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Embryology Glossary

Egg Retrieval Day

Oocyte/Egg: female gamete cell provided by the ovary.

  • MII Oocyte/Egg: “Mature” egg stage characterized by the display of the Polar Body (PB) in the Perivitelline space. These oocytes are ready to perform ICSI or Vitrification.

 

The following egg stages, we cannot perform ICSI or cryopreserve them and they will be discarded.

  • GV Oocyte/Egg: immature stage which presents an intracytoplasmic nucleus called the ‘Germinal Vesicle’. This type of eggs is not used in the ICSI process since they are not nuclear nor cytoplasmatic matured.
  • MI Oocyte/Egg: immature stage of the eggs that have neither a visible GV nor a Polar Body (PB). It is in the process of maturation, but still, the polar body is not extruded, thus the ICSI cannot be performed.
  • EZ: “Empty Zona” when there is no cytoplasm within the egg, this phenomenon is rare, but it can happen and shows that some cells tried to create an oocyte but failed in the creation of cytoplasm.
  • BZ: “Broken Zona” when the Zona pellucida “shell” is broken, meaning there is no egg inside of it. The reason is attributed to the poor quality of the egg.
  • DEG: it is the Degenerated form of the oocyte/egg in which the cell has undergone apoptosis.
  • Parthenogenetic Egg: This egg has already started the mitotic divisions by itself without use of sperm. These embryos need to be discarded.

What to Expect on Day 1 - Fertilization Check

DAY 1 = Zygote stage

  • 2PN: This means Normal fertilization and is assessed by the presence of two Pronuclei (PN) and two Polar bodies (PB). One PN comes from the mother and the other from the father. This stage is meant to continue the culture and see if they continue to the next mitotic divisions.

 

The following eggs/embryo stages are not optimal to see during the fertilization check. However, we keep them in culture to monitor future cell division and see if they reach the blastocyst stage. It is highly recommended to perform PGT-a testing on these embryos to confirm their euploidy, as we could not observe normal fertilization of the 2PN.

  • PB: This stage shows a Polar Bodies (PB) but no Pronuclei (PN). This could occur due to the accelerated/late fusion of the Pronuclei (PN). Also, it may be a lack of fertilization. This stage is kept under culture to see if they divide. PGT-a is highly recommended to do in these embryos to confirm the euploidy of it.
  • 1PN: When the fertilized egg shows just one Pronuclei (PN), it can result from errors in the fertilization process with asynchrony in PN formation or PN fusion. This type of fertilization is kept in culture to see if they divide. PGT-a is highly recommended to do in these embryos to confirm the euploidy of it.
  • 2 Cell: This is an embryo stage of two cells that could be derived from an accelerated fusion of the Pronuclei and division of the zygote. In this situation, the embryos can continue their development and see if they reach the blastocyst stage. PGT-a is highly recommended to do in these embryos to confirm the euploidy of it.

 

The following eggs or embryos are discarded because they are not compatible with life if fertilized or due to degradation.

  • 3PN: It is abnormal fertilization and known as a Triploid zygote due to the presence of three Pronuclei (3PN), which differs in origin when fails in extrusion of the second Polar Body (digenic) or two spermatozoon entry into the oocyte (diandry) if conventional insemination was performed.
  • >3PNs: It is abnormal fertilization where more than 3 Pronuclei are shown, and the causes can come from failed cytokinesis (cell division) or binucleate spermatozoon. In this kind of fertilization and polyploidy found, the genetic abnormalities are higher and not compatible with life.
  • DEG: Some fertilized eggs (zygotes) after ICSI can undergo degeneration. In this case, the Degraded eggs must be discarded. Most of the cases can be related to poor egg quality.

What to expect on day 5

The ideal stage for day five is to have the blastocyst or at least the early stages of it. Between 45-55% of the embryos that showed a 2PN fertilization on day one reach the blastocyst stage. This stage can be reached between days five, six, or day seven.

The earlier an embryo reaches the blastocyst stage, its chances of successful implantation are higher. However, the blastocyst grade and euploidy also influence the implantation rate. For instance, a blastocyst on day 5 has a better implantation rate than one on day 6, which is better than one on day 7. It is crucial to consider these factors when evaluating the likelihood of successful implantation.

On your embryology report for day five, you can have the following stages.

Embryos with good prognosis on Day 5:

  • >10 cells: in this stage, the embryo is still dividing blastomeres and is near the compacting stage. This type of embryo can be kept in culture to see if it continues its process of forming a blastocyst in the next few days.
  • COMP: Also known as “Morula”, is the Compaction (COMP) process in which the cell’s junctions become tighter and is linked to the activation of the embryonic genome. This is one step before the next stage, called Cavitation. This form is also maintained in culture to see if it continues the development.
  • CAV: The cavity or cavitation (CAV) process is the stage that will lead to the blastocyst formation. This stage is when the embryo raises the salt concentration within the embryo, attracting water through osmosis. This increased water pressure gradually increases the size of the cavity, which continues throughout the blastocyst stages.
  • Early Blastocyst: The Early blastocysts are those in which the blastocoel cavity is less or half of the volume of the embryo, and since their cells have not created tight junctions, the Inner cell mass (ICM) and Trophectoderm (TE) are hardly differentiated, are not ready for performing biopsy and tend to stay in culture to see if they develop the structures mentioned.
 

If the blastocyst is grade 3-4-5-6 and meets our grade standards, we will perform a biopsy and cryopreserve it. The definitions of grades A, B, and C are at this glossary’s end.

The following embryo stages with poor prognosis will be kept in culture if observed on day five but discarded if observed on day six or day seven.

  • Embryos in the cleavage stage, between 2 – 10 cells. In this case, the embryos are considered to stay or not in culture since their lack of division or slow division rate indicates arrested embryos, and their prognosis is poor.
  • Poor Blastocyst: The poor-quality blastocyst starts showing some death cells or has very few cells in the inner cell mass (ICM) or Trophectoderm (TE). These embryos are kept in culture to follow their fate that could be improved by mitosis, the number and quality of the structures mentioned, or the Degradation by Necrosis/apoptosis.
 

The following stages will be discarded on day five and day six.

  • PB: If cell division is not observed by day five, the egg will be discarded and considered a failed fertilization, and we discarded them.
  • DEG: If cell death occurs due to necrosis or apoptosis, it can cause the embryo to degrade. This involves swelling and rupture of cells, as well as irreversible damage, cellular shrinkage, and condensation of the cytoplasm. The nuclear and cytoplasmic membranes also become indented. Since this embryo faces cellular death, it cannot biopsy or cryopreserve and is discarded.

What to expect on day 6

Embryos that remain in culture should exhibit signs of cellular division. The likelihood of reaching the blastocyst stage decreases if they remain in the same stage. The optimal stage for this day is a blastocyst graded 3-4-5-6. The embryos in the cleavage stage, as well as degenerated embryos, will be discarded.

What to expect on day 7

If embryos do not reach the blastocyst stage, they are discarded because IVF culture media can only support them until this point.

Blastocyst Grading:

The grade defines the progression of the blastocyst and the quantity and quality of the cells within it.


There are two types of cells in the blastocyst. One cell type forms the Inner Cell Mass (ICM). These cells will eventually grow into the fetus. The second cell type is the Trophectoderm Epithelium (TE), these cells will go on to make the placenta and surrounding structures.


PROGRESSION: Higher number means more advanced stage of the blastocyst.


Stages 1 and 2 will be reported in our laboratory as Early Blastocyst. We cannot biopsy embryos at this early stage.


Stage 3 Full Blastocyst: This type of blastocyst commonly fills more than 50 % of the blastocoel with fluid, and the ICM and TE are well differentiated. The Zona pellucida (ZP) is still a bit thick. Embryo biopsy could be performed depending on the cell junctions the embryos show.


Stage 4 Expanded Blastocyst: Blastocysts in which the blastocyst cavity is now greater than the original volume of the embryo and the ZP is thinned.


Stage 5 Hatching Blastocyst: Blastocysts or hatching blastocysts in which the blastocoel cavity is greater than the original volume of the embryo and the TE is herniating through a natural breach in the ZP


Stage 6 Completely Hatched Blastocyst: Blastocysts or hatched blastocysts are those in which the blastocyst has completely escaped from a natural breach in the ZP.
The ICM and TE are visible from grade 3 to grade 6 blastocysts, the following classification you can find in the report with the Letters: A, B, or C, (i.e., Blastocyst 3AA, the number denotes the cavity expansion grade, the first A will denote the ICM quality and the second A will classify the TE quality).

The ICM and TE are visible from grade 3 to grade 6 blastocysts, the following classification you can find in the report with the Letters: A, B, or C, (i.e., Blastocyst 3AA, the number denotes the cavity expansion grade, the first A will denote the ICM quality and the second A will classify the TE quality).

ICM

  • Grade A: contains many cells that are tightly packed together.
  • Grade B: is composed of several cells that are loosely grouped.
  • Grade C: describes an ICM that contains very few cells that are loosely bound.

TE

  • Grade A: contains many cells that form a cohesive epithelium
  • Grade B: is composed of few cells forming a loose epithelium
  • Grade C: describes a TE that contains very few, large cells that struggle to form a cohesive epithelium

 References

  • Romeu, A., Veeck, L.L., Oehninger, S. et al. Significance of the recovery of fractured-zona oocytes in an in vitro fertilization program. J Assist Reprod Genet 5, 216–224 (1988). https://doi.org/10.1007/BF01131125.
  • “Atlas of Human Embryology”, https://atlas.eshre.eu/es/14555410245416920.
  • Gardner DK, Schoolcraft WB. In vitro culture of human blastocysts. In: Jansen R, Mortimer D, editors. Toward Reproductive Certainty: Fertility and Genetics Beyond 1999. UK: Parthenon Publishing London; 1999. p. 378-388.
  • Richter KS, Harris DC, Daneshmand ST, Shapiro BS. Quantitative grading of a human blastocyst: optimal inner cell mass size and shape. Fertil Steril 2001; 76:1157-1167.
  • Hardy K, Handyside AH, Winston R. The human blastocyst: cell number, death, and allocation during late preimplantation development in vitro. Development 1989; 107:597-604.
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