Embryo Culture Media for Human IVF

By MD, PhD, MSc (gynecologist), MSc, PhD (senior clinical embryologist), BSc, MSc (embryologist), BSc, MSc (clinical embryologist), BSc, MSc (embryologist) and (invitra staff).
Last Update: 03/18/2020

Embryo culture is one of the most important phases of in vitro fertilization (IVF) treatments. It is performed in the IVF laboratory and the conditions of the laboratory directly influence the quality of the eggs and embryos.

When is the embryonic culture performed?

Culture in the in vitro fertilization laboratory begins after obtaining the eggs by means of follicular puncture, also called ovum pick-up. Initially, they will be cultured without removing the cell layer of the surrounding granulose (decumulation).

The day of the puncture is what is known as day 0 of the embryonic development, as this is when fertilization (union of the egg and sperm) occurs.

Depending on the IVF technique applied to fertilize the eggs, the next step will be:

Classic IVF
the undecumulated eggs and sperm are brought together.
the eggs are decumulated and the intracytoplasmic microinjection of the sperm is performed.

The culture of the embryos obtained after fertilization will be extended until the day the embryos are:

  • transferred to the mother's uterus, which is usually on day 3 or in the blastocyst stage (day 5-6).
  • Vitrified.
  • Are discarded.

Stages of embryonic development

Next we will briefly see which stages embryologists evaluate in order to select the embryo most likely to be implanted:

Day 1
is the day after the follicular puncture and is when it is evaluated, 16-18h after fertilization, whether it has occurred correctly. At this stage, the embryos are called zygotes. Its metabolic needs are very similar to those of the oocytes.
Day 2
the embryos have already made the first divisions and have 2-4 cells, also known as blastomeres.
Day 3
at this point, the embryos have about 6-8 blastomeres. The embryo has already begun to express its own genes, so its energy needs are gradually changing.
Day 4
the embryo is in a morula stage and compacts.
Days 5 and 6
these days, the embryo reaches the blastocyst stage. Embryos that reach this stage with good morphological characteristics have a high probability of implantation.

How should the cultivation be?

There may be some variations in the characteristics of the embryo culture between different clinics or even be customized for each patient. However, they always agree that the conditions have to be perfectly regulated, since any alteration can affect the embryonic development and compromise the probabilities of success of the treatment.


Depending on the duration of the culture, a distinction is made between

  • Short cultur: until day 2 or 3 of development.
  • Long culture: up to the blastocyst stage You can use the same type of media throughout the development (single media) or use several different ones (sequential media).

Whether one type of culture is carried out or another depends on several factors, such as the quantity of eggs obtained, the quality of the embryos on day 3 or the need to carry out a preimplantation genetic diagnosis (PGD).

In some cases, co-culture is also performed, which consists of cultivating the embryos with endometrial cells to improve the conditions of the environment and, thus, development. However, it is a rare technique in the clinic and is used more in the field of research.

Necessary conditions

The eggs and embryos should be in an environment that is as stable as possible. For this purpose, they are kept inside incubators for most of the time, as this way the culture conditions are not altered. The following aspects are controlled:

  • Temperature
  • Carbon dioxide and oxygen concentration
  • Humidity

In addition, it is also necessary for assisted reproduction laboratories to have highly controlled conditions. Some of the risk factors are:

  • Air purity (with very few suspended particles)
  • Light
  • Temperature
  • Gases present and in what percentage

Embryo culture media

The embryos develop in plates with culture media that provide them with the necessary nutrients to evolve correctly. As mentioned above, their energy needs vary at different stages. Depending on its composition, we distinguish:

Sequential media
contain the specific nutrients that embryos need at certain stages of their development. There is usually a medium for the first two days of embryonic development and another for use from the third day onwards.
Unique media
contain all the nutrients the embryo needs from day one of development to the blastocyst stage. The embryo will consume what it needs at each moment.

FAQs from users

By Cristina Duque Royo MSc, PhD (senior clinical embryologist).

If it were possible, the ideal would be to always perform the blastocyst culture. Over 5- days of culture, we will assess all the embryonic events that must happen (cleavage, compaction, blastulation etc), these allow a better embryo selection, which translates into a higher rate of implantation and pregnancy. If we also apply time-lapse technology, we will obtain more information for an even better embryo selection.

How do I know the chances of my embryos becoming blastocysts?

By Carmen Ochoa Marieta MD, PhD, MSc (gynecologist).

From the clinical point of view, young women without pathology are more likely to have embryos that reach blastocyst and from the laboratory point of view, those embryos of good quality that correctly comply with cell division times and maintain a perfect morphology in their early development, are also more likely to reach blastocyst.

Which is better, the short cultur or the long one?

By Rebeca Reus BSc, MSc (embryologist).

It depends on each case, since each type of culture offers a series of advantages and disadvantages. On the one hand, it is easier to carry out the short culture and, if there are not many embryos available, it is more likely that some will reach the day of the transfer.

On the other hand, by means of the long culture we can better select which is the embryo with more probabilities of implanting, since only those that are of good quality reach this stage.

Therefore, each patient will be more interested in one culture or another depending on the characteristics of their case.

Do all laboratories have the same conditions?

By Rebeca Reus BSc, MSc (embryologist).

No, because the parameters that are best for one clinic may not be the best for another. For example, some laboratories obtain better results using low oxygen concentrations in incubators, while others see no significant difference.

Are the eggs cultivated in the same way as the embryos?

By Rebeca Reus BSc, MSc (embryologist).

There are some differences in the composition of the culture media used in some and in others, but in general the conditions are very similar.

Despite the fact that both are very delicate, the eggs are extremely sensitive to changes in temperature, so sudden changes must be avoided when handling them from the moment of follicular puncture.

Suggested for you

Embryo culture allows us to evaluate the evolution of the embryos, which is a key aspect in achieving pregnancy through IVF. Did you know that the evaluation of the zygotes (day 1 of development) is fundamental to discard some embryos with abnormal chromosome load? You can find more information on this subject at the following link: Classification of the embryos according to their quality.

In recent years, there has been a very important improvement in the field of embryo evaluation thanks to the application of time-lapse technology in culture. It allows us to visualize the embryonic development continuously and without altering the culture conditions, providing a lot of new information. If you want to read more about this technique, we recommend you read this article: What is time-lapse?

Despite this progress, not all embryos are able to reach the blastocyst stage. Therefore, deciding whether the transfer takes place on day 3 or in the blastocyst stage remains a controversial issue. You can learn more about the advantages and disadvantages of each of these options at the following link: Embryo transfer on day 3 or day 5?

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Authors and contributors

 Carmen Ochoa Marieta
Carmen Ochoa Marieta
MD, PhD, MSc
Bachelor's Degree in Medicine from the Basque Country University. PhD in Medicine & Surgery from the University of Murcia. Currently, she is the director of the Assisted Reproduction Unit of Centro de Estudios para la Reproducción (CER SANTANDER) in Santander, Spain, as well as the director of the Diagnostic Unit of Human Assisted Reproduction in Bilbao. More information about Carmen Ochoa Marieta
License: 484805626
 Cristina  Duque Royo
Cristina Duque Royo
MSc, PhD
Senior Clinical Embryologist
Specialist in Assisted Reproduction with a Master's Degree in Clinical and Experimental Biology of Human Reproduction from the University Hospital la Fe of Valencia ans PhD from the Unisersity of Zaragoza More information about Cristina Duque Royo
 Laura Garrido
Laura Garrido
BSc, MSc
Bachelor's Degree in Biotechnology from the Pablo de Olavide University (UPO) of Seville, Spain. Master's Degree in Biotechnology of Human Assisted Reproduction from the University of Valencia (UV) and the Valencian Infertility Institute (IVI). Experience at IVF, andrology, and general analysis laboratories. Embryologist specialized in Assisted Reproduction. More information about Laura Garrido
 María de Las Heras Martínez
María de Las Heras Martínez
BSc, MSc
Clinical Embryologist
Bachelor's Degree in Biology from the Pompeu Fabra University and Master's Degree in Biology of Reproduction & Assisted Reproductive Technologies from the Autonomous University of Barcelona, in collaboration with Instituto Universitario Dexeus. Master's Degree in Biochemical Research from the Basque Country University. Clinical Embryologist by the ESHRE. More information about María de Las Heras Martínez
 Rebeca Reus
Rebeca Reus
BSc, MSc
Degree in Human Biology (Biochemistry) from the Pompeu Fabra University (UPF). Official Master's Degree in Clinical Analysis Laboratory from the UPF and Master’s Degree about the Theoretical Basis and Laboratory Procedures in Assisted Reproduction from the University of Valencia (UV). More information about Rebeca Reus
Adapted into english by:
 Romina Packan
Romina Packan
inviTRA Staff
Editor and translator for the English and German edition of inviTRA. More information about Romina Packan

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