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Study Question: Do embryos with delayed blastulation have inferior reproductive potential or poorer outcomes due in part to embryo and endometrial synchrony?, Summary Answer: Diminished outcomes in embryos with delayed blastulation undergoing fresh embryo transfer (ET) may be attributed to a loss of embryonic-endometrial synchrony., What Is Known Already: Embryos that blastulate slowly have lower sustained implantation rates (SIR) than those which blastulate normally on Day 5 (D5). Traditionally this has been attributed to reduced embryo quality; however, dyssynchrony with the endometrium is also a possibility and has not been fully described. This convenient cohort composed of groups that resulted from a practice wide change in laboratory protocol allows for evaluation of embryo and endometrial synchrony as it related to blastocyst expansion., Study Design Size Duration: A retrospective cohort analysis was carried out from January 2009 to February 2013. Three cohorts were identified: D5 ET, D6 ET and frozen ET that comprised 822 patients, 763 patients and 718 patients, respectively. Each of these cohorts had slowly blastulating and normally blastulating embryos identified., Participants/materials Setting Methods: The study setting was academic affiliated private practice. All first fresh or cryopreserved ETs from 2010 to 2013 were studied. Non-biopsied embryos were classified into two groups on D5: slowly blastulating (Morula-Gardner 1) or normally blastulating (Gardner 2-6). Only single ETs or transfer of two embryos within the same classification group were included. Outcomes were compared between classification groups in embryos undergoing transfer on D5, D6, or after cryopreservation. This assesses the impact of transfer timing in fresh cycles as well as isolating a pure embryonic factor in frozen ET cycles. Sustained implantation was defined as heart beat detection at discharge sonogram at 8 weeks gestation. SIR was defined as the number of embryos transferred meeting criteria for sustained implantation divided by the total number of embryos transferred., Main Results and the Role of Chance: In total, 3391 embryos were transferred to 1966 patients. On D5, SIRs were significantly lower with slowly blastulating embryos (44% versus 64% in women <35 years of age ( P < 0.001) and 18% versus 56% in women ≥35 years of age ( P < 0.001)). Fresh D6 ETs also had significantly lower SIRs for embryos that were slowly blastulating on D5 (52% versus 63% in <35 years of age ( P < 0.05) and 32% versus 48% in ≥35 years of age ( P < 0.005)) despite continued development to full blastocysts and being morphologically equivalent at the time of ET, suggesting dyssynchrony. However, when slowly blastulating embryos underwent vitrification and then ET, they had SIRs which were equivalent to their normally blastulating counterparts (57% versus 60% in <35 years of age ( P = 0.5) and 37% versus 42% in ≥35 years of age ( P = 0.3)). An intraclass correlation and a generalized estimating equation corrected for patient age was performed which confirmed these findings. The normalization in cryopreserved ETs indicates that dyssynchrony may be a major adverse factor limiting outcomes with late blastulating embryos in fresh cycles., Limitations Reasons for Caution: This is a retrospective study comprising cohorts from a convenient sample chosen due to a uniform change in embryology laboratory protocol regarding ET day, however, this was done independent of the management of embryo and endometrial synchrony. Although strict ultrasound and serum progesterone criteria were utilized to make endometrial receptivity uniform, pathologic states of pre-receptive and post-receptive endometrium cannot be ruled out., Wider Implications of the Findings: The data surrounding embryo and endometrial synchrony should be considered in patients undergoing IVF and attention to the variations in blastulation rates can be applied to any patient undergoing extended embryo culture., Study Funding/competing Interests: None.

Objective: To determine whether sequential or monophasic media is the more optimal formulation for blastocyst development and sustained implantation rates (SIR) in IVF., Design: Paired randomized controlled trials., Setting: Academic., Patient(s): Infertile couples (N = 192) with female partner ≤42 years old and normal ovarian reserve., Intervention(s): Fertilized zygotes from each patient were randomly divided into two groups: [1] cultured in sequential media and [2] cultured in monophasic medium. Sequential media consisted of Quinn's Advantage Cleavage Medium (SAGE) followed by Blast Assist (Origio). The monophasic medium used was Continuous Single Culture (Irvine Scientific). Paired ETs were accomplished by transferring the best euploid blastocyst from each media group. DNA fingerprinting was used to link outcomes., Main Outcome Measure(s): The primary outcome measure was the proportion of blastocysts suitable for clinical use. Secondary outcome measures included timing of blastulation, aneuploidy rates, and SIR. Sustained implantation rate is defined as the number fetal heart beats at 8-9 weeks of gestation, divided by the number of embryos transferred., Result(s): A total of 192 patients had their 2PN embryos (N = 2,257) randomized to each culture system. Sequential media had higher blastulation rate than monophasic medium (55.2% vs. 46.9%). No differences were found in the day of blastulation or aneuploidy rate. Of the 168 patients who had euploid blastocysts suitable for transfer, 126 completed a paired ET. Among the double ETs, there was no difference in implantation between groups., Conclusion(s): This is the first randomized controlled trial to examine paired euploid transfers of sibling zygotes cultured in sequential versus monophasic media. This study demonstrates that the usable blastocyst rate is greatest after culture in the sequential media tested in comparison with the monophasic formulation selected for study. However, no difference exists in timing of blastulation, aneuploidy, or SIR. Whether these observations are generalizable to other media systems remains to be determined., Clinical Trial Registration Number: NCT01917240., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: To compare monozygotic twinning (MZT) rates in patients undergoing blastocyst or cleavage-stage ET., Design: Retrospective cohort., Setting: Academic research center., Patient(s): Autologous, fresh IVF cycles resulting in a clinical pregnancy from 1999 to 2014., Intervention(s): None., Main Outcome Measure(s): Monozygotic twin pregnancy in blastocyst-stage transfer vs. cleavage-stage transfer when controlling for patient prognosis and embryo cohort quality factors., Result(s): There were a total of 9,969 fresh transfer cycles resulting in a pregnancy during the study period. Of these pregnancies, 234 monozygotic twin pregnancies were identified (2.4%). Of all transfers, 5,191 were cleavage-stage and 4,778 were blastocyst-stage transfers. There were a total of 99 MZT identified in the cleavage-stage group (1.9%) and 135 MZT in the blastocyst ET group (2.4%), which was significant. Multivariable logistic regression revealed that increasing age was associated with a significant reduction in MZT, regardless of transfer order. Embryo cohort quality factors, including the number and proportion of six- to eight-cell embryos and availability of supernumerary embryos, were also significant. When controlling for patient age, time period during which the cycle took place, the number and proportion of six- to eight-cell embryos, and availability of supernumerary embryos, there was no longer a difference in MZT rate between blastocyst and cleavage transfer., Conclusion(s): Patient prognosis and embryo cohort quality seem to be major factors in MZT rate in women undergoing blastocyst transfer. Although technology-based effects cannot be excluded, patient and embryo characteristics play an important role., (Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Objective: To determine whether endometrial hCG infusion at the time of human blastocyst transfer impacts implantation rates., Design: Randomized double-blinded placebo-controlled trial., Setting: Academic., Patient(s): Infertile couples with the female partner less than 43 years old (n = 300) undergoing fresh or frozen ET of one or two blastocysts., Intervention(s): Patients undergoing ET were randomized into either a treatment or a control group. The treatment group received an infusion of 500 IU of hCG diluted in ET media. The control group received a sham infusion of ET media. Infusions were done using a separate catheter less than 3 minutes before actual ET., Main Outcome Measure(s): Sustained implantation rate: ongoing viable gestation (primary outcome) and ongoing pregnancy rate (secondary outcome)., Result(s): A total of 473 blastocysts were transferred into 300 patients. There were no differences between the two groups in sustained implantation rate (48.1% in the hCG group, 44.2% in the control group) or ongoing pregnancy rate (58.8% in the hCG group, 52.0% in the control group)., Conclusion(s): Endometrial infusion of hCG at the time of blastocyst ET does not improve sustained implantation rates., Clinical Trial Registration Number: NCT01643993., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: To characterize each chromosome's risk for being involved in embryonic aneuploidy., Methods: This is a retrospective cohort study conducted at a single, academic center. The cohort consisted of 15,169 consecutive trophectoderm biopsies which then underwent comprehensive chromosome screening utilizing validated real-time polymerase chain reaction (RT-PCR) or single nucleotide polymosphism (SNP) array platforms. Analysis was done to determine probability of aneuploidy by chromosome, changes in that risk with increasing maternal age, and in relationship of aneuploidy to chromosomal structure as classified by prior cytogenetic literature., Results: The highest prevalence of imbalances leading to aneuploidy was seen for chromosomes 13, 15, 16, 18, 19, 21, and 22. While elevated in all age groups, there was a disproportionate rise in aneuploidy rates for these chromosomes with increasing maternal age. When classic cytogenetic karyotype groups were compared, the overall smaller groups D, E, and G were associated with the highest rates. Similarly, when grouped based upon structure, acrocentric chromosomes exhibited the highest rates of aneuploidy, followed by the metacentric chromosomes, with the lowest prevalence of error in those with submetacentric structures., Conclusions: The highest rates of chromosomal aneuploidy were found in chromosomes known to be involved in clinically detectable, abnormal pregnancies, not just simply implantation failure. The rate of aneuploidy in these chromosomes rises disproportionately with age when compared to the other chromosomes which may provide information about chromosomal susceptibility to aging. The biological structure groupings did show varied aneuploidy rates which may provide insight into the biology of aneuploidy.

Objective: To determine whether culture at a more physiologically cooler temperature, as suggested by limited human and animal data, would improve blastulation and pregnancy rates in human clinical IVF., Design: Paired randomized controlled trial., Setting: Academic., Patient(s): Infertile couples (n=52) with a female partner less than 42 years old with eight or more mature oocytes retrieved., Intervention(s): Mature oocytes obtained from a single cohort of oocytes were randomly divided into two groups; one was cultured at 37°C and the other at 36°C from the time of ICSI to the time of embryo transfer or vitrification. Paired embryo transfers were accomplished by transferring one euploid embryo from each group. DNA fingerprinting was used as needed to determine the outcome for each embryo., Main Outcome Measure(s): Rate of development of expanded blastocysts suitable for transfer or vitrification (primary outcome), fertilization, aneuploidy, and sustained implantation., Result(s): A total of 805 mature oocytes were cultured; 399 at 36°C and 406 at 37°C. Paired analysis demonstrated a higher rate of usable blastocyst formation per zygote at 37°C (48.4%) vs. at 36°C (41.2%). Rates of fertilization, aneuploidy, and sustained implantation were equivalent., Conclusion(s): IVF culture at 36°C does not improve clinically relevant parameters of embryo development or sustained implantation rates., Clinical Trial Registration Number: NCT01506089., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: To determine if comprehensive embryology training for clinical Reproductive Endocrinology fellows could be completed to a level of proficiency equivalent to that of experienced embryologists., Method: Clinical fellows were integrated into the clinical embryology team and were trained to perform all the various procedures utilized in clinical embryology. The fellows were trained to the same standards as the clinical embryology staff and underwent the same certification and sign off procedures. To determine if inclusion of clinical fellows on the embryology team impacted outcomes, outcomes for individual oocytes/embryos and the clinical cases where the fellows perform embryology procedures were compared to the outcomes of those oocytes/embryos and cases performed by the full time embryology staff., Results: Clinical procedures performed by the fellows included isolation and processing of oocytes following retrieval, loading catheters for embryo transfer, and vitrification (N = 823 cases). Micromanipulation procedures compared included ICSI and assisted hatching (N = 650 cases). For each procedure, the outcomes in those cases performed by the RE fellows were equivalent to those done by the fully trained clinical embryology staff., Conclusions: When fellows are trained to perform embryology procedures as an integral part of their fellowship curricula, laboratory efficiencies and clinical outcomes are fully maintained. This experience provides valuable insight into the ART process critical to this subspecialty. It also empowers fellows to fully participate in research relating to the viability of gamete and embryos and optimization of the clinical ART laboratory.

Objective: To determine the relationship between the age of the female partner and the prevalence and nature of human embryonic aneuploidy., Design: Retrospective., Setting: Academic., Patient(s): Trophectoderm biopsies., Intervention(s): Comprehensive chromosomal screening performed on patients with blastocysts available for biopsy., Main Outcome Measure(s): Evaluation of the impact of maternal age on the prevalence of aneuploidy, the probability of having no euploid embryos within a cohort, the complexity of aneuploidy as gauged by the number of aneuploid chromosomes, and the trisomy/monosomy ratio., Result(s): Aneuploidy increased predictably after 26 years of age. A slightly increased prevalence was noted at younger ages, with >40% aneuploidy in women 23 years and under. The no euploid embryo rate was lowest (2% to 6%) in women aged 26 to 37, was 33% at age 42, and was 53% at age 44. Among the biopsies with aneuploidy, 64% involved a single chromosome, 20% two chromosomes, and 16% three chromosomes, with the proportion of more complex aneuploidy increasing with age. Finally, the trisomy/monosomy ratio approximated 1 and increased minimally with age., Conclusion(s): The lowest risk for embryonic aneuploidy was between ages 26 and 30. Both younger and older age groups had higher rates of aneuploidy and an increased risk for more complex aneuploidies. The overall risk did not measurably change after age 43. Trisomies and monosomies are equally prevalent., (Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Objective: To determine if cleavage- or blastocyst-stage embryo biopsy affects reproductive competence., Design: Paired randomized clinical trial., Setting: Academic-assisted reproduction program., Patient(s): Attempting conception through IVF., Intervention(s): After selecting two embryos for transfer, one was randomized to biopsy and the other to control. Both were transferred within shortly thereafter. The biopsy was submitted for microarray analysis and single-nucleotide polymorphism (SNP) profiling. Buccal DNA obtained from the neonate after delivery had microarray analysis and SNP profile compared with that of the embryonic DNA. A match confirmed that the biopsied embryo implanted and developed to term, whereas a nonmatch indicated that the control embryo had led to the delivery., Main Outcome Measure(s): Paired analysis of the delivery rates of the transferred embryos. Either twin delivery or failure to deliver represents equivalent outcomes for the biopsied and control embryos. In contrast, singletons were determined to be from the biopsied or the control embryo., Result(s): Blastomere biopsy on day 3 of development resulted in a significant reduction in sustained implantation. Only 30% of biopsied embryos had sustained implantation and ultimately developed into live-born infants versus 50% of unbiopsied controls, a relative reduction of 39%. In contrast, sustained implantation rates were equivalent (51% vs. 54%) for biopsied and control blastocysts., Conclusion(s): Cleavage-stage biopsy markedly reduced embryonic reproductive potential. In contrast, trophectoderm biopsy had no measurable impact and may be used safely when embryo biopsy is indicated., Clinical Trial Registration Number: NCT01219504., (Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Objective: To determine how often trophectoderm biopsy and rapid, real-time quantitative polymerase chain reaction (PCR)-based comprehensive chromosome screening (CCS) alters clinical management by resulting in the transfer of a different embryo than would have been chosen by traditional day 5 morphology-based criteria., Design: Prospective., Setting: Academic center for reproductive medicine., Patient(s): Infertile couples (n = 100; mean age 35 ± 4 years) with at least two blastocysts suitable for biopsy on day 5., Intervention(s): Prior to trophectoderm biopsy for CCS the embryologist identified which embryo would have been selected for traditional day 5 elective single ET., Main Outcome Measure(s): The risk of aneuploidy in the embryos that would have been selected on day 5 was calculated and compared with the aneuploidy rate of the cohort of all embryos that underwent CCS testing. The aneuploidy risk was compared between age groups., Result(s): After quantitative PCR-based CCS, 22% (95% confidence interval 15%-31%) of the embryos selected by day 5 morphology were aneuploid, which was lower than the 32% aneuploidy rate of the cohort. Patients ≥35 years had a higher risk of an aneuploid blastocyst being selected by morphology than those <35 years old (31% vs. 14%). Among patients who had selection altered by CCS, 74% (14/19) delivered, including 77% (10/13) after elective single ET. Most patients (77%) had an additional euploid blastocyst vitrified for future use., Conclusion(s): The CCS results alter embryo selection due to the presence of aneuploidy in embryos with optimal day 5 morphology. Excellent outcomes were obtained when CCS-based selection was different than morphology-based selection., (Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Objective: To determine whether blastocyst biopsy and rapid quantitative real-time polymerase chain reaction (qPCR)-based comprehensive chromosome screening (CCS) improves in vitro fertilization (IVF) implantation and delivery rates., Design: Randomized controlled trial., Setting: Academic reproductive medicine center., Patient(s): Infertile couples in whom the female partner (or oocyte donor) is between the ages of 21 and 42 years who are attempting conception through IVF., Intervention(s): Embryonic aneuploidy screening., Main Outcome Measure(s): Sustained implantation and delivery rates., Result(s): We transferred 134 blastocysts to 72 patients in the study (CCS) group and 163 blastocysts to 83 patients in the routine care (control) group. Sustained implantation rates (probability that an embryo will implant and progress to delivery) were statistically significantly higher in the CCS group (89 of 134; 66.4%) compared with those from the control group (78 of 163; 47.9%). Delivery rates per cycle were also statistically significantly higher in the CCS group. Sixty one of 72 treatment cycles using CCS led to delivery (84.7%), and 56 of 83 (67.5%) control cycles ultimately delivered. Outcomes were excellent in both groups, but use of CCS clearly improved patient outcomes., Conclusion(s): Blastocyst biopsy with rapid qPCR-based comprehensive chromosomal screening results in statistically significantly improved IVF outcomes, as evidenced by meaningful increases in sustained implantation and delivery rates., Clinical Trial Registration Number: NCT01219283., (Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)