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In vitro fertilization | Online Biotech Notes

In vitro fertilization | Online Biotech Notes


What is in vitro fertilization, In vitro fertilization, studybiotech4u, IVF
IN- Vitro Fertilization


What is in vitro fertilization (IVF)?

  • The term in vitro refers to the outside of the body, and the term fertilization states to the condition where sperm has attached to and entered the egg.
  • Overall, in vitro fertilization is defined as the condition where the fertilization takes place in a laboratory dish (i.e. outside the body).
  • IVF is one of the techniques of assisted reproductive technology (ART).
  • The IVF is mainly employed to assist with fertility and avoid genetic problems.
  • While performing IVF at the lab, firstly mature eggs are retrieved from the ovaries and are fertilized by sperm.
  • Then, the fertilized eggs are placed in a uterus.
  • One complete cycle of IVF may take about 3 weeks.
  • The first successful test-tube baby was Louise Brown, born in the year 1978 performed by Dr. Robert G.

What are the steps for In vitro fertilization (IVF)?

  • The IVF comprises of basic 5 steps. They are enlisted as follows:
    • a. Stimulation
    • b. Egg retrieval
    • c. Insemination and fertilization
    • d. Embryo culture
    • e. Embryo selection and embryo transfer

Step I: Stimulation

  • It is also termed superovulation.
  • In a normal condition, a woman produces one egg during each menstrual cycle.
  • However, IVF has a requirement of multiple eggs because the use of multiple eggs increases the probability of developing viable embryos.
  • Fertility drugs are prescribed to the woman, so that the egg production boosts up.
  • The fertility drugs consist of exogenous gonadotropins and similar substances, that cause hormonal stimulation of the ovary causing the production of a large number of eggs per cycle.
  • On the third day of menstruation, treatment cycles are generally started, following the use of fertility medicines to trigger the development of multiple follicles of the ovaries.
  • During this step, females are required to undergo regular transvaginal ultrasounds and blood tests to check hormone levels.
  • The stimulation of the ovary can be performed by 2 major protocols:
    • 1.  Lengthy protocol is the one where the suppression (down-regulation) of the pituitary ovarian axis is performed by the prolonged use of a gonadotropin-releasing hormone (GnRH) agonist.
    • Once the process of down regulation is accomplished, usually after 10-14 days, subsequent ovary hyperstimulation generally using follicle-stimulating hormone (FSH) starts.
    • 2.  Short protocol is the one where the down regulation part is neglected and consists of prescription of injectable gonadotropins under regular monitoring in order to trigger the development of multiple follicles of the ovaries.
    • The frequent monitoring checks the level of estradiol, and the follicular growth is checked via gynecologic ultrasonography.
    • Usually, ten days of injection is required.
  • During the last days of stimulation, the use of GnRH antagonists usually prevents the spontaneous ovulation during the cycle.
  • It blocks the natural surge of luteinizing hormones (LH) facilitating the start of the ovulation process by the use of injectable human chorionic gonadotropins.

Step II: Egg retrieval

  • It is also termed as follicular aspiration.
  • It is a minor surgery performed for the removal of eggs from the woman’s body.
  • After the ovarian follicles reach a certain level of degree of development, final maturation is induced by an injection of human chorionic gonadotropin (hCG).
  • hCG hormone plays a role as that of luteinizing hormone(LH).
  • After a single hCG injection, ovulation would take place between 38 and 40 hours.
  • However, the eggs are retrieved between 34 and 36 hours after hCG injection, which is, just before the rupture of follicles.
  • This assists in scheduling the process of egg retrieval at a time when the eggs are completely matured.
  • A technique called transvaginal oocyte retrieval is used to retrieve eggs.
  • In this process, the woman is given anesthesia, prior to surgery.
  • The health care provider by using ultrasound images as a guide, inserts a thin needle through the vagina into the ovary and sacs (follicles).
  • Then, the needle is connected to a suction device, that pulls the egg and fluid out of each follicle, one at a time.
  • The same process is repeated for other ovary.
  • Generally, 10-30 eggs are removed.

Step III: Insemination and fertilization

  • The best quality of embryos that are potent for a successful pregnancy is selected.
  • It is also termed as oocyte selection.
  • Along with it, the process termed as sperm washing is also conducted.
  • In this process, the inactive cells and seminal fluids are removed from semen to prepare it for fertilization.
  • In the case where semen is supplied by a sperm donor, the preparation for treatment takes place before being frozen and quarantined, then it will be thawed ready for use.
  • For about 18hrs, the incubation of sperms and egg (at the ratio of about 75000:1) is done in the culture media.
  • In the majority of the cases, the egg will be fertilized by that time and the fertilized egg shows two pronuclei.
  • In specific cases such as low sperm count or motility, intracytoplasmic sperm injection(ISCI) can be used to inject a single sperm directly into the egg.
  • Now, the fertilized egg is transferred to a special growth medium and left for about 48hrs until the egg reaches the 6-8 cell stage.

Step IV: Embryo culture

  • After, the fertilized egg reaches the 6-8 celled stage, embryos are cultured usually 3 days after retrieval.
  • Embryo culture can be performed either in an artificial culture medium or in an autologous endometrial co-culture.
  • Embryo culture in artificial culture medium:
    • In this type of culture, there can be either the same culture medium throughout the process or an embryo can be sequentially placed in different media by use of a sequential system. Ex. One medium can be used for culture to day 3, and the second medium is employed for culture after it, when culturing to blastocyst stage.
    • For the culture of human embryos to the blastocyst stage, both the single and sequential medium are equally effective.
    • The media for artificial culture usually contain glucose, pyruvate, and energy supplying components.
    • However, the addition of nucleotides, amino acids, vitamins, and cholesterol enhances the performance of embryonic growth and development.
  • The techniques that allow dynamic embryo culture along with the fluid flow and embryo movement are also present.
  • A new technique in development where the embryos are encapsulated in the permeable intrauterine vessel.

Step V: Embryo selection and embryo transfer

  • 1. Embryo selection:
    • Based on the number of cells, evenness of growth, and degree of fragmentation, embryos are graded by embryologists.
    • For the selection of embryos, the morphological scoring system is considered the best strategy that optimizes pregnancy rates as well.
    • If it is to be chosen between embryos of morphologically equal quality, the presence of soluble human leukocyte antigen-G (HLA-G) is regarded as a second parameter.
    • Embryos that have reached the 6-8 celled stage are then transferred 3 days after retrieval.
    • It has been seen that blastocyst stage transfer results in higher pregnancy rates.
  • 2. Embryo transfer:
    • The number of embryos that are to be transferred depends on the number available, the age of the woman, and other health and diagnostic factors.
    • Most of the clinics and country regulatory bodies tend to reduce the risk of pregnancies carrying multiples.
    • The best embryos are transferred to the patient’s uterus utilizing a thin plastic catheter that goes through the cervix.

Summaries of steps of IVF:

  • The follicle maturation along with ovulation is promoted by ovarian hormonal stimulation.
  • To achieve fertilization by assisted reproductive technology (ART) several fertilization methods are used.
  • Under the cultured conditions, the re-implantation embryo spends a certain time that will influence its further development.
  • Preimplantation embryo biopsies can be used during this period of time.
  • Finally, the embryo is transferred to a recipient female.

Risks of In vitro fertilization (IVF):


  1. Multiple births:
    • There are chances of multiple births if more than one embryo is transferred to the uterus. A multi-fetus pregnancy carries a higher risk of early labor and low birth weight than a single-fetus pregnancy.
  1. Premature birth and low body weight:
    • It is suggested that IVF causes the birth before the normal delivery time with the low weight of the baby.
  1. Ovarian hyperstimulation syndrome:
    • The use of fertility drugs like human chorionic gonadotropins (hCG) to induce ovulation can result in ovarian hyperstimulation syndrome. In this condition, the ovaries get swollen and become painful.
  1. Miscarriage:
    • For women who conceive using IVF with fresh embryos, the incidence of miscarriage is close to that of women who conceive naturally, i.e. around 15% to25%, however, the rate rises with maternal age.
  1. Complications of the egg-retrieval process:
    • Bleeding, inflammation, or damage to the intestines, bladder, or blood vessels could be caused by the use of an aspirating needle to collect eggs. Sedation and general anesthesia, if used, are also associated with risks.
  1. Ectopic pregnancy:
    • About 2% to 5% of women who use IVF may have an ectopic pregnancy. Ectopic pregnancy is the condition in which the fertilized egg implants outside the uterus, generally in a fallopian tube. Outside the uterus, the fertilized egg can’t survive, and there’s no way to continue the pregnancy.
  1.  Birth defects:
    • The mother’s age is the main risk factor, no matter how the baby is conceived, in the development of birth defects. To decide if babies conceived using IVF might be at increased risk of certain birth defects, further research is required.

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