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In Vitro Fertilization (IVF)
EGG COLLECTION FERTILIZATION EMBRYO TRANSFER CRYOPRESERVATION

The test-tube baby technique

The fertilization of an egg by sperm.

With in vitro fertilization, a doctor uses a needle inserted through the wall of the vagina to remove eggs from the ovary. The needle is guided by ultrasound to reach the fluid-filled ovarian follicles that contain the eggs (A). Once retrieved from the ovary, the eggs are combined with sperm in a petri dish (B) and placed in an incubator (C). If fertilization occurs, the embryos are transferred by means of a fine, ultrasound-guided cannula (D) to the uterus 3 to 5 days later (E).

Egg collection

This is usually performed in the morning or early afternoon, approximately 36 hours after the hCG injection. Some clinics allow the male partner to attend the egg collection procedure if their wives have the operation under sedation. A monitor connected to the microscope will allow you to view the eggs when collected.

Methods of egg collection

Several methods can be employed to collect the eggs, these include:

Vaginal ultrasound guided egg collection

This is the most common technique; it is a minor and safe surgical procedure usually performed under sedation or a general anesthetic. Sedation is a safe and ecceptable method of providing pain relief for egg collection.

A vaginal ultrasound probe with a fine hollow needle attached to it, is inserted into the vagina. Under ultrasound guidance, the needle is then advanced from the vaginal wall into the ovary to suck out the fluid from the follicle which contains the egg. Each egg is removed in turn through the needle by a suction device. Follicle flushing is not associated with improvement in pregnancy rates or the number of eggs collected, but does increase the duration of the procedure and associated pains. The whole procedure takes about 20-30 minutes.

You may experience some mild discomfort following the procedure, but this will be relieved with painkillers. Antibiotic is usually given to prevent infection.

Abdominal ultrasound guided egg collection

Occasionally, egg collection is performed by passing a needle through the abdominal wall into the ovaries under ultrasound guidance. This is usually performed if the ovaries are abnormally placed.

Laparoscopy

Originally, eggs were always collected laparoscopically. This method of egg collection is hardly ever used nowadays, as it requires a general anesthetic, in addition to the risks of laparoscopy.

The average number of eggs collected is about 12 depending upon the number of follicles present. Not every follicle contains an egg. Occasionally, no eggs are collected, so-called “empty follicle syndrome”. The reported incidence is about 1%.  Sometimes, giving another dose of hCG and scheduling another egg collection 24 hours later could salvage the cycle. The cause of empty follicle syndrome is unknown, but it is possible that it is a drug related rather than a clinical problem.

What happens after egg collection?

Pain

The patient may experience pain. It is not unusual for women to experience some abdominal or pelvic pain. A hot water bottle or painkillers are often helpful. However, if the pain is severe, or persists, then you should consult your doctor.

Bleeding

The patients may have to wear panty liners for a day or two following egg collection. Any bleeding should be minimal and dark or brown in colour. If bleeding is severe or bright red then consult your doctor.

Nausea and vomiting

The patient may experience nausea or vomiting for the first 24 hours following egg collection. This is usually a side-effect of the drugs given (anaesthetics or pain killers). If the nausea or vomiting persists, the you should consult your doctor.

Activities

You are advised to rest during the first 24 hours following surgery and must not operate any machinery such as a car or cooker. Furthermore, a responsible adult must be available to look after you during this period.

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Fertilization (Fertilisation)

The term in-vitro fertilization is derived from Latin in-vitro meaning glass because the eggs are fertilized in laboratory glassware. On the day of egg collection, the male partner is asked to provide a semen sample. He collects his semen by masturbation unless there has been other arrangements.

The sperm is washed in a culture medium and prepared in order to separate the sperm from seminal plasma.  If a donor sperm is to be used, the sample will then taken from the freezer, thawed and prepared.

Sperm are seperated from seminal plasma.

The eggs are collected into a specially prepared culture medium and once collected, they are examined under the microscope and each is graded for maturity, the maturity of an egg will determine when the sperm will be added to it. The eggs are then placed in the incubator for a period of time (about 3-8 hours) before it is mixed with the selected sperm.

Between 20000-30000 sperm are mixed with each egg in a drop of specially prepared culture medium. This medium is prepared in a labeled dish that is kept in the incubator to allow fertilization to occur. The first sign of fertilization is the presence of two small dots inside the egg so-called “two- pronucleate” and the fertilized egg is called zygote. The two pronuclei, one came from the sperm and one came from the egg. Occasionally the egg is fertilized abnormally with more than two pronuclei. Although this embryo may go on to divide, they are not viable and should not be replaced. Usually about 60-70% of the eggs collected will be fertilized, but this can vary from 0% to 100%. Regrettably, some 5-10% of couples will not achieve fertilization of any eggs. This could be due to, sperm lacking the fertilizing capacity, or poor egg quality or poor culture medium. However, in about 50% of the cases there is no obvious cause.

It takes about 18 hours for the egg to be fertilized, about 12 hours later the fertilized egg start to divide into two cells, and subsequently into four and so on. After about 48-72 hours from the egg collection, the embryos will usually consist of 4-8 cells each, and ready for replacement into the woman ‘s uterus.There is no significant difference in the expected pregnancy rates if the embryos are replaced on day two or three after egg collection. Sometimes, the embryos fail to develop even though they have fertilized normally and in this case; a transfer would not be made.

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Embryo Transfer

The embryo replacement (embryo transfer) procedure is quite simple and usually pain free. It may cause minimal discomfort and no anesthetic is used, although some women may need sedation or occasionally a general anesthetic. The male partner is usually invited to attend the procedure. The couple may also be able to view the embryos through a monitor before the embryos are replaced.

Some couples are concerned that their eggs, sperm or embryos may mix up with that of other couples. The probability of this happening in a good centre is very low.

The patient lies on a table or bed, usually with her feet in stirrups, some times the embryo transfer is performed with the patient in the knee-chest position. Using a vaginal speculum, the doctor exposes the cervix. The cervix is then cleaned with a little of culture medium or sterile water. One or more embryos suspended in a drop of culture medium are loaded in a fine plastic catheter so-called “embryo transfer catheter” with a syringe on one end. Gently and carefully, the doctor guides the tip of the catheter through the vagina and cervix, and deposits the embryos into the uterine cavity. The procedure may be guided by ultrasound scan to check the position of the catheter. The sue of ultrasound scan during embryo transfer appears to increase pregnancy rates. After the catheter is removed, it is handed over to the embryologist who will check it to ensure that no embryos remains. All the embryos replaced are transferred at the same time. Implantation begins three to four days later.

Successful pregnancy is related to the ease with which the embryos are transferred into the womb. Occasionally the position of the womb can make the transfer difficult. This may be overcome , to an extent, by a full bladder. A tenaculum may be applied to the cervix to straighten the uterus. If this fails, the doctor may use a stylet to negotiate the cervical canal. Very rarely, the cervix is to tight to allow the embryo catheter to pass through. In this case the doctor may resort to transferring the embryos through the muscle of the uterus (transmyometrial) or through the Fallopian tube if the tubes are healthy (TET).

Embryo replacement through embryo transfer catheter.

Once the embryos have been replaced, you may be asked to rest for a short while before going home. Prolonged bed rest of more than 20 minutes following embryo transfer has not been shown to improve pregnancy rates.

Occasionally, your doctor may advice you against having a fresh embryo transfer instead recommend freezing all embryos for later transfer. This may occur if you have one of the following:

  • If you are at a high risk of developing a severe ovarian hyperstimulation syndrome as shown from the scan and blood hormone levels.
  • If you have vaginal bleeding around the time of embryo transfer.
  • If your endometrium is not well developed (less than 5mm thickness) or there are polyps, you are unlikely to conceive as a result of fresh embryo transfer.
  • If the doctor was unable to transfer the embryos fresh because of narrowing of the cervix.

Embryo grading

Embryos are assessed by their appearance under the microscope for the number of cells, the characteristic of cells and the presence or absence of fragmentation. Good quality embryos divide rapidly, have equal cells with clear cytoplasm, and have only few fragments. Some IVF clinics classify the embryos into grade one, two, three and four. Grade one are the best quality embryos, these have a higher chance of implantation than those of grade 4. Research have shown that up to one third of embryos are genetically abnormal. There is no guarantee that a normal looking embryo will be genetically normal. How embryos attach and implant into the womb remains a mystery.

How many embryos to transfer?

At present, you have the best chance of success if more than one embryo is replaced.  The number of embryos transferred is crucial to your risk of a triplet pregnancy or higher.

There is a trend to transfer only two embryos especially in younger women.You are very unlikely to have triplets if only two embryos are transferred.

General advice after embryo transfer

  • Take it easy for a few days, avoid any strenuous exercise such as aerobics and horse riding.
  • Avoid douching, tampons and swimming in order to avoid undue contamination of the vagina.
  • Avoid unnecessary exposure to solvents and paints containing lead.
  • Avoid carrying or lifting heavy objects.
  • You should eat sensible and healthy diet, avoid consumption of excess alcohol, and avoid non-pasteurized milk and soft cheeses and blue cheeses because of the risk of listeria infection.
  • Avoid handling cat litter and soil.
  • Stop smoking.
  • Continue taking folic acid tablet.
  • Refrain from taking medication or drugs that are not necessary, and only after checking with your doctor.
  • Avoid contact with anyone who has a "flu-like" illness.
  • Avoid intercourse for two weeks.
  • Do not stop luteal phase support until you have both a negative pregnancy test and a period.

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Freezing of spare embryos (cryopreservation of supernumerary embryos)

Following embryo transfer, any remaining viable embryos are then cryopreserved (frozen) for later transfer. Not all embryos are suitable for freezing. The embryos can be frozen at the pronucleate, early cleavage or blastocyst stage. Overall about 50% of frozen embryos survive the thawing process. In the United Kingdom, the embryos can be frozen for up to 10 years with the intent of thawing and transferring them at a later date.

  • Cryopreservation is accepted in some countries but banned in others.
  • Freezing spare embryos allows multiple embryo transfers from a single egg collection and improves the chance of live birth. Freezing is very cost effective, since transferring is much less expensive than starting a new IVF treatment cycle.
  • Frozen embryos can be thawed and replaced in either natural or artificial cycles. In women who ovulate regularly, the likelihood of a live birth after replacement of frozen thawed embryos is similar whether natural or artificial cycles are used.
  • The overall success rates after thawing and replacing frozen embryos is lower than that for fresh embryos. However, in some IVF clinics, the transfer of frozen embryos results in similar pregnancy and live birth rates to fresh embryo transfers.
  • Babies born following embryo freezing have the same risks of abnormalities as those born following IVF, i.e. the same as those conceived normally.
  • If you decide that you no longer require the frozen embryos, they may be allowed to perish, be used for research or donated to infertile couple(s) according to your wishes and the current legislation in your country.
  • Not all clinics are able to offer freezing facilities.
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Success rate of IVF treatment
(pregnancy rate)

The success rate of IVF treatment has steadily increased with time.

Since the birth of Louise Brown in 1978, there has been a steady improvement in success rates of IVF treatment. The pregnancy rates vary between IVF clinics, and from time to time within the same clinic. The success of individual IVF clinics may be found in league tables published by HFEA. The overall live birth rate per treatment cycle is about 20%. There are many factors that may affect success rates.

The overall live birth rate per treatment cycle was 20% (HFEA patient report for the year 2000) and 24% in the USA (SART registry for the year 2000). This represents an increase of about 1-2% compared with the success rate for the last year.

Approximate chances of success of IVF treatment in women aged 38 years or younger in relation to the main steps of the IVF treatment are as follows:

Ovarian stimulation 90-95%
Collection of the eggs 99-99.7%
Fertilization of the eggs 90-97%
Normal embryo cleavage (division) 94-98%
Implantation rate per embryo 10-25%
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