Gamete and Embryo Culture
The term “culture” in the context of IVF refers to maintenance of eggs, sperm, or embryos in a nutrient solution (or culture medium) outside the body in the IVF laboratory. In our laboratory, we culture embryos in small drops of culture medium under a protective layer of mineral oil, in GPS dishes. Each dish is identified using the female patient’s name and the sponsor’s last 4, etched into the bottom of the dish. The dishes are always kept at body temperature in special incubators in the IVF laboratory, except when the embryos need to be evaluated or micromanipulated (for brief periods) under a microscope. Day of fertilization is considered Day 1 of development and Day 1 of embryo culture. Embryos are typically cultured in the laboratory for 3 days or 5-6 days before they are transferred to the uterus. Culture to Day 5 or 6 is referred to as “extended culture.”
The sequence of events following insemination of the eggs is as follows: Approximately 18-20 hours after insemination, eggs are checked for evidence of fertilization. Normally fertilized eggs (zygotes) are placed in GPS dishes and the dishes are placed in the incubator. At this point, the zygote is a single cell. Approximately 4 to 6 hours after the fertilization check, the zygotes are checked a second time for evidence of cleavage (or division into 2 cells). Division at this point is considered “early” and may be a positive indicator of embryo quality. The embryos are examined again carefully but briefly on Days 2 and 3 of culture. The assessment on the morning of Day 3 is used to determine whether the patient will have an embryo transfer (ET) on Day 3 or the embryos will continue in culture for a Day 5 transfer. There are a number of factors that influence this decision, including embryo development profile, patient’s age, cause of infertility and number of previous IVF attempts.
If a decision is made to transfer Day 3 embryos, the best embryo(s) are selected for transfer and they are transferred to the patient later that day. Good to fair quality extra embryo(s) remaining after a Day 3 transfer are placed in extended culture. Embryos with multiple critical morphological/development abnormalities are discarded according to protocol since such embryos have not been proven to fair well in extended culture. When embryo quality cannot be determined with confidence on Day 3, embryos are given the advantage of being placed in extended culture. All embryos in extended culture are reassessed every 24-hours until Day 6 of development. Those that reach the blastocyst stage and meet the criteria for cryopreservation are vitrified on Day 5 or 6 with the informed consent of the patient. Embryos that do not reach the blastocyst stage or show multiple critical morphological/development abnormalities are discarded according to protocol.
If on Day 3 the decision is made to transfer embryos on Day 5, all embryos are moved into extended culture on Day 3. For these cases, the best embryo(s) are selected on Day 5 and transferred to the patient. Extra embryo(s) that remain after a Day 5 transfer and meet the criteria for cryopreservation are vitrified with the informed consent of the patient. Any remaining embryos with morphological/development abnormalities are discarded according to protocol.