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Embryo Biopsy

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Embryo Biopsy Embryo Biopsy Caryn Costa Trainee Center for Reproductive Medicine Cleveland Clinic Summary What is Embryo Biopsy. What is the need for Embryo Biopsy. How is embryo biopsy performed. What is embryo biopsy Procedure which involves removal of one or more nuclei from early embryos (Blastomere) or pre- implantation blastocyst (Trophectoderm). Biopsy steps 1. Opening the zona pellucida 2. Removal of the cellular material PGT – Pre Implantation Genetic Testing Biopsied samples are used for PGT. Identify mutations in a gene sequence or aneuploidy. Unaffected/euploid embryos can be selected. Decrease pregnancy terminations and births with genetic abnormalities. What are the different types of pre- implantation genetic testing? Pre-implantation genetic diagnosis (PGD) One or both parents carry genetic abnormalities, diagnosis is performed on the resultant embryo. Pre-implantation genetic screening (PGS) Genetic parents are known or presumed to be genetically normal and their embryos are screened for aneuploidy. Indications PGD PGS Single gene disorders Women of advanced maternal age Triple repeat disorders History of recurrent pregnancy loss Human leukocyte antigen Repeated IVF failure (HLA) matching Carriers of structural Severe male infertility chromosomal abnormalities Female carriers of x-linked Sex selection disorders PGT - Flowchart Ovarian hyper stimulation Egg retrieval In-vitro fertilization/Intracytoplasmic sperm injection Embryo Biopsy Embryo testing Embryo transfer When to perform biopsy? Polar body. Genetic analysis of the polar body extruded from the oocyte. Gives an accurate chromosomal evaluation of the oocyte. This approach gives no clue on the paternal contribution to the embryo, and so it is unable to specify whether the participating sperm presents any genetic abnormality. Day 3 (cleavage). Requires the extraction of one of the theoretical eight blastomeres of the embryo. The embryo has the ability to compensate this loss, but it also presents a higher risk from the technical point of view. When to perform biopsy? Day 5 (blastocyst). o Once the embryo has reached the blastocyst stage, mechanical manipulation is safer, since the inner cell mass (ICM) can be left aside while extracting several cells form the trophectoderm (TE). o Also, potential genetic abnormalities or aneuploidies which occurred during meiosis are likely to have been resolved at this point. Zona Opening Procedures Mechanical Partial Zona Dissection(PZD) The embryo is held with a holding pipette while a microneedle is used to pierce the zona pellucida. The embryo is released from the holding pipette, which is used to cut the ZP against the microneedle. Also useful for Assisted Hatching. Chemical Zona Drilling Acid Tyrode (pH 2.3) Larger, rounder hole than with PZD Two separate pipettes are usually used (double holder). Drilling pipette with an inner ø of 5-7 μm plus the aspiration pipette. Target site: between two blastomeres. Embryo wash after AT exposure is recommended. Chemical Zona Drilling Limitations Size cannot be controlled Limiting the extent and duration of AT exposure is necessary to avoid acidification of medium and cell lysis Laser-assisted zona opening Microdissection of zona pellucida using a 1.48 μm diode laser beam. Easily adapted to the microscope. Laser is transmitted trough a 45X objective. Laser-assisted zona opening Minimal, Superficial damage. A direct relationship between the hole diameter (μm) and the exposure time (ms). Safe, with no mechanical, thermal or mutagenic effects. Increased implantation rate. Early Embryo (Blastomere) biopsy Methods of blastomere removal Aspiration The blastomere is aspirated with the biopsy pipette and pulled out of the embryo Methods of blastomere removal Displacement Two holes are made, media is injected through one hole and the blastomere is displaced out of the other hole Methods of blastomere removal Extrusion The embryo is pushed, causing the blastomere to come out of the hole Advantages and limitations of blastomere biopsy Pre-implantation embryo (Blastocyst) Biopsy Preimplantation embryo – Blastocyst biopsy strategies Day 3 The hole is made on the third day of development and the herniated cells are removed on day five. Day 5 The hole is made on day 5 at the pole opposite to the inner cell mass. The blastocysts are then left for 4-24 hours until some trophectoderm herniates through the slit. When herniation involves about 25% of the blastocyst, the trophectoderm is excised. Advantages and limitations of blastocyst biopsy Conclusion Embryo Biopsy is the removal of one or more nuclei from embryos for genetic testing. The genetic testing can help prevent the births with genetic abnormalities and pregnancy terminations. Microdissection of zona pellucida is usually made using either acid or a laser beam and removal of cells can be performed either at D3 or D5. Choice between biopsy on early embryos (D3) or on pre- implantation embryos (D5) should be made considering the ET strategy and personnel experience. Thank You !.
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