During follicle puncture (aspiration), the embryologist receives cumulus-oocyte complexes. Immediately after the puncture, the maturity of the collected cumulus-oocyte complexes (COCs) is assessed.
COC with a translucent cumulus usually contains a mature oocyte. The COCs data do not appear in the embryological protocol, but can be revealed at the request of the patient.
1-2 hours after aspiration, the embryologist cleans (denudates) the oocytes. The puncture may result in obtaining mature, immature, as well as degenerative and destroyed eggs.
Accurate assessment of the oocyte is possible only after its purification before ICSI. In mature, ready-to-fertilize oocytes, a polar body can be visualized. In the embryological protocol, the mature oocyte is recorded as MII.
If the process of maturation of the oocyte in the follicle is disrupted, then there is a high probability of obtaining immature cells, recorded as MI and GV. Complete degeneration of the oocyte is also possible (Deg).
18-20 hours after the addition of sperm or ICSI or pICSI (1st day) during normal fertilization, two pronuclei are formed.
With proper fertilization, both pronuclei are clearly visible. In this case, they are assigned a score of 2pN.
If pronuclei are not visible, which is usually associated with lack of fertilization, they are recorded as 0pN (mpN) in the protocol.
With improper fertilization, the appearance of several pronuclei is possible, which is reflected in the record, such as 3pN, 6pN, etc.
“Incorrectly” fertilized oocytes are not suitable for further work and are discarded. Further development of the embryo, fragmentation, occurs within 5-6 days. Embryo quality is assessed every 23-25 hours from the time of fertilization testing.
On the second day of development, the embryo consists of 2, 3 or 4 cells (blastomeres). At this stage, the quality of the embryo can be assessed by the degree of fragmentation (the volume of the embryo occupied by non-nuclear fragments of the cytoplasm). The more fragments, the lower the potential of the embryo for implantation and further development.
In addition to fragmentation, the shape and relative size of blastomeres are evaluated. The generally accepted classification of embryos for quality is A-B-C-D, where A is the best and D is the worst.
Uneven fragmentation (the presence of blastomeres of different sizes) reduced the potential of the embryo before implantation. The presence of fragments of the cytoplasm is marked fr. The presence of vacuoles is also estimated (visible vacuoles are marked vac).
From the 2nd to the 6th day of development, embryos can be transferred into the uterine cavity (embryo transfer). In vivo, the embryo at this stage moves down the fallopian tube towards the uterus.
On the third day, the embryo normally consists of 6-8 blastomeres, but 4 blastomeres are also accepted if it has been a 2-cell embryo on Day 2. Assessment and recording of embryo quality on the third day is carried out on the same principles as on the second day.
By the 8-cell stage, all human embryonic cells are totipotent, i.e. each of them can give rise to a whole organism.
On the fourth day of development, the human embryo usually consists of 10–16 cells, the intercellular contacts are gradually compacted and the surface of the embryo is smoothed (compaction process) This is the beginning of the morula stage (from the Latin ‘morulae’ ̶ silk berry). It is at this stage that the embryo enters the uterine cavity from the fallopian tube in vivo.By the end of the fourth day of development, a cavity is gradually formed inside the morula. Thus, the process of cavitation begins.
On the 5th day, approximately 120 hours after fertilization, the embryo forms a blastocyst (Bl). From the time the cavity inside the morula reaches 50% of its volume, the embryo is called a blastocyst.
The blastocyst consists of two cell populations – a trophoblast (a single layer of epithelium surrounding the cavity) and an inner cell mass (a dense clump of cells). Trophoblast cells will later give rise to all outside the embryonic membranes of the fetus, and from the inner cell mass tissues and organs of the baby will form.
Blastocyst quality is evaluated in terms of size, which is graded from 1 to 5, the inner cell mass (ICM) (from A to C) and the surrounding cells – trophoblast cells (graded from a to c).
Blastocysts graded 3-5 with multicellular ICM and Bl 4Aa, Bl 4Ab trophoblast cells will be optimal for transfer.
The larger the blastocyst cavity and the better developed the inner cell mass and trophoblast, the greater the potential for implantation. When the blastocyst cavity reaches a significant size, the glossy shell thinned by stretching breaks and the process of hatching (exit) of the embryo from the glossy shell begins. Only after this process is the blastocyst able to implant (attach) to the endometrium of the uterus. Implantation usually occurs on day 6-7 of embryonic development, assuming the day of fertilization is zero.
Thus, using the above data, the couple can independently assess the quality of embryos and their potential for implantation based on the data of the embryological protocol.