During IVF, the embryos are cultured for up to six days and receive quality grades each day.
Egg Retrieval and Insemination Day 0
Egg maturity is important because a mature egg has the best chance of being fertilized. There are three different stages of egg maturation:
- Germinal vesicle (GV): The egg has not begun meiosis yet, so it is considered immature.
- Metaphase I (MI): The egg is in the first phase of meiosis; however, it is still not completely mature because it has not entered the second phase of meiosis. This kind of immature egg may mature after a couple of hours of temperature-controlled incubation.
- Metaphase II (MII): The egg is in the second phase of meiosis and is mature. Eggs at this stage of maturity are ready for fertilization.
- Clear cytoplasm/normal shape
- Single distinct polar body
- Clear/thin zona pellucida
- Slightly grainy cytoplasm/misshapen
- Fragmented/abnormal polar body
- Slightly pigmented/amorphous zona
- Cytoplasmic bodies
- PV debris
- Dark/grainy cytoplasm/misshapen
- >1 polar body structure
- Pigmented/thickened zona
- PV debris
Fertilization Check Day One
Fertilization can be seen 16 to 22 hours post insemination. Normal fertilization is identified by exactly two pronuclei in the center of the single cell zygote. Fertilization is considered abnormal when there is only one pronucleus or when there are more than two pronuclei.
Multicell Grading Day Two/Three
On day two the single cell zygote should divide into an embryo (approx. two to four cells). On day three the embryo should continue to divide (four to eight cells).
- Good: cells are symmetrical with clear cytoplasm
- Fair: cells are slightly asymmetrical and/or have slight cytoplasmic irregularities
- Poor: cells are significantly asymmetrical and/or have dark, grainy cytoplasm
- A = No fragmentation
- B = <10% fragmentation
- C = 10–35% fragmentation
- D = >35% fragmentation
On day four, embryos begin their transition from a multicell embryo to a more advanced developmental stage. Embryos should begin compacting and forming morulae. Cells of a morula-stage embryo are not as distinct as in previous days; therefore, these embryos do not receive quality grades.
Day Five/Six Blastocyst Stage
A blastocyst is a highly developed embryo that is composed of two different cell types: one group of cells, called the inner cell mass, leads to fetal tissue and another group of cells, called the trophoectoderm, forms the placenta. Blastocysts are graded on their expansion (early, expanding, expanded, and hatching) as well as the quality of the two different cell types (graded on a good-fair-poor scale). Blastocysts that are good to fair quality meet freeze criteria.