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While every clinic's protocol will be slightly different and treatments are adjusted for individual needs, here is a summary of what generally takes place during an IVF treatment cycle.
Sometimes birth control pills are used before IVF treatment begins.
During the cycle before your IVF treatment is scheduled to start, you may be put on birth control pills. This may seem awkward – because you’re trying to get pregnant. Using birth control pills before starting a treatment IVF cycle has been shown to decrease your risk of Ovarian Hyper Stimulation Syndrome and ovarian cysts and may even improve the rate of success.
Another possibility, if you rarely get cycles on your own, progesterone is given, to bring on your period.
The doctor may ask you to take an injection about 4 – 5 days before the last birth control pill, depending on the protocol of ovarian stimulation he’ll use later. Always follow the doctor's instructions.
The first official day of your treatment cycle is the day you get your period. Spotting is not considered as day 1. On the second day of your period, the doctor will likely order blood exams and a vaginal ultrasound. (A vaginal ultrasound during your period isn't exactly pleasant, but it’s necessary) This is referred to as your baseline blood exam and your baseline ultrasound.
The ultrasound is to check the size of your ovaries, and look for ovarian cysts. If there are cysts, the doctor may just delay treatment, as most cysts will resolve by their own with time. In other cases, your doctor may aspirate the cyst with a needle. The doctor will check also the thickness or the lining of the endometrium to evaluate its receptivity and ability for embryo implantation. If everything looks OK, treatment moves on to the next step.
The next step is ovarian stimulation with fertility drugs. Depending on your treatment protocol, this may mean injections either IM (intra muscular) or subcutaneous for approximately 8 – 14 days. The staff of our clinic will teach you how to make the injections before your treatment begins.
During the ovarian stimulation, the patient has to visit our clinic several times because the physician will monitor the growth and development of the follicles. This will include blood tests to measure Estradiol, LH, and progesterone levels and vaginal ultrasounds, to monitor the oocyte growth and size. Monitoring the cycle is important, as it helps the doctor decide whether or not the medications need to be increased or decreased in dosage. Once your largest follicle is 16 to 18mm in size, the doctor will start a new phase.
The next step in your IVF treatment is triggering the oocytes to go through the last stage of maturation, before they can be retrieved. This last growth is triggered with human chorionic gonadotropin (hCG). Timing this shot is important. If it's given too early, the eggs will not be mature enough. If given too late, the eggs may be empty, because they already ovulated. The daily ultrasounds at the end of the last step are meant to time this trigger shot just right. Usually, the hCG injection is given when four or more follicles have grown to be 18 to 20mm in size and your estradiol levels are not less than 1,000pg/ML.
The timing of the shot will be based both on your ultrasounds and blood tests and when the doctor schedules your egg retrieval at the clinic. The egg retrieval takes place approximately 35 - 37 hours after the trigger shot is done.
If not enough follicles grow or if you're at risk for severe ovarian hyper stimulation syndrome, your treatment cycle may be canceled and the hCG shot will not be given. If the treatment is canceled because your ovaries didn't respond well to the medications, the doctor may recommend different medications to be tried on the next cycle.
Because of the risk of Ovarian Hyper Stimulation Syndrome, when you experience unusual cramping + difficulty in breathing, be sure to tell Dr. Najib Dagher right away.
The egg retrieval or pickup is done under a quick anesthesia (light sedation) in the Operation room at the clinic. Usually, a light sedative is used, which will make you "sleep" through the procedure. This isn't the same as general anesthesia, which is used during surgery. Side effects and complications are less common.
The egg retrieval will take place about 35 to 37 hours after doing the triggering shot. It's normal to be nervous about the procedure, but all women go through it without any trouble or pain.
The doctor will use a trans-vaginal ultrasound to guide a needle through the back wall of your vagina, up to your ovaries. He will then use the needle to aspirate the follicular fluid gently from the follicles in to the needle to a tube. There is one oocyte per follicle. These oocytes will be transferred to the embryology lab for fertilization.
The number of oocytes retrieved varies but can usually be estimated before retrieval via ultrasound. The average number of oocytes is 5 to 15, with more than 95% of patients having at least one oocyte retrieved.
After the retrieval procedure, you'll be kept for a few hours to make sure all is well. Light spotting is common, as well as low abdominal cramping, but most women feel better in a day or 2 after the procedure. You'll also be told to watch for signs of ovarian hyper-stimulation syndrome, a side effect from fertility drug use during IVF treatment.
While you’ll be recovering from the retrieval, the follicles that were aspirated will be searched for oocytes, or eggs. Not every follicle will contain an oocyte.
Once the oocytes are found, they'll be evaluated by the embryologists. If the eggs are overly mature, fertilization may not be successful. If they are not mature enough, the embryology lab may be able to stimulate them to maturity in the lab.
The eggs will be at different stages of maturity. Not all the stages are good for fertilization:
The eggs are classified into: M2 (Metaphase 2), M1 (Metaphase 1), GV (Germinal Vesicle). The M1 can be developed to M2 in the lab. Also the GV oocyes can undergo IVM (In Vitro Maturation) in the lab. Only the M2 eggs can be fertilized.
Before release from the ovary, oocytes are stopped at an early stage of the first meiotic division as a primary oocyte (primordial follicle). During each menstrual cycle, pituitary gonadotrophin stimulates completion of meiosis 1, the day before ovulation. Early oocytes are also classified as immature (germinal vesicle (GV) or metaphase I (MI) stage). The breakdown of the germinal vesicle indicates the restart of meiosis and the extrusion of the first polar body (1 PB) indicates completion of the first meiotic division in human oocytes.
The released oocyte is surrounded by a thick extracellular matrix, the zona pellucida, which is covered by layers of cells, the granulosa layer.
In an adult human female, the development of a primordial follicle containing an oocyte to a pre-ovulatory follicle takes about 120 days.
In females, the total number of eggs produced is present in the newborn female initially stopped at the diplotene stage of the meiosis I from fetal life through childhood until puberty. During puberty, when the Lutenizing hormone (LH) surges it stimulate the resumption of meiosis.
1. All eggs are stopped at an early stage (prophase I) of the first meiotic division as a primary oocyte (primordial follicle). Following puberty, during each menstrual cycle, pituitary gonadotrophin stimulates completion of meiosis 1 the day before ovulation.
2. In meiosis 1, a diploid cell becomes 2 haploid (23 chromosomes) daughter cells, each chromosome has two chromatids. One cell becomes the secondary oocyte the other cell forms the first polar body.
3. The secondary oocyte then commences meiosis 2 which stops at metaphase and will not continue without fertilization.
4. At fertilization, meiosis 2 completes, forming a second polar body. Note that the first polar body may also undergo this process forming a third polar body.
Human oocyte at metaphase II showing polar body at top
The breakdown of the germinal vesicle indicates a resumption of meiosis and the extrusion of the first polar body (1 PB) indicates completion of the first meiotic division in human oocytes. The polar body is a small cytoplasmic exclusion body formed to enclose the excess DNA formed during the oocyte (egg) meiosis and following sperm fertilization. There are 2-3 polar bodies derived from the oocyte present in the zygote, the number is dependent upon whether polar body 1 (the first polar body formed during meiosis 1) divides during meiosis 2. This exclusion body contains the excess DNA from the reductive division (the second and third polar bodies are formed from meiosis 2 at fertilization). These polar bodies do not contribute to the future genetic complement of the zygote, embryo or fetus.
Fertilization of the oocytes must happen with 12 to 20 hours. Your partner will provide his sperm sample the same morning you have the retrieval. After the eggs and sperm are collected, sorted, and prepared, the Embryology team begins the fertilization process.
Once the semen sample is given, we’ll wait until it will liquefy, then a special washing process will take place (Simple wash, Swim up, Density gradient centrifuge, swim out) which separates the sperm from the other materials that are found in semen.
There are two ways that fertilization can occur:
Conventional Fertilization: Is frequently used in cases such as blocked fallopian tubes or unexplained infertility. The embryologist isolates the healthy sperm which are then exposed to each egg inside the embryology laboratory where fertilization occurs naturally.
Intra-cytoplasmic Sperm Injection (ICSI): The embryologist selects a single best looking healthy sperm and injects it directly into the center of each egg. ICSI is used in cases when the quantity or quality of sperm is poor and therefore unable to effectively penetrate the egg on its own. Since fertilization only requires one healthy sperm, ICSI has become one of the most incredible advances in treating severe male factor infertility.
The decision about which method to use is based on the quality of the sperm. Sperm analysis is completed as part of the initial basic fertility work-up for diagnosis before the treatment and again when the semen sample is provided on the day of the egg retrieval. If the results from this analysis do not meet the parameters required for conventional fertilization, the embryologist will make the decision to switch to ICSI so that the cycle can still produce embryos and increase your chances of success. The clinical team will let you know if an unplanned ICSI procedure is recommended.
Once fertilization occurs, the embryos begin to develop. Every morning for the next 3 to 6 days, an embryologist examines the developing embryos and adds notes in your record regarding each individual embryo. A daily follow-up concerning your fertilization will take place updating you on the status of each growing embryo and the recommendations for either a day 3 or a day 5 embryo transfer.
Indications of different ART technologies:
IVF+ICSI: The embryologist will choose a healthy-looking sperm and fertilize the oocyte with the sperm using a special thin needle. One healthy sperm is injected into each egg.
IVF +IMSI: The sperm is selected under a high power light microscope (enhanced by digital imaging) to magnify the sperm sample over 6000 times. One healthy sperm is injected into each egg.
IVF+PGD (Preimplantation Genetic Diagnosis) or gender selection (sex selection):
The embryos in the lab are created from the couple's eggs and sperm, on the third day after fertilization, when embryos have reached 6- to 8-cells, one of the cells is removed from the embryo, to be analyzed for its chromosomal makeup. (The removal of the cell at this stage does not negatively affect the embryo's growth competency.) This chromosomal analysis allows us to determine whether the embryo is male or female. Then, only the embryos of the desired gender are transferred to the uterus. PGD gender selection is the most reliable method for gender or sex selection, 99% accuracy.
Reasons for Preimplantation Genetic Diagnosis (PGD):
For Medical Reasons
Preimplantation Genetic Diagnosis (PGD) can be used to prevent and avoid genetic diseases.
Elective Gender Selection
Gender selection can be "non-medical" or "elective." In such cases, a child of a specific gender is desired without obvious medical indications. The most frequent indication for such gender selection is "family planning," when one gender is already represented in the family and the other gender is desired.
Surgical sperm collection:
PESA, MESA, TESA andTESE
Overview of the male reproductive system
The function of the male reproductive system is to produce, store and transport the sperm outside the body. The organs that produce sperm are the testes. Sperm production begins with immature sperm cells that grow and develop within the seminiferous tubules. These tubes are very tiny and the sperm inside them are not fully mature. As a result they are unable to move on their own. As they travel along the length of the epididymis they mature and become motile. During ejaculation they are carried from the epididymis to the penis along the vas deferens.
Until recently there was no treatment available for men who have a complete absence of sperm in the ejaculate (azoospermia), and it has been estimated that about 10–15% of cases of male infertility are due to azoospermia. Azoospermia has many causes; some of the causes are called “obstructive” meaning that there is a blockage in the sperm delivery system. Other causes are “non-obstructive” meaning that there is an absence or a very marked reduction of sperm production in the testes.
Obstructive azoospermia accounts for about 40% of azoospermia cases. Obstruction may result from defects in any of the ducts (passage ways) involved in the sperm delivery system. The obstruction may be either congenital (you were born with it) or acquired (you were not born with it). Vasectomy is a common form of male contraception. With this the vas deferens is cut forming an acquired obstruction. It is the most common cause of obstruction. Another cause is infection, which can scar the epididymis. Congenital obstruction can be due to either a malformation or the absence of a ductal structure. Congenital absence of the vas (CAV) is a genetic disorder associated with cystic fibrosis and with this the vas deferens is either absent or malformed. If CAV has been diagnosed your doctor will advise you on the correct course of action. In obstructive azoospermia the reason for the absence of sperm in the ejaculate is physical and in general, does not involve the process of sperm production. Therefore in most cases surgically retrieved sperm are normal in their function and fertilization rates and pregnancy rates are similar to those obtained using ICSI on ejaculated sperm.
The three major causes for reduced sperm production are hormonal problems, testicular failure and varicocele. A significant proportion of men with non-obstructive azoospermia have testicular failure caused by chromosomal abnormalities such as Klinefelter’s syndrome or abnormalities of the Y chromosome. Your doctor will discuss in detail what the problem is and how best it might be treated. If azoospermia is due to chromosomal abnormalities the concern is that male offspring could inherit the disorder, and therefore the implications of inheritance of the genetic disorder need to be considered.
Surgical Sperm Collection
Micro Epididymal Sperm Aspiration (MESA)
MESA involves aspiration of sperm from the epididymis with a fine needle (Figure 1). It is a surgical procedure and is carried out under a general anesthetic. Sperm collected using this procedure are often of poor quality but are usually suitable for cryostorage. One aspiration may provide enough sperm for several attempts at IVF using ICSI. MESA can be performed well in advance of any proposed IVF procedure.
Testicular Sperm Extraction (TESE)
TESE involves taking a small piece of tissue from the testis and isolating the sperm from the seminiferous tubule (Figure 2). The number of sperm isolated is often very small (usually less than with MESA). The procedure is thus performed typically shortly prior to the egg collection procedure. Originally TESE was only performed in cases of non-obstructiveazoospermia, however because the procedure can be performed under local anesthetic using a biopsy needle (TESA), it has become the method of choice for all types of azoospermia in some clinics. A surgical biopsy may be less damaging to the testis than a needle biopsy, and is probably less painful.
In some cases live sperm will not be obtained. Your options are:
1. IVF oocyte (egg) collection may be cancelled, or
2. Oocytes collected can be frozen.
3. Sperm donation.
Non-use of immature sperm
In some cases of non-obstructive azoospermia only immature sperm are obtained. Fertilization rates with immature sperm are often quite poor. Even if fertilization does occur and pregnancy follows an embryo transfer, the rate of miscarriage is two to three times higher than in pregnancies obtained using mature sperm. Recent studies have shown that this result may be linked to an increase in the level of a chromosomal disorder called Mosaicism, which is itself linked to sperm immaturity. For this reason we do not inject immature sperm or sperm that are immotile. If mature motile sperm cannot be located then the procedure will be abandoned.
A consent form requesting the above techniques must be signed before commencing a surgical sperm collection.
Percutaneous Epididymal Sperm Aspiration (PESA)
PESA is a simple technique to obtain sperm for Intra Cytoplasmic Sperm Injection (ICSI) in men who have an obstruction of the vas deferens, either due to vasectomy or other obstruction. To minimize scarring and damage, PESA usually is attempted on one side only. It is sometimes necessary to aspirate from both sides. Sufficient sperm for ICSI is obtained in 80% of attempts. In 10% of cases enough suitable sperm is found for cryopreservation.
The procedure is performed in the IVF Centre rooms (Figure 3). After the procedure the man will be asked to wear a very tight pair of underpants to provide support to the scrotum. There is no other special preparation for the patient.
PESA is performed under local anaesthesia. This means that an anaesthetic is injected into the scrotum by the specialist to make the area numb. When this has been achieved the doctor will swab the scrotum with a warm antiseptic. The doctor will examine the testes to locate the vas deferens by gently feeling the scrotum. A small needle will be inserted into the vas deferens and the doctor will instruct the nurse assisting to draw back on the plunger in order to aspirate seminal fluid. When fluid is obtained it is passed to the embryologist to be examined for motile (moving) sperm. The procedure may need to be attempted again until motile sperm have been found.
The procedure is usually performed just prior to the woman’s oocyte collection (on the same day). If no sperm is retrieved the oocyte collection may be cancelled.
What is assisted hatching?
For the first 5 to 7 days of development, the embryo is surrounded and protected by an outer shell called the zona pellucida. In normal circumstances, when the embryo reaches the uterus, this zona partially dissolves and the embryo ‘hatches’ out, allowing it to implant in the uterus.
In some patients it is thought that infertility may be caused by a hardening of the zona, which makes it difficult for the embryo to hatch and implant.
Assisted hatching is a laboratory micromanipulation technique carried out before the embryos are replaced in the uterus that helps the embryo to hatch from the zona.
What does assisted hatching involve?
Assisted hatching is carried out in the laboratory by experienced embryologists. Using a very high powered microscope, a small slot is made in the zona using a very fine needle. Assisted hatching is carried out before the embryo transfer on those embryos that have been chosen for transfer.
Once it is returned to the uterus, the embryo can hatch through this opening and implant naturally.
Who is assisted hatching suitable for?
Assisted hatching is generally recommended in the following circumstances:
The woman is aged 35 years or older
The woman has high FSH levels
Couples who have failed to get pregnant following previous IVF cycles
Couples where a distinct thickening of the zona is noted by the embryologist.
Frozen embryo replacements.
The embryo transfer is an easier procedure than the egg retrieval.
About three to five days after the retrieval, the fertilized eggs will be transferred. The procedure for embryo transfer is just like IUI treatment. You don’t need anesthesia.
During the embryo transfer, a thin tube, or catheter, will be passed through your cervix from the vagina. You may experience very light cramping but nothing more than that. Through the catheter, they will transfer the embryos, along with a small amount of fluid.
The number of embryos transferred will depend on the quality of the embryos and previous discussion with the doctor. Depending on your age, from one to five embryos may be transferred.
After the transfer, you'll stay lying down for a couple hours and then head home.
If there are "extra" high-quality embryos left over, you may be able to freeze them. This is called "embryo cryopreservation." They can be used later if this cycle isn't successful.
You’ll need bed rest for 3 days after the embryo transfer.
How is embryo quality assessment and grading done on day 3 of development?
We assess the "quality" of embryos from in vitro fertilization by carefully evaluating and scoring some aspects of their appearance.
Embryos should be at 2 to 4 cells at 48 hours after egg retrieval and preferably about 7 to 10 cells by 72 hours. The cells in an embryo are also referred to as "blastomeres".
Cell regularity - degree of regularity of size of blastomeres
It is generally better if the size of the individual cells (blastomeres) in the embryos is similar in size. If they are not, it is better if they are close to the same size.
Degree of fragmentation
Fragmentation, also called blebbing, is a process where portions of the embryo's cells have broken off and are now separate from the nucleated portion of the cell. It is preferable to have little or no fragmentation.
However, fragmentation in human embryos is quite common and many beautiful babies have resulted from implantation of embryos with fragments. Many IVF labs, including ours estimates and records the percentage of fragmentation in each day 3 embryo. Embryos with more than 25% fragmentation have a low implantation potential.
Presence of multi-nucleation
An embryo is multinucleated if more than one nucleus can be seen in any individual cell on either day 2 or day 3. After day 3 it is extremely difficult to identify the presence of multi-nucleation. The great majority of multinucleated embryos have been shown to be chromosomally abnormal in PGD, preimplantation genetic screening studies.
However, sometimes multinucleated embryos implant and lead to a healthy pregnancy and birth of a normal baby. These embryos are generally only transferred to the uterus if they are the only ones available.
Additional factors involved with embryo grading and selection for transfer
Other aspects of the microscopic appearance of the embryos are also noted, including the presence of vacuoles, granularity, and thickness of the outer shell (or zona pellucida) of the embryo, etc.
Usually, the embryo "quality" is determined not until at least 48 hours after the egg retrieval.
By 48 hours ("day 2"), the embryos must be to at least 2 cells - or they have "stopped". It’s better to have some of them at the 3 or 4 cell stage by then.
By 72 hours ("day 3"), better to have at least 6 cells - and preferably some at about 8 cells. We have seen babies that came from 4 cell embryos on day 3, but chances for pregnancy increase significantly with increasing cell number.
Embryos with higher cell numbers, regular appearing cells and little or no fragmentation have a higher overall chance of implanting than do other embryos with less cells, more irregularity and significant fragmentation.
Embryo quality as we see it under the microscope in the IVF lab gives us some reasonable ability to predict the chances for pregnancy after the embryo transfer procedure. However, because there are many other contributing factors involved that we cannot see or measure, the generalizations about "quality" made from grading embryos are often inaccurate.
We see some cycles fail after transferring 3 perfect looking embryos, and we also see beautiful babies born after transferring only one "low grade" embryo. The true genetic potential of the embryo to continue normal development is impossible to measure accurately with current technology.
An important variable that is often overlooked is the embryo transfer technique. A smooth transfer with no trauma to the endometrial lining is essential to give the embryos the best chance for implantation and continuation of normal development.
Most IVF clinics "grade" each embryo using one of many scoring systems. Unfortunately, there is no agreement at all as to which system to use.
Patients often ask whether embryos that were given a "low grade" by the embryologist would result in a problem with the baby. As far as we know, the children born from low grade embryos are just as cute, intelligent, strong, etc. as those born from high grade embryos. The only difference seems to be with the chance for the embryo(s) to result in a pregnancy.
Embryo quality, to a great extent, is determined by the quality of the egg from which it started. If we could improve the quality of the eggs that we start with in IVF, we would have a better chance of having a successful outcome. However, for the most part egg quality is predetermined and "we get what she gives".
Most of the egg quality is determined by the chromosomal and genetic competence of the individual egg. There is no treatment that can fix chromosomal or genetic defects - so we do our best with the eggs that we retrieve.
By getting numerous eggs, we have a better chance of having one or more with "high egg quality" - in other words, an egg that is fully competent and capable of fertilizing, developing normally, implanting and going on to become a healthy baby.
Progesterone support and the two week of waiting the result of blood test beta Hcg to know if you’re pregnant or not.
On the next day after your retrieval, and before the embryo transfer, you'll start taking progesterone supplements. Usually, the progesterone during IVF treatment is given as intramuscular plus pills, or vaginal gel or vaginal suppositories.
At the end of the two week wait, you’ll do a blood test for pregnancy outcome.
If the test is positive, you may need to keep taking the progesterone supplementation for another several weeks. About 10 – 14 days after the positive pregnancy test the doctor will follow up with occasional blood tests and ultrasounds to monitor the pregnancy and watch for miscarriages or ectopic pregnancies.
The doctor will also monitor whether the IVF has led to a multiple pregnancy. If it's a multiple pregnancy (3 or more babies), the doctor may discuss the option of reducing the number of fetuses in a procedure called a "multifetal pregnancy reduction." This is done to increase the chances of having a healthy and successful pregnancy.
If the pregnancy test is negative 12 to 14 days post-transfer, you have to stop taking the progesterone, and book an appointment with the doctor to discuss why the IVF failed and how to improve the outcome of a possible next IVF, and you'll wait for your period to start. The next step will be decided among you, your partner and the doctor.
The failure of a treatment cycle is never easy. It's heartbreaking. It's important, however, to keep in mind that having one cycle fail doesn't mean you won't be successful if you try again.