Blastocyst Culture

IVF, In Vitro Fertilization with Blastocyst Culture and Day 5 Transfer

• Blastocyst culture and day 5 embryo transfer for in vitro fertilization allows selection of the best quality embryos for transfer
• Embryo quality is important with infertility
• With blastocyst transfer, we transfer fewer embryos - reducing multiple pregnancy risks, and maintain high IVF success rates

Definition of a blastocyst

• An embryo that has developed for five to seven days after fertilization and has 2 distinct cell types and a central cavity filled with fluid (blastocoel cavity)
• The cells in a blastocyst have just started to differentiate
• The surface cells that surround the cavity (just under the outer shell) are called the trophectoderm and will later develop into the placenta
• A more centrally located group of cells - the inner cell mass, will become the fetus

Blastocyst development

The most commonly used indications for assisted hatching with an in vitro fertilization case are:

• The blastocyst usually forms on day 5 as fluid builds within the compacted morula
• A healthy blastocyst often begins hatching from its outer shell, called the zona pellucida between day 5 to day 7 after fertilization
• Within 24 hours after hatching, embryo implantation after IVF (or a “natural” pregnancy) begins as the embryo invades into the uterine lining
• The blastocyst releases HCG hormone (the pregnancy test hormone) which leaks into the mother's blood as the embryo implants

Blastocyst formation rate

• The goal of in vitro fertilization and embryo culture is to provide high quality embryos which are capable of continued development and result in live births
• However, under standard IVF culture conditions, only about 25 to 60% of human embryos progress to the blastocyst stage after 5 days of culture
• The low rate of embryo development has 2 main causes
  • A less than optimal culture environment in the lab dish
  • The inherent “weakness” of human embryos

Therefore, in the past embryos were transferred to the uterus after 2 or 3 days.

Blastocyst Development and Blastocyst Quality Grading (or scoring)

• There are 3 distinct parts to a blastocyst - the two cell types and the fluid cavity
• As development of blastocysts progresses, cells in the 2 components divide and the fluid cavity enlarges
• With continued blastocyst development the embryo expands, the shell thins, and it hatches from its shell
• Blastocyst implantation into the lining of the uterus can then begin

Our IVF clinic and many others that do blastocyst culture and transfer use a blastocyst scoring system originally developed and published by Dr. David Gardner in the late 1990’s. Dr. Gardner is well known for being first to develop a commercially available embryo culture media system that supported high rates of viable blastocyst development. There are other grading systems for blastocysts, but this one is most widely used.

The Gardner blastocyst grading system assigns 3 separate quality scores to each blastocyst embryo:

• Blastocyst development stage - expansion and hatching status
• Inner cell mass (ICM) score, or quality
• Trophectoderm (TE) score, or quality

Blastocysts are given a quality grade for each of the 3 components and the score is expressed with the expansion grade listed first, the inner cell mass grade listed second and the trophectoderm grade third.

For example, a blastocyst quality grade of 4AB means that the blastocyst is expanded (grade 4), has many tightly packed cells in the inner cell mass (grade A), and has a trophectoderm with few cells forming a loose epithelium (grade B). See the examples of blastocyst grading pictures below.

There are other blastocyst grading systems, including the grading that we use in our IVF program (see below). Some use more letter grades than the Gardner system. For example, some clinics give component cells grades of D or even E in addition to the A, B and C grades described above.

We have seen records from some labs that give blastocysts a single letter grade, and no number at all. For example the blasts are simply graded as A's or B's or C's, etc. This system is easier for lab personnel to deal with but at the cost of being less descriptive. There is also some loss of correlation between grading of the blastocysts and expectations for successful pregnancy outcome.

Blastocyst embryo grading is helpful. However, the potential of the embryo to continue normal development, implant and result in a healthy baby born is difficult to predict based on grading alone.

Assessment of an embryo's chromosomal status using preimplantation genetic testing for aneuploidy (PGT-A) along with standard morphology grading is the current gold standard for selection of the “best” embryo for transfer.

• A pretty embryo is good, but pretty and chromosomally normal is much better

Advantages of blastocyst transfer for IVF

• One problem with this is that 2 to 3-day-old embryos are normally n the fallopian tubes, not in the uterus. The embryo gets to the uterus about 80 hours after ovulation.
• Embryo implantation process begins about 3 days later - after blastocyst formation and hatching out of the embryonic shell have occurred.
• Therefore, if in vitro culture conditions are maximized so healthy blastocysts form at a high rate, then day 5 blastocyst embryo transfer can be done.
• The uterine lining on day 5 should be receptive to the arriving embryo - this a more “natural” time for the embryos to be in the uterus. It is the same timing as with a natural pregnancy.
• The transfer is done shortly before the time for actual invasion and implantation

Pregnancy rates and reduce risks for multiples

• Transferring blastocysts following IVF also provides another potential benefit - reducing possibility for multiple pregnancy
• Many 2 or 3-day-old embryos do not have the capacity to become high quality blastocysts and make a viable pregnancy. However, on day two or three of culture we don't have methods to determine which embryos will be viable long-term, and which will soon arrest their development.
• By culturing embryos to day 5 we will find that some of them have not become blastocysts - allowing us an opportunity to choose the most competent embryos for transfer.
• We can then transfer fewer embryos and still obtain high pregnancy success rates - with very little risk for having high order (triplets or higher) multiple pregnancies.

Extended in-vitro culture systems

• In the past, it was difficult to get high quality blastocysts with in vitro culture systems - unless “feeder” cells were utilized - called coculture.
• However, since 1998 more advanced culture media have been commercially available that (if used properly) can yield high blastocyst formation rates.
• Now blastocyst embryo transfer is a viable IVF treatment option for many couples

Disadvantages of blastocyst transfer

• If the IVF clinic is proficient at blastocyst culture and has excellent quality control in the laboratory, there is potentially no disadvantage to day 5 blastocyst transfer.
• However, if the culture environment is suboptimal, delayed embryo development and even embryonic arrest will occur in some cases.
• Therefore, if the culture system and laboratory quality control are inconsistent - good results will not be obtained with extended culture to day 5.
• Such programs will do better with day 3 transfers - putting back embryos earlier, before they are “stressed” excessively by the weak culture environment.
• Another possible disadvantage - some clinics charge more for day 5 transfer