As an embryologist, I often find that patients are not well informed on the actual things that go on in the lab. So here, I will provide an overview of what a complete IVF cycle looks like in the lab. First and foremost, record keeping and identification of specimens is the most crucial aspect of embryology which needs to be up to date and perfectly implemented. Different labs have different procedures in place to ensure identification is exact yet easy to implement. Embryologists need to be fully trained in this before they can handle any specimens. The start off with more simple procedures such as sperm analysis, sperm freezing, thawing, and dish preparation until they can move onto more difficult procedures such as IVF.
For an IVF cycle, the day before the egg retrieval, culture media dishes are prepared and incubated overnight to equilibrate. Each patient requires several different kinds of media dishes along with a sperm test tube containing a gradient. The culture media has to be checked regularly to ensure expiration dates have not passed and the quality is optimal. Incubators also have to be checked and calibrated regularly to maintain temperature and pH.
On the day of egg retrieval, the first step is to prepare the sperm sample. The sample can either be collected at home or produced on site. Once we receive the sample, a simple sperm analysis is completed to ensure that enough healthy looking sperm is present. This is usually not an issue because the patient has previously undergone a sperm analysis and abnormalities have been dealt with. Once the sperm sample is sufficient, a gradient is used to separate debris and non-motile sperm from the sample. At the end of the preparation, the sample contains progressive motile sperm. A microscopic amount of the sample is added to the ICSI dish and placed in the incubator.
During the egg retrieval, an embryologist is stationed at a microscope close to the operating room. At the same time that the specialist is performing the retrieval, each test tube is examined thoroughly under the microscope and eggs are picked up and transferred to the egg retrieval dish. This is repeated for each tube passed on from the operating room. Once the egg retrieval is completed, the eggs are stripped of the surrounding cells. This is done by a very fine tipped pipette. The time of denuding/stripping varies between labs and is dictated by the lab director. After denuding, the egg is ready for ICSI. The timing of ICSI also varies between labs and is determined by the lab director. Once the timing is right for ICSI, the eggs are placed onto drops on the same dish as the sperm droplet.
ICSI is performed on the eggs under an inverted microscope. This is a delicate procedure and requires high degree of focus and expertise. One sperm is injected into each egg. Once all eggs have been injected, they are placed into another culture dish which is placed into the incubator. The next day, approximately 16-18 hours after ICSI, fertilization check is completed. Each egg is examined under the microscope to check for fertilization and the number is noted on the patient's chart. Once this is done, the eggs are again placed into the incubator. In some labs, developmental checks are completed every day, on day 3 and 5 or just on day 5.
Depending on whether fresh transfer is chosen or freezing of all embryos, the procedures vary slightly. For a fresh transfer cycle, on day 5, the embryos are checked, the best looking embryo (blastocyst at this stage) or in some cases 2 embryos are chosen for transfer. If the patient has additional embryos, they are frozen for future use. If a freeze-all cycle is chosen, the embryos are checked on day 5 and those that have developed properly are frozen for future transfers. In some cases, the embryo development may take an extra day to reach the blastocyst stage. In these cases embryos freezing is completed on day 6 of development.
Before transfer, the embryos can be artificially hatched using a laser under the microscope. The transfer of embryos, whether fresh or frozen, requires the embryos to be placed in a transfer dish with culture media, half an hour or so before transfer. Once the specialist/nurse has cleaned out the cervix and vagina, the outer sheath of the transfer catheter is placed through the cervix and inside the uterus with a determined distance away from the fundus or end of the uterus. The embryo is then loaded, under the microscope, by the embryologist into the inner sheath of the catheter and is passed on through the outer sheath into the uterus. The embryo is deposited a certain distance away from the fundus. The inner sheath is removed and checked again under the microscope to ensure the embryo has been deposited.
The freezing procedure requires the use of a few different freezing media each with different concentrations. the embryo is moved through the solutions for a set amount of time and loaded onto a freezing straw which is immediately plunged into liquid nitrogen. Once the embryo is frozen, it can stay in liquid nitrogen fro prolonged periods of time. Every few days, the embryologist is responsible for topping off each storage tank with liquid nitrogen to ensure levels do not drop and embryos/egg/sperm stay frozen.
The embryos can be thawed at any point the patient wants to have a transfer. The thawing procedure is usually completed a few hours before the transfer to allow the embryo time to recover before it is transferred. The embryo is moved under the microscope through a few different solutions of varying concentration for fixed amounts of time. After the last thaw solution it is placed in a recovery culture media dish and put into the incubator. Half an hour or so before the transfer, the embryo is moved to the transfer dish and placed back into the incubator. The transfer is the same as the fresh transfer procedure described above. All of the dishes are prepared the night before.
If a patient has elected to undergo preimplantation genetic screening/diagnosis, the embryos are lasered on day 4 of development to allow a small portion of the embryo to hatch. On day 5 of development, the portion of the embryo that has hatched in biopsied. It is important to keep track of which biopsy belongs to which embryo and be able to correctly identify them once the results have returned from the genetic lab. The biopsy procedure itself is also very delicate and requires focus. The embryo has to be biopsied from the trophectoderm which grows to form the placenta. If the inner cell mass (ICM) is biopsied, this can cause problems with development as the ICM grows to form the fetus. Each of the biopsied cluster of cells is washed in media and placed in special kits from genetic labs. The kits are prepared prior to starting the biopsy. Each kit is then shipped to the genetic lab. Once the results are provided, we are able to transfer the normal embryos at the time the patient is ready for transfer. All embryos are kept frozen until the patient and partner sign a consent for disposal or donation.
There are also other procedures that an embryologist is responsible for that I will go into more detail in later posts.
