What is embryo grading?

After successful fertilization of retrieved eggs, the embryo or embryos will start to develop in a petri dish in an IVF lab. Embryo grading can take place at different stages of the culture process, and it is used as a tool to assess quality and development.  Embryo grading allows for a standardized measurement of an embryo's quality — good, fair, or poor — based on its morphology (structure and appearance).  The grade can then be used to assess the likelihood of a transfer being successful and to prioritize the highest quality embryo for transfer.ⁱ

There are many variations in criteria for embryo grading, as well as the way embryo grades are assigned. These differences can make comparisons between clinics or between studies extremely difficult.ⁱⁱ

At what stage are embryos graded?

Embryo development is assessed at different stages by the embryology team, and the timing of assessments vary depending on the protocols used by the clinic. Most often, grading is performed at the cleavage stage (Day 3) and blastocyst stage (Days 5 to 7).

When embryos are assessed on the third day after egg retrieval, the embryologist is looking at cell number and embryo morphology under a high-powered microscope. Day 3 embryos have cells that are actively dividing but the embryos themselves are not growing in size or volume. Day 3 embryos should ideally have six to 10 cells, with eight cells being optimal to develop into viable blastocyst embryos.ⁱⁱⁱ

Rarely, embryos are graded at the morula stage (Day 4) when a Day 4 transfer is planned. But far more often, embryos are graded at the blastocyst stage (Days 5 to 7), if they reach that point, to assess the quality of their development after embryonic genes have been activated. Blastocyst-stage grading also allows for the examination of the inner cell mass and trophectoderm. The grading of the trophectoderm (TE) will reflect the embryo’s potential to attach and implant in the uterine lining, as it will later become a large part of the placenta. Grading of the inner cell mass (ICM) will assess the potential development of the embryo (and later fetus) itself.^iv  

Some IVF clinics perform the embryo-grading process at other stages (i.e., zygote, morula), but grading at the 8-cell cleavage stage and blastocyst stage are the most common.

What do the different embryo grades mean?

The early blastocyst grading system was first introduced by Gardner and Schoolcraft in 1999 and was quickly adopted by most IVF laboratories. Although this system did not cover all features of embryo morphology, it was useful to classify the quality of blastocyst development, assessing the blastocoel (fluid-filled cavity), the inner cell mass, and trophectoderm under a microscope. Grading the expansion was important, since formation of the fluid-filled cavity requires the embryo to produce a lot of energy and therefore reflects embryo quality. The Gardner-Schoolcraft system also assigns grades of A, B, or C separately for the structural appearance of the inner cell mass and trophectoderm.^v

The Society for Assisted Reproductive Technology (SART) in the U.S. then developed a simple and widely used embryo grading guideline in 2006. Using this methodology, embryos are graded into three broad categories of good, fair, or poor using morphologic features. By identifying the embryos as good, fair, or poor, the SART system helps patients and embryologists select which embryo(s) to transfer.  However, it does not further narrow down or rank the “good” embryos.^vi

In 2011, experts from the European Society of Human Reproduction and Embryology (ESHRE) and Alpha Scientists in Reproductive Medicine issued updated guidelines to standardize embryo grading systems. Their grading classifies embryo quality based on cell number, the degree of fragmentation (uneven division of cells), the symmetry of the cells (even cell size), the presence of multiple cell nuclei, and compaction status.^vii

Embryo grading chart: Cleavage stage

Cleavage stage embryos are assessed under the microscope to determine the number of cells (seven to nine is optimal), the cell size and symmetry, the presence of one nucleus (structure that contains DNA) in each cell, low vacuolization (fluid-filled sacs), as well as degree of fragmentation.^viii

Fragmentation measures the number of small membrane-bound cell pieces that have broken off. These are usually less than 40 μm in size. Less fragmentation is associated with better outcomes. Experts agree that large vacuoles (larger than 14 μm in diameter) can be associated with fertilization failure, but that a few smaller vacuoles generally have little effect. There should be only one nucleus present and multi-nucleation is often associated with poor outcomes.^ix

Below is a table of a typical grading system used to score a Day 3 cleavage-stage embryo. However, it should be noted again that many clinics have variations in their scoring systems.^x

How is the table read?

The typical scoring format for Day 3 embryos includes the grade and number of cells. The grade is assigned as explained in the table and is followed by the number of cells that the embryo has when examined under the microscope. For example, a 2/7 is graded fair (2) and has 7 cells. Some embryology labs may report other details, such as “15 percent fragmentation” to the patient.

Ideally, a Day 3 embryo (approximately 68 hours post-insemination) would have 8 equally sized cells (called blastomeres) with less than 10 percent fragmentation, only one nucleus, and no large vacuoles visible.

Embryo grading chart: Blastocysts

Grading systems vary among embryology labs, but many use some type of adaptation of the Gardner and Schoolcraft grading system.^xii

Below is a table of a typical grading system used to score a blastocyst.

How is the table read?

During grading, the embryos are viewed by an embryologist under a microscope. The grade will include a number corresponding to the stage of blastocyst expansion, as well as two letter grades for the inner cell mass and trophectoderm, respectively.

Example: 5AA

This grading indicates a hatching blastocyst (5), which refers to the embryo emerging, or hatching, out of the zona pellucida shell. The embryo has a tightly packed inner cell mass (A) and a good quality trophectoderm (A) forming a cohesive covering layer. If a clinic uses a numerical scale instead, 5AA would be graded as 511.

AA is the highest quality score an embryo can have at each stage of development.

How does embryo grade impact success rates?

The number of well-graded embryos resulting from an IVF patient’s oocyte retrieval, particularly when graded at the blastocyst stage, may predict how many embryos continue to implantation.

For blastocyst grading, many studies show a correlation between better-graded blastocysts and success rates. One 2021 study showed a significant difference in implantation rates for good-quality Day 5 blastocysts (79.8 percent) versus poor-quality blastocysts (48.1 percent).^xiv However, not all studies observe this relationship of a “good” grading providing better outcomes than a “poor” grading. A 2019 study of over 600 frozen embryo transfers observed similar live birth rates between good-quality Day 5 embryos (48.65 percent) and poor-quality Day 5 embryos (47.50 percent).^xv

Despite reported correlations, even poorly graded blastocysts still commonly result in live births, while the highest graded embryos may not implant.^xvi

With Day 3 embryos, grading appears to be much less consistent in predicting implantation and pregnancy rates with some studies showing that Day 3 grading does not correlate with success, while others show it is useful in predicting implantation. For example, a 2018 study found that ongoing pregnancy rates were not significantly different between grade 1, 2, and 3 embryos, with clinical pregnancy rates of 58.0 percent, 57.9 percent and 55.6 percent, respectively.^xvii

In contrast, other studies have reported that when Day 3 embryos are transferred, live birth rates improved with increasing cell number (up to the eight-cell stage): 16.2 percent when there are more than eight cells versus 2.9 percent when there are less than six cells.^xviii And a study of 426 Day 3 fresh embryo transfers showed that grade 1 embryos had a higher implantation rate (41.8 percent) than grade 3 embryos (18.4 percent).^xix

While embryo grading may help to predict implantation, its main purpose is to be a useful tool to decide which embryo(s) to transfer first. There remain inconsistencies in the predictive value of embryo grading, particularly at the Day 3 cleavage stage.

What is the difference between grading and PGT?

Embryo grading and preimplantation genetic testing (PGT), previously called preimplantation genetic diagnosis/screening (PGD/PGS), are both commonly used to select which embryos should be transferred. While embryo grading visually evaluates the quality of an embryo’s morphology, it does not provide information about whether the embryo is genetically competent (euploid, meaning it has the correct number of chromosomes). In contrast, PGT assesses an embryo for genetic abnormalities (such as aneuploidy, or incorrect number of chromosomes) by analyzing the chromosomes via embryo biopsy. Embryos with an abnormal number of chromosomes are more likely to stop growing and fail to implant.

A 2019 study showed that embryos with higher grades were more likely to be euploid than lower-graded embryos.^xx Nonetheless, morphological grading should not be used to make assumptions about whether an embryo is euploid.

Overall PGT is better able to predict likelihood of clinical pregnancy than embryo grading. Morphological grading can still be used in selecting embryos for transfer, but only when there are multiple euploid embryos to choose from.^xxi In other words, once an embryo is determined to be euploid by PGT-A, the embryo grading is less important in the embryo selection process than the fact that the embryo is euploid.^xxii,xxiii

Conclusion

Grading embryos at different stages gives the embryology team the opportunity to determine which embryos may be most likely to implant after embryo transfer. While embryos are graded based on their quality in relation to cell division and appearance, even a poor embryo with a lesser grade may implant and result in pregnancy and a live birth. It is also important to remember that embryo grading does not predict the health of an embryo or of a future child. Nevertheless, the process gives doctors one more tool to help achieve better outcomes with IVF treatment.