IVF SUCCESS DEPENDS ON FERTILIZATION

The fundamental process in IVF is fertilization itself. The fact that the human egg can be fertilized by human sperm in a laboratory is a scientific miracle without equality. Technologies for optimizing sperm function and fertilization rates in vitro are now widely available. These techniques are important for the routine handling of sperm and even more important in cases of sperm deficiency. This is important since 45% of cases involve male factor issues, and male factor is the exclusive diagnosis in about 1/3 of cases.

Our specialists at Fertility & Lifespan Medical Institute in San Diego, California, led by reproductive endocrinologist PhD, utilize the latest techniques to optimize fertilization to save and maintain your eggs when you’re younger, making successful pregnancy more likely.

How Do Sperm Cells Fertilize Eggs?

Sperm samples must be prepared carefully prior to IVF. The male partner produces the sample by masturbation before or just after the egg retrieval. I recommend abstaining from sex for 2–5 days. Longer periods of abstinence may increase the number of dead sperm or cellular artifacts. If a frozen semen sample is used, the processing of thawing the sample will occur later that morning. The andrologist will evaluate the semen analysis for count, motility, and morphology. After that, the goal is to separate the sperm cells, which are most likely to fertilize an egg.

After the semen analysis is complete, the semen sample is filtered by being placed in a column in a centrifuge tube. This is done to isolate the sperm cells with the best motility and morphology. The ‘swim-up’ method takes advantage of the fact that the more viable sperm cells will swim up into a separate population from the media. Another technique is to prepare the sperm sample by centrifugation. In this process, we use a density gradient, which consists of small particles that can separate one sperm population from another.

Can Sperm Be Selected Without Prior Processing?

An interesting technique for sperm preparation with IVF is called ZyMot. The technical term for this is a microfluidic chip. It is best used to select sperm cells for ICSI. Remarkably, the sperm sample does not need to be processed before it is applied to the chip. This avoids the need for the sperm sample to undergo centrifugation, which can stress or damage the sperm cells. However, there must be at least 1 million total motile sperm cells per cc.

With the ZyMot system, the sperm cells that are most motile swim through the membrane of the device, mimicking the natural way that the body selects sperm. This procedure may be particularly useful in men with abnormal DNA fragmentation on a test called the sperm chromatin assay.

What Methods Are Used?

The day of egg retrieval is called Day 0. The laboratory personnel will inseminate the eggs 4–6 hours after egg retrieval. All the eggs are placed in aliquots of fluid consisting of 100,000 sperm cells per egg. The eggs in the petri dish are then incubated overnight in a Petrie dish. This is ‘routine IVF.’

With routine IVF, the sperm penetrates the outer shell surrounding the egg, called the zona pellucida, on its own. It then fuses with the membrane of the egg, called the oolemma. The genetic material enters the cytoplasm of the cell, called the ooplasm. The sperm cell goes through a fertilizing reaction in its head called the acrosome reaction.

Penetration into the egg activates the final genetic stage of egg maturity. The chromosomes of the mother and father line up. The egg sends off its final extra set of 23 chromosomes into a structure called the second polar body, just outside of the main cell membrane. Once initial sperm cell penetration occurs, the zona pellucida becomes even thicker, preventing additional sperm cells from penetrating.

How Is Fertilization Confirmed?

The embryologist checks the culture medium about 16–20 hours after insemination for evidence of fertilization. A 400x inverted microscope is used. The determination of fertilization is confirmed by the appearance of two round structures in the center of the cytoplasm. Each round structure contains the genetic material from the mother and father, respectively. These round structures are called pronuclei because they occur ‘before’ the formation of the single nucleus of the embryo.

Both pronuclei look identical; you cannot tell which is from the male or which is from the female. One circle represents the nucleus of the egg, and one circle represents the nucleus of the sperm cell. The initial appearance of the fertilized egg is called a zygote.

Fertilization is transient, so the pronuclei are only visible for a few hours. Within the first day, the two pronuclei will come together and join in forming a single nucleus, constituting a single-cell embryo. The process of fertilization is completed when the two pronuclei have come together and combine their chromosomes into a single nucleus.

The nucleus will contain 23 chromosomes from the egg and 23 from the sperm. These 46 chromosomes will assemble into 23 pairs. Only the eggs with normal fertilization will continue to grow in the laboratory and form normal embryos.

Another feature of fertilization is the formation of a second polar body. It is the small round or oval structure just outside of the fertilized egg itself but inside the thick shell of the zona pellucida. In the illustration, there are two small blue circular dots. Each one is a polar body. Each one contains a discarded set of chromosomes from the egg as it matures.

What Is Genetic Maturity?

There are two stages of meiosis: the genetic process in the egg leading to maturity. At the end of meiosis I, the egg discards an extra set of chromosomes and forms the first polar body. The presence of the first polar body confirms that the egg is mature and ready for fertilization. At the time of fertilization, the egg discards its final extra set of chromosomes and forms the second polar body. Subsequently, the round nucleus of the egg and the round nucleus of the sperm cell fuse and form a single nucleus. The fertilized egg is now a one-cell embryo.

Mitosis is the genetic process by which the embryo continues multiplying and forming additional cells. Mitosis becomes apparent about 24 to 28 hours after the sperm cells breach the eggs. Some eggs will not fertilize, either due to a sperm problem or an egg problem. After fertilization is confirmed, the embryologist will transfer the fertilized embryos into a new culture medium that is designed to aid the growth of multicellular embryos.

Schedule a consultation with Dr. Brody at Fertility & Lifespan Medical Institute or call our office at 858-216-2096 to get the best, and most personalized, fertility care.