To study the developmental impact of embryo biopsy on the progression of embryos from day 3 to day 5. Retrospective analysis of IVF cycles from January 2006 through December 2006. During this time period, 105 patients (Group 1) had embryo biopsy for preimplantation genetic diagnosis (PGD) and 253 patient cycles were cultured to day 5 for blastocyst transfer (Group 2) without biopsy. Indications for PGD were: aneuploidy screening, chromosome rearrangements, family balancing, and various single gene defects. Ovarian stimulation and IVF were done with standard methods. Quinn's sequential media (Sage) was used throughout the study. Embryos were cultured in individual 20 μl droplets under oil in 5% CO2 and 5% O2. Biopsy was done either by physical slit or laser heat ablation of the zona pellucida. A single cell was removed from each embryo that had progressed to at least 5 cells on the morning of day 3 of embryo development. Embryos were grouped and compared by their cleavage stage of development on Day 3 and their progression to blastocyst. An embryo was considered a blastocyst if it formed a blastoceol and an inner cell mass (ICM) could be clearly delineated. Chi-squared analysis was done comparing the outcome of the various groups and p values are displayed in Table 1. A p value of < 0.05 was considered statistically significant. 781 embryos were cultured and followed in Group 1 (Biopsy) and 1,959 embryos were cultured and followed in Group 2 (No Biopsy). The results of the study can be seen in Table 1 below: TableDay 5 Group 1 (Biopsy)Day 5 Group 2 (No Biopsy)Stage D3NBlasts% BlastsNBlasts% BlastsP5 Cells117762656725<0.0016 Cells139312236713236<0.057 Cells191743943020648N.S.8 Cells2421174868641360N.S.9+ Cells77476121113563N.S.Total78127335195995349<0.001 Open table in a new tab Performing single blastomere embryo biopsy on Day 3 embryos clearly had a negative impact on progression to the blastocyst stage on Day 5. However, the effects were more pronounced on embryos that were already progressing slowly. For embryos at later cell stages (at least 7 cells or higher) the impact was minimal and did not reach statistical significance. This information may be used counseling patients considering embryo biopsy.

In some situations where embryo biopsy and Pre-Implantation Genetic Diagnosis (PGD) are being performed, it is necessary to freeze some or all of the embryos created in the fresh IVF cycle. These clinical situations are: 1) Thin endometrial lining, 2) Ovarian Hyper Stimulation Syndrome (OHSS), or 3) Excessive embryos are created. Our purpose is to determine if cryopreserving embryos prior to biopsy and PGD is a feasible clinical alternative, and whether developmental stage has an impact on the outcome. A retrospective study comparing the survival rates of embryos and the pregnancy rates of patients in which embryos were cryopreserved at either the pronucleate stage or at early cleavage stages and then later biopsied for PGD. All embryos were cryopreserved using standard controlled rate freezing and slow thawing methods. Embryos were individually cryopreserved and stored in straws. Mechanical biopsy was performed on day 3 on all embryos with ≥5 blastomeres. When possible, 2 blastomeres were removed for genetic diagnosis. A total of 171 pronuclear zygotes and 214 cleaved embryos were utilized from 23 and 29 patients respectively. Table 1 shows the number of thaw cycles, numbers of biopsies and transfers and the outcomes of the thaws at each developmental stage. There was no significant difference in any of these parameters among the 3 groups. Table 1Tx = Transfer Bx = Biopsy Clinical Pregnancy = Fetal Heart Beat on Ultrasound Tx = Transfer Bx = Biopsy Clinical Pregnancy = Fetal Heart Beat on Ultrasound Embryos from day 1, day 2 and day 3 were all used successfully. The most effective and efficient method from a logistical standpoint is to freeze embryos on day 1 or day 2, then thaw on the equivalent day in a subsequent frozen embryo transfer cycle prior to doing biopsy and genetic diagnosis. We conclude that embryos can be successfully frozen, thawed and biopsied when it is clinically necessary or advisable to do so; with very reasonable results.

Marfan syndrome (MFS) is an autosomal dominant disease that affects the skeletal, ocular and cardiovascular systems. Defects in the gene that codes for fibrillin (FBN-1) are responsible for MFS. Here we report the world's first use of preimplantation genetic testing (PGT) to achieve a clinical pregnancy and live birth of a baby free of a Marfan mutation. One or two blastomeres from each embryo were tested for a CA repeat within the FBN-1 gene. The prospective mother is homozygous for the CA repeat (2/2) and has two normal copies of the FBN-1 gene, while the prospective father is heterozygous for the CA repeat (1/2), and is affected with the Marfan syndrome. In the father's family, allele 2 segregates with the mutated FBN-1 gene. For PGT, any embryo diagnosed as heterozygous for the CA repeat (1/2) would be presumed to have inherited normal FBN-1 genes from the father and the mother and be unaffected. One in-vitro fertilization (IVF) cycle yielded 12 embryos for preimplantation testing; six of the embryos were heterozygous for the CA repeat (1/2) and presumed to be free of the Marfan mutation. Five of the six embryos were subsequently transferred into the uterus. The fetus was tested by chorionic villus sampling and found to be free of the Marfan mutation by the same linkage analysis, had a normal fetal echocardiogram, and was normal at birth.