Embryology/IVF

Several important procedures in IVF involve sperm and egg micromanipulation under microscopic observation. PZD, SUZI, and ICSI are aimed at assisting penetration of sperm through the outer membrane of the egg (the zona pellucida) and in the case of ICSI (Intracytoplasmic Sperm Injection), the oolemma as well. Partial dissection of the zona pellucida is also used in assisted hatching as this is thought to enhance the chances of implantation in the uterus. Implantation rates may also be improved by cytoplasmic transfer where donor cytoplasm is used to enhance the quality of the egg.

Pre-implantation diagnosis for the presence of inherited gene mutations involves removing one cell from the developing embryo for testing (usually at the eight cell stage). The embryo biopsy involves either immobilising the embryo with a holding pipette and then disrupting part of the zona pellucida to allow aspiration of a cell, or direct insertion of a needle through the zona pellucida followed by needle aspiration. Embryos are examined under high magnification (400x) to ensure that the remaining cells have not been damaged and that the embryo is capable of developing normally.

The two most commonly used techniques for pre-implantation genetic diagnosis are FISH /mFISH and PCR. PCR is used mainly to detect single gene abnormalities and requires IVF using ICSI to avoid contamination by sperm DNA. FISH is used to detect chromosomal abnormalities.

Key microscopy techniques for observing embryos include brightfield, darkfield, phase contrast, DIC and time-lapse imaging. Microscopes used for IVF must be exceptionally stable and vibration free with space to accommodate micromanipulation equipment. High N.A objectives with long working distances, heated stages and environmental control are an advantage.