Mouse Embryo Cryopreservation by Slow Freezing

2018 ◽  
Vol 2018 (5) ◽  
pp. pdb.prot094540 ◽  
Author(s):  
Robert Taft
Keyword(s):  
Mouse Embryo ◽  
Slow Freezing ◽  
Embryo Cryopreservation

Effect of genotype on the efficiency of mouse embryo cryopreservation by vitrification or slow freezing methods

1995 ◽  
Vol 40 (4) ◽  
pp. 429-435 ◽  
Author(s):  
A. Dinnyés ◽  
G. A. Wallace ◽  
W. F. Rall
Keyword(s):  
Mouse Embryo ◽  
Slow Freezing ◽  
Embryo Cryopreservation

132 SUCCESSFUL EMBRYO TRANSFER OF IN VIVO-PRODUCED RED DEER (CERVUS ELAPHUS) EMBRYOS AFTER CRYOPRESERVATION BY SLOW FREEZING OR VITRIFICATION

2007 ◽  
Vol 19 (1) ◽  
pp. 183
Author(s):  
J. P. Soler ◽  
G. G. Kaiser ◽  
N. Mucci ◽  
L. B. Ferre ◽  
R. H. Alberio
Keyword(s):  
Liquid Nitrogen ◽  
Embryo Transfer ◽  
Cervus Elaphus ◽  
Red Deer ◽  
Slow Freezing ◽  
Alternative Methods ◽  
Embryo Cryopreservation ◽  
Chi Square ◽  
Cryopreserved Embryos

Multiple ovulation and embryo transfer (MOET) programs for red deer (Cervus elaphus) have been established commercially over the last decade, with embryo cryopreservation being a related practice necessary to enhance the use of valuable genetic information. The aim of this work was to establish alternative methods for red deer embryo cryopreservation by using slow freezing with ethylene glycol (SF–EG) and vitrification by open pulled straw (OPS) methods. After surgical flushing of 18 superstimulated donors, 54 transferable embryos were recovered; 28 were transferred fresh to synchronized recipients and the others were cryopreserved by SF–EG (n = 11) or OPS (n = 15), respectively thawed or warmed, and transferred to recipients. Fresh embryos were maintained in Dulbecco's PBS + 20% cow serum (holding medium, HM) until transfer (maximum 3 h after collection). SF–EG cryopreserved embryos were suspended in HM + 1.78 M EG + 0.1 M sucrose + 4 mg mL−1 BSA. After a 10-min equilibration, embryos were loaded individually into 0.25-mL plastic straws and placed into a −7°C methanol bath chamber. After seeding (5 min later), the straws were cooled from −7 to −35°C at a rate of 0.5°C min. Straws were plunged into and stored in liquid nitrogen. Thawing was performed by placing the straws in a 30°C water bath for 30 s; their contents were drained into HM until transfer. Embryos were vitrified using the OPS method with minor modifications. They were first incubated in HM + 1.78 M EG + 1.3 M DMSO for 3 min and then transferred for 25 s into a vitrification solution of HM + 3.56 M EG + 2.6 M DMSO + 0.5 M sucrose. Each embryo was loaded by touching a 1-µL drop with the straw, which was immediately submerged into and stored in liquid nitrogen. Warming was done by placing the narrow end of the straws into HM + 0.25 M sucrose for 5 min. Embryos were then transferred into HM + 0.15 M sucrose for 5 min and finally to HM until transfer. Both types of cryopreserved embryos were transferred a few hours after collection, immediately after thawing or warming. Before embryo transfer, the presence of corpus luteum (CL) of recipients was confirmed by laparoscopic examination. Each embryo was surgically transferred into the apical extreme of the uterine horn ipsilateral to the CL of one recipient. Pregnancy was determined by ultrasonography 41 days after embryo transfer. The pregnancy rate between groups was compared with the chi-square test (P < 0.05). No statistical differences were found between groups (Table 1). Our results show that both vitrification and slow freezing methods with EG are suitable to cryopreserve red deer embryos. Table 1. Pregnancy rates in recipient hinds after transfer of fresh, vitrified, or frozen red deer embryos

scholarly journals Perinatal outcomes after ART with transfer of cryopreserved embryos

2013 ◽  
Vol 62 (1) ◽  
pp. 48-54
Author(s):  
Yana Nikolayevna Kravchuk ◽  
Alla Stanislavovna Kalugina
Keyword(s):  
Perinatal Outcomes ◽  
Slow Freezing ◽  
Embryo Cryopreservation ◽  
Number Of Children ◽  
Freezing Method ◽  
Article Analysis ◽  
Cryopreservation Method ◽  
Cryopreserved Embryos

The number of children born after ART, which includes cryopreservation methods, is steadily rising worldwide. Data on perinatal outcomes after transfer of cryopreserved embryos is presented in the article. Analysis of the influence of fertilization method — IVF/ICSI, embryo cryopreservation method — slow freezing method and vitrification is composed.

scholarly journals Cryopreservation of in vitro-produced bovine embryos by vitrification: In pursuit of a simplified, standardized procedure that improves pregnancy rates to promote cattle industry use

2020 ◽  
Vol 36 (3) ◽  
pp. 251-270
Author(s):  
Van Do ◽  
Andrew Taylor-Robinson
Keyword(s):  
Slow Rate ◽  
Survival Rates ◽  
Slow Freezing ◽  
Embryo Cryopreservation ◽  
Human Embryos ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Cattle Industry

The goal of cryopreservation is to retain the original stage of gametes and embryos after they have endured cooling and warming. Slow freezing is a standard method for in vivo-derived bovine embryo cryopreservation, threefifths of such embryos being frozen by this method globally. However, it is evident that slow freezing is not efficient for cryopreserving in vitro-produced bovine embryos. Hence, only one-third of in vitro-produced bovine embryos are cryopreserved. Vitrification is a preferred method for storage of human embryos; consequently, it has been explored as a novel means to store in vitro-produced bovine embryos, for which it shows considerable promise as an alternative to slow freezing. This is due to several reasons: vitrification is often less time-consuming than slow freezing; it does not need expensive slow rate freezing machines; and it has been proven to have comparatively higher survival rates. Yet, in the cattle industry vitrification continues to present shortcomings, such as possible toxicity of vitrification solutions and failure to standardize methods, which pose a challenge for its application to in vitro-produced bovine embryos. Therefore, determining the most suitable procedure is crucial to make vitrification more practical in commercial settings.

Comparison of the vitrification and slow freezing protocol for day 3 embryo cryopreservation

2011 ◽  
Vol 96 (3) ◽  
pp. S213
Author(s):  
J. Liu ◽  
G. Huang
Keyword(s):  
Slow Freezing ◽  
Embryo Cryopreservation ◽  
Day 3 Embryo

Embryo cryopreservation: is vitrification ready to replace slow freezing?

2008 ◽  
Vol 3 (4) ◽  
pp. 455-463
Author(s):  
Mohamed Youssry ◽  
Yasser Orief ◽  
Vassilis Palapelas ◽  
Safaa Al-Hasani
Keyword(s):  
Slow Freezing ◽  
Embryo Cryopreservation

scholarly journals Impact of the obtained embryos’ quality on the cryopreservation programs outcome in comparison slow freezing method with vitrification in IVF cycles

2012 ◽  
Vol 61 (4) ◽  
pp. 48-54
Author(s):  
Alla Stanislavovna Kalugina ◽  
Yana Nikolaevna Kravchuk ◽  
Svetlana Aleksandrovna Shlykova ◽  
Olga Vladimirovna Bystrova ◽  
Julia Kazimirovna Kamenetskaya ◽  
...  
Keyword(s):  
Controlled Ovarian Hyperstimulation ◽  
Gonadotropin Releasing Hormone ◽  
Clinical Pregnancy ◽  
Slow Freezing ◽  
Embryo Cryopreservation ◽  
Ovarian Hyperstimulation ◽  
Embryo Survival ◽  
Freezing Method ◽  
The Impact

The objective of this study was to investigate the impact of protocols with agonists and antagonists of gonadotropin-releasing hormone (GnRH) on the quality of obtained embryos and also to compare the efficacy of different methods of embryo cryopreservation — vitrification and slow freezing method. The results revealed, that vitrification method provides better embryo survival rate, higher clinical pregnancy rate. No significant differences between both protocols of controlled ovarian hyperstimulation (COH) were found in the number of embryos of top and good quality

70 ONE-STEP CRYOPROTECTANT DILUTION FOLLOWING VITRIFICATION OF IN VITRO-PRODUCED BOVINE EMBRYOS

2013 ◽  
Vol 25 (1) ◽  
pp. 182
Author(s):  
R. Morató ◽  
T. Mogas
Keyword(s):  
Embryo Transfer ◽  
Statistical Significance ◽  
Survival Rates ◽  
Blastocyst Stage ◽  
Slow Freezing ◽  
Embryo Cryopreservation ◽  
Dilution Method ◽  
Bovine Embryo ◽  
One Step

Although slow freezing continues to be the most widely used technique of cryopreservation for bovine in vivo- and in vitro-produced embryos, vitrification has been tested in different species with good results, especially when dealing with in vitro-produced embryos. Vitrification represents a minor expense in time and equipment associated with cryopreservation compared with conventional slow freezing. However, vitrification, which is the most common method for human embryo cryopreservation, has not been widely adopted by embryo-transfer practitioners for commercial use in cattle. In general, vitrification requires gradual cryoprotectant dilution in a laboratory setting, and it is difficult to perform in the field. The objective of this study was to develop a one-step dilution method suitable for one-step bovine embryo transfer using the cryotop vitrification method. Embryos produced in vitro by standard procedures were vitrified at the blastocyst stage at Day 7 post-insemination in a mixture of 15% ethylene glycol + 15% dimethyl sulfoxide + 0.5 M sucrose using cryotop devices. Embryos were randomly assigned to 1 of 3 warming methods: (1) W3: warming was carried out following the cryotop method (1 M sucrose for 1 min, 0.5 M sucrose for 3 min, and 0 M sucrose for 6 min); (2) W1/0.5: embryos were warmed directly in 0.5 M sucrose for 3 min; and (3) W1/0: embryos were warmed directly in 0 M sucrose for 5 min. Survival rates were assessed in terms of blastocyst re-expansion, hatching, and hatched status at 3 and 24 h after warming. Data were analyzed using the statistical analysis systems package (SAS, v9.1). Data from at least 3 replicates were collected. Comparisons of vitrified–warmed blastocyst survival rates between groups were performed using the chi-squared test. The level of statistical significance was set at P < 0.05. When embryo survival was evaluated at 3 h postwarming, embryos warmed using the 3-step dilution protocol and those warmed directly in 0.5 M sucrose showed higher percentages of survival (W3: 89.8%, n = 98; W1/0.5: 87.5%, n = 64; P < 0.05) than those blastocysts that were warmed directly in 0 M sucrose (W1/0: 66.4%, n = 146). However, similar rates irrespective of the warming procedure were observed at 24 h postwarming (W3: 85.7%, W1/0.5: 88.2%, W1/0: 70.5%). Warmed in vitro-produced embryos exposed to W3 (47.6%) and W1/0.5 (35.6%) achieved higher percentages of embryos developing to the hatched blastocyst stage after 24 h of culture than those embryos warmed in W1/0 (20.4%; P < 0.05). Our results indicate that direct warming and dilution of cyotop-vitrified embryos in 0.5 M sucrose for 3 min may enable one-step bovine embryo transfer without requirement of a microscope or other laboratory equipment, simplifying the embryo-transfer procedure of vitrified embryos on farm at the same level of complexity as carrying out AI. Support came from Spanish MEC (RZ2010-00015-0-00; AGL2010-19069) and Generalitat de Catalunya (2009 SGR 621).

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