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An Annual Journal of Dept. of Biotechnology
Vol: 3, No. 1, Year : 2010 – 11; Page: 91-96 When Nature Fails, Test Tube Baby Prevails
Jyotika Sharma, Kh. Beauty Singha,
Raima Das and Shaheen Laskar
One of the greatest boons for infertile couples is the development of Test Tube Baby. This
incredible work was performed by Dr. Patrick Steptoe and Dr. Robert Edwards and they developedthe first test-tube baby, Louise Brown born on the 25th July, 1978.
Test tube baby is the one which is conceived outside the woman’s womb. The sperm of man and the egg taken from the woman’s ovaries are fertilised within a test tube in the laboratory.
Once successful fertilisation is completed, the embryo is then transplanted into the woman’s
uterus to complete the pregnancy. The embryo won’t survive within the test tube and needs to be
transplanted to the woman’s uterus for its normal growth. The first In-Vitro fertilization was
done in the test tube and that is why it is popularly known as Test tube Baby.
Normally this procedure is a part of the IVF (In-vitro Fertilisation) technique and is done in
patients who have problems conceiving normally and when fertilization of egg with sperm
occurs outside the body it is called “In Vitro Fertilization”.
Technique of In Vitro Fertilization
In vitro fertilisation or fertilization (IVF) is a process by which egg cells are fertilised by sperm outside the woman’s womb, in vitro. IVF is a major treatment in infertility when othermethods of assisted reproductive technology have failed. The process involves hormonallycontrolling the ovulatory process, removing ova (eggs) from the woman’s ovaries and lettingsperm fertilise them in a fluid medium. The fertilised egg (zygote) is then transferred to thepatient’s uterus with the intent to establish a successful pregnancy.
Preliminary testing
This includes first consultation, blood tests culture if necessary, hysteroscopy, D21 visit i.e.
mock transfer trial, semen freezing and serum progesterone.
Firstly,a history and physical examination of both partners are completed. Transvaginal sonography of female partner and semen examination of male partner and culture of it is done,if necessary.
B.Sc 3rd Year, Department of Biotechnology, Gurucharan College, Silchar, Assam BIOTECH
An Annual Journal of Dept. of Biotechnology
Vol: 3, No. 1, Year : 2010 – 11; Page: 91-96 This includes D3 FSH, LH, PRL, TSH. Major preoperative profile and other special blood investigations if necessary, are performed.
Hysteroscopy test must be scheduled to be completed on days 5-11 of the menstrual cycle.
Menstruation must be absent for this test to be done.
D21 visit- Semen freezing- Mock transfer trial- Serum progesterone- TVS Semen freezing: It is necessary to freeze a semen specimen before IVF cycle starts.It is preferred
to inseminate the eggs with fresh semen if possible.
Transvaginal sonography (TVS) will determine the thickness of endometrium on D21.
Serum progesterone level is estimated.
Mock transfer trial: Embryo transfer is very important procedure and it is usually done without
anesthesia so, on day 21 we will do mock embryo transfer to anticipate any difficulty and solution
far it before actual transfer. It is painless procedure.
Development of oocytes (Eggs)
It is necessary to stimulate the ovary to produce multiple follicle (the sac that contains the
egg), in order to improve the chances of a successful outcome from an IVF cycle. The developmentof ooctyte is based on factors such as age, infertility history, a past response to these medications,and a base line FSH level of the female patient. The fertility medications that are necessary tostimulate the ovary are unfortunately all injectable medications.
If there are more than one egg then there is better chance of pregnancy of in IVF cycle. To get more eggs the ovaries are stimulated with stimulation drugs called gonadotrophins.
Gonadotrophins can be manufactured from urine or by recombinant DNA technology. Appropriatetype of gonadotrophins and dose of administration are selected for individual patients. Availablegonadotrophins include HMG, FSH and recombinant FSH manufactured by differentpharmaceuticals.
Side effect of Fertility edication
Over stimulation of the ovary, It can occur to varying degrees: mild, moderate or severe.
Multiple births: The risk of conceiving a multiple pregnancy during and IVF cycle isdependent on age, response to the medications, the quality of the embryos, the numberof embryos replaced into the uterus and other unforeseen factors are considered whenjudging your specific risk of multiple births.
Common complaints: Pain at the injection site, headaches and fatigue.
An Annual Journal of Dept. of Biotechnology
Vol: 3, No. 1, Year : 2010 – 11; Page: 91-96 GnRH analog (Lupride): This medication is administered by subcutaneous injection. Itis given to prevent premature release of the oocytes (eggs). Side effects may include:localized skin reaction, allergic reaction, headaches, hot flashes and mood swings HCG (Human Chorionic Gonadotropin): It is generally taken 34-36 hours prior to theegg retrieval. This medication should be injected into the muscle. This medicationcompletes the maturation of the egg.
This medication, assist the embryo to attach to the uterus. The side effects that have been reported include: breast tenderness, headache, nausea, fluid retention, fatigue, mood swings,depression, and pain at the site of injection (in the case of injectable progesterone).
Oocyte (egg) retrieval
The egg collection process (retrieval) is usually accomplished using the Ultrasound-guided trans vaginal method. Other methods of retrieving oocyte that are rarely utilized in our practicebut are sometimes necessary, include Laparoscopy or a Trans-Abdominal approach.
This ultrasound-guided trans vaginal method of egg retrieval allows this procedure to be done in an out patient setting. A vaginal ultrasound allows for visualization of both ovaries. Aneedle is inserted through the vaginal wall in the ovary. Each follicle is punctured individuallyand the fluid containing the egg is examined by the embryologist under the microscope until theegg is found. The duration of this procedure is usually less than 45 minutes.
This procedure is done under general anesthesia Sperm collection and insemination
The specimen will be processed in the laboratory and prepared for egg insemination. Frozen specimen collected will be used only in case of emergency (illness, inability to procedure aspecimen etc.) Incubation and fertilization of oocytes (eggs)
The eggs and sperm will be placed together in a special culture fluid and kept in incubators in our laboratory. This procedure is called insemination.
Formation and cleavage of embryos
The eggs will be examined 16-20 hours after insemination for signs of fertilization. If fertilization occurs, the fertilized eggs are now described as pre-embryos or zygotes. When theydivide to at least 2 cells they are called embryos. The laboratory environment is conducive forfertilization to occur, however, it cannot be guaranteed that fertilization will occur. Typically,60% of the eggs retrieved will be fertilized. This percentage may be higher or lower dependingon each couple.
Embryo transfer
Embryo transfer (ET) usually occur forty eight to seventy two hours post retrieval. The time of transfer will be designated by the IVF staff. The embryo(s) that is (are) assessed to bedeveloping normally will be considered for transfer. Although a recommendation (3-5 embryos) BIOTECH
An Annual Journal of Dept. of Biotechnology
Vol: 3, No. 1, Year : 2010 – 11; Page: 91-96 will be made regarding the number to transfer, the final decision resides with the couple and thedoctor. Transferring multiple embryos may result in the growth of more than one foetus. If youhave extra embryos after the transfer, they will be cryopreserved if they have demonstratedappropriate development.
The method used for transferring embryos is similar to that of the mock transfer. ET is performed by inserting a small catheter through the cervical opening into the uterine cavity. Theembryo transfer is usually a painless procedure. There is a recommended rest period after thetransfer.
Within 13 days post-transfer, hormonal levels and a pregnancy test will be done. If a pregnancy has occurred, further blood testing blood work and ultrasound will be required toassess normal progression.
Some problem in fertilization during test tube baby process we need to asses the fertilization
to help fertilization we take out of the egg from the wife of egg from the woman’s ovary and incontrol lab and environment we fertilized egg with the help of sperm of her male partner. Afterfertilization the fertilized egg are cultured into the control laboratory environment to form pre-embryos and this pre-embryos are then transferred back into the uterus of the woman. If theyimplant and grow become the pregnancy, and hence test tube baby is most convenient process.
The First Test Tube Baby
The incredible work performed by Dr. Patrick Steptoe and Dr. Robert Edwards, the first test-tube baby, Louise Brown was born on the 25th July 1978, marked hope for other infertilecouples to have a baby through this procedure. However, many others were concerned about theethical issues surround this. One major area of concern was the fact that as the egg is locatedoutside the womb for a few days while the cells multiplied before being put back inside theuterus, the health issues affecting this baby is unknown. Indeed, research has been shown thattest-tube babies have a higher chance of birth defects and low birth weight, and researchers stillhave not been able to determine the reason for this.
Success Rates
Due to advancement in reproductive technology, the IVF success rates are substantially better today than they were just a few years ago. The most current data available in the UnitedStates a 2009 summary complied by the Society for Reproductive Medicine (SART) whichreports the average national IVF success rates per age group using non-donor eggs (see tablebelow).
The live birth rates using donor eggs are also given by the SART and include all age groups Fresh Donor Egg Embryos Thawed Donor Egg Embryos
An Annual Journal of Dept. of Biotechnology
Vol: 3, No. 1, Year : 2010 – 11; Page: 91-96 In 2006, Canadian clinics reported an average pregnancy rate of 35%. A French study estimated that 66% of patients starting IVF treatment finally succeed in having a child (40%during the IVF treatment at the center and 26% after IVF discontinuation). Achievement ofhaving a child after IVF discontinuation was mainly due to adoption (46%) or spontaneouspregnancy (42%).
Live birth rate
The live birth rate is the percentage of all IVF cycles that lead to a live birth. This rate does not include miscarriage or stillbirth and multiple-order births such as twins and triplets are countedas one pregnancy. The live birth rate is calculated as follows: Number of live births ÷ Number of procedures performed (cycle start, egg retrieval or
embryo transfer).

Success or Failure Factors
The main potential factors that influence pregnancy (and live birth) rates in IVF have been suggested to be maternal age, duration of infertility or subfertility, bFSH and number of oocytes,all reflecting ovarian function. Optimal woman’s age is 23–39 years at time of treatment.
In a few cases, laboratory mix-ups (misidentified gametes, transfer of wrong embryos) have occurred, leading to legal action against the IVF provider and complex paternity suits.
Another concern is that people will screen in or out for particular traits, using preimplantation Although menopause is a natural barrier to further conception, IVF has allowed women to be pregnant in their fifties and sixties. Women whose uterus has been appropriately preparedreceive embryos that originated from an egg of an egg donor. Therefore, although these womendo not have a genetic link with the child, they have an emotional link through pregnancy andchildbirth. In many cases the genetic father of the child is the woman’s partner. Even aftermenopause the uterus is fully capable of carrying out a pregnancy.
Same-sex couples, single and unmarried parents A 2009 statement from the ASRM found no persuasive evidence that children are harmed or disadvantaged solely by being raised by single parents, unmarried parents, or homosexualparents. It did not support restricting access to assisted reproductive technologies on the basis ofa prospective parent’s marital status or sexual orientation. Ethical concerns include reproductiverights, the welfare of offspring, nondiscrimination against unmarried individuals, homosexual,and professional autonomy.
The Catholic Church opposes all kinds of in vitro fertilisation because, as with contraception, it separates the procreative purpose of the marriage act from its unitive purpose.
An Annual Journal of Dept. of Biotechnology
Vol: 3, No. 1, Year : 2010 – 11; Page: 91-96 Legal status
Government agencies in China passed bans on the use of IVF in 2003 by unmarried women or by couples with certain infectious diseases. Costa Rica is the only country in the westernhemisphere that forbids this technique. According to the rule, Costa Rica had to enact legislationallowing the use of this type of procedure. The Court set a first dateline on February 2011 for areport on the question by the Costa Rican government. Federal regulations in the United Statesinclude screening requirements and restrictions on donations, but generally do not affect sexuallyintimate partners. However, doctors may be required to provide treatments due tonondiscrimination laws, as for example in California.
Reichman DE, Jackson KV, Racowsky C (May 2009). “Incidence and development of zygotes exhibitingabnormal pronuclear disposition after identification of two pronuclei at the fertilization check”. Fertilityand Sterility Vol- 94 (3): 965–970.
Schmidt L, Holstein BE, Christensen U, Boivin J (November 2005). “Communication and coping aspredictors of fertility problem stress: cohort study of 816 participants who did not achieve a delivery after12 months of fertility treatment”. Human Reproduction Vol-. 20 (11): 3248–3256.
Xella S, Marsella T, Tagliasacchi D, et al. (April 2010). “Embryo quality and implantation rate in twodifferent culture media: ISM1 versus Universal IVF Medium”. Fertiity and. Steriity. Vol-.93 (6): 1859–1863.
Hammoud AO, Gibson M, Stanford J, White G, Carrell DT, Peterson M (2009). “In vitro fertilizationavailability and utilization in the United States: a study of demographic, social, and economic factors”.
Fertility and Sterility 91 (5): 1630–1635.
Swain JE, Smith GD (2011). “Advances in embryo culture platforms: novel approaches to improvepreimplantation embryo development through modifications of the microenvironment”.HumanReproduction .Vol- 17 (4): 541–557.
Kyono K, Uto H, Nakajo Y, Kumagai S, Araki Y, Kanto S (January 2007). “Seven pregnancies and deliveriesfrom non-mosaic Klinefelter syndrome patients using fresh and frozen testicular sperm”. J. Assist. Reprod.
Genet. Vol 24 (1): 47–51 Chambers GM, Sullivan EA, Ishihara O, Chapman MG, Adamson GD (June 2009). “The economic impactof assisted reproductive technology: a review of selected developed countries”. Fertility and Sterility.Vol-91 (6): 2281–2294.
Okada H, Goda K, Muto S, Maruyama O, Koshida M, Horie S (November 2005). “Four pregnancies innonmosaic Klinefelter’s syndrome using cryopreserved-thawed testicular spermatozoa”. Fertility andSterility.Vol- 84 (5) 1508.
Gleicher, Norbert; Weghofer, Andrea, et al. (2010). Mailund, Thomas. ed. “FMR1 Genotype withAutoimmunity-Associated Polycystic Ovary-Like Phenotype and Decreased Pregnancy Chance”. PLoWONE.Vol- 5 (12):153-155.
Hammoud AO, Gibson M, Stanford J, White G, Carrell DT, Peterson M (2009). “In vitro fertilizationavailability and utilization in the United States: a study of demographic, social, and economic factors”.
Fertility and Sterility. Vol- 91 (5): 1630–1635.


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