Freezing Egg | Sperm | Embryo

EGG FREEZING

Egg freezing, also known as mature oocyte cryopreservation, is a method used to preserve reproductive potential in women.

Eggs are harvested from the ovaries, frozen unfertilized and stored for later use. A frozen egg can be thawed, combined with sperm in a lab and implanted in the uterus (in vitro fertilization).

Why it’s done

Egg freezing might be an option if you’re not ready to become pregnant now but want to try to ensure your ability to get pregnant or to have a biological child in the future.
Unlike with fertilized egg freezing (embryo cryopreservation), egg freezing doesn’t require sperm because the eggs won’t be fertilized before they’re frozen. However, you’ll need to use fertility drugs to induce ovulation so that you’ll produce multiple eggs for retrieval.

You might consider egg freezing if:
  • You’re about to undergo treatment for cancer or another illness that may affect your future fertility potential. Certain medical treatments — such as radiation or chemotherapy — can harm your fertility. Egg freezing before treatment might allow you to have biological children at a later date.
  • You’re undergoing in vitro fertilization. If your partner isn’t able to produce sufficient sperm on the day you have your eggs retrieved, egg freezing might be needed.
  • You wish to preserve younger eggs now for future use. Freezing eggs at a younger age may help you preserve your ability to reproduce when the time is right in the future.

You can use your frozen eggs to try to conceive a biological child with sperm from a partner or a sperm donor. A donor can be known or anonymous. The embryo can also be implanted in the uterus of another person to carry the pregnancy (gestational carrier).

How you prepare

Before beginning the egg-freezing process, you’ll likely have some screening blood tests, including:
Ovarian reserve testing. To determine the quantity and quality of your eggs, your doctor might test the concentration of follicle-stimulating hormone (FSH) and estradiol in your blood on day three of your menstrual cycle. Results can help predict how your ovaries will respond to fertility medication. Other blood tests and an ultrasound of the ovaries may be recommended to get a more complete assessment of ovarian function.

  • Infectious disease screening. You’ll be screened for certain infectious diseases, such as HIV. Potentially infectious eggs are stored differently than are other eggs.

Egg freezing has multiple steps — ovulation induction, egg retrieval and freezing.

Ovulation induction

At the beginning of your menstrual cycle, you’ll begin treatment with synthetic hormones to stimulate your ovaries to produce multiple eggs — rather than the single egg that normally develops each month. Several different medications might be needed, including:

    • Medications for ovarian stimulation. To stimulate your ovaries, you might inject medication such as follicle-stimulating hormone or human menopausal gonadotropins
    • Medications to prevent premature ovulation. To prevent premature ovulation, you might inject medication such as a gonadotropin-releasing hormone agonist (Lupron) or a gonadotropin-releasing hormone antagonist.
      During treatment, your doctor will have you return for follow-up visits. These visits will include having vaginal ultrasounds — a procedure that uses sound waves to create an image of the inside of your ovaries — to monitor the development of fluid-filled ovarian cysts where eggs mature (follicles). Blood tests also will be used to measure your response to ovarian-stimulation medications. Estrogen levels typically increase as follicles develop and progesterone levels remain low until after ovulation.
      When the follicles are ready for egg retrieval — generally after eight to 14 days — injections ,human chorionic gonadotropin or other medications can help the eggs mature.

Egg retrieval

Egg retrieval is done under sedation, typically in your doctor’s office or a clinic. A common approach is transvaginal ultrasound aspiration. During this procedure, an ultrasound probe is inserted into your vagina to identify the follicles. A needle is then guided through the vagina and into a follicle. A suction device connected to the needle is used to remove the egg from the follicle. Multiple eggs can be removed from your follicles in about 15 to 20 minutes.
After egg retrieval, you might experience cramping. Feelings of fullness or pressure might continue for weeks because your ovaries remain enlarged.

Freezing

Shortly after your unfertilized eggs are harvested, they’re cooled to subzero temperatures to stop all biological activity and preserve them for future use.
Vitrification. In this method, high initial concentrations of cryoprotectants are used in combination with cooling so rapid that intracellular ice crystals don’t have time to form.

After the procedure

Typically, you can resume normal activities within a day or two of egg retrieval. Avoid unprotected sex to prevent unwanted pregnancy.
Contact your clinician if you have:
• A fever higher than 101.5 F (38.6 C)
• Severe abdominal pain
• Weight gain of more than 2 pounds (0.9 kilograms) in 24 hours
• Heavy vaginal bleeding — filling more than two pads an hour
• Difficulty urinating

Results

ICSI

When you choose to use your frozen eggs, they’ll be thawed, fertilized with sperm in a lab, and implanted in your or a gestational carrier’s uterus.
Intracytoplasmic sperm injection (ICSI). In ICSI, a single healthy sperm is injected directly into each mature egg.
About 90 percent of eggs survive freezing and thawing, and about 75 percent will be successfully fertilized. The chances of becoming pregnant after implantation are roughly 30 to 60 percent, depending on your age at the time of egg freezing. The older you are at the time of egg freezing, the lower the likelihood that you’ll have a live birth in the future.
Keep in mind that pregnancy rates might be lower when frozen eggs are used, compared with fresh or frozen embryos.

Ovarian Tissue Freezing

Ovarian tissue cryopreservation (freezing) is an experimental method of fertility preservation in which the outer layer of an ovary, which contains a large number of immature eggs, is taken out of the body and frozen for future use. In ovarian tissue freezing process, a part of an ovary or a whole ovary is surgically removed, usually by laparoscopy. In the laboratory, the ovary’s outer layer (called ovarian cortex) is cut into small strips and frozen. An experimental procedure, ovarian tissue freezing is for the most part performed for medically indicated fertility preservation in cancer patients.When the patient is cured of the primary disease and ready to have children, the ovarian strips are thawed and transplanted back into her body, either on the remaining ovary or elsewhere. In most cases reported in the literature, transplanted ovarian strips regain normal function of producing hormones and eggs. So far, more than 30 live births have been reported worldwide after ovarian tissue freezing and re-transplantation.

Benefits of Ovarian Tissue Freezing

A major advantage of ovarian tissue freezing is the number of eggs that can be frozen in “one shot.” Embryo freezing and egg freezing typically result in approximately 10 eggs or embryos per freezing attempt. In contrast, ovarian tissue freezing lets women freeze a vastly larger number of immature eggs—hundreds to thousands—for future use. When frozen ovarian tissue is re-implanted into the body and regains its functions, immature eggs that were frozen within the tissue start developing normally, and become retrievable in an IVF procedure.

Unlike egg and embryo freezing, where just one cycle can take a few weeks to complete, ovarian tissue freezing can be performed on very short notice. Even women who must undergo chemotherapy or radiation almost immediately after diagnosis may be able to preserve their fertility using this method.

Ovarian tissue freezing is also an important method of fertility preservation for young girls. When a young girl is diagnosed with cancer or other conditions requiring life-saving, ovary-toxic treatments, the option of egg freezing may not yet be feasible because her eggs have not started the maturation process, which starts with menarche (first menstrual period). While still considered an “experimental” procedure, ovarian tissue freezing is utilized frequently for young prepubescent girls.

Counseling and Informed Consent

Patients considering ovarian tissue freezing should be thoroughly informed about risk-benefit considerations, the procedure’s experimental nature and available alternative treatments. Since ovarian tissue freezing is typically performed for patients facing life-threatening illnesses and ovary-toxic therapies, close communication and coordination among patients, physicians treating the primary disease (oncologists, oncologic surgeons, etc.), fertility specialists performing the fertility preservation procedure and family members is essential.

The Potential of Ovarian Tissue Freezing with In Vitro Maturation

Reproductive medicine is rapidly evolving, and one of the possibilities in this area is to in vitro mature the primordial follicles (very immature eggs) contained in the cryopreserved ovarian tissue. If successful, in vitro maturation (IVM) of primordial follicles would eliminate the need to surgically re-transplant the ovarian tissue back into the body. While IVM is currently not able to mature eggs in such an early stage of development, successful IVM of primordial follicles would make hundreds to thousands of eggs in each small strip of frozen ovary available to cancer survivors later in life, rather than the few dozens currently banked from egg or embryo banking cycles.

Embryo Freezing

The ability to freeze & thaw embryos successfully is one of the greatest advancements in assisted reproductive technology. There are two methods of freezing embryosnamely Slow freezing &Vitrification.
Vitrification is a flash or fast freeze technology & the advent of this technique greatly improved outcomes from cryopreserved embryos. Couples who opt for the embryo freezing now experience success rates equal to or better thanthose from fresh embryo transfer cycles.Embryos can be frozen at any stage like from Day 2 (4 celled stage) to Day 5(~150-250 celled blastocyst stage)

Advantages of Frozen Embryos

1) Surplus or Supernumerary embryos

Surplus or Supernumerary embryos can be frozen for later use. In case the fresh embryo transfer fails, these embryos provide opportunity of trying again without having to undergo ovarian stimulation medication & egg retrieval. Development of vitrification technologies has led to outstanding outcomes, with more & more patients achieving live birth from a single stimulated cycle.

2) Frozen embryo transfer (FET)

(A) More Economical

Expenses of ovarian stimulation medications, blood work & ultrasound study, egg retrieval, insemination, embryo culture etc can be evaded.

(B) Less strenuous & more convenient for the couple

Simple estrogen supplementation during the proliferative phase of endometrial (uterine) preparation & later on addition of progesterone injections to prepare uterus for embryo implantation. Frozen embryos are thawed, allowed to equilibrate & expand in culture for few hours, prior to transferring them to the uterus. The hormonal levels can be maintained at desired levels, unlike the fresh transfer cycles, thereby the success rates are infact more when compared to the fresh transfer following ovarian stimulation.

3) Freezing all the embryos for future (FET)’s

(A) FET allows reducing the risk of OHSS (ovarian hyperstimulation syndrome), while maintaining excellent success rates.
At DRIFF we carefully monitor patients for OHSS & use stimulation protocols designed according to individual needs to prevent this from occurring. But sometimes in few patients say less than 5% of the patients where our clinicians see potential warning signs that a woman may have risk of OHSS like high estrogen levels more number of follicles, fluid in the pelvis etc. Then the couple is are recommended for freezing of all the available embryos rather than proceeding with fresh transfer, as pregnancy may further increase the OHSS risk. The embryos thus frozen can be safely transferred in a FET cycle. The pregnancy outcomes will also be far better in FET in such scenario.

(B) To overcome the negative effect that elevated progesterone levels have on pregnancy outcomes. During ovarian stimulation, few women ~ 5- 10% of the women may have raised levels of progesterone which maybe detrimental to the endometrium (uterine lining) making it less receptive for embryo implantation. Thus the pregnancy rates are rates are reduced. If progesterone rises above a critical threshold, the clinician recommend to freeze all the available embryos, rather than proceeding with the fresh transfer. An FET can then be performed in a cycle without the stimulation medications.
Our data demonstrates that transferring embryos via FET is associated with greater chance of success than the fresh transfer in the setting of an elevated progesterone level. Women who undergo “Freeze – all” cycle experience the same excellent live-birth rates as do women who have a fresh transfer (with normal progesterone & estrogen levels) in their FET’s.

4) Frozen embryos allow for genetic testing.

For couples with a risk of passing certain genetic conditions, onto their children, we can test the embryos for certain genetic mutations. This is called as pre-implantation genetic diagnosis (PGD). After the clinician retrieves the eggs & they are fertilized the embryologist will biopsy each suitable embryo on day5 or 6 of development. The embryologist then freezes each embryo individually while awaiting results from the biopsied cells. The clinician & embryologist then transfer only those embryos after thawing that are free of the genetic mutation (corresponding medical condition).
In addition to testing for genetic mutations (PGD), embryo freezing has allowed for pre-implantation genetic screening (PGS). PGS looks for abnormalities in chromosome number, such as trisomy 21, which causes Down’s syndrome and many others that are likely to result in implantation failure or miscarriage. Likely candidates for this testing include patients with recurrent pregnancy loss and older women, whoare at higher risk for chromosomal abnormalities. In appropriately selected candidates, this testing may result in improved live-births.

5) Frozen embryos offer the potential to grow family at a later date.

Vitrified embryos maintain reproductive potential far into the future, giving our patients time to make decision to expand their families. Frozen embryo transfer can suspend proverbial biological clock, since the embryos thawed for FET maintain the reproductive potential associated with age of the egg at the time it was fertilized. For example, if a woman has her first child at the age of 35 years through IVF & has the remaining embryos frozen at,she can comeback to have embryos transferred via FET at a time when conceiving with her own eggs would otherwise be unlikely. Evidence –based medicine & years of research.
Evidence based medicine & years of research performed at DRIFF has allowed our patients to have better treatment options than ever before, with frozen embryos at the forefront of this technology revolution.

FREEZING / CRYOPRESERVATION

Introduction: Cryopreservation or Cryobiology is branch of science which deals with the process of freezing biological material at extreme low temperature, most commonly at 1960C / -3210F in liquid nitrogen.
At these low temperatures, all the biological activity comes to a standstill including the biochemical reactions that leadto cell death andDNA degradation.This preservation method makes it possible to store living cells as well as other biological material unchanged for centuries.
The challenge of cryopreservation is to help cells to survive both cooling to extreme temperatures and thawing back to physiological conditions. Intracellular ice formation in particular is a critical issue that has to be controlled to keep the cell membrane intact and the cells alive. The crucial elements to prevent this are the freezing rate (degrees per minute) and the composition of the freezing medium used. The freezing medium generally consists of a diluter, (sometimes) a protein source, as well as a cryoprotectant compound. The choice of most suitable cryprotectant will influence to preservation result and will be different between different cells and different species.

SEMEN FREEZING

Sperm freezing is a process of collecting, analyzing, freezing and storing of a man sperm. The frozen semen samples can be used for assisted reproductive procedures, such as IVF(in-vitro fertilization),IUI (intrauterine insemination) or for sperm donation.

The cryopreservation process involves.
  • Routine screening for STD’S (sexually transmitted diseases) HIV, HbsAg (Hepatitis) and VDRL
  • Providing a semen sample or undergoing a sperm extraction.
  • Lab analysis of sperm quality and quantity.
  • Freezing of viable sperm.
  • Storage of sperm indefinitely

Semen sample is typically collected through masturbation.Men are generally asked to abstain from sexual activity for 3-5 days to allow best possible specimen. If there are no sperms in the sample or if the man is unable to ejaculate, a surgical retrieval to remove sperm directly from the epididymis or testicles.

Common reasons for choosing to freeze sperm.
  • Advancing age
  • Deteriorating sperm quality or low quantity
  • Cancer are other medical reasons
  • Prevasectomy patients
  • Career and lifestyle choices, such as those with high risk occupations.
  • Nonavailability at the time of ovulation during treatment
  • As a backup sample incase not able to provide ejaculate on day of IUI or IVF.

Special cryoprotectants are used, mixed well with the semen sample, aliquoted into vials or straws properly labeled and frozen. When required these samples are thawed, washed and tested for motility, viability prior to use in IUI or IVF.
There are no risks or side effects to collecting semen samples naturally (through masturbation) if surgical extraction is required, there are small risks as with any surgery, such as minute bleeding or discomfort.
Sperm freezing has been successfully used since 1953 to help individuals conceive healthy babies. The process is safe, standardized and continues to improve as technology advances.
The primary concern with sperm freezing is that not all sperm survive the freezing and thawing process. However, as most semen ejaculations contain an appropriate number of sperm, the chances of having enough healthy sperm for fertility treatments is very high. Moreover the advantage of frozen sperm sample over fresh is that unhealthy, defective, DNA fragmented spermatozoa become prone to cryodamage and are thus naturally eliminated,leaving healthy, fertile mature, spermatozoa to form healthy embryo and thus offspring.The capability of the surviving sperm cells to fertilise an egg & form an embryo is not jeopardized during the freezing or thawing process.

Cryopreservation is considered to have no time limit and stored sperm as old as 20 years have been used to create healthy babies.

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