Cleavage Stage Embryo Biopsy Research Articles

P-773 No impact of cleavage-stage or blastocyst-stage embryo biopsy on growth and health in children up to 2 years of age

Abstract Study question Does embryo biopsy for preimplantation genetic testing (PGT) followed by fresh or vitrified-warmed embryo transfer affect children’s health up to 2 years of age? Summary answer Cleavage- or blastocyst-stage embryo biopsy followed by fresh or vitrified-warmed-embryo transfer had neither impact on anthropometry at birth, infancy or childhood nor on health outcomes. What is known already Literature data regarding neonatal outcomes after embryo biopsy for PGT show mixed results due to small sample sizes and/or the heterogeneity in terms of embryo biopsy (cleavage- or blastocyst-stage) and type of embryo transfer (fresh or frozen-thawed). Even fewer data exist on the impact of embryo biopsy on children’s health beyond infancy, including growth. Study design, size, duration This single-center cohort-study compared outcomes in singletons conceived after cleavage- or blastocyst-stage embryo biopsy either in a fresh or vitrified-warmed transfer cycle with results after non-biopsied embryo transfer between 2014 and 2018. Pregnancies after IVM, oocyte vitrification or oocyte/embryo donation were excluded. Eligible singletons living in Belgium were invited for examination at 3-6 months and 2 years. Anthropometric measurements at birth, infancy and childhood and health outcomes including surgeries, medication use and hospitalisations are reported. Participants/materials, setting, methods Birth characteristics were available for 630 and 222 children after cleavage-stage and blastocyst-stage embryo biopsy and for 1532 children after transfer of a non-biopsied embryo. Follow-up data were available for 426, 131 and 662 children, respectively. The impact on children’s health following embryo biopsy in vitrification and fresh cycles was the primary outcome. Other outcomes were the impact of timing of biopsy and of vitrification. Subgroup analysis according to infertility background was additionally performed. Main results and the role of chance Regarding the impact of embryo biopsy, either at the cleavage or blastocyst stage, no differences in anthropometrics were found at birth, infancy or childhood in vitrified-warmed transfer cycles compared to outcomes in non-biopsied vitrified-warmed transfer cycles, even after adjustment for neonatal, treatment and maternal characteristics. Likewise, no impact of embryo biopsy (cleavage stage) was found in fresh transfer cycles. The timing (day 3 or day 5/6) of embryo biopsy did not affect anthropometrics at birth, infancy or childhood. However, children born after cleavage-stage embryo biopsy followed by a vitrified-warmed transfer cycle had larger birth sizes than children born after cleavage-stage biopsy followed by a fresh transfer. Weight and height gain from birth to infancy and from infancy to early childhood were comparable in all biopsied and non-biopsied groups. Reassuringly, comparable rates of major congenital malformations, (severe) developmental problems, hospital admissions, surgical interventions and of chronic medication use up to the age of 2 years were observed in biopsied and non-biopsied groups. Subgroup analysis in children born to parents with an infertility diagnosis showed that birth parameters were not different in children born after embryo biopsy. Limitations, reasons for caution As the majority of the PGT cycles in our center are performed for a genetic indication only, the number of children born after embryo biopsy in infertile parents is rather limited and the results should therefore be interpreted cautiously. Wider implications of the findings Our findings add reassurance that embryo biopsy does not adversely affect the health of offspring up to 2 years of age. This is of particular importance given the widespread shift to blastocyst-stage embryo biopsy in PGT. Trial registration number Not applicable

Pre-implantation genetic testing: Past, present, future.

BackgroundPre‐implantation genetic testing (PGT) has been performed worldwide since it was first used by Handyside et al in the United Kingdom to sex embryos in 1990. Until about 2010, cleavage stage embryo biopsy and fluorescent in situ hybridization (FISH) were mainstream; however, in 2012, blastocyst biopsy (trophectoderm; TE biopsy) became mainstream. In addition, array comparative genomic hybridization (aCGH) was used for analysis and further evolved to next‐generation sequencing (NGS), which is used worldwide.MethodsPGT for reciprocal balanced translocation and Robertsonian translocation (PGT‐SR) was approved in Japan for habitual abortion to reduce pregnancy loss, and since 2008, we have been performing PGT‐SR using cleavage stage embryos and FISH. In 2014, we performed TE biopsy and NGS analysis.Main findingsIn this paper, I separately described the details of our methods and clinical results of FISH and NGS. NGS is superior to FISH because it can detect all chromosomes.ConclusionTE biopsy and NGS, which have recently become mainstream, have stable outcomes, because TE biopsy yields more cells and fewer mosaics than the cleavage stage. As a result, diagnoses are more reliable, resulting in higher pregnancy rates and lower abortion rates.

Blastocyst and cleavage stage embryo biopsy for preimplantation genetic testing of the sickle cell gene in carrier couples: the experience of an IVF clinic in a developing country: a retrospective study

Introduction: Embryo biopsy is a prerequisite for preimplantation genetic testing (PGT). Although cleavage stage biopsy is very common, trophectoderm biopsy at blastocyst stage has become increasingly popular in recent times. This study describes the clinical success of PGT cycles for sickle cell anemia using both cleavage and trophectoderm biopsy in a developing country. Materials and Methods: All patients undergoing in vitro fertilization and PGT for sickle cell anemia from April 2011 to February 2017. Embryos were biopsied either on day 3 (blastomere) or day 5/6 (trophectoderm). Laser pulses (ZILOS-tk Laser) perforating the zona pellucida were followed by either blastomere aspiration from a day 3 cleavage stage embryo or trophectoderm biospy from a day 5/6 blastocyst. Embryos were vitrified awaiting subsequent thaw and transfer. After excluding homozygous hemoglobin SS embryos, frequencies of positive human chorionic gonadotrophin, clinical pregnancy, implantation rate and live birth rate were recorded for day 3 cleavage stage embryos (group A) and day 5/6 blastocysts (group B). Results: Of the 34 patients undergoing in vitro fertilization PGT for sickle cell anemia, embryos from 18 underwent day 3 blastomere aspiration (group A) whereas embryos from 16 underwent day 5/6 trophectoderm biopsies (group B). The mean patient age was 34.4 years for group A and 34.1 years for group B. A total of 131 embryos were biopsied in group A and 106 in group B. Percentages of unaffected embryos (ie, HB AA and AS) in groups A and B were 40.4% and 68.0%, respectively. Positive human chorionic gonadotrophin rates were 7.7% and 60%, clinical pregnancy rates 7.7% and 20%, implantation rates 3.7% and 32.1%, and live birth rates 3.7% and 20%, respectively. Conclusions: In this developing country, the use of trophectoderm biopsy for interrogating embryos at risk for sickle cell anemia appeared superior to blastomere aspiration at the cleavage stage for the purpose of PGT.

The interval transfer of a frozen-thawed embryo is more successful than a fresh embryo transfer for women undergoing IVF with recurrent implantation failure after cleavage stage embryo biopsy.

Recurrent implantation failure (RIF) is repeated unsuccessful embryo transfers (ETs). To identify predictive embryonic markers of implantation in RIF, following pre-implantation genetic screening (PGS) of cleavage stage embryos, after accounting for male and female factors. Retrospective analysis of RIF patients undergoing PGS after correction of modifiable causes. Eighty-four patients underwent 140 in vitro ferilisation cycles. Forty-one cycles were excluded: 12 (no embryo for transfer), four (double ETs) and 25 (no biopsy). Sixty-three patients underwent 99 single euploid ETs (48 fresh, 51 frozen) resulting in 11 biochemical pregnancies, 36 clinical pregnancies (CP), and six miscarriages and 30 live births (LB). Frozen ET was more successful than fresh; respective live birth rate (LBR) and clinical pregnancy rate (CPR), 39.2% versus 20.8%, (P=0.02), 45.1% versus 27.1% (P=0.04). LBR and CPR were lower when 5-6 blastomeres were present at embryo biopsy, compared to embryos with ≥7 blastomeres: 15.4% versus 32.6% (P=0.185) and 15.4% versus 39.5% (P=0.074) respectively. Serum β human chorionic gonadotropin (βhCG) concentration was greater when a more developed embryo was biopsied (r=0.448, P=0.017 and r=0.476, P=0.118, fresh and frozen transfers, respectively). Embryo morphokinetic analysis demonstrated faster development to blastocyst stage when more cells were present at biopsy: mean 103.3, 102.2 and 96.0h for biopsy at the 5-6, 7-8 or ≥9 cell stage respectively (P=0.040 for difference between 7-8 cells vs ≥9). After cleavage stage biopsy, frozen ET was more successful than fresh ET. Chance of conception and serum βhCG concentration correlated with number of cells present at time of biopsy.

Automatic segmentation of zona pellucida and its application in cleavage-stage embryo biopsy position selection.

A very important step of Pre-implantation genetic diagnosis (PGD) is embryo biopsy, in which process the zona pellucida (ZP) is cut open partially and a part of cellular material is extracted from the embryo. Recognition of the ZP is necessary not only for embryo biopsy, but also for other applications such as zona pellucida thickness variation (ZPTV), embryo dissection, etc. The ZP opening position is closely related to the cell survival rate after the biopsy. Selection of an unsuitable position may cause blastomere lysis after the ZP opening. Normal procedures of ZP recognition and biopsy position selection involve a skilled human embryologist. In order to make the process automatic, we introduce an automatic segmentation method for ZP recognition by using edge detection and ellipse fitting with a value adjustment algorithm in this paper. An application of ZP recognition in embryo biopsy position selection is also introduced. Our ZP recognition algorithm was able to correctly segment 43 out of 45 sample embryo images, achieving a success rate of 96%. Its application in embryo biopsy position selection achieved a success rate of 93%.

Blastocysts Re-analysis after Cleavage Stage Biopsy with 24 Chromosome Array Comparative Genomic Hybridization (CGH). How Reliable Is the PGS Report?

With improvements in culture conditions, blastocyst development rates have increased. It is clear that some embryos diagnosed as aneuploid from cleavage stage biopsy will progress to blastocysts following extended culture. The objective of this prospective study was to determine the accuracy of the diagnosis of aneuploidy after cleavage stage embryo biopsy using CGH with confirmatory re-biopsy at the blastocyst stage.

Does First Polar Body Array CGH Analysis Correlate with a Cleavage Stage Embryo Biopsy in IVF-ICSI Cycles and Preimplantation Genetic Screening

To correlate first polar body(PB1) biopsy array comparative genomic hybridization(aCGH) technology with cleavage stage(D3) biopsy results analyzed with either FISH or SNP array. Prospective clinical trial. 15 IVF patients underwent PB1 biopsy followed by a D3 biopsy. Between 09/11-04/12, 27 patients consented and 15 met the study's criteria and underwent PB1 biopsy and D3 biopsy. 11 patients completed their cycle with a total of 101 PB1 and corresponding D3 biopsies. All mature metaphase II (MII) oocytes were fertilized by ICSI 4-6 hours following oocyte retrieval. 2-4 hours later, PB1 biopsies were performed and frozen. D3 biopsy was then performed and analyzed using either 9-probe FISH(13,15,16,17,18,21,22,X,Y) or SNP 24 CGH(physician/patient preference). Clinical decisions were based solely on the results of D3 analysis.ET was performed on day-5. Concordance between PB1 and D3 results, as well as ongoing pregnancy rates and IVF cycle outcome. 83% of MII oocytes fertilized normally using ICSI. Concordance analysis of the first 96(5 embryos had incomplete results) PB1 and corresponding D3 revealed 67.33% partial or complete concordance and 32.7% discordance. When the 9-probes FISH was used partial concordance was expected. PB1 aCGH had a sensitivity of 82.8% and specificity of 55.7% compared to D3. The kappa statistics was 0.35. Implantation rate was 31% (average 2.3 embryos per transfer). 36% pregnancy rate per cycle started and 27% ongoing pregnancy rate (57% and 42%, respectively, in patients who had ET).Tabled 1PB1 biopsyBlastomere biopsy%NormalNormal28.71AbnormalAbnormal - identical6.93AbnormalAbnormal - partial concordance8.91AbnormalAbnormal - different17.82NormalAbnormal5.94AbnormalNormal26.73N/A4.95100%PB1-Clinically correct diagnosis67.33%PB1-Clinically incorrect diagnosis32.67% Open table in a new tab Tabled 1D3 biopsy + (Normal)D3 biopsy - (Abnormal)Polar body + (Normal)292756Polar body - (Abnormal)63440356196KAPPA = 0.35 (Fair agreement) with C.I. (0.14, 0.54) p value = 0.0001 Open table in a new tab KAPPA = 0.35 (Fair agreement) with C.I. (0.14, 0.54) p value = 0.0001 PB1 biopsy can not completely replace embryo biopsy for PGS as it doesn't appraise 2nd meiosis error and mitotic errors. Our study suggests, that PB1 aCGH can accurately predict oocyte chromosomal status in 2/3 of cases and completed within 12 hours. Aneuploidy(partly due to mosaicism) was the predominant cause of non-viability in our study group. PB1 biopsy should be used in certain clinical scenarios and selected patients.

Does first polar body array CGH analysis correlate with a cleavage stage embryo biopsy in IVF-ICSI cycles and preimplantation genetic screening

OBJECTIVE: To correlate first polar body(PB1) biopsy(Bx) aCGH technology with cleavage stage(D3) Bx results analyzed with FISH or SNP-array. DESIGN: Prospective clinical trial. Main Outcome: Concordance between PB1 and D3 results, ongoing pregnancy rates and IVF outcome. MATERIALS AND METHODS: Between 09/11-04/12, 27 patients(Pts) consented and 15 enrolled and underwent PB1 Bx followed by a D3 Bx.11 Pts completed their cycle with a total of 101 PB1 and corresponding D3 Bxs.All mature oocytes(MII) were fertilized by ICSI 4-6h post retrieval with PB1 Bx performed and frozen 2-4h thereafter.D3 Bx was then performed and analyzed using 9 probe FISH(13,15,16,17,18,21,22,X,Y) or 24 chromosomes SNP array(physician/Pts preference).Clinical decisions were based solely on D3 analysis.ET was performed on D5. RESULTS: 83% of MIIs fertilized normally.Concordance analysis of PB1 and corresponding D3 revealed 67.3% partial/complete concordance and 32.7% discordance(6% normal embryos possibly discarded or 26.7% abnormal embryos possibly transferred).When 9-probe FISH was used partial concordance was expected.PB1 had a 82% sensitivity and 55% specificity when compared to D3.Kappa statistic was 0.35[fair agreement], P1⁄40.0001(95% CI1⁄40.14,0.54) Implantation rate was 31%(average of 2.28 fresh embryos per ET). Pregnancy rate/cycle started was 36% with 27% ongoing(57% and 42% per cycle transferred).

Cleavage stage embryo biopsy significantly impairs embryonic reproductive potential while blastocyst biopsy does not: a novel paired analysis of cotransferred biopsied and non-biopsied sibling embryos

Recent advances in molecular genetics provide the opportunity to more accurately assess the reproductive potential of embryos. These technologies require embryonic DNA which necessitates embryo biopsy. Amongst the most critical questions surrounding these technologies are the risks associated with embryo biopsy itself, and whether biopsy is safest at the cleavage or blastocyst stage. This study uses a novel paired design to accurately assess the impact of biopsy on day 3 and day 5 of development. Paired randomized controlled trial. Patients received routine care inclusive of embryo selection. Once the 2 best embryos were selected for transfer, one was randomly selected for biopsy and the other to control. The embryos were then transferred together. With resulting pregnancies, DNA fingerprinting of the conceptus was used to determine whether it was derived from the biopsied (self) or non-biopsied (sibling) embryo. Analysis was done using a paired test of proportions. 103 patients were studied - 32 at the cleavage stage and 71 at the blastocyst stage. There was a dramatic reduction in implantation rates amongst biopsied cleavage stage embryos compared to the non-biopsied controls (31% vs 53%;P=0.035) - a loss of reproductive potential of more than one third. In contrast, the implantation rate of biopsied and non-biopsied blastocysts was equivalent (52% vs 54%; P=0.80). Trophectoderm (TE) biopsy of blastocysts does not adversely impact embryonic reproductive potential. In contrast, biopsy at the cleavage stage was clearly detrimental with a 22% reduction in implantation rate. Optimizing clinical outcomes following PGD might be best realized by performing biopsies at the blastocyst stage. Potential explanations as to why blastocyst biopsy is safe when cleavage stage biopsy is not include the removal of a smaller proportion of the embryo, certainty that only TE cells are biopsied, and better embryonic tolerance of manipulation after genomic activation.

Genetic Normalization of Differentiating Aneuploid Human Embryos

AbstractEarly embryogenesis involves a series of dynamic processes, many of which are currently not well described or understood. Aneuploidy and aneuploid mosaicism, a mixture of aneuploid and euploid cells within one embryo, in early embryonic development are principal causes of developmental failure.^1,2^ Here we show that human embryos demonstrate a significant rate of genetic correction of aneuploidy, or "genetic normalization" when cultured from the cleavage stage on day 3 (Cleavage) to the blastocyst stage on day 5 (Blastocyst) using routine in vitro fertilization (IVF) laboratory conditions. One hundred and twenty-six human Cleavage stage embryos were evaluated for clinically indicated preimplantation genetic screening (PGS). Sixty-four of these embryos were found to be aneuploid following Cleavage stage embryo biopsy and single nucleotide polymorphism (SNP) 23 chromosome molecular karyotype (microarray). Of these, 25 survived to the Blastocyst stage of development and repeat microarray evaluation was performed. The inner cell mass (ICM), containing cells destined to form the fetus, and the trophectoderm (TE), containing cells destined to form the placenta were evaluated. Sixteen of 25 embryos (64%) [95% CI: 44-80%] possessed diploid karyotypes in both the ICM and TE cell populations. An additional three Blastocyst stage embryos showed genetic correction of the TE but not the ICM and one Blastocyst stage embryo showed the reverse. Mosaicism (exceeding 5%), was not detected in any of the ICM and TE samples analyzed. Recognizing that genetic normalization may occur in developing human embryos has important implications for stem cell biology, preimplantation and developmental genetics, embryology, and reproductive medicine.1)Hassold, T. et al. A cytogenetic study of 1000 spontaneous abortions. Ann Hum Genet. 44, 151-78 (1980).2)Menasha, J., Levy, B., Hirschhorn, K., & Kardon, N.B. Incidence and spectrum of chromosome abnormalities in spontaneous abortions: new insights from a 12-year study. Genet Med. 7, 251-63 (2005).

Trophectoderm Cells Derived From Blastocyst Biopsy Are Suitable For Array CGH Analysis of 24 Chromosomes

BACKGROUND: Recent studies raise concern that use of blastomeres derived from cleavage stage embryo biopsy may compromise PGD results due to high incidence of mosacism. Use of trophectoderm cells derived from blastocyst biopsy may become the choice for chromosomal screening with array CGH.

Impact of cleavage-stage embryo biopsy in view of PGD on human blastocyst implantation: a prospective cohort of single embryo transfers

Human embryo biopsy is performed for preimplantation genetic diagnosis (PGD). The impact of 1- or 2-cell removal at cleavage-stage on future embryo development and implantation capacity is highly debated. In order to explore this issue further, a cohort of Day 5 single embryo transfers was analysed prospectively for embryological and clinical outcome. All transferred embryos resulted from 8-cell embryos on Day 3, from which subsequently either one cell (group I, n = 182) or two cells (group II, n = 259) were removed, or on which no invasion by means of embryo biopsy was performed (group III, control group, n = 702). RESULTS Blastocyst formation was significantly better in group III compared with group II, and similar to group I. Group I and group II did not differ in Day 3 nor in Day 5 embryo development. The overall live birth rate was significantly higher in group I (37.4%, CI 29.0-47.4%) than in group II (22.4%, CI 17.0-28.9%), and comparable to the reference ICSI population (35.0%, CI 30.8-39.7%). The clinical outcome of 1-cell biopsy was significantly better than that of 2-cell biopsy, even when adjusted for availability of genetically transferable embryos.

Apoptosis is not induced by slow cryopreservation or vitrification in biopsed cleavage stage embryos

OBJECTIVE: Cleavage stage embryo biopsy makes embryos more vulnerable to development failure. Cryopreserved biopsied embryos are at a higher risk for developmental or implantation failure. The aim of our study was to 1) examine if apoptosis is induced in cleavage stage embryos after biopsy and 2) evaluate the extent of apoptosis following vitrification or slow cryopreservation of biopsied cleavage stage embryos.

Pregnancies and live births after trophectoderm biopsy and preimplantation genetic testing of human blastocysts

To compare multiple-cell trophectoderm biopsy for preimplantation genetic diagnosis (PGD) from day-5 blastocysts with previously published experience with day-3 cleavage-stage embryos. Retrospective review of laboratory and clinical experience. Sydney IVF, a private clinic in Australia. Preimplantation genetic diagnosis (PGD) patients age < 44 years with at least one IVF blastocyst suitable for biopsy, recruited from January 2002 through August 2004. Biopsy of trophectoderm from blastocysts on day 5 or 6, with same-day PGD for mutation testing, translocation testing, aneuploidy screening or sex selection. Spare, normal biopsied blastocysts were cryostored for possible later transfer. Fetal heart-positive pregnancy rate and accumulating live birth rate after adding results from biopsied fresh and frozen blastocysts for particular couples. In 231 started PGD treatment cycles, unambiguous results were obtained from 974 of 1,050 biopsied blastocysts (93%); all blastocysts survived the biopsy procedure by reconstitution of their blastocele. One hundred nineteen women (median age, 36 years) have had 127 blastocysts transferred fresh (fetal heart-positive implantation rate, 41%). Of 146 blastocysts cryostored, 27 have been thawed (all with > 50% cell survival) and 24 transferred (implantation rate, 26%). To date, 53 pregnancies have been delivered or are ongoing, with an additional 4 clinical miscarriages (7%) and 6 subclinical miscarriages (total miscarriage rate, including biochemical pregnancies, 16%). There were no twin pregnancies. With technically appropriate blastocyst culture and freezing, blastocyst biopsy and cryostorage and later transfer of biopsied blastocysts is shown to be a practical and probably preferable path to preimplantation genetic testing of embryos compared with cleavage-stage embryo biopsy, being accompanied by a high implantation rate (and hence more conducive to elective single embryo transfer) and by a low rate of twinning and miscarriage.

Specific detection of deleted and non-deleted dystrophin exons together with gender assignment in preimplantation genetic diagnosis of Duchenne muscular dystrophy.

We have developed a preimplantation genetic diagnosis (PGD) strategy for Duchenne muscular dystrophy (DMD) allowing the simultaneous amplification of four exons (6, 8, 28 and 32) of the dystrophin gene together with ZFX/ZFY genes for gender determination. Preliminary experiments were carried out on 215 single lymphocytes from male and female individuals. Amplification rates ranged from 90.2% for exon 6 to 96.7% for exons 8 and 32. At least four of the five sequences were successfully amplified in 95.8% of single cells, and sexing was possible in 98.5%. This 5-plex assay was found to be robust enough to be used in a PGD clinical procedure and was therefore applied to a family whose female partner was a heterozygous carrier of a large deletion extending from exon 21 to exon 34 of the dystrophin gene. We have thus analysed two exons located in the deleted region of the gene, two non-deleted exons used as intrasample controls, and ZFX/ZFY genes. Cleavage stage embryo biopsy followed by PCR resulted in transfer of three unaffected embryos. The advantage of the present approach is to identify and subsequently transfer unaffected male embryos in addition to female embryos, and is now applicable to all families displaying a deletion involving at least one of these exons.

Diagnosis failure in preimplantation genetic diagnosis (PGD): poor technique or bad embryos?

Objective: To investigate the causes of diagnosis failure observed in the course of PGD. Design: In the course of 41 PGD cycles performed by the Béclère-Necker hospitals in Paris within a one year period, we correlated the efficiency of diagnosis with embryo grades and their developmental potential. Materials/Methods: We started our newly created PGD program in November 1999 and have performed 41 cases in just over a one year period. Patients were hyperstimulated using GnRh agonist and Recombinant FSH. The embryos were cultured in IVF (MediCult) until day 2 and placed in M3 (MediCult) until day 3. One or two cells were removed from each embryos by cleavage stage embryo biopsy on day 3 using a fertilase laser for zona drilling in G2.2 medium (JCD). The embryos were then placed in G2.2 until the transfer procedure on day 4. PGD was performed by PCR (23 cases) for a range of single gene defects or by FISH for X-linked recessive diseases (5 cycles) and translocations (13 cycles). Embryo grades on day 3 and signs of compaction on day 4 were recorded for each embryo and correlated with the availability of diagnosis. Grade 1 embryos had cells of even size with no or little fragmentation. Grade 2 had cells of even size with less than 20% fragments and grade 3 were highly fragmented embryos associated or not with cells of different size. Statistical significance of the results was calculated using a χ2 test. Results: Overall, a diagnosis could be obtained for 187 of the 237 embryos analysed (79%). From the 50 embryos for which no diagnosis could be obtained, 40 were of grade 3 (80%) on day 3 and only 7 showed signs of compaction (14%) on day 4. These data were significantly different from those obtained from the group of embryos for which a diagnosis was obtained in which 40% were of grade 3 and 40% subsequently compacted. Embryo grade on day 3 and signs of compaction on day 4 depending on whether a diagnosis was obtained. ∗TotalNumber of embryos on day 3 (%)Number of embryos on day 4 (%)Grade 1Grade 2Grade 3CompactedArrestedDiagnosis18741 (22)71 (38)75 (40)∗p < 0.001.75 (40)∗p < 0.001.112 (60)No diagnosis503 (6)7 (14)40 (80) ∗p < 0.001.7 (14) ∗p < 0.001.43 (86)Total23744 (19)78 (33)115 (48)82 (35)155 (65)∗ p < 0.001. Open table in a new tab Conclusions: No diagnosis could be obtained from 21% of the embryos biopsied, however our study demonstrates that most of these embryos had a poor morphology and a poor developmental potential. We can speculate that the blastomeres analysed from theses embryos were arrested with degraded DNA which was not longer a suitable target for PCR or FISH analysis. We conclude that most untyped embryos would not have been good candidates for transfer. Supported by: This work was supported in part by the AFM.

Single intragenic microsatellite preimplantation genetic diagnosis for cystic fibrosis provides positive allele identification of all CFTR genotypes for informative couples.

This study is part of a strategy aimed at using fluorescent polymerase chain reaction (PCR) on informative genetic microsatellite markers as a diagnostic tool in preimplantation genetic diagnosis (PGD) of severe monogenic disease. Two couples, both of whom had previously had children who were compound heterozygote for severe cystic fibrosis mutations, were offered PGD using fluorescent PCR of the highly polymorphic cystic fibrosis transmembrane conductance regulator (CFTR) intragenic microsatellite marker IVS17bTA. Cleavage-stage embryo biopsy followed by PCR resulted in transfer of one unaffected carrier embryo for each couple. This approach eliminates the need for single cell multiplex PCR strategies to detect CF compound heterozygotes. It also provides a control of chromosome 7 ploidy in the blastomeres and a selection against allele dropout by positive detection of each CFTR copy of all genotypes in preimplantation embryos from genetically informative families.

Preimplantation genetic diagnosis for couples at high risk of Down syndrome pregnancy owing to parental translocation or mosaicism

The population risk for trisomy 21 is 1 in 700 births but some couples are at a much higher risk owing to parental translocation or mosaicism. We report on the...

Isolated Blastomere Development After Cleavage Stage Embryo Biopsy is a Perfect Model to Predict Embryo's Potential Future Development and to Select for Transfer

Isolated Blastomere Development After Cleavage Stage Embryo Biopsy is a Perfect Model to Predict Embryo's Potential Future Development and to Select for Transfer

PCR Amplification of X- and Y- Chromosome-Specific Regions From Trophectoderm Stage Blastomeres That Divided In Vitro, After Cleavage Stage Embryo Biopsy

PCR Amplification of X- and Y- Chromosome-Specific Regions From Trophectoderm Stage Blastomeres That Divided In Vitro, After Cleavage Stage Embryo Biopsy