Associated Procedures
In certain cases, manipulation of the embryo can enhance implantation and subsequent pregnancy. Common practice is to use acidified Tyrode’s solution (pH 2.35) to thin the zona pellucida. An increase in implantation from 18 % to 25 % is achievable for oocytes with poor prognosis with this intervention, refeffed to as assisted hatching. Assisted hatching is not applicable only for male factor infertility, in fact, it is a treatment for specific defects in embryo development or oocyte abnormalities. However, it is a micromanipulation technique that is routinely performed during IVF at many centers.
The application of micromanipulation techniques in the IVF laboratory has allowed the development of analysis and selection of embryos with specific genetic, chromosomal or biochemical characteristics prior to transfer of those embryos. An embryo at the four or eight cell stage is isolated and an individual cell is extracted for evaluation. Chromosome-specific sequences can be identified using fluorescent hybridization probes or, alternatively, polymerase chain reaction (PCR) amplification of individual alleles on the chromosomes themselves may be applied to identify the genotype of the “biopsied” embryo. These techniques can allow identification of embryos at high risk of carrying X-linked diseases such as hemophilia A or von Willebrand’s disease. In addition, specific genetic defects such as the homozygous delta-F508 mutation of the CFTR gene, associated with the development of a severe form of cystic fibrosis, can also be identified. These techniques have been applied for couples known to be at high risk of having children with specific genetic diseases. “Biopsied” embryos have been successfully transferred resulting in pregnancies and live births’. These micromanipulation techniques are highly labor intensive and carry some potential pitfalls. For example, if both parents in a couple are heterozygous for the delta-F508 CFTR mutation, then an individual embryo has a one-in-four chance of being homozygous for that gene mutation. PCR has the potential for identifying the homozygous condition since the delta-F508 mutation is a deletion of 3 base pairs in the normal allele. The normal CFTR gene is 3 base pairs longer than the delta-F508 mutated allele, and a heterozygous carrier would be expected to have both alleles detectable with PCR. However, PCR involves magnification of the single signal present on both chromosomal alleles; if one allele is preferentially bound, read and amplified with PCR, then the results of PCR analysis are significantly confounded.
For sex chromosomal analysis, this evaluation is more accurately performed (up to 95.5 % efficiency) using different colored fluorescent probes”. A test for both X and Y-specific sequences is possible and provides further confirmation of the results of these tests. Given the extensive manpower needed for single-day biopsy and evaluation of the results of embryo biopsy, this technique is limited to those cases where life threatening genetic defects can be reliably detected prior to embryo transfer to prevent the potential termination of a fetus later in development.
