1,721,009 research outputs found
Vitrification of mammalian spermatozoa in the absence of cryoprotectants: from past practical difficulties to present success
No full textUltra-rapid freezing and storage of rat embryos in an electric refrigerator at -130 degree C without liquid cryo-agents, with ultra-short exposure in the freezing medium and direct rehydration after thawing
No full textAn ultra-rapid freezing technique for embryos has been developed. This procedure involves: ultra-short exposure to cryoprotectants; freezing of embryos in a metallic powder pre-cooled in an electric refrigerator at -130 degree C; storage of embryos ina refrigerator at -130 degree C; direct rehydration after thawing
Congenitally caused fused labia in the common marmoset (Callithrix jacchus)
No full textIn this paper, the occurrence of an external genital abnormality in female marmoset monkeys (fused labia) is discussed. This malformation was detected, for the first time, in a group of animals at the German Primate Center (GPC), Goettingen. The malformed vulva was completely sealed except for an opening of 1.5-2.5 mm around the urethra sufficient for urination. Because of this defect the animals were not able to copulate. As a consequence, the affected females were functionally infertile although they had a normal genital tract and a regular cycle. This vulvar abnormality was found in 12 females, offspring of 10 pairs in which either one or both came to the German Primate Center from two genetically related colonies in Munich, Germany, and one colony in Basel, Switzerland. The abnormality appeared to be recessive and inheritable from either parent. In pairs in which both animals were from one of the mentioned colonies, 45% of the female offspring were affected. In pairs where only one partner came from these colonies, 26% of female offspring had the malformation. These results indicate that avoidance of inbreeding, which is frequently performed in primate colonies, may reduce, but not eliminate the expression of abnormalities of genetic origin. Therefore selective breeding is required, and, in colonies where these recessive mutations are widespread, the development of genetic screening tests would be advantageous
DNA integrity and motility of human spermatozoa after standard slow freezing versus cryoprotectant-free vitrification
No full textAbstract
BACKGROUND:
In contrast to the technique of conventional freezing, the vitrification of spermatozoa requires high cooling rates (720 000 degrees K/min), which could be damaging for spermatozoa. The aim of our study was to compare slowly frozen and vitrified spermatozoa in terms of their post-thaw DNA integrity and motility.
METHODS:
Semen samples were prepared according to the routine swim-up technique and divided into aliquots for comparison of fresh, conventionally frozen and vitrified spermatozoa from the same ejaculate in the presence or absence of cryoprotectants. Spermatozoa motility and DNA integrity were determined.
RESULTS:
The motility of spermatozoa conventionally (slowly) frozen with a cryoprotectant was similar to that recorded for spermatozoa vitrified in the absence of cryoprotectant (47 versus 52%). The DNA integrity was unaffected by the cryopreservation method or presence of cryoprotectants.
CONCLUSION:
The vitrification of human spermatozoa in the absence of conventional cryoprotectants is indeed feasible. The DNA integrity of vitrified sperm is comparable with that shown by standard slow-frozen/thawed spermatozoa, yet the method is quick and simple and does not require special cryobiological equipment
Comparison of different vitrification protocols on viability after transfer of ovine blastocysts in vitro produced and in vivo derived
No full textDevelopmental rate and ultrastructure of vitrified human pronuclear oocytes after step-wise versus direct rehydration
No full textAbstract
BACKGROUND:
This study compared the viability of human pronuclear oocytes subjected to vitrification followed by post-thaw step-wise removal of cryoprotectants versus direct rehydration, in terms of their subsequent in vitro survival and ultrastructural features.
METHODS:
A total of 115 three-pronuclei stage oocytes were cryopreserved in super-open-pulled straws by vitrification in 40% ethylene glycol + 0.75 mol/l sucrose for either 1 min or 10 s at 38 degrees C, followed by direct plunging into liquid nitrogen. After thawing, oocytes vitrified for 1 min (group 1) or 10 s (group 2) were expelled into a graded series of sucrose solutions (1.0, 0.75, 0.5, 0.25 and 0.12 mol/l) for removal of the cryoprotectant in five 2.5 min steps. A second batch of oocytes vitrified for either 1 min (group 3) or 10 s (group 4) were directly expelled into culture medium at 38 degrees C after thawing. Finally, the ultrastructural changes occurring in oocytes in each of the treatment groups were evaluated.
RESULTS:
Oocyte development (division to two-blastomere stage) rates after in vitro culture were 82, 83, 0 and 0% for groups 1, 2, 3 and 4, respectively. The harsh osmotic process involved in direct rehydration provoked ultrastructural changes, including the disruption of cytoplasmic and pronuclear membranes as well as intracellular organelles.
CONCLUSION:
The direct post-thaw rehydration of human pronuclear oocytes has lethal osmotic effects, such that protocols for vitrifying human pronuclear oocytes should include the step-wise removal of the cryoprotectant
Ultra-rapid freezing and storage of rat embryos in an electric refrigerator at-130 degrees C without liquid cryo-agents, with ultra-short exposure in the freezing medium and direct rehydration after thawing
No full textAn ultra-rapid freezing technique for embryos has been developed. This procedure involves: ultra-short exposure to cryoprotectants; freezing of embryos in a metallic powder pre-cooled in an electric refrigerator at -130 degrees C; storage of embryos in a refrigerator at -130 degrees C; direct rehydration after thawing
Vitrification of sheep blastocysts: the choice of protocol does appear to affect the pregnancy and lambing rate
No full textCryoprotectant-free cryopreservation of human spermatozoa by vitrification and freezing in vapor: effect on motility, DNA integrity, and fertilization ability
No full textAbstract
Human spermatozoa can be successfully cryopreserved avoiding the use of cryoprotectants through vitrification at very high cooling rates (up to 7.2 x 10(5) degrees C/min). This is achieved by directly plunging a copper cryoloop loaded with a sperm suspension into liquid nitrogen. After storage, vitrified spermatozoa are instantly thawed by melting in an agitated, warm medium. The goal of the present study was to compare the quality of spermatozoa cryopreserved using this rapid vitrification method with that of spermatozoa cooled relatively slowly by preexposure of the loaded cryoloop to liquid nitrogen vapor (-160 degrees C) with speed in the range 150-250 degrees C/min) before immersion into liquid nitrogen. Both cooling modes led to comparable results in terms of the motility, fertilization ability, and DNA integrity of the warmed spermatozoa. In both cases, instant thawing by melting in a warm medium was essential for successful cryopreservation. Our findings suggest that optimal regimes for the cryoprotectant-free cryopreservation of spermatozoa need not be restricted to very fast cooling before storage in liquid nitrogen, a wide range of cooling rates being acceptable. Herein, we discuss the implications of this finding in the light of the physics of extra- and intracellular vitrification
ASEPTIC VITRIFICATION OF HUMAN GERMINAL VESICLE OOCYTES USING DIMETHYL SULFOXIDE AS A CRYOPROTECTANT
No full textOBJECTIVE: To evaluate the viability of vitrified human germinal vesicle (GV)-oocytes to mature to metaphase II (MII) stage after "rapid" cooling directly in liquid nitrogen in comparison with "slow" cooling in a closed 0.5-mL straw (aseptic system), with or without dimethyl sulfoxide (DMSO) in vitrification solution. The possibility of avoiding parthenogenesis of the oocytes after vitrification using DMSO was investigated.
DESIGN:
In vitro maturation after vitrification.
SETTING:
Assisted reproduction centers.
PATIENT(S):
Patients undergoing standard superovulation treatment and having GV-oocytes after follicular puncture.
INTERVENTION(S):
The GV-oocytes were vitrified with long/short exposure to DMSO using slow or rapid cooling, then warmed and matured in vitro.
MAIN OUTCOME MEASURE(S):
Maturation after warming.
RESULT(S):
Oocyte development up to MII stage after vitrification with DMSO was 71% in the group with "rapid" cooling, and in groups with "slow" cooling, 68% and 72% for long and short exposure to DMSO, respectively. The maturation rate of GV-oocytes after slow cooling without DMSO was 51%. In the vitrification with long-term contact of oocytes with DMSO group, a high rate of parthenogenesis was observed. When vitrification with short-term contact of oocytes with DMSO at room temperature was used, no parthenogenesis was observed.
CONCLUSION(S):
Cryopreservation of human GV-oocytes in open-pulled straws OPS) using an aseptic slow cooling method gives high maturation rates but only in combination with DMSO. To avoid spontaneous parthenogenesis, the exposure to DMSO must occur for a reduced time and at room temperature
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