with different glycerol concentrations, to reach a final concentration of 50–100 × 106 spermatozoa ml?1, 20% egg yolk, 0.5% Equex and 4% and 5% glycerol, respectively. Mean values of total sperm concentration, sperm viability and the percentages of intact acrosome and abnormal spermatozoa were not significantly different between experimental groups, and therefore, the anaesthetic protocols assessed did not affect sperm parameters mentioned above. However, our study confirmed a detrimental effect of the use of thiopental (TG) over the total sperm motility (p < 0.05) and progressive sperm motility (p < 0.05) of the fresh and chilled epididymal sperm samples. The anaesthetic protocols including the application of propofol or ketamine–dexmedetomidine can be used to recover sperm in domestic canids without significant changes in sperm quality compared when semen is collected routinely and these techniques could be applicable to endangered wild canids.  相似文献   

13.
The present study was undertaken to elucidate the effect of non‐luteal oviductal proteins on sperm characteristics in Murrah buffaloes. Oviducts from healthy buffaloes were collected immediately after slaughter and the oestrous cycle phase was determined as either luteal or non‐luteal based on ovarian morphology. Non‐luteal oviducts (n = 80) were flushed from the isthmic end of the oviduct with PBS, fluid was centrifuged at 10 000 g at 4°C for 20 min and then dialysed and clarified. The supernatant obtained was lyophilized to concentrate the protein and stored at ?20°C till use. Sixteen good quality ejaculates from four Murrah buffalo bulls were collected using an artificial vagina. After fresh semen analysis, each ejaculate was split into two parts and extended in Tris–citrate–egg yolk glycerol dilutor. Part I of the split ejaculate was treated with non‐luteal oviductal proteins at the dose rate of 1 mg/ml of diluted semen, while part II remained as control. The extended semen was equilibrated for 4 h at 5°C, filled in 0.5 ml French straws, exposed to LN2 vapour, plunged into LN2 and then stored at ?196°C. The equilibrated and frozen–thawed semen was evaluated for sperm motility, viability, acrosomal integrity, cervical mucus penetration test and hypo‐osmotic sperm swelling test (HOST). In frozen–thawed semen, the percentage of sperm motility, viability and acrosomal integrity was significantly (p < 0.05) higher in the treatment group compared to the control group. The incorporation of non‐luteal oviductal proteins in the extender increased the ability of sperm to penetrate cervical mucus both after equilibration and the freeze‐thaw process. Similarly, the proportion of sperm with intact plasma membrane, as revealed by HOST values, was also significantly (p < 0.05) higher in the treatment group (32.6%) than the control group (27%) in frozen–thawed semen. It was inferred that incorporation of non‐luteal whole oviductal fluid proteins improved the sperm quality in frozen–thawed semen in Murrah buffaloes.  相似文献   

14.
The magnitude of damage to buffalo spermatozoa during incubation with different levels of H2O2 was assessed. A total number of 24 ejaculates from four Murrah buffalo bulls were analysed in the study. Each ejaculate was split into two parts (part I and II). Part I was extended in Tris–egg yolk–citrate extender (20% egg yolk:7% glycerol), equilibrated (4 h at 5°C) and cryopreserved in 0.5‐ml French straws and stored in liquid nitrogen. The other part was utilized for fresh semen studies. The sperm in fresh, equilibrated and frozen–thawed semen was separated by centrifugation (1500 g ; 15 min) and were washed with sperm TALP. The sperm cells were re‐suspended in incubation TALP at the rate of 108 sperm cells per millilitre and incubated with 0, 10, 25, and 50 μm H2O2 per ml at 37°C. Sperm motility, viability and intact acrosome percentages were assessed at 15‐min intervals up to 60 min of incubation. Lipid peroxidation levels of sperm were assessed at 0 and 60 min of incubation. The results of the experiment revealed that sperm motility decreased drastically during incubation with H2O2. Among the different levels of H2O2, the 50‐μm H2O2‐incorporated group had significantly (p < 0.05) higher malonaldehyde (MDA) level than the other groups. In the 50‐μm H2O2‐incorporated group, the MDA levels in fresh, equilibrated and frozen–thawed semen after incubation for 60 min were 961.6 ± 12.7, 991.8 ± 10.3 and 1234.9 ± 9.6 nm per 109 spermatozoa respectively. An inverse relationship was observed between sperm motility, viability, intact acrosome percentages and concentration of H2O2 and duration of incubation. The decrease in sperm functions with duration of incubation and concentration of H2O2 was significantly (p < 0.05) higher in frozen–thawed than fresh and equilibrated spermatozoa.  相似文献   

15.
To improve the Boer goat semen quality during cryopreservation process, three experiments were carried out to investigate the effect of (i) different concentration of ascorbic acid supplementation (ii) rate of cooling with chilled semen characteristics and (iii) method of freezing on post‐thaw Boer goat sperm using Tris‐based extender. Ascorbic acid at 8.5 mg/ml improved the sperm parameters (motility, integrity of membrane and acrosome, morphology and viability), compared to control in cooled samples (p < 0.05). With regard to other concentrations and post‐thawed parameters, ascorbic acid at 2.5–8.5 mg/ml led to higher percentages of sperm motility and integrities of membrane and acrosome when compared to control (p < 0.05). Slow cooling rises to higher percentages of sperm motility, acrosome integrity and viability, in comparison with fast cooling, in terms of cooled and frozen samples (p < 0.05). Programmable freezing method produced the higher percentages of sperm motility, integrities of membrane and acrosome and viability when compared to the freezing method of polystyrene box during goat sperm freezing (p < 0.05). In conclusion, chilled and post‐thawed sperm quality of Boer goat was improved when a Tris‐based extender supplemented with ascorbic acid was used at stages of different cooling rates and freezing methods.  相似文献   

16.
Dairy bull sperm may be sex‐sorted, frozen and used to artificially inseminate heifers with acceptable fertility if the herd is well‐managed. One drawback to the technology is that donor bulls must be located within a short distance of the sorting facility in order to collect semen, which limits the number of bulls from which sorted sperm are available. A successful method used to overcome this limitation in sheep is sex‐sorting from frozen–thawed semen and refreezing for artificial insemination. This technique is attractive to the dairy industry, and therefore a series of three experiments was designed to investigate the optimal methods to prepare, sex‐sort and re‐freeze frozen–thawed bovine sperm. Sperm were prepared for sorting by density gradient separation in either PureSperm® or BoviPure?, followed by staining in one of three diluents (Androhep®, Bovine Sheath Fluid + 0.3% BSA or TALP buffer). Sperm were sorted and collected into Test yolk buffer, and frozen in an extender containing 0, 0.25, 0.375 or 0.5% Equex STM Paste. Frozen–thawed sperm were better orientated (p = 0.006) and had fewer damaged membranes (8.7 ± 0.6% vs 19.5 ± 2.4%; p = 0.003) after centrifugation in PureSperm® rather than BoviPure? gradients. Sperm orientation (p < 0.05) and motility (69.9 ± 3.0 vs 55.6 ± 4.0; p < 0.001) were highest after staining in Androhep® rather than in TALP buffer. Sperm were more motile (58.2 ± 4.7 vs 38.7 ± 3.5; p < 0.001) and had better acrosome integrity (74.3 ± 2.9 vs 66.8 ± 2.0; p < 0.001) after freezing in an extender containing 0.375% Equex STM Paste than in extender without Equex. Hence, a protocol has been developed to allow frozen–thawed bull sperm to be sex‐sorted with high resolution between the sexes, then re‐frozen and thawed with retention of motility and acrosome integrity.  相似文献   

17.
Fertility after insemination of cryopreserved boar semen is currently below that of fresh semen. In an attempt to improve the post-thaw motility and acrosome integrity of boar sperm, semen was frozen using an adapted Westendorf method in which the chicken egg yolk was replaced by either duck or quail egg yolk. The different composition of the yolk types, particularly the amount of cholesterol, fatty acids and phospholipids, were thought to potentially afford a greater level of protection to sperm against damage during freezing and thawing. Sperm frozen in medium containing chicken egg yolk displayed higher motility immediately after thawing, but there was no difference in the motility of sperm frozen with different types of egg yolk 3 or 6 h after thawing and maintenance at 37 degrees C. Sperm frozen in media containing chicken or duck egg yolk had a higher proportion of intact acrosomes immediately after thawing than sperm frozen in medium containing quail egg yolk, but 6 h after thawing and maintenance at 37 degrees C the sperm that had been frozen in medium containing chicken egg yolk had a higher proportion of intact acrosomes than the sperm frozen in media containing duck or quail egg yolk. Analysis of the composition of the different yolk types showed that the basic components of the yolks were similar, but the ratios of fatty acids and phospholipid classes differed. Duck egg yolk had more monounsaturated fatty acids (MUFA) than chicken egg yolk, which had more MUFA than quail egg yolk. Duck egg yolk contained more phosphotidylinositol (PI) than chicken or quail egg yolks and quail egg yolk contained more phosphotidylserine than either chicken or duck egg yolks. The differences in post-thaw motility and acrosome integrity of boar sperm when frozen in media containing the different types of egg yolk may be due to the variation in composition.  相似文献   

18.
Evaluation of glucose as a cryoprotectant for boar semen   总被引：2，自引：0，他引：2  
Fertility parameters of boar spermatozoa were evaluated in vitro, after freeze-thawing the semen in three different extenders containing permeable and non-permeable cryoprotectants: A (111.0 mM Tris, 31.4 mM citric acid, 185.0 mM glucose, 20 per cent egg yolk, 3 per cent glycerol and 100 iu/ml penicillin G); B (200 mM Tris; 70.8 mM citric acid, 55.5 mM glucose, 20 per cent egg yolk, three per cent glycerol and 100 iu/ml penicillin G); C (200 mM Tris, 70.8 mM citric acid, 55.5 mM fructose, 20 per cent egg yolk, 3 per cent glycerol and 100 iu/ml penicillin G). The freeze-thawing techniques were the same for each extender. Eight ejaculates from four boars were obtained; the sperm-rich fraction of each ejaculate was extended in each of the three media at a final concentration of 400 x 106 sperm/ml, loaded into 0.5 ml straws and frozen at a rate of 30 degrees C/minute to -196 degrees C. The straws were thawed at 60 degrees C for eight seconds. Sperm motility, acrosomal integrity and in vitro sperm penetration through the zona pellucida of gilt oocytes matured in vitro were evaluated. The motility of unfrozen spermatozoa was 93.1 per cent compared with 60.7 per cent, 48.2 per cent and 35 per cent for sperm frozen in extenders A, B and C respectively; these values were all significantly different (P<0.05). There was no significant decline in sperm motility after incubation for 30 minutes in extender A, but there were significant decreases in sperm motility after 30 minutes of incubation in B and C. The percentage acrosomal integrities were 97.2 per cent for the control and 45.5 per cent, 30.3 per cent and 16.8 per cent for the frozen-thawed spermatozoa in extenders A, B and C respectively. The results of the in vitro penetration assay were 80.7 per cent when using control spermatozoa, and 42.2 per cent, 18.4 per cent and 3.3 per cent when using frozen-thawed spermatozoa in extenders A, B and C respectively  相似文献   

19.
The addition of 0.5% (v/v) of Equex STM Paste (Nova Chemical Sales, Scituate Inc., MA, USA), whose active ingredient is sodium dodecyl sulphate (SDS), to a Tris–egg yolk extender was demonstrated to improve the longevity of frozen–thawed dog spermatozoa during in vitro incubation at 38°C. The aim of the first experiment was to compare the effects of two SDS‐containing compounds, Equex STM Paste and Equex Pasta (Minitüb, Tiefenbach, Germany), when added to a Tris–egg yolk based extender, on the post‐thaw longevity of dog spermatozoa, as well as on the intracellular Ca2+ concentration of spermatozoa, during post‐thaw incubation at 38°C. The post‐thaw sperm survival and longevity, as well as the quality of the sperm movement, were significantly better when using Equex STM Paste. Such prolonged sperm longevity, however, was associated to a higher intracellular Ca2+ concentration in a large subpopulation of the live spermatozoa. A second experiment was aimed to evaluate the effects of sperm dilution immediately post‐thaw with a Tris buffer containing glucose or fructose. The two Tris buffers were no different for any of the sperm parameters studied. The aim of a third experiment was to evaluate the sperm longevity, motility patterns and intracellular Ca2+ concentration of cryopreserved dog spermatozoa during post‐thaw incubation in capacitating conditions [canine capacitating medium (CCM) with or without 5 μg/ml of heparin]. Heparin had no significant effects on any of the sperm parameters evaluated. During the first 8 h of incubation, the majority of the live spermatozoa had a high intracellular Ca2+ content. However, after 8–10 h of incubation, it had significantly declined. The highest proportion of fast motile sperm, and the highest curvilinear velocity, average path velocity and amplitude of lateral head displacement for the total motile sperm were observed during the 2–4‐h incubation period. It was concluded that: (a) the addition of 0.5% (v/v) of Equex STM Paste to a Tris–egg yolk based extender significantly improved the post‐thaw longevity of dog spermatozoa, but the same concentration of Equex Pasta had no significant beneficial effects; (b) sperm dilution after thawing with a Tris buffer containing glucose or fructose made no difference in post‐thaw sperm longevity; (c) the addition of 5 μg/ml of heparin to CCM had no significant capacitating effects on frozen–thawed dog spermatozoa.  相似文献   

20.
To improve the results obtained with a reference cryopreservation extender (control extender: Triladyl® + 20% (v/v) egg yolk + 6.4% (v/v) glycerol) for freezing caprine semen, glutamine was added to 18 split ejaculates at concentrations of 0, 20, 40, 80 and 120 m m (experiment 1). In experiment 2, glutamine was added to 18 split ejaculates at concentrations of 20, 25, 30, 35 and 40 m m . In the third experiment, the egg yolk was replaced with the low-density lipoprotein (LDL) fraction of egg yolk. The quality of frozen then thawed spermatozoa in each extender was compared using computer-assisted semen analysis. In experiment 1, glutamine at concentrations of 20 m m and 40 m m significantly improved sperm motility compared with the control extender. However, at 120 m m , a significant decrease in motility and velocity was observed. In experiment 2, motility, curvilinear velocity and amplitude of lateral head displacement (ALH) were improved in glutamine at 25 m m compared with the control. In experiment 3, 8% LDL and 25 m m glutamine significantly improved sperm motility, straight line velocity and ALH. In the fourth experiment, the quality of the previously defined freezing extender (Triladyl® + 8% (v/v) LDL + 25 m m glutamine + 6.4% (v/v) glycerol) was tested by comparing acrosome, tail membrane, plasma membrane and DNA integrity in 18 split ejaculates of frozen then thawed spermatozoa with spermatozoa that had been frozen then thawed in the control extender, and with spermatozoa from fresh, unfrozen sperm. The percentage of spermatozoa with intact acrosomes and tail membranes was significantly higher with the newly defined extender than that observed with the control extender. There was no significant difference in the percentage of spermatozoa with intact DNA between the frozen and fresh semen.  相似文献   

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