Vitrification Of Human Embryos Research Articles

Effect of repeated vitrification of human embryos on pregnancy and neonatal outcomes

BackgroundRepeated cryopreservation of embryos should occasionally be considered when embryos were not suitable for transfer. The effect of re-cryopreservation on embryos remains contentious.MethodsThis retrospective cohort study aimed to evaluate the pregnancy and neonatal outcomes of twice vitrificated blastocyst derived from once vitrified embryos. Total 410 vitrified-warmed blastocyst transfer cycles were divided into two groups according to the times of embryo vitrification: (1) vitrified blastocysts derived from fresh blastocysts (control group, n = 337); (2) twice vitrified blastocysts derived from once vitrified embryos (n = 73). The primary outcome was live birth rate. Multivariable logistic or linear regression analysis model was performed to describe the association between embryo cryopreservation times and clinical outcomes.ResultsNo difference was observed in female age at retrieval and transfer, infertility period, body mass index (BMI), infertility type, endometrial thickness, and embryo transfer numbers between the two groups. The pregnancy outcomes of embryos in repeated cryopreservation group were comparable to those of embryos in control group, including implantation rate, clinical pregnancy rate, and live birth rate. In multivariate logistic regression analysis, the cryopreservation times did not affect the outcomes of biochemical pregnancy, clinical pregnancy, and live birth. Moreover, there was no difference in gestational age, birthweight and sex ratio of singleton newborns between groups. After correcting several possible confounding variables, no significant association was observed between cryopreservation times and neonatal birthweight.ConclusionIn conclusion, pregnancy and neonatal outcomes achieved with twice vitrified blastocyst transfer were comparable to those achieved with vitrified blastocyst transfer in control group.

Three live births after human embryo vitrification with the use of aluminum oxide as an intermediate cooling agent: a case report

To study the possibility of increasing the cooling rates of the vitrification procedure in a closed system with the use of aluminum oxide as an intermediate coolant. Case report. Six patients undergoing procedures for assisted reproduction. Comparative studies of cryopreservation of donor embryos with aluminum oxide as an intermediate cooling agent (experimental group) and without it (control group) have been performed. After thawing, the embryo morphology and its potential to develop to the blastocyst stage have been assessed. The methodology was then applied to clinical practice. Twenty embryos of 6 patients have been vitrified on day 4 after fertilization with the use of aluminum oxide as an intermediate coolant. Fourteen of them have been thawed. All have displayed normal morphology and 10 have formed blastocysts after 24 hours of culture. Four of the patients received embryo transfer with 2 embryos and the other 2 with single embryos. After preliminary comparative studies of embryos frozen with aluminum oxide and a control group, the results showed no statistically significant difference between their quality and potential to reach to blastocyst stage. That gave us ground to apply the methodology in clinical practice. After the embryo transfer, 3 clinical pregnancies with successful live births have been obtained. Our experience shows that preimplantation embryos can be cryopreserved aseptically, in closed systems, with the help of aluminum oxide as an intermediate coolant.

Specifics of vitrification of in vitro-produced cattle embyos at various development stages

Producing embryos in vitro is an important technology used to improve the genetic potential of cattle and perfect the programs of their breeding. Regardless of the way they are produced, all embryos that had not been used for transplantation to recipients must be conserved. Because of significantly increased interest in the problem of cryoconservation of embryos, both coming from scientists and businesses, there are emerging new commercial environments that allow the facilitation of cryoconservation and the increase in the embryo survival. Oocyte-cumulus complexes obtained from the ovaries of slaughtered clinically healthy cows matured in 22–24 h in in vitro conditions. The oocytes were co-cultured with spermatozoids in Fertilization medium, and the obtained zygotes were cultured in Culture medium with Sodium-Pyruvate for 4 or 7 days up to the stage of morula or blastocyste, respectively. For the vitrification of cow embryos, we used a commercial kit for the vitrification of human embryos, having compared the duration of equilibration. According to the results of the studies, we observed high efficiency of cryoconservation of cow embryos using the commercial kit for vitrification of human embryos. The results revealed the significant effect of equilibration on survival and further development of embryos. In addition, we described the dependence of development stage of cattle embryo on the duration of the contact of embryo with equilibration solution. Therefore, optimal time of contact of cattle embryos at the morula stage with equilibration solution was 12 minutes. On the 24th h after thawing, 46.7 ± 3.3% of the embryos were observed to undergo blastulation, and on 48th h, this parameter increased to 96.7 ± 3.3%, which corresponded to the parameters in the group of embryos that had not been subjected to cryoconservation. In the conditions of further cultivation, the percentage of blastocystes that hatched in the experimental group was no different from that of the control. At the same time, the highest efficiency of vitrification of blastocystes of cows was seen after the contact with the equilibration solution for 15 min, since the percentage of hatched blastocystes was the same as in the control group. Therefore, using the commercial kit for vitrification of human embryos is beneficial, for it promotes the parameters of cow embryos after vitrification/thawing that are similar to such of intact embryos (without freezing). The data we analyzed and presented in the paper could help to increase the efficiency of cryoconservation of cattle embryos for both scientific and commercial purposes.

P-140 A retrospective study on combination of vitrification/warming protocols from different manufacturers

Abstract Study question What is the embryo survival rate when combining freezing and warming media from different manufacturers. Summary answer Vitrification protocol with DMSO in freezing medium and 4-step warming at room temperature resulted in highest implantation and high embryo survival rates. What is known already Cryopreservation media containing 7,5% and 15% DMSO in a two-steps protocol are widely used for vitrification of human embryos. Studies with the data on safety, survival rates, cellular structures integrity and the warmed embryo's implantation potential confirm efficiency of DMSO as cryoprotectant. However, questions remain on whether DMSO have any negative impact, thus interest to DC. Likewise with the choice of cryoprotectant, more data is needed to determine which temperature corresponds to better survival rates. Study design, size, duration This retrospective cohort study covered the period from January 2019 till November 2021. A total of 834 consecutive cycles of frozen/warmed embryo transfers were included in the study. Inclusion criteria were: women's age below 40 years, cryopreservation on the day 5 of embryo culture. Exclusion criteria were: donor oocyte cycles, cycles with pre-implantation genetic testing for aneuploidies and cycles with severe male factor after surgical sperm retrieval. Participants/materials, setting, methods Following groups were compared using Rapid-i carrier device: 1 - Kitazato vitrification/warming protocol, 2 - Kitazato vitrification and Vitrolife's Omni warming, 3 - Vitrolife's Omni vitrification/warming, 4 - Vitrolife's Omni vitrification and Kitazato warming, 5 - Kitazato vitrification/Vitrolife's Omni warming media used in accordance with Kitazato protocol (ambient temperature, Reproplate dish). Efficiency was evaluated comparing embryo survival, implantation, clinical pregnancy and early pregnancy termination rates. Mann-Whitney and Pearson's Chi-square tests were used for statistical analysis. Main results and the role of chance The average patient age in assigned groups was: group 1: 32,5±3,6 years (n = 569); 2:32,2±3,5 years (n = 47); 3:30,4±3,5 years (n = 15); 4:32,3±3,5 years (n = 74); 5:32,7±3,4 years (n = 129). Average number of transferred embryos: 1,3±0,5; 1,2±0,4; 1,1±0,3; 1,3±0,4 and 1,2±0,4 per group, respectively. Embryo survival rates were: 97%, 96%, 100%, 97% and 100% in each group, respectively. Group 5 was significantly different from other groups (p < 0,05; group 3 had too small sample size for an adequate comparison). Implantation rates per group: 43%, 57%, 44%, 40% and 49%, respectively (p > 0,05). Clinical pregnancy rates were not different between groups. The early pregnancy termination rates were: 18,1%, 20,7%, 33,3%, 14,7% and 8,9% per group, respectively (p > 0,05). Therefore, group 2 resulted in a best clinical outcomes. Limitations, reasons for caution The present study is a retrospective cohort study and it has a limited number of patients in compared groups. Additionally, live births and health of newborn children was not taken into account. Wider implications of the findings Freezing medium from one manufacturer can be combined with warming medium from another manufacturer and result in a good clinical outcome. Furthermore, this study sheds the light that there is still a room for the improvement of existing cryopreservation media and protocols. Trial registration number not applicable

Metabolic Activity of Human Embryos after Thawing Differs in Atmosphere with Different Oxygen Concentrations.

The vitrification of human embryos is more and more frequently being utilized as a method of assisted reproduction. For this technique, gentle treatment of the embryos after thawing is crucial. In this study, the balance of amino acids released to/consumed from the cultivation media surrounding the warmed embryos was observed in the context of a cultivation environment, which was with the atmospheric oxygen concentration ≈20% or with a regulated oxygen level—hysiological (5%). It is the first time that total amino acid turnover in human embryos after their freezing at post compaction stages has been evaluated. During this study, progressive embryos (developed to blastocyst stage) and stagnant embryos (without developmental progression) were analyzed. It was observed that the embryos cultivated in conditions of physiological oxygen levels (5% oxygen) showed a significantly lower consumption of amino acids from the cultivation media. Progressively developing embryos also had significantly lower total amino acid turnovers (consumption and production of amino acids) when cultured in conditions with physiological oxygen levels. Based on these results it seems that a cultivation environment with a reduced oxygen concentration decreases the risk of degenerative changes in the embryos after thawing. Therefore, the cultivation of thawed embryos in an environment with physiological oxygen levels may preclude embryonal stagnation, and can support the further development of human embryos after their thawing.

Survival Rates and Clinical Effectiveness of Vitrified Human Blastocysts in the Practice of Embryological Laboratories

Objective – to carry out comparative analysis of embryo survival and pregnancy rate after the vitrification of human embryos at different stages of the preimplantation development. Material and methods . A total of 1311 embryos were analyzed. For vitrification of embryos, the Irvine Scientific media (USA) and the CryoTop open-type carriers (Japan) were used. The vitrification and the defrosting of embryos were carried out according to the manufacturer's recommendations. The statistical processing of the results was performed on a computer in the statistical computing environment R (R V. 3. 5. 3, RStudio V. 1. 1. 463), the primary data entry was performed using MS Excel spreadsheets. We used methods of descriptive statistics, tests for comparing proportions, including an accurate binomial test for small samples and a single-sample test of proportions with continuity correction for large samples. Results . The range of the embryo survival is quite wide and ranges from 75% to 100%. When conducting a detailed analysis of the dependence of the pregnancy rate on the stage of cryopreserved blastocysts, we found a direct link between the clinical indicators and the structure of the embryos of 5–6 days of the preimplantation development. The blastocysts with the highest degree of the internal cell mass and the trophoblast formation in combination with a high degree of the expansion (4) give the best results of the clinical pregnancy rate. Conclusions. When performing vitrification we found out that the stage of the blastocyst development influences their survival and pregnancy rates. The phenomenon of hatching in the embryos of 5–6-day preimplantation development should be regarded as a factor that reduces the success of vitrification.

Clinical performance and newborn data of a newly developed closed vitrification device, cryotop CL for human embryo vitrification

Clinical performance and newborn data of a newly developed closed vitrification device, cryotop CL for human embryo vitrification

Retrospective study of vitrification of human embryos at 4-cell stage in assisted reproduction

Retrospective study of vitrification of human embryos at 4-cell stage in assisted reproduction

Витрификация эмбрионов человека после манипуляций на zona pellucida

Витрификация эмбрионов человека после манипуляций на zona pellucida

March 2017: This Month in JoVE: Human Embryo Vitrification, Paper Microfluidic Devices, Virtual Hand/Face Illusions, and Quantification of Amyloid-Beta for Alzheimer's Diagnosis

Here's a look at what's coming up in the March 2017 issue of JoVE: The Journal of Visualized Experiments. As the start of spring brings new life, we begin the month investigating exciting new developments in embryo storage for in-vitro fertility treatment. In JoVE Developmental Biology, our authors present a modified technique for vitrification and preservation of human embryos. This aseptic technique uses all FDA-compliant materials including ionomeric resin embryo straws and sterile flexipettes, and allows fast, safe, and repeatable embryo storage independent of operator variation. In addition, with a greater than 95% embryo survival-rate the method contributes to excellent pregnancy outcomes. Paper cards...printers, it's all so 1990's, right? Well, our next video highlight might make you reconsider. In JoVE Bioengineering this month, our authors reveal 3-dimensional paper microfluidic devices for immunoassays. By patterning paper with hydrophobic barriers, the transfer of fluids can be controlled and directed. Stacking multiple layers of patterned paper creates sophisticated 3-D microfluidic networks that can be utilized in cheap and simple bioanalytical assays, such as the detection of Human Chorionic Gonadotriphin, or hCG - the pregnancy hormone. So don't throw out those old Valentine's Day cards yet - upcycle! In JoVE Medicine this month, our authors reveal an accurate procedure for quantifying the plaque-forming amyloid-beta protein in cerebral spinal fluid to provide diagnoses in suspected Alzheimer's cases. Current diagnosis is mostly based on mental state and cognitive testing, with some studies beginning to examine amyloid-beta levels, but this is hampered by variability and reproducibility issues. Here, our authors use a combination of solid-phase extraction and liquid chromatography tandem mass spectrometry to quantify amyloid-beta levels to a high degree of accuracy, quantifying the deposition of the protein in plaques in the brain and enhancing the power for accurate diagnoses. Our final video highlight this month comes from JoVE Behavior, and showcases a new technique to create human body illusions. Here, our authors demonstrate a computer-based virtual hand and face that move in synchrony with the operator's action, creating an illusion that the model is an extension of the participant's own body. This is a useful tool in the study of self-representation, and is an advance over traditional studies requiring demonstrators to mimic actions on participants in time with actions performed on a physical model body part. Ah, just one small step until I realize my dream of becoming a super-fast blue hedgehog. You've just had a sneak peek of the March 2017 issue of JoVE. Visit the website to see the full-length articles, plus many more, in JoVE: The Journal of Visualized Experiments.

Appendix D: Irvine Scientific® Vitrification System.

This chapter will describe the use of the Irvine Scientific® vitrification and warming solutions (Vit Kits®) along with a detailed protocol for the correct use of the CryoTip® vitrification device for human embryos and oocytes. Successful pregnancies have been reported after carrying out rapid vitrification methods of oocytes, cleavage stage embryos, and blastocysts using the CryoTip® (Popwell et al. Fertil Steril, 101:e20, 2014; Kuwayama et al. Reprod Biomed 11:608-614, 2005; Kuwayama et al. Fertil Steril 84:S187, 2005; Kuwayama et al. Vitrification of human embryos using the CryoTip TM method. Reprod Biomed 11:608-614). Compared to other vitrification carrier devices, the CryoTip® is considered a closed carrier for vitrification, thereby eliminating the theoretical risk of disease transmission through contaminated liquid nitrogen during cooling and storage (Bielanski et al. Cryobiology, 40:110-116, 2000). The CryoTip® is cleared by the FDA and has CE mark approval as a closed device to carry gametes or embryos in a specialized medium during cryopreservation procedures and subsequent long-term storage in liquid nitrogen. The CryoTip® has been shown to be a safe and reliable vitrification device, and when compared to other open system vitrification devices, it has provided similar results (Martino et al. Reprod Biol Endocrinol 11:27, 2013, Valbuena et al. Fertil Steril 97:209-217, 2012; Kuwayama et al. Reprod Biomed 11:608-614, 2005). The CryoTip® has also been shown to be suitable for use as a vitrification device for the cryopreservation of small volumes of sperm (Tanaka et al. Fertil Steril 90: S292, 2008).

Successful vitrification of human embryos using equilibration solution

Successful vitrification of human embryos using equilibration solution

Shortened equilibration time can compromise clinical outcomes in human embryo vitrification

Vitrification is an important way to cryopreserve human embryos and the recommended time of embryo exposure to the vitrification solution is 1 min. However, practically speaking, the duration of embryos exposure to equilibration solution can vary from 5 to 15 min. The purpose of this study was to investigate the effect of different equilibration times on the outcomes of frozen-thawed embryo transfer cycles. The data were collected from our medical records from January 2012 to June 2013 and a total of 517 cycles were included. These cycles were divided into four groups according to the equilibration time: (i) 5–6 min; (ii) 7–8 min; (iii) 9–10 min and (iv) 11–12 min. The results show that there were no differences in terms of survival rate and fully intact embryo rate among the four groups. However, lower clinical pregnancy, embryo implantation and live birth rates were observed in the 5–6 min exposure group (54.6%, 31.9% and 48.2%, respectively) compared with the three other groups. The corresponding rates in the 9–10 min group (73.5%, 47.6% and 64.7%) were the highest. This study indicated that different equilibration times influenced the clinical outcomes of human embryo vitrification and vitrification with shortened equilibration time compromised the clinical outcomes. Appropriate prolongation of the equilibrium time would probably improve the clinical outcomes.

Neonatal outcomes after the implantation of human embryos vitrified using a closed-system device.

Closed vitrification poses a risk of adversely affecting embryo development, while it may minimize the risk of contamination. We assessed the effects of closed-system human embryo vitrification on fetal development after implantation, neonatal outcome, and clinical safety. This was a retrospective cohort study conducted at a private fertility clinic. A total of 875 vitrified-warmed blastocysts that were single-transferred under hormone-replacement cycles between November 2011 and December 2013 were randomly divided into two groups (closed vitrification, n 313; open vitrification, n 562) after receiving the patients' consent forms. Developmental competence after implantation, including gestational age, birth weight, sex, Apgar score, and anomalies of newborns, after the transfer of blastocysts vitrified by closing vitrification was compared with that obtained in the case of open vitrification. There were no significant differences between the use of closed and open vitrification systems in embryo development after implantation, gestational age, birth weight, sex ratio, Apgar score, and congenital anomalies of newborns. Human embryos can be vitrified using a closed vitrification system without impairment of neonatal development.

Efficiency of an automatic dehydrated carrier for the vitrification of human embryos derived from three pronuclei fertilized zygotes

The efficiency of a new vitrification technique using embryos obtained after abnormal fertilization was assesed. A total of 100 embryos were vitrified, 48 with the automatic dehydrated carrier (ADC) technique and 52 with cryotop. The survival rates in ADC and cryotop groups were similar (85.4% versus 92.3%). The loss rates of 10.4% for the ADC group and 1.9% for the cryotop group were not significantly different. The intact rate was significantly lower in the ADC group (26.8% versus 64.6%; P = 0.001). After 48 h of culture, blastocyst formation rates in the ADC and cryotop groups were 14.6% and 25.0%, respectively, which were not significantly different. In conclusion, the practicability of ADC vitrification was demonstrated. The study has some limitations, which necessitates further work.

Perinatal outcomes for transfer of blastocysts vitrified and warmed in defined solutions with recombinant human albumin: 374 babies born after 898 embryo transfers

To assess the efficacy of a novel, defined vitrification procedure using recombinant human albumin (rHA) for cryopreservation of human blastocysts. Retrospective study. Private IVF clinic. 1,496 patients received vitrified/warmed embryo transfer (ET). Surplus blastocysts, and blastocysts from patients undergoing elective embryo cryopreservation, were vitrified/warmed using Cryotop carriers in homemade solutions containing either human serum albumin (HSA) or rHA. Clinical and neonatal outcomes regarding the vitrified/warmed ET procedures. The HSA and rHA groups had a total of 1,163 and 898 vitrified/warmed cycles, respectively. Embryo survival rates (98.7% vs. 98.9%, respectively) and the number of embryos transferred (1.08 ± 0.01 vs. 1.06 ± 0.01, respectively) were similar in the HSA and rHA groups. Clinical pregnancy rates/ET were higher (P < 0.05) in the rHA group (56.0%) than in the HSA group (51.5%). The HSA and rHA groups had similar live delivery rates/pregnancy (72.2% vs. 72.3%, respectively) and perinatal outcomes, including birth weight (2,988 ± 28 vs. 3,046 ± 26 g, respectively). Birth defects occurred in 0.9% and 1.6% of neonates in the HSA and rHA groups, respectively. rHA effectively replaced HSA for human embryo vitrification procedures, and yielded high rates of pregnancy and live births after vitrified/warmed ET. This new approach will support the development of defined ART systems, which will eliminate the variation and risks associated with the use of blood-derived products.

Cryopreservation of human oocytes, zygotes, embryos and blastocysts: A comparison study between slow freezing and ultra rapid (vitrification) methods

Abstract Preservation of female genetics is currently done primarily by means of oocyte and embryo cryopreservation. The field has seen much progress during its four-decade history, progress driven predominantly by research in humans. It can also be done by preservation of ovarian tissue or entire ovary for transplantation, followed by oocyte harvesting or natural fertilization. Two basic cryopreservation techniques rule the field, slow-rate freezing, the first to be developed and vitrification which in recent years, has gained a foothold. The slow-rate freezing method previously reported had low survival and pregnancy rates, along with the high cost of cryopreservation. Although there are some recent data indicating better survival rates, cryopreservation by the slow freezing method has started to discontinue. Vitrification of human embryos, especially at early stages, became a more popular alternative to the slow rate freezing method due to reported comparable clinical and laboratory outcomes. In addition, vitrification is relatively simple, requires no expensive programmable freezing equipment, and uses a small amount of liquid nitrogen for freezing. Moreover, oocyte cryopreservation using vitrification has been proposed as a solution to maintain women’s fertility by serving and freezing their oocytes at the optimal time. The aim of this research is to compare slow freezing and vitrification in cryopreservation of oocytes, zygotes, embryos and blastocysts during the last twelve years. Therefore, due to a lot of controversies in this regard, we tried to achieve an exact idea about the subject and the best technique used.

A closed system supports the developmental competence of human embryos after vitrification : Closed vitrification of human embryos.

Closed-system vitrification may enable the risk of contamination to be minimised. We performed three studies to compare the developmental competence of human embryos vitrified using either a closed vitrification system (CVS; Rapid-i®) or an open vitrification system (OVS; Cryo-top®). The first study was performed in vitro using 66 zygotes previously vitrified at pronuclear stage. These were warmed and randomised 1:1 to revitrification using either the OVS or the CVS. After re-warming, embryo development and blastocyst cell number were assessed. For the second study, also performed in vitro, 60 vitrified-warmed blastocysts were randomised 1:1:1 into three groups (OVS or CVS revitrification, or no revitrification). The proportion of dead cells was assessed by staining. The third study was performed in vivo, using 263 high-grade blastocysts randomly assigned to vitrification using either the CVS (n = 100) or the OVS (n = 163). After warming, single blastocyst transfer was performed. There were no differences between the CVS and the OVS in survival rate (100% vs. 97%), blastulation rate (96h: 50% vs. 50%; 120h: 68% vs. 56%), proportion of good blastocysts (96h: 32% vs. 22%, 120h: 47% vs. 41%), or mean number of cells (137 vs. 138). The proportion of dead cells in blastocysts re-vitrified by CVS (31%) was similar to that for OVS (38%) and non-revitrification (32%). In vivo, the implantation rate for blastocysts vitrified using the CVS (54%) was similar to that with the OVS (53%). Our studies consistently indicate that human embryos may be vitrified using a CVS without impairment of developmental competence.

The new Rapid-i carrier is an effective system for human embryo vitrification at both the blastocyst and cleavage stage

BackgroundThe Rapid-i is a new FDA cleared closed carrier for embryo vitrification. The cooling rate of - 1220°C/min is far lower than that reported with open vitrification systems such as the cryoloop (−15,000°C/min). Little published data is currently available on this device. This study presents our initial clinical data, as well as live birth outcomes, with the Rapid-i. The efficacy of this device for the cryopreservation of cleavage, as well as blastocyst stage human embryos is also analyzed. We further compare outcomes to those achieved with the cryoloop, an “open” vitrification system routinely used in our laboratory.MethodsHuman embryos were vitrified at either the 8–10 cell stage or else the blastocyst stage. The vitrification protocol was: 7.5% DMSO/7.5% ethylene glycol (EG) (2–3 min) followed by incubation in 15% DMSO /15% EG (45 sec) before loading on the vitrification carrier. Cryoprotectant was removed during warming by sequential washes in 0.25 M and 0.125 M sucrose in culture medium. Clinical outcome data for frozen cycles between January 2011 and August 2012 were stratified according to carrier and cell stage. The student t-test and chi square test were used to compare results. P value of < 0.05 was considered significant.ResultsA total of 486 vitrified-warmed embryos were assessed and 92% of them were transferred. The clinical pregnancy rate (CPR) and implantation rate (IR) with Rapid-i vitrified blastocysts were 59% and 49%, versus 47% and 37%, respectively for cleavage stage embryos. This was not statistically different from results with the cryoloop vitrified blastocysts (CPR 46%, IR 38%) nor the cleavage stage vitrified embryos (CPR 49%, IR 35%). To date, there have been 31 deliveries of 34 healthy infants from Rapid-i vitrified embryos, with another 12 pregnancies still on-going.ConclusionsThe Rapid-i offers an excellent alternative to existing open vitrification devices for embryo cryopreservation at the 8–10 cell stage as well as the blastocyst stage. Use of this type of “closed” sealed system that prevents direct contact between the embryos and liquid nitrogen reduces the potential risk of sample cross-contamination or infection. These preliminary data and live birth outcomes have paved the way toward transitioning to a closed vitrification system in our own IVF program.

Vitrification of human embryos previously cryostored by either slow freezing or vitrification results in high pregnancy rates

Occasionally, clinical scenarios arise where embryos, previously cryostored and warmed, need to be recryopreserved. The outcome of 30 such transfer cycles from 25 women where embryos were recryopreserved is detailed. In 16 cases, embryos were initially cryopreserved by slow freezing and in 14 cases by vitrification. The cryopreservation stages were the pronuclear stage (n=16), day-3 cleavage stage (n=12), blastocyst (n=1) and oocytes (n=1). All recryopreservation was by Cryotop-based vitrification. From this mixed source, 30/31 twice-cryopreserved embryos survived warming and were transferred, resulting in 13 pregnancies, 11 deliveries with normal gestational age and birthweight, one pre-term birth at 33 weeks and two miscarriages. There were no malformations reported for the live births. Recryopreservation using vitrification by CryoTop has been used in a variety of clinical scenarios to preserve surplus cryopreserved embryos. The current study, although limited in numbers, resulted in high survival rates, clinical pregnancy rates similar to once-cryopreserved embryos and healthy live births independently of the initial stage and cryopreservation method. The technique may increasingly be applicable to elective single-embryo transfer and blastocyst transfer to maximize the pregnancy rate while minimizing the number of cryopreserved embryo transfers.Freezing of embryos has significantly influenced the likelihood of pregnancy after an IVF stimulation cycle. Since it is often unclear which embryos have the potential to continue developing, more embryos may be thawed than transferred. If, in such cases, the surplus embryos remain viable, the ability to refreeze them is an important laboratory tool. In this study, we report on 30 cycles from 25 women where, for a variety of clinical scenarios, surplus embryos previously cryopreserved by either slow freezing or more recently by vitrification were recryopreserved using the Cryotop vitrification protocol. The report details 13 pregnancies, including 11 live births. The study group included one case where oocytes rather than embryos were initially vitrified. When these recryopreserved embryos were subsequently warmed, they exhibited a high survival rate and, after embryo transfer, a pregnancy rate similar to once-cryopreserved embryos. While recryopreservation has to date demonstrated that the embryos retain viability, leading to healthy children, and is, as such, an additional tool for embryologists, its application should proceed with caution.