Abstract Study question Due to the rising trend for oocyte cryopreservation and increasing demand, there is an urgency to make oocyte vitrification and warming protocols more efficient. Summary answer Oocyte vitrification and warming can be done successfully in 4 minutes, reducing the time of commonly used current protocols by up to 25 minutes. What is known already Vitrification and warming of oocytes is widely used in many IVF clinics worldwide and routinely utilized as a dominant technique for fertility preservation and freezing of donor as well as autologous oocytes in many clinical settings. Outcome data using cryopreserved oocytes is comparable to fresh oocytes. However, vitrification and warming of human oocytes can be time consuming. Usually, vitrification of oocytes is done in about 11-16 minutes, and warming requires another 11 minutes. Also, it is a common practice to load not more than 2 oocytes per carrier at time of vitrification. Study design, size, duration From November 2022 to November 2023, 504 germinal vesicles (GV) and 207 metaphase I (MI) oocytes, which did not convert into mature oocytes at day of oocyte retrieval, were vitrified in Vit Kit Freeze (FujiFilm Irvine Scientific, USA), and RapidVit Oocyte (Vitrolife, Sweden). Their survival rates were compared, and as an additional viability indicator, post-warming developmental capabilities were evaluated after 24 and 48hrs of culture in Global High Protein (LifeGlobal, USA). Participants/materials, setting, methods A group of 4 GV or MI oocytes was vitrified at room temperature for 1 minute in Equilibration solution (ES) and 1 minute in Vitrification solution (VS), loaded on a S-Cryolock (Biotech, USA) and then plunged into liquid nitrogen. Warming was done at 37ºC in 0.5M sucrose for 1 minute and Wash solution (WS) for another minute. Post warming, GV and MI oocytes were cultured for developmental evaluation. Statistical significance was defined as P < 0.05. Main results and the role of chance There was no statistically significant difference in the survival rate of GV oocytes between Irvine (209/219; 95.4%), and Vitrolife media (270/285; 94.7%; P = 0.72). The survival of MI oocytes was also comparable and not significantly different between both media; Irvine: 103/106; 97.2%, vs Vitrolife: 96/101; 95.0%; P = 0.43). Because of the similar survival rates of oocytes in both groups, for the developmental potential and conversion rates, oocytes of both groups were combined. Of the 504 GV, 479 survived (95.0%) and after 24hrs of culture, 441 converted to MI (92.1%). After a culture of 48hrs, 56.5% (249/441) converted to metaphase II (MII) oocytes. Combining the vitrified and warmed MI oocytes from both groups, 96.1% (199/207) survived the warming, and after 24hrs of culture, 168 (84.4%) converted to MII oocytes. Vitrification and warming of oocytes at different developmental stages can be done with a short exposure time to hypertonic (ES/VS) and hypotonic (0.5M sucrose) solutions in a total of 4 minutes. Limitations, reasons for caution Immature oocytes were used as study material to evaluate a new protocol with significantly reduced vitrification and warming times. At this moment we cannot provide data on fertilization, embryo development, and blastocyst formation. Wider implications of the findings Oocytes are exposed to less suboptimal temperatures during dehydration and rehydration and help to maintain their developmental potential. Loading 4 oocytes per carrier combined with the decrease in time resulted in increased efficiency. Warming in 0.5M instead of 1M sucrose contributes positively to survival and allows earlier resumption of functionality. Trial registration number not applicable

RESEARCH QUESTIONCan immature oocytes vitrified and warmed using a short protocol survive and resume meiosis?DESIGNThis study examined modifications of oocyte vitrification and warming protocols that shorten the time of exposure to vitrification and warming solutions. A total of 561 germinal vesicles (GV), and 218 metaphase I (MI) oocytes that were immature at oocyte retrieval were vitrified at room temperature in 2 minutes. Warming was done at 37°C in 2 minutes. Resumption of meiotic activity was evaluated after 24 and 48 hours of culture. Two different commercially available vitrification and warming kits were used for comparison.RESULTSNinety five percent of GV oocytes survived with no difference observed between kits. The survival of MI oocytes was on average 95.4% and not significantly different between the two kits. Of the 533 GV that survived, 491 converted to MI (92.1%). After culture for 48 hours, 54.4% converted to MII oocytes. In addition, of the 208 MI that survived the warming, 84.1% converted to MII oocytes after culture for 24 hours.CONCLUSIONSVitrification and warming of oocytes at different nuclear maturation stages can be done with a 2-minute exposure time to hypertonic and a 2-minute exposure time to hypotonic solutions. With this approach, the oocytes are less exposed to room temperature during dehydration and rehydration. Warming in 0.5M sucrose helps to maintain and support the potential to resume nuclear meiotic activity of oocytes and conversion from GV to MI and to MII oocytes.

Today, preimplantation genetic screening (PGS) is performed at the blastocyst stage by removing trophectoderm cells. The biopsy can be done at the blastocyst stage on day 5 or day 6. Elective single embryo transfer (eSET) is key to healthy singleton pregnancies. This outcome can be further improved by transferring a single euploid embryo. One would expect that a euploid embryo regardless of the day of biopsy should have an equal potential to implant. Therefore, we were looking into single euploid transfers using day 5 blastocysts and compare it to single euploid day 6 blastocysts.

After more than 3 decades since the birth of Louse Brown, the definition of success in infertility treatment has changed dramatically. Today, quality of IVF is measured on three criteria; 1) term birth, 2) birth weight and 3) singleton pregnancy. These goals can only be achieved by performing embryo transfer of one selected embryo at a time, preferably at the blastocyst stage. For those reasons, it is utterly important to have well defined criteria in place to determine which patients should be pushed to day 5 for an eSET. Therefore, we examine if the number of good quality cleavage stage embryo available on day 2 has on impact on the baby-take-home rate after eSET. Retrospective analysis. A total of 1545 eSETs were performed from February 2007 to February 2016. The average maternal age was 32±3.2yrs. Oocytes had undergone ICSI and cultured in single step media (Global) for extended culture under low oxygen tension (5%). eSETs were divided in four groups based on the number of available good cleavage stage embryos on day 2 defined as 4-cell stage embryos without fragmentation. The 4 groups were compared in terms of baby-take-home rate. Chi square test was used for statistical analysis of the data. There was no significant difference among the groups in regards to patient's age, clinical pregnancy, implantation or baby-take-home rate. Regardless of the mean number of day 2 embryos, in all groups about 50% were vitrified at the blastocyst stage. Even the ratio between day 5 and day 6 vitrified blastocysts was similar in all groups.Tabled 1No. of good cleavage stage embryos on day 21-56-89-12>13No. of eSETs245430468402Age (Mean±SEM)32.4±3.232.3±3.331.8±3.231.5±3.1Mean number of day 2 embryos471017Mean number of blastocysts vitrified2358Total number of blastocysts vitrified433125122503159% Ratio of day 5 vs. day 6 vitrified68 vs. 3267 vs. 3366 vs. 3471 vs. 29Clinical pregnancy/eSET (%)54.3a55.1a56.5a52.5aImplantation rate (%)62a62.1a63.2a62.2aBabe-take-home rate (%)54.3a55.1a55.5a52aSets of Monozygotic twins (n)7345Live births (n)133237260209aχ2, P<0.05 Open table in a new tab aχ2, P<0.05 Our study found that when there are 5 or less day 2 good morphologic embryos, the patient can be confidently pushed to a day 5 eSET. The cryopreservation utilization rate was similar among the groups. Overall, stimulation toward lower egg numbers and in turn to less cleavage stage embryos provides a safe opportunity for the embryology laboratory to push patients under age 35 for an eSET. Along with the low number of multiple births, this approach delivers a high pregnancy rate, moving towards the endpoint of a single, safe and healthy live birth.

Elective single embryo transfer (eSET) is the preferred recommendation towards the optimum outcome of a single, healthy live birth. Evidenced based medicine supports this strategy from both reproductive and obstetric viewpoints. The potential to reduce multiple pregnancies has the greatest impact in good prognosis patients with age <37. A successful embryo cryopreservation program can offer the patient the opportunity to transfer one blastocyst in the fresh cycle, and if negative outcome, the patient can subsequently transfer frozen blastocysts. We examined patients with failed fresh day 5 eSETs, and the likelihood of pregnancy in the following frozen embryo transfer (FET). The primary endpoint was the rate of ongoing pregnancies (oPR) from fresh eSETs compared with FET. Retrospective analysis A total of 701 FETs with one previously failed fresh eSET were analyzed. The average maternal age was 32.4±3.1yrs. Blastocysts were collapsed prior to vitrification and subsequently hatched using a laser (Hamilton Thorne) 1-2 hrs prior to transfer. Both natural and hormone replacement cycles were used to increase receptivity of the endometrium. Progesterone was supplemented on day 15 of the cycle and blastocysts were warmed on day 5 of progesterone supplementation. The oPR of those FETs was then compared to 1847 fresh day 5 eSETs. Chi square test was used for statistical analysis. There was no significant difference between the 2 groups in regards to patient’s age, or oPR. There was a lower implantation rate (IR) and higher twin pregnancy rate in the FET group due to 60% of patients who transferred 2 blastocysts with FET when the fresh eSET failed.Tabled 1FET groupeSET groupχ2Number of embryo transfers (n)7011847-Age (Mean±SEM)32.4±3.132.0±3.1nsMean number of embryos being transferred (n)1.61.0-Clinical pregnancy/transfer (%)62.961.9nsImplantation rate (%)50.6a62.0a<0.05Ongoing pregnancy rate/transfer (%)58.556.1nsPatients transferring TWO embryos (%)60--Twin pregnancy rate (%)30.1b1.9b<0.001Denotes statistical significance aP<0.05; bP<0.001 Open table in a new tab Denotes statistical significance aP<0.05; bP<0.001 The findings demonstrated a high oPR with FET when the fresh eSET is not successful. The higher number of embryos transferred in FET due to the first unsuccessful fresh eSET resulted in lower IR and high rate of twin pregnancies. The next step will be to recommend transfer of single blastocyst with FET even after a failed eSET due to the high rate of pregnancy.

Elective single embryo transfer (eSET) in a highly selected population offers excellent clinical pregnancy rates with low rates of multiple gestation. Preimplantation genetic screening (PGS) is increasingly utilized, but whether it is superior to eSET in a good prognosis population has yet to be determined. Our objective was to compare pregnancy outcomes in good prognosis patients who utilized PGS versus those who underwent a fresh IVF cycle with eSET. Retrospective matched cohort study. IRB approval was obtained. Women <38yo undergoing autologous IVF with single embryo transfer between 2012-2015 were included. eSET was offered to women <38yo with <2 prior failed cycles and >1 high quality blastocyst on day 5. Outcomes with eSET were compared to outcomes with PGS followed by frozen transfer of a single euploid embryo (PGS/sFET). eSET patients were matched to PGS/sFET patients by age and date of embryo transfer with a 3:1 ratio. The primary outcome was ongoing pregnancy rate (OPR) at 20 weeks gestation. Odds ratios (OR) used eSET as the reference group. 237 women were included: 172 (72.6%) eSET and 65 (27.4%) PGS/sFET. There were no significant demographic differences between groups (Table 1). There was no significant difference in clinical pregnancy rate (CPR) between PGS/sFET and eSET, 58.5% vs. 61.6%, OR 0.88 (95% CI 0.49-1.57). There was also no significant difference in OPR between PGS/sFET and eSET, 44.6% vs 55.8%, OR 0.64 (95% CI 0.36-1.13). There was a significantly higher spontaneous abortion rate with PGS/sFET compared to eSET, 23.7% vs. 7.5%, OR 3.80 (95% CI 1.34-10.74). In patients <35yo, there was a trend toward a lower OPR in PGS/sFET compared to eSET, 44.7% vs 62.3%, P=0.09. In patients 35-37 years old, the OPR was similar between groups, 44.4% vs 43.1%, P=0.92. Outcomes with PGS/sFET were not superior to eSET in this population. In women <38yo who underwent fresh IVF with careful visual selection of the single best embryo on day 5, ongoing pregnancy rates were similar to women who underwent PGS/sFET with euploidy as guidance for embryo selection. Surprisingly, there was a significantly higher miscarriage rate in women <38 undergoing PGS/sFET, which needs to be examined in a larger study. While the use of PGS may confer other benefits, it was not found to increase ongoing pregnancy rates in a good prognosis population.Tabled 1Table 1. Group Characteristics (N=237)PGS/sFET (n=65)eSET(n=172)p-valueMean Age yrs (SD)33.4 (3.0)33.1 (2.8)0.47Mean BMI kg/m2 (SD)25.1 (5.7)25.5 (6.4)0.66Mean Day 3 FSH (SD)8.4 (3.2)8.2 (4.1)0.72Smoking2/65 (3.1%)13/172 (7.6%)0.25Clinical Pregnancy Rate (CPR)38/65 (58.5%)106/172 (61.6%)0.76Ongoing Pregnancy Rate (OPR)29/65 (44.6%)96/172 (55.8%)0.16Biochemical Loss Rate8/46 (17.4%)12/118 (10.2%)0.29Spontaneous Abortion Rate9/38 (23.7%)8/106 (7.5%)0.01 Open table in a new tab

The Eeva Test is an automated time-lapse enabled test that generates predictions based on cell division timings. As blastocyst development was used to build the model, Eeva Test has been shown to improve Day 3 embryo selection (1). However, there is evidence suggesting by incorporating key cell division timings Eeva results reflect underlying molecular health of embryos (2,3). The objective of the study is to assess whether Eeva Test can be applied to blastocyst selection. Retrospective multi-center study. Total of 342 patients underwent blastocyst transfers from 7 centers consented to use Eeva Test in IVF. Patients were separated into three groups, at least 1 Eeva High (H) transferred, at least 1 Medium (M) transferred and only Low (L) transferred, and clinical pregnancy and implantation rates were compared. Spearman rank were used to check correlations among the variables of interest (age, #eggs, #2PN, morphology, Eeva results, #embryos transferred). Univariate logistic regression was used to assess predictive power of the variables against pregnancy. Multivariate logistic regression with backward selection was used to test the predictive value of Eeva results with other key pregnancy predictors included. Patients with at least 1 Eeva H blastocyst transferred had significantly higher pregnancy and embryo implantation rates than patients with only L blastocysts transferred (54% vs 34%, p=0.002 and 46% vs 29%, p=0.002), while patient age and #embryos transferred was similar between groups. Univariate logistic regression showed that age, #2PN and Eeva results (H vs L) are significantly correlated with pregnancy (p=0.008, 0.02, 0.02), while Spearman correlations showed Eeva results were not correlated with the other variables. Multivariate model revealed that Eeva results remain as significant predictor of pregnancy after adjusting for age and blastocyst morphology while the other variables were not significant.Tabled 1Logistic regression result.VariableOdds ratioP-value95% CI (odds ratio)Eeva results (H vs. L)2.150.007(1.23,3.74)Age0.920.005(0.86,0.97)Blastocyst morphology0.460.002(0.29,0.75)#embryos transferred1.410.14(0.88,2.42)#eggs0.980.69(0.92,1.05)#2PN1.030.49(0.94,1.12) Open table in a new tab It is well known that age and blastocyst morphology are predictors of pregnancy. The current study is the first to show early cell division timing parameters have independent predictive value for blastocyst transfer outcome. Not only does it indicate Eeva Test may be used to improve blastocyst selection, but it also supports the hypothesis that normal embryo development may follow programmed timings at early stage (2).

Our aim was to examine instances in which no sperm could be identified after thaw among cryopreserved samples of rare or very low counts of sperm. Retrospective review. We performed a retrospective review of our institutional fertility clinic database from 2010 until 2015. We identified couples that underwent intracytoplasmic sperm injection (ICSI) with either ejaculated or testicular cryopreserved-thawed sperm. Inclusion criteria was limited to men with ejaculated cryptozoospermia or severe oligozoospermia (< 100,000 total sperm per specimen) or men with azoospermia due to spermatogenic defects who underwent microsurgical testicular sperm extraction (mTESE) with similarly low pre-cryopreservation total sperm counts. Sperm were cryopreserved in 1 mL vials using liquid nitrogen. Pre-cryopreservation specimens were categorized as rare sperm only (Group 1) or less than 100,000 total sperm (Group 2). The primary outcome was the availability of any post-thaw sperm for use at the time of ICSI. The relationship between successful post-thaw sperm retrieval and pre-cryopreservation parameters was examined using Chi-square statistical analysis for ordinal variables. We identified 34 couples who met inclusion criteria and with sufficient follow-up data, encompassing 49 ICSI cycles (Table 1). There were three ICSI cycles (6.1% of all ICSI cycles) among three different couples in which no sperm were able to be identified post-thaw. All three instances (n=1 ejaculated source, n=2 testicular source) were noted to have pre-thaw parameters of rare sperm only (Group 1) (8.6% of ICSI cycles utilizing cryopreserved rare sperm only). Chi-square analysis of the ability to locate sperm post-thaw and pre-cryopreservation total sperm count revealed a significant association (Pearson χ2=49.0, df=2, p=<0.001). Failure to retrieve any sperm after thawing of rare or very low quantities of cryopreserved sperm is an infrequent event and limited to those patients with rare quantities of sperm prior to cryopreservation. Although this study is limited in size, the results may influence recommendations to have back-up donor sperm available or to consider use of fresh sperm in further mTESE procedures for future ICSI cycles among men with cryopreserved rare sperm.Tabled 1Table 1. Post-Thaw Outcomes of Rare or Very Low Counts of Cryopreserved Sperm.Number of ICSI cycles in which sperm were identified post-thaw (n=46 ICSI cycles)Number of ICSI cycles in which sperm were not identified post-thaw (n=3 ICSI cycles)Pre-cryopreservation rare sperm only (Group 1)353Pre-cryopreservation total count110Ejaculated sperm source131Testicular sperm source332Number of vials of sperm thawed697 Open table in a new tab

AMH is a widely used for assessment of ovarian reserve prior to Assisted Reproductive Technology (ART). As AMH is cycle independent, it is easy to implement. Recent data has questioned its reliability in women who fall in extreme ranges of body mass index (BMI). We sought to determine if AMH is a reliable predictor of a patient's ovarian reserve in obese women. A retrospective chart review was conducted on all patients who underwent in vitro fertilization (IVF) at a large clinical practice after Institutional Board Review approval. Data was collected from January 1, 2012 through December 31, 2013. All patients received an infertility evaluation in preparation for their upcoming cycle of IVF. Patient characteristics were reviewed, including: age, BMI, AMH, number of oocytes retrieved and cycle outcome. Analysis was performed to determine if AMH predictability for oocyte yield was maintained in obese women compared to non-obese. Subsequent analysis used subcategories of BMI. All of these variables were analyzed for various associations using version 19 of SPSS software. 5558 patients underwent IVF. 2965 patients were included due to availability of AMH testing. 2963 patients were included due to BMI reporting. In the population of patients that were normal weight: mean age was 35.9 years, mean AMH was 2.30, mean BMI was 22.9 mg/kg2 and mean number of retrieved oocytes was 13.9 oocytes. In the patients that met criteria for obesity with a BMI greater than 30 mg/kg2: mean age was 36.7 years, mean AMH was 2.29, mean BMI was 35.5 mg/kg2 and mean number of retrieved oocytes was 13 oocytes. AMH and number of retrieved oocytes were similar in both populations, no difference of statistical significance was found. Subsequent analysis compared patients with BMI less than 19 mg/kg2, BMI from 20 to 25 mg/kg2, BMI 25 to 29 mg/kg2, and BMI greater than 30 mg/kg2. No significant differences were noted between these various subsets of BMI; in terms of AMH and number of oocytes retrieved. AMH is a reliable predictor of ovarian reserve in both obese and non-obese women; it is an accurate predictor regardless of BMI.

The Eeva Test combines automated time-lapse analysis with statistical modeling to predict continued embryo development. It automatically captures quantitative image features that the human eye cannot detect, and incorporates statistical modeling of dynamic parameters to maximize the test’s predictive power. A new predictive algorithm includes parameters of patient prognosis, morphology, early cleavage timings and quantitative image features to generate a 5-category score for developmental potential. The objective of this multi-center study was to validate this new algorithm against embryo implantation following blastocyst transfer. Retrospective multi-center study The study included a total of 151 patients from 9 centers who consented to use the Eeva Test. Embryos were transferred on Day 5, and embryos with known implantation data were analyzed. Images and clinical data were processed by the Eeva Test’s new Xtend algorithm, which utilizes automated image analysis software to classify embryos into five categories by incorporating key cell division timing parameters, Day 3 cell#, age and quantitative image features reflective of activity during the third cell cycle. The overall known implantation rate (IR) for the study population was 39% (84/216). The 5-category output of the extended algorithm correlated positively with blastocyst IR: 51% (36/71), 45% (33/74), 25% (10/40), 24% (5/21), 0% (0/10). In patients younger than 35yo, the overall known IR was 50% (62/123); however, the IR for “Category 1” embryos was 60% (32/53). In the same patient group, IR for the good morphology blastocysts was 56% (54/97); however, embryos with good morphology and Category 1 achieved a 61% (31/51) IR. The novel aspect of the new algorithm is including computer-extracted quantitative image attributes, patient characteristics and traditional morphology parameters in the predictive model. Although the model was built using blastocyst as outcome variables, in current study we were able to demonstrate that 5-category output in the new algorithm correlated positively with blastocyst implantation. Specifically, in young patients who are more likely in need of embryo selection, Category 1 identified a subset of embryos with 60% implantation rate, which approaches the implantation rate of euploid blastocysts (Forman et al, 2013). The algorithm also identified embryos with better implantation potential among good morphology blastocysts from young patients. Combining extended algorithm and traditional morphology may present a non-invasive approach assisting embryologists to select the best blastocyst(s) for transfer, and encourage the practice of elective SET.

There are a number of commercially available storage devices used for the vitrification of oocytes. The increased cooling and warming rates of an "open" device are favored over that of a closed system. The thickness of the loading region of the storage device may affect the kinetics of rapid temperature change. This study is designed to evaluate three open devices that differ in thickness by comparing cryo-survival, fertilization, cleavage and clinical pregnancy rates, as well as the number of cycles with surplus blastocysts available post embryo transfer. Retrospective study. We reviewed donor oocyte warming cycles performed by multiple IVF laboratories using vitrified oocytes from a commercially available donor egg bank. Mature donor oocytes were vitrified on one of three different cryodevices labeled A-C. The thickness of each device varied from 0.012'' (A), 0.009'' (B) to 0.005''(C). A proven oocyte vitrification protocol was used (7.5% EG+DMSO followed by 15% EG+DMSO+0.5M Suc) with commercially available media (Irvine Scientific). Each recipient received between 5 to 7 oocytes and they were inseminated by ICSI. Surplus blastocysts were vitrified for future use. Oocyte survival, fertilization, embryo cleavage and clinical pregnancy rates (presence of a fetal heart beat) are comparable between the groups and do not appear to be affected by the thickness of the cryo device. Of note is a higher percentage of cycles with surplus blastocysts available for vitrification with cryodevice C, the thinnest device, as compared to device A, the thickest device (49% vs 41% respectively P=0.02, Fisher's Exact test). There was no significant difference observed in SAB or cancellations between devices. The rates of oocyte cryo survival, fertilization, embryo cleavage, clinical pregnancy and IR by FCA are not significantly affected by thicknesses of the cryo device. However, more cycles resulted in surplus blastocysts that were vitrified and available for future cycles using the thinnest cryo device (C) as compared to the thickest device. While all labs received training in carrying out an identical protocol, it is impossible to completely control for any differences between labs as cryodevices segregated exactly with the laboratory staff and environment.Tabled 1ABCThickness0.012"0.009"0.005"Cycles325609578% Survival89%85%84%% Cleavage95%96%98%% Clinical50%49%51%% SAB7%6%7%IR by FCA37%36%38%% cycle with surplus blast41%45%49%Canceled cycles*21 (6%)49 (8%)36 (6%) Open table in a new tab

The automated, time-lapse enabled Eeva Test provides test scores that predict blastocyst formation (1) and correlate to implantation (2). Specifically, positive correlation was demonstrated between Eeva Test results and implantation/pregnancy following Day 3/Day 5 transfer (2). However, the relationship between Eeva Test results and blastocyst implantation has not yet been examined. Furthermore, there is no data to inform the implication for patients in different age groups. Retrospective multi-center study. The study included a total of 342 blastocyst transfer patients from 7 centers (2012-2014) who consented to have embryos imaged using the Eeva System, a platform technology that automatically measures P2 (time between 2- and 3- cell) and P3 (time between 3- and 4- cell) and generates an Eeva Test score of High, Medium, or Low to indicate the embryo developmental potential. Embryos with known implantation data were included in this study. P-values were calculated using χ2 or student's t-test (SAS 9.3). The overall known implantation rate for the study was 38% (148/386). Eeva High blastocysts had a relative 40% higher implantation rate than Eeva Low blastocysts (p=0.002), while the age was similar between the two groups. Further analysis revealed that in older patients (age≥35), Eeva High blastocysts had over twice the implantation rate than Eeva Low blastocysts (p=0.002).Tabled 1Eeva resultsImplantation rateAgeImplantation rate (Age<35)Implantation rate (Age≥35)High45% (86/191)34.1 ± 4.351% (52/101)37% (34/90)Medium40% (27/67)34.5 ± 3.551% (18/35)28% (9/32)Low27% (35/128)34.7 ± 4.240% (25/62)15% (10/66)P-value0.0020.20.20.002 Open table in a new tab This study demonstrated that Eeva High blastocysts implant at a higher rate than Eeva Low blastocysts, which suggest that the automated Eeva Test is informative for Day 5 blastocyst assessment. Improving Day 5 implantation will give clinics and patients more confidence to perform single blastocyst transfer and thereby reduce the risk of multiple pregnancies.

ObjectiveThroughout the three-decade history of in-vitro-fertilization (IVF) treatment, elective single embryo transfer (eSET) has been the exception, rather than norm. To compensate the low rate of implantation for individual embryos and to achieve pregnancy rates higher than 50%, the general practice is to systematically transfer multiple embryos in the vast majority of patients. Because eSET is the only truly effective means by which to avoid multiple pregnancy in IVF cycles, we looked at our eSET program implemented in 2007 and the cumulative pregnancy rate after transfer of fresh and vitrified-warmed blastocysts (VET) in patients treated with eSET, to evaluate the total potential of a stimulated cycle. In addition, the augmentation potential of blastocyst vitrification was reviewed.DesignA retrospective analysis.Materials and MethodsA total of 1017 autologous eSET (mean age 32.0±3.3) and 320 consecutive VET’s (mean age 32.1±3.1) after failed fresh eSET on day 5 between 2007 and 2013 were reviewed. Oocytes had undergone ICSI and then were cultured under low oxygen tension (5%) using a single step media (Global (IVF Online) for extended culture. For cryopresevation, laboratory protocols were followed including vitrification and warming of the blastocysts. To prepare patients for VET, both natural and hormone replacement cycles were used to increase the receptivity of the endometrium. Statistical significance was evaluated by Chi-Square. A probability of <0.05 was considered significant.ResultsBoth cPR and oPR were not different between eSET compared to VET (61.7%; 625/1017 vs. 62.8%; 201/320, p>0.05; and 55.9%; 568/1017) vs. 59.0%; 189/320, p>0.05). This leads to a cumulative ongoing PR per oocyte retrieval of 74.4% (757/1017), an added value of 18.5% through the cryopreservation. The occurrence of twin pregnancy in the eSET population was less than 3% (16/525). In contrast, a twin rate of 36.3% was noted in the VET population because 77% of these patients (247/320) decided to transfer 2 embryos.ConclusionOur data indicates that eSET in selected patients provides excellent delivery rates with one healthy baby in more than 97% of all pregnant patients after fresh eSET. Furthermore, cryopreservation at the blastocyst stage can increase the reproductive potential of one stimulated cycle by up to 20%. ESET combined with cryopreservation can help to reduce the occurrence of multiple pregnancies, which overall will be beneficial for ART and our patients. ObjectiveThroughout the three-decade history of in-vitro-fertilization (IVF) treatment, elective single embryo transfer (eSET) has been the exception, rather than norm. To compensate the low rate of implantation for individual embryos and to achieve pregnancy rates higher than 50%, the general practice is to systematically transfer multiple embryos in the vast majority of patients. Because eSET is the only truly effective means by which to avoid multiple pregnancy in IVF cycles, we looked at our eSET program implemented in 2007 and the cumulative pregnancy rate after transfer of fresh and vitrified-warmed blastocysts (VET) in patients treated with eSET, to evaluate the total potential of a stimulated cycle. In addition, the augmentation potential of blastocyst vitrification was reviewed. Throughout the three-decade history of in-vitro-fertilization (IVF) treatment, elective single embryo transfer (eSET) has been the exception, rather than norm. To compensate the low rate of implantation for individual embryos and to achieve pregnancy rates higher than 50%, the general practice is to systematically transfer multiple embryos in the vast majority of patients. Because eSET is the only truly effective means by which to avoid multiple pregnancy in IVF cycles, we looked at our eSET program implemented in 2007 and the cumulative pregnancy rate after transfer of fresh and vitrified-warmed blastocysts (VET) in patients treated with eSET, to evaluate the total potential of a stimulated cycle. In addition, the augmentation potential of blastocyst vitrification was reviewed. DesignA retrospective analysis. A retrospective analysis. Materials and MethodsA total of 1017 autologous eSET (mean age 32.0±3.3) and 320 consecutive VET’s (mean age 32.1±3.1) after failed fresh eSET on day 5 between 2007 and 2013 were reviewed. Oocytes had undergone ICSI and then were cultured under low oxygen tension (5%) using a single step media (Global (IVF Online) for extended culture. For cryopresevation, laboratory protocols were followed including vitrification and warming of the blastocysts. To prepare patients for VET, both natural and hormone replacement cycles were used to increase the receptivity of the endometrium. Statistical significance was evaluated by Chi-Square. A probability of <0.05 was considered significant. A total of 1017 autologous eSET (mean age 32.0±3.3) and 320 consecutive VET’s (mean age 32.1±3.1) after failed fresh eSET on day 5 between 2007 and 2013 were reviewed. Oocytes had undergone ICSI and then were cultured under low oxygen tension (5%) using a single step media (Global (IVF Online) for extended culture. For cryopresevation, laboratory protocols were followed including vitrification and warming of the blastocysts. To prepare patients for VET, both natural and hormone replacement cycles were used to increase the receptivity of the endometrium. Statistical significance was evaluated by Chi-Square. A probability of <0.05 was considered significant. ResultsBoth cPR and oPR were not different between eSET compared to VET (61.7%; 625/1017 vs. 62.8%; 201/320, p>0.05; and 55.9%; 568/1017) vs. 59.0%; 189/320, p>0.05). This leads to a cumulative ongoing PR per oocyte retrieval of 74.4% (757/1017), an added value of 18.5% through the cryopreservation. The occurrence of twin pregnancy in the eSET population was less than 3% (16/525). In contrast, a twin rate of 36.3% was noted in the VET population because 77% of these patients (247/320) decided to transfer 2 embryos. Both cPR and oPR were not different between eSET compared to VET (61.7%; 625/1017 vs. 62.8%; 201/320, p>0.05; and 55.9%; 568/1017) vs. 59.0%; 189/320, p>0.05). This leads to a cumulative ongoing PR per oocyte retrieval of 74.4% (757/1017), an added value of 18.5% through the cryopreservation. The occurrence of twin pregnancy in the eSET population was less than 3% (16/525). In contrast, a twin rate of 36.3% was noted in the VET population because 77% of these patients (247/320) decided to transfer 2 embryos. ConclusionOur data indicates that eSET in selected patients provides excellent delivery rates with one healthy baby in more than 97% of all pregnant patients after fresh eSET. Furthermore, cryopreservation at the blastocyst stage can increase the reproductive potential of one stimulated cycle by up to 20%. ESET combined with cryopreservation can help to reduce the occurrence of multiple pregnancies, which overall will be beneficial for ART and our patients. Our data indicates that eSET in selected patients provides excellent delivery rates with one healthy baby in more than 97% of all pregnant patients after fresh eSET. Furthermore, cryopreservation at the blastocyst stage can increase the reproductive potential of one stimulated cycle by up to 20%. ESET combined with cryopreservation can help to reduce the occurrence of multiple pregnancies, which overall will be beneficial for ART and our patients.

The objective was to describe clinical pregnancy, live birth, and multiple gestation rates resulting from blastyocyst transfers in gestational carriers using anonymous donor oocytes. In 2013, the American Society of Reproductive Medicine's committee opinion on the number of embryos to transfer encouraged elective single embryo transfer (eSET) in donor oocyte recipients. Many intended parents using gestational carriers were hesitant to transfer a single embryo. Retrospective analysis. A review of gestational carrier cycles in which anonymous donor oocytes were used from January 2009 to June 2013 (N=108). Only fresh cycles with single or double blastocyst transfers were included for analysis. Fisher's Exact and Chi squared were utilized where appropriate. Twenty-four (22.2%) eSET cycles and 84 (77.8%) DET cycles were identified. The clinical pregnancy rate (CPR) was 14/24 (58.3%) for eSET and 60/84 (71.4%) for DET, which was not significantly different with p=0.223. The live birth rate was 10/24 (41.7%) for SET and 56/84 (66.7%) for DET which was significantly different with p=0.027. Of the 56 live births in the DET group, there were 29 (48.3%) singletons, 26 (46.4%) twins, and 1 (1.7%) triplets. In the eSET group there was 1 set of twins. Gestational carriers utilizing anonymous donor oocytes have both excellent prognosis embryos as well as an optimal uterine environment. Both CPR and live birth rates are less in the eSET group compared to the DET. However, a 48.1% multiple gestation rate in the DET group is clinically important. Interestingly, approximately 78% of cases still underwent DET. The dramatically high multiples rate should be considered when determining eSET versus DET in this population with an excellent prognosis.

The two-step protocol is thought to mimic the physiological interaction between specific developmental stages of the embryo within the oviduct and uterus. The opposing system is based on the ‘let the embryo choose’ principal that is analogous to a single step culture medium. By culturing sibling oocytes in both media under the same culture conditions, we compared embryo development until day 6, as well as blastocyst formation and utilization. A retrospective analysis. Group embryo culture was done in 4-well dishes under oil and 5% oxygen. On day-1, following 2pn-check, sibling zygote cohorts from 181 patients (average age of 33.5±4.5) were randomly equally allocated into the SAGE (Trumbull, CT) or GLOBAL (IVFonline, Guilford, CT) culture media. A combined total of 2002 embryos were cultured in groups of up to five in 0.5ml volumes of their assigned medium. Embryos cultured in the SAGE were transferred from cleavage to blastocyst medium early on day-4 of culture. Embryos cultured in GLOBAL were transferred in fresh media on day-4 culture also. Chi square test was used for statistical analysis. Day 3 embryo development (≥6 blastomeres) was not significant between both media. On day 4, a significant higher number of compacting embryos was observed in the GLOBAL (63.5% vs. 51.5%; P

Today, vitrification of human blastocysts provides an excellent outcome although a big water filled cavity called the blastocoele needs to be dehydrated prior to the cooling steps of vitrification. The replacement of water by the cryoprotectant seems to be the supported by artificially collapsing of the blastocoel. Therefore we started to apply artificial collapsing on human blastocysts using a laser pulse prior to vitrification, and then compared outcome data with frozen embryo transfers (FET), where no artificial collapse was applied. Retrospective analysis. A total of 1395 FET (average age 35.1±5.0) including day 5/6 blastocysts without artificial collapsing (group A) were performed. The study group with artificial collapsing (group B) consisted of 143 FET (day 5/6) with an average patient age of 35.3±4.9. Prior to vitrification, the junction between two trophectoderm cells in the blastocyst was located and one laser shot at 100% power with a 500ms pulse (Hamilton Thorne) was released. The High Security Vitrification Kit (CryoBioSystem, France) was used as a closed carrier, with a mixture of 15% Ethylene Glycol/Dimethyl Sulfoxide (v/v)+0.5M sucrose. Statistical significance was evaluated by Chi-Square. No significant difference in the survival between group A and B (98.5% vs. 98.5%) as well as in the mean number of embryos transferred was observed (1.8 vs. 1.8). However, group B had a significant higher clinical pregnancy and implantation rate than group A (63.6% vs. 45.0%; P<.001) and 49.0% vs. 33.0%; P<.001, respectively. The overall ongoing pregnancy in group A was significant lower than in group B (38.5% vs. 58.7%; P<.001). This study demonstrates that artificially collapsing of day 5/6 blastocysts prior to the steps of vitrification is beneficial for all outcome parameters including clinical and ongoing pregnancy rate as well as implantation rate. Using a laser, the procedure is fast, simple and precise, with the benefit of improvement in outcome.

ObjectiveThe culture of embryos has been approached by two culture protocols. The two-step protocol is thought to mimic the physiological interaction between specific developmental stages of the embryo within the oviduct and uterus. The opposing system in based on the ‘let the embryo choose’ principal that is analogous to a single-step culture medium. By culturing sibling oocytes in both media under the same culture conditions, we compared both groups with respect to outcome and implantation.DesignRetrospective analysis.MATERIALS AND METHODSA single-step medium (GLOBAL, IVFonline, Guilford, CT) was compared to a two-step media from SAGE (Trumbull, CT). Embryo culture was performed as group culture in 4-well dishes under oil and 5% oxygen. On day-1, following 2pn-check, sibling zygote cohorts from 181 patients (average age of 33.5±4.5) were randomly allocated into the SAGE or GLOBAL culture media. 953 embryos were cultured in the SAGE, whereas 1049 embryos were cultured in GLOBAL. 58 transfers from the SAGE system were compared to 123 transfers from the GLOBAL system. Clinical pregnancies showing FCA activity as determined by ultrasound. Chi square test was used for statistical analysis.RESULTSFrom both culture systems an average of 1.6±0.5 embryos were transferred. Almost 70% of the transfers (123/181) had embryos selected from the GLOBAL culture system. Overall, the total number of embryos cryopreserved and transferred was significantly higher in the GLOBAL group (57.3%; 601/1049; P<.04) compared to the SAGE group (47.6%; 454/953). Clinical pregnancy was slightly, but not significantly higher in SAGE (58.6%; 34/58; P<.92) than in GLOBAL (56.1%; 69/123). Furthermore, implantation was marginal, but not significant higher in SAGE (46.1%; 42/91; P<.75) compared to GLOBAL (42%; 82/196).CONCLUSIONThis trial demonstrates that embryos selected from a single-step media provide the same potential in achieving good clinical pregnancy and implantation rates compared to their sibling embryos cultured in a two-step media. ObjectiveThe culture of embryos has been approached by two culture protocols. The two-step protocol is thought to mimic the physiological interaction between specific developmental stages of the embryo within the oviduct and uterus. The opposing system in based on the ‘let the embryo choose’ principal that is analogous to a single-step culture medium. By culturing sibling oocytes in both media under the same culture conditions, we compared both groups with respect to outcome and implantation. The culture of embryos has been approached by two culture protocols. The two-step protocol is thought to mimic the physiological interaction between specific developmental stages of the embryo within the oviduct and uterus. The opposing system in based on the ‘let the embryo choose’ principal that is analogous to a single-step culture medium. By culturing sibling oocytes in both media under the same culture conditions, we compared both groups with respect to outcome and implantation. DesignRetrospective analysis. Retrospective analysis. MATERIALS AND METHODSA single-step medium (GLOBAL, IVFonline, Guilford, CT) was compared to a two-step media from SAGE (Trumbull, CT). Embryo culture was performed as group culture in 4-well dishes under oil and 5% oxygen. On day-1, following 2pn-check, sibling zygote cohorts from 181 patients (average age of 33.5±4.5) were randomly allocated into the SAGE or GLOBAL culture media. 953 embryos were cultured in the SAGE, whereas 1049 embryos were cultured in GLOBAL. 58 transfers from the SAGE system were compared to 123 transfers from the GLOBAL system. Clinical pregnancies showing FCA activity as determined by ultrasound. Chi square test was used for statistical analysis. A single-step medium (GLOBAL, IVFonline, Guilford, CT) was compared to a two-step media from SAGE (Trumbull, CT). Embryo culture was performed as group culture in 4-well dishes under oil and 5% oxygen. On day-1, following 2pn-check, sibling zygote cohorts from 181 patients (average age of 33.5±4.5) were randomly allocated into the SAGE or GLOBAL culture media. 953 embryos were cultured in the SAGE, whereas 1049 embryos were cultured in GLOBAL. 58 transfers from the SAGE system were compared to 123 transfers from the GLOBAL system. Clinical pregnancies showing FCA activity as determined by ultrasound. Chi square test was used for statistical analysis. RESULTSFrom both culture systems an average of 1.6±0.5 embryos were transferred. Almost 70% of the transfers (123/181) had embryos selected from the GLOBAL culture system. Overall, the total number of embryos cryopreserved and transferred was significantly higher in the GLOBAL group (57.3%; 601/1049; P<.04) compared to the SAGE group (47.6%; 454/953). Clinical pregnancy was slightly, but not significantly higher in SAGE (58.6%; 34/58; P<.92) than in GLOBAL (56.1%; 69/123). Furthermore, implantation was marginal, but not significant higher in SAGE (46.1%; 42/91; P<.75) compared to GLOBAL (42%; 82/196). From both culture systems an average of 1.6±0.5 embryos were transferred. Almost 70% of the transfers (123/181) had embryos selected from the GLOBAL culture system. Overall, the total number of embryos cryopreserved and transferred was significantly higher in the GLOBAL group (57.3%; 601/1049; P<.04) compared to the SAGE group (47.6%; 454/953). Clinical pregnancy was slightly, but not significantly higher in SAGE (58.6%; 34/58; P<.92) than in GLOBAL (56.1%; 69/123). Furthermore, implantation was marginal, but not significant higher in SAGE (46.1%; 42/91; P<.75) compared to GLOBAL (42%; 82/196). CONCLUSIONThis trial demonstrates that embryos selected from a single-step media provide the same potential in achieving good clinical pregnancy and implantation rates compared to their sibling embryos cultured in a two-step media. This trial demonstrates that embryos selected from a single-step media provide the same potential in achieving good clinical pregnancy and implantation rates compared to their sibling embryos cultured in a two-step media.