Embryo biopsy consists of the removal of one or two blastomeres for the pre-implantation diagnosis. For this technique to be succesful it is neccessary to be sure of certain points:

The removed blastomere must be intact and appropriate for diagnostic procedure.

The removed blastomere must be intact and appropriate for diagnostic procedure.

The number of blastomeres removed depends on the embryo developmental stage. The more cells present in the embryo, the more cells can be removed in the biopsy. Embryos most indicated for pre-implantation diagnosis are the ones containing 8 cells. Many studies show that na 8 cell biopsied embryo has the same potential to arrive at the blastocyst stage as the non biopsied embryo, after two or three days of in vitro culture. Three different biopsy procedures have been shown for the removal of one or two blastomeres out of na 8 cell embryo. 1) chemical thinning of the zona pellucida (zona drilling) followed by blastomere aspiration; 2) Partial dissection of zona pellucida (PZD), blastomere removal by pressioning the embryo and aspiration of the exposed blastomere; 3) introduction of needle direcly into the embryo and blastomere aspiration.

The method which combines chemical thinning of the zona pellucida (zona drilling) and blastomere aspiration has been the most used. Using micromanipulation technique, the embryo is immobilized with the holding pipette. A hole approximately 35 mm large is pierced into the zona pellucida by spraying Tyrode’s acidic solution through a micropipette having approximate 10 mm in diameter. Once the hole is done, the pipette containing the acid is removed and the blastomeres are aspirated through the hole by a 35mm diameter micropipette. After the biopsy procedure, the embryos are transferred to the culture medium containing G2.2 or HTF + 15% SSS and cultured in vitro at a 37oC atmosphere containing 5% CO2.

Embryo checking is made 1 hour after the biopsy to inspect their survival, in na inverted microscope at 400X magnification. The embryo is considered intact if the other blastomeres remain intact.

MOST COMMON DIAGNOSIS METHODS

Technically, the best efficiency in analysing DNA or chromosomes can be obtained with just one cell. The most common methods for pre-implantation diagnosis are PCR (Polymerase Chain Reaction) and FISH (Fluorescent In Situ Hybridization).

PCR is a frequently used technique to analyze DNA at the gene level. It uses the DNA of a single cell, because it is amplificated “in vitro”, which permits the theoretical diagnosis of any genetic disease having a known gene responsible for causing the disease.

FISH is a technique that permits to analyze DNA at the chromosome level. It has the advantage of providing results in just 2 to 4 hours. Chromosomal diseases most commonly inspected nowadays are those ones involving trisomies of pairs 13, 18 and 21. Through FISH we can determine for sure the embryo sex, and thus infer about sex linked genetic diseases.

BLASTOMERE BIOPSY TECHNIQUE (sequenced photos)