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ASSISTED REPRODUCTIVE 
TECHNIQUES 
PROF. M.C. BANSAL 
M.B.B.S , M.S. , M.I.C.O.G, F.I.C.O.G. 
Founder Principal& Controlle...
All technique involving direct manipulation of 
oocyte/sperm outside the body
History of ART 
• 1978- first successful 
birth using In Vitro 
Fertilization 
• 1984- first successful 
birth using Gamet...
Patrick step toe Robert edwards 
Nobel prize in2010
• The world’s second and India’s first IVF baby, 
Kanupriya, alias Durga, was born 67 days later 
on October 3, 1978, thro...
Definition of Infertility & ART 
• Infertility is defined classically as the inability 
to conceive after 1 year of unprot...
Definition of Infertility & ART Cont’d 
• ART refers to all techniques involving direct 
retrieval of oocytes from the ova...
Causes of infertility
Indications of ART 
• Tubal factor infertility 
• Endometriosis 
• Male factor infertility 
• Unexplained infertility 
• O...
MAJOR ASSISTED REPRODUCTIVE TECHNOLOGIES (ART) 
• In vitro fertilization and embryo transfer (IVF-ET) 
• Direct intra-uter...
Artificial Insemination 
• Sperm is collected and 
placed into a woman’s 
vagina, cervical canal or 
in the uterus. 
• Spe...
DONATED SPERM 
(INTRAUTERINE INSEMINATION) 
• Doctor puts donated 
sperm in woman 
• Seminal fluid washed 
from sperm 
• D...
Natural cycle 
• First birth form IVF is from oocyte collected 
from natural cycle 
• Cycle cancellation are high (25%-75%...
STIMULATION PROTOCOLS 
• Protocol A (suppression and stimulation) 
• Protocol B(flare up) 
• Protocol C(cc+ HMG) 
• Protoc...
PROTOCOL A 
• Suppression /down regulation 
• GnRH analogue Inj suprefact(Buserelin 
acetate) 0.5 cc s/c BD(i.e 20 units o...
On D1 do E2 and LH estimation 
a. If E2 is <26 and LH<4 down regulation has been 
done 
b. If either of the values are mor...
The initial dose of exogenous gonandotrophins used 
to stimulate ovarian follicular development 
depends on individual nee...
• Continue with the same dose .If <2 follicles are 
leading then stop suprefact and HMG both 
• If size <7 mm increase the...
PROTOCOL B 
• FLARE UP 
• This is suitable for aged patients:where 
ovarian functions are poor 
• D1 & D2 Inj buserelin 0....
PROTOCOL C 
• Clomiphene citrate 150-250 mg from D1-D5 
• TVS ON D5/6 
• HMG started on D5,follicle 5 mm in each ovary 
• ...
Protocol D 
• CC from D2-D6 
• Follicular study USG from D9/D10 
• If endometrium is very thin(<5mm) (tab lynoral 
0.05 mg...
Prognostic Factors of ART 
• Maternal age 
• Ovarian reserve
Maternal age 
• Young good result 
• Previous live birth carries better result
Ovarian reserve 
• No of follicle decreases with age 
• At higher age inhibin B decreases due to shrinkage of 
follicular ...
Ovarian reserve 
• Cycle day 3 Serum Estradiol 
> 75-80pg/ml 
• Clomiphene citrate challenge test 
Provocative test. It in...
Evaluation Before IVF 
• Ovarian reserve 
• Male factor 
• Infectious disease 
chlamydial ,HIV,HBV,HCV, 
• Mock embryo tra...
Typical ART cycle 
• COH 
• Monitoring with TVS and Serum E2 
• Prevention of premature LH surge and Ovulation 
• Oocyte m...
In Vitro Fertilization (IVF)
Oocyte retrival 
• 36 h after hCG 
• Laproscopically 
• TVS 
• I.V. sedation propofol/Midazolam/Fentanyl. 
• Prophylactic ...
• All folicle > 10mm aspirated 
• Empty follicle syndrome 
• Complications 
haemorrhage 
pelvic infection 
rupture of a cy...
Oocyte maturation 
• 20-30% of retrieved are immature 
• hCH triggers resumption of meosis
Oocyte maturity 
• Expansion of cumulus 
• Radiance of corona 
• Size and cohesivness of granulosa cells 
• Shape and colo...
Mature oocyte 
• Cumulus cells are expanded Luetinised 
• corona cells –sunburst appearance
Metaphase 1 oocyte 
• No polar body 
• Dense cumulus cells 
• Germinal vesicle and nucleous faded 
Immature oocyte
Fertilization 
• Semen is collected by mastrubation 
• Sperm preparation 
swim up 
density gradient 
• Incubation in high ...
• Conventional ivf – 50-60% fertilization 
• Achieves second meiotic division 
• Extrude second polar body 
• Look for pol...
Intracytoplasmic sperm injection 
ICSI 
• Zona drilling (micropipette and acidified 
tyrode solution) 
• Partial zona diss...
Intracytoplasmic sperm injection 
• Intracytoplasmic sperm injection (ICSI, 
pronounced "eeksee") is an in vitro fertiliza...
Procedure 
• Single sperm is immobilized 
• Drawn in to pipette 
• Oocyte is stabilized 
• Polarbody is 6/12 o’ clock posi...
Indication of ICSI 
• Male factor 
• Oligospermia <5% 
• Asthenospermia <5% 
• Teratospermia <4% 
• PGD 
• Poor IVF/failed...
Embryo culture 
• 4-7% CO2 conc. 
• Incubation volume 10-50microL 
• Embryo group size 1-4 
• Protein supplement 
High ser...
Co-culture system 
• Efforts to create optimal culture led to development of 
co-culture system 
• Human tubal fluid 
• Ma...
• Many of the large programs have attempted 
blastocyst culture, but returned to day 3 
transfers because they had trouble...
Extended (Blastocyst) culture 
• First human birth 
• Now many use cleavage stage embryo(2-3day) 
With knowledge of physio...
Advantage of blastocyst culture 
• True viability assesment is better 
• Excludes embryos that have limited 
Devolopmental...
Disadvantage 
• Doesn’t improve the quality of embryo 
• Lesser quality embryo may fail to grow 
• Multiple pregnancy 
• N...
Preimplantation Genetic Diagnosis 
• It offers couples who carry serious genetic 
disordes the opportunity to have healthy...
• Equipment and procedure is same as ICSI 
• 1-2 cells are extracted 
• Polar bodies 
first polar body 
second polar body ...
PREIMPLANTATION GENETIC DIAGNOSIS 
(PGD)
Embryo transfer 
Can transfer zygote to blastocyst 
Day 3 cleavage embryo 
6-8cells equal size 
no cytoplasmic fragmentati...
Day 5 blastocyst 
• Blastocoel cavity less than half the volume of 
the embryo 
Many cells, tightly packed
Transfer technique 
• Cervical mucus plug aspirated slowly 
• No blood on catheter tip 
• Catheter tip examined microscopi...
Fertilization 
• After the fertilization, 
embryos are transferred 
into uterus, anywhere from 
one to six days later, 
• ...
STEPS
Assisted hatching 
• In vivo zona dissolves on zona- endomet interface 
• In vitro embryo make opening in zona and escapes...
Embryo cryopresevation 
• SUCCESS WITH FROZEN EMBRYO CYCLE significantly increases the cumulative 
pregnancy rate/retrival...
Frozen Embryos 
• Embryos may be taken 
from an individual and 
stored for later use. 
• Once ready to use, they 
can be t...
• All stage of Embryo can be frozen 
• For indefinite time 
• Embryo survival 50-90% 
• Better for zygote than clevage sta...
• Embryo can be refrozen 
• Embryo can be transferred 
natural cycle 
artificial cycle 
GnRHa down regulation 
E2 (microni...
Results of IVF 
• Measured by % of pregnancy,live birth 
• 18 % of pregnancy- miscarriage 15% 
induced abortion 0.9% 
stil...
ART outcome over years 
Age of patients < 35y 35-37y 38-40y 41-42 
ART out come 
in 1996 
(live birth/ ET) 
33.6% 29% 21.6...
• Results are better with 
<35 year 
previous live birth 
previous success IVF 
• poor result 
diminished ovarian reserve ...
Multiple pregnancy 
35% of ART are multiple pregnancy 
30.7% are twins 
4.3% are triplets 
3% in general population
Risks of IVF 
• Ectopic pregnancy 
• OHSS 
Earlier worries of possible link b/n Ovarian 
Ca and OI drugs has declined but ...
Ectopic pregnancy 
• Two time more 
• Risks 
tubal factor infertility 
ET placed high in tube 
larger volume of media 
dif...
Risks of OHSS 
• High exogenous Gn 
• High /rapidly increase in E2 
• Higher /repeated dose of hCG 
• Multiple pregnancy
Prevention of OHSS 
• Elevated/rapidly rising E2 ,then coasting 
• Use low dose hCG(5000iu) 
• Use GnRHant in PCOD 
• Use ...
Offspring from IVF 
• Prematurity 
• LBW 
• Delayed neurological devolopment 
• Congenital bith diffects- two time higher ...
ICSI 
Genetic/epigenitic abnormalities 
Sex chromosomal abnomalities 
1. angelman syndrome 
MR 
Delayed motor development ...
2.Beckwithian syndrome 
Macrosomia 
Macroglossia 
Midline abdominal wall defect 
predisposition to embryonal Ca
Oocyte donaton 
• First reported in 1983 
• Achieved by IVF 
• Recipients partners sperm 
• Transferred to synchronized ut...
Indication 
• Ovarian failure 
• Genetically transmitted disease 
• Diminished ovarian reserve 
• Inaccessible ovaries
Evaluation of recipients 
• Similar in IVF 
• Psychological counseling 
• Turner syndrome 
• Marfans syndrome 
cardiac dis...
Controlled endometrial devolopment 
Endogenous hormones are suppressed-GnRHa 
Follicular phase 7days-3 weeks by E2 
Window...
Donor screening 
• 21-34 y 
• History and examination 
rule/out STD,Genetic disease 
preconception testing 
Blood group Rh...
Embryo endometrial synchronization 
• Progesterone therapy started on day the 
donor undergo retrieval 
• Day 2 embryo – t...
Luteal support 
• 5-7week 
• 10 week for added support
Results of oocyte donation 
• Age is important 
• 35 y 
• 47% live birth with avg 2.9 ET
Gestational surrogacy 
Indication 
Absence of uterus 
Irreparable uterus 
Congenital 
Ascherman Syndrome 
life threatening...
Gamete intrafallopian transfer 
• In gamete intrafallopian transfer (GIFT), eggs 
are removed from the woman, and placed i...
Zygote intrafallopian transfer 
• Zygote intrafallopian transfer (ZIFT) is an 
infertility treatment where a blockage in t...
Gamete Intra-Fallopian Transfer 
(GIFT) 
• A mixture of a 
woman’s eggs and 
sperm are placed 
into the fallopian 
tube du...
GIFT,ZIFT 
• GIFT oocyte and sperm are transferred 
• Zygote in ZIFT 
• Embryo transfer is done by laparoscopy 
4 cm insid...
Surrogacy 
• Two types: 
– Egg donor surrogacy 
– Gestational surrogacy 
• Surrogate may be relative, friend, or paid 
str...
Applications of ART 
• Endanger species 
• Assisted reproductive technology, allows in 
vitro fertilized (IVF ET) embryos ...
Ovarian tissue cryopresevation 
Indication 
pt on chemo/radiotherapy 
Orthotropic transplantation 
Heterotropic transplant...
Oocyte cryopreservation 
• No surgery 
• Few Ca patient have time for stimulation 
• Vitrifaction 
high concentration of c...
Sperm retrieval technique 
• Sperm retrival is done in cases of - 
• Ejaculatory failure 
• Obstructive/Nonobstructive azo...
Testicular sperm aspiration (TESA) 
Testicular sperm aspiration (TESA) is a procedure 
performed for men who are having sp...
Testicular sperm extraction (TESE)/Testis biopsy/Testis 
mapping 
TESE/testis biopsy/testis mapping are procedures perform...
Microepididymal Sperm Aspiration (MESA) 
MESA is a procedure performed for men who have vasal or epididymal 
obstruction (...
Thank you!!
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
Assisted reproductive techniques
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Assisted reproductive techniques

Assisted reproductive techniques

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Assisted reproductive techniques

  1. 1. ASSISTED REPRODUCTIVE TECHNIQUES PROF. M.C. BANSAL M.B.B.S , M.S. , M.I.C.O.G, F.I.C.O.G. Founder Principal& Controller; Jhalawar Medical College and Hospital , Jhalawar. Ex. Principal & Controller; Mahatma Gandhi Medical college And Hospital, Sitapura, Jaipur.
  2. 2. All technique involving direct manipulation of oocyte/sperm outside the body
  3. 3. History of ART • 1978- first successful birth using In Vitro Fertilization • 1984- first successful birth using Gamete Intra Fallopian Transfer • 1986-first successful birth using Zygote Intra Fallopian Transfer
  4. 4. Patrick step toe Robert edwards Nobel prize in2010
  5. 5. • The world’s second and India’s first IVF baby, Kanupriya, alias Durga, was born 67 days later on October 3, 1978, through the efforts of Dr. Subhas Mukherjee and his two colleagues in Kolkata.
  6. 6. Definition of Infertility & ART • Infertility is defined classically as the inability to conceive after 1 year of unprotected intercourse. This definition is based on the cumulative probability of pregnancy: Abdulkareem Sultan Al-Olama 6
  7. 7. Definition of Infertility & ART Cont’d • ART refers to all techniques involving direct retrieval of oocytes from the ovary • ART procedures include IVF, GIFT, ZIFT, and ICSI. • The simplest ART procedure, IVF has been around for over 20 years and is perhaps the most commonly recognized ART of all procedures. Abdulkareem Sultan Al-Olama 7
  8. 8. Causes of infertility
  9. 9. Indications of ART • Tubal factor infertility • Endometriosis • Male factor infertility • Unexplained infertility • Ovarian failure and diminished ovarian reserve • Pelvic malignancy • Mullerian anomaly • Genetic risk
  10. 10. MAJOR ASSISTED REPRODUCTIVE TECHNOLOGIES (ART) • In vitro fertilization and embryo transfer (IVF-ET) • Direct intra-uterine insemination • Gamete intra-fallopian transfer (GIFT) • Zygote intra-fallopian transfer (ZIFT) • Intracytoplasmic sperm injection (ICSI)
  11. 11. Artificial Insemination • Sperm is collected and placed into a woman’s vagina, cervical canal or in the uterus. • Sperm can come from partner or an anonymous donor. •Insemination is when sperm is collected and processed. The sperm is then placed into a woman’s vagina, cervical canal or directly into the uterus. •Insemination may be used if the mucus around cervix is not compatible with partner’s sperm, or may have problems with immune system. This can cause sperm to be killed before egg is fertilized •Artificial Insemination is when the sperm used comes from partner.
  12. 12. DONATED SPERM (INTRAUTERINE INSEMINATION) • Doctor puts donated sperm in woman • Seminal fluid washed from sperm • Donor selection possible
  13. 13. Natural cycle • First birth form IVF is from oocyte collected from natural cycle • Cycle cancellation are high (25%-75%) • Low pregnancy rate • Indication -poor response to stimulation • Advantage -Less monitoring Less costly
  14. 14. STIMULATION PROTOCOLS • Protocol A (suppression and stimulation) • Protocol B(flare up) • Protocol C(cc+ HMG) • Protocol D
  15. 15. PROTOCOL A • Suppression /down regulation • GnRH analogue Inj suprefact(Buserelin acetate) 0.5 cc s/c BD(i.e 20 units of insulin syringe)=0.5 mg from Day 22 of previous cycle (i.e day 7 of post ovulation) • If the cycle is anovulatory/totally irregular then,E2+progesterone along with buserelin
  16. 16. On D1 do E2 and LH estimation a. If E2 is <26 and LH<4 down regulation has been done b. If either of the values are more than that ,continue the same dose of down regulation for another 2 days.if only marginally high OD x 2 days If marginally low no need to repeat Start the stimulation along with 4 units of insulin syringe i.e 0.1 cc OD Suprefact
  17. 17. The initial dose of exogenous gonandotrophins used to stimulate ovarian follicular development depends on individual need • Typical starting dose of HMG (1ampule= 75IU) 3-4 amp I/M daily Suprefact (0.1 cc) I/M or S/C Age 35 yrs,wt 50 kg,FSH-10,previous h/o surgery for endometriosis/PCOD then increase the dose by 1 amp for each cause but not>than 6 amp daily After 3rd day i.e from 4th day onward,start USG for follicular study.We should have a cohort 4-5 follicles on each side and size of >7-8 mm each and about E2>100
  18. 18. • Continue with the same dose .If <2 follicles are leading then stop suprefact and HMG both • If size <7 mm increase the dose by 1 amp of HMG • Growth should be 2mm/day.If <2 mm/day,then increase the dose by 1 amp step by step.When the leading follicle is 18-20 mm give HCG 10000 iu IM • CALL after 36 hrs for ovum pick up(confirm availability of embryologist and other preparations)
  19. 19. PROTOCOL B • FLARE UP • This is suitable for aged patients:where ovarian functions are poor • D1 & D2 Inj buserelin 0.5cc s/c • D3 onwards in morning inj buserelin 0.5 cc s/c and in evening inj HMG 3 amp • D5 onwards start follicular study other steps as before
  20. 20. PROTOCOL C • Clomiphene citrate 150-250 mg from D1-D5 • TVS ON D5/6 • HMG started on D5,follicle 5 mm in each ovary • Next all steps as protocol A
  21. 21. Protocol D • CC from D2-D6 • Follicular study USG from D9/D10 • If endometrium is very thin(<5mm) (tab lynoral 0.05 mg x 5 days/progynova 2 mg x 5 days) may be continued throughout the cycle • When the follicle size >18-20 mm then give inj HCG IM • If 1 follicle-inj HCG 5000 IU • If >2 follicle –inj HCG 10000 IU
  22. 22. Prognostic Factors of ART • Maternal age • Ovarian reserve
  23. 23. Maternal age • Young good result • Previous live birth carries better result
  24. 24. Ovarian reserve • No of follicle decreases with age • At higher age inhibin B decreases due to shrinkage of follicular pool n FSH progressively rises • Ovarian reserve tests even when grossly abnormal should be used to Guide rather than to deny Rx • Tests are Cycle Day 3 Serum FSH <10-15iu/L peak E2 level ie <75-80pg/ml No of Oocytes Pregnancy Live Birth
  25. 25. Ovarian reserve • Cycle day 3 Serum Estradiol > 75-80pg/ml • Clomiphene citrate challenge test Provocative test. It includes day 3 Serum FSH & E2 Clomiphene 5-9 day day 10 serum FSH (>2SD then abnormal)
  26. 26. Evaluation Before IVF • Ovarian reserve • Male factor • Infectious disease chlamydial ,HIV,HBV,HCV, • Mock embryo transfer • Evaluation of uterus HSG,Hysteroscopy,Sonohysterography.
  27. 27. Typical ART cycle • COH • Monitoring with TVS and Serum E2 • Prevention of premature LH surge and Ovulation • Oocyte maturing with HCG • Oocyte retrival • Fertilization by IVF/ICSI • Invitro embryo culture • Luteal support • Transfer of fresh embryo/cryopreserved • First trimister preg monitoring
  28. 28. In Vitro Fertilization (IVF)
  29. 29. Oocyte retrival • 36 h after hCG • Laproscopically • TVS • I.V. sedation propofol/Midazolam/Fentanyl. • Prophylactic antibiotic • No antiseptic ,clean with NS • 5-7 MHz, TVS 16G needle,vaccum100- 200mmhg,the follicle wall rapidly collapse but donot obstruct the needle lumen
  30. 30. • All folicle > 10mm aspirated • Empty follicle syndrome • Complications haemorrhage pelvic infection rupture of a cyst laceration of sacral vein lumbosacral osteomyelitis
  31. 31. Oocyte maturation • 20-30% of retrieved are immature • hCH triggers resumption of meosis
  32. 32. Oocyte maturity • Expansion of cumulus • Radiance of corona • Size and cohesivness of granulosa cells • Shape and color of oocyte • first polar body • Germinal vesicle
  33. 33. Mature oocyte • Cumulus cells are expanded Luetinised • corona cells –sunburst appearance
  34. 34. Metaphase 1 oocyte • No polar body • Dense cumulus cells • Germinal vesicle and nucleous faded Immature oocyte
  35. 35. Fertilization • Semen is collected by mastrubation • Sperm preparation swim up density gradient • Incubation in high protien media for 0.5-4hr • Each oocyte incubated with 50-100 thousand motile sperm in 5% CO2 in air, 98%humidity,37°C for 12-18hr
  36. 36. • Conventional ivf – 50-60% fertilization • Achieves second meiotic division • Extrude second polar body • Look for polyploidy.it can be observed in 5- 10% embryos
  37. 37. Intracytoplasmic sperm injection ICSI • Zona drilling (micropipette and acidified tyrode solution) • Partial zona dissection • Subzonal insertion This all requires sperm to interact with oolemma and did not prevent polyspermic fertilization
  38. 38. Intracytoplasmic sperm injection • Intracytoplasmic sperm injection (ICSI, pronounced "eeksee") is an in vitro fertilization procedure in which a single sperm is injected directly into an egg. • This procedure is used to overcome male infertility problems, although it may also be used where eggs cannot easily be penetrated by sperm, and occasionally as a method of in vitro fertilization, especially that associated with sperm donation.
  39. 39. Procedure • Single sperm is immobilized • Drawn in to pipette • Oocyte is stabilized • Polarbody is 6/12 o’ clock position • Oocyte is entered 3 o’clock • Pipette pierce zona and oolemma • 50-70% fertilization
  40. 40. Indication of ICSI • Male factor • Oligospermia <5% • Asthenospermia <5% • Teratospermia <4% • PGD • Poor IVF/failed IVF
  41. 41. Embryo culture • 4-7% CO2 conc. • Incubation volume 10-50microL • Embryo group size 1-4 • Protein supplement High serum albumin Recombinant albumin Synthetic serum substitute
  42. 42. Co-culture system • Efforts to create optimal culture led to development of co-culture system • Human tubal fluid • Maternal serum/protien substitute • Autologus endometrial cells Risk of infection FDA has not approved Used in failed IVF
  43. 43. • Many of the large programs have attempted blastocyst culture, but returned to day 3 transfers because they had trouble getting the embryos to grow the blastocyst stage. Growing the embryos to the blastocyst stage requires great attention to detail - a luxury not afforded in a big program performing dozens of procedures a day. • Programs performing less than 200 cycles per year have had the best luck with blastocyst culture and transfer with reported (but unverified) pregnancy rates in the 50-70% range for younger patients and egg donor cycles.
  44. 44. Extended (Blastocyst) culture • First human birth • Now many use cleavage stage embryo(2-3day) With knowledge of physiologic requirement,there is development of sequential media that varies in composition with stage of embryo development Precompaction embryo(morula) need pyruvate and non essential a a Post compaction embryo(blastocyst) glucose and essential AA
  45. 45. Advantage of blastocyst culture • True viability assesment is better • Excludes embryos that have limited Devolopmental potential • Synchronize the stage of development • Reduce the abnormal endometrial milieu • Reduce risk of expulsion • Allows PGD • Few embryo can be transformed
  46. 46. Disadvantage • Doesn’t improve the quality of embryo • Lesser quality embryo may fail to grow • Multiple pregnancy • Need extended culture
  47. 47. Preimplantation Genetic Diagnosis • It offers couples who carry serious genetic disordes the opportunity to have healthy child • Aneuploidy • Structural abnomalites (translocations,invertions) • Inherited gene disorder Cystic fibrosis Thalassemia Haemophelia Duchnemuscular dystrophy
  48. 48. • Equipment and procedure is same as ICSI • 1-2 cells are extracted • Polar bodies first polar body second polar body paternal chromosome are not assesed • The most common aproach to PGD is Cleavage stage embryo biopsy typically performed on day 3 of fertization before embryo starts to compact
  49. 49. PREIMPLANTATION GENETIC DIAGNOSIS (PGD)
  50. 50. Embryo transfer Can transfer zygote to blastocyst Day 3 cleavage embryo 6-8cells equal size no cytoplasmic fragmentation
  51. 51. Day 5 blastocyst • Blastocoel cavity less than half the volume of the embryo Many cells, tightly packed
  52. 52. Transfer technique • Cervical mucus plug aspirated slowly • No blood on catheter tip • Catheter tip examined microscopically after transfer • Soft catheter/ stiff catheter • Volume of media < 50 micro L • Catheter tip does not touch fundus and transfer occus at level of 0.5 cm below fundus.whenever possible ,mucus ,blood ,ut.contractions should be avoided. • A preliminary trial transfer is done to identify women who may benefit from cervical dilatation before treatment begins
  53. 53. Fertilization • After the fertilization, embryos are transferred into uterus, anywhere from one to six days later, • but usually this happens between 2 to 3 days after the egg retrieval. • During this time, the fertilized egg separates to become a 2-4 cell embryo.
  54. 54. STEPS
  55. 55. Assisted hatching • In vivo zona dissolves on zona- endomet interface • In vitro embryo make opening in zona and escapes ,leaving behind an empty zona • Methods Zona drilling with acidic tyrode’s solution partial zona dissection with micro glass needle Laser photo ablation Enzymatic Hatching Use of Piezo -micromanipulator Indication failed IVF poor prognostic factor Disadv.-Hatching may cause embryo demage and the risk of monozygotic the risk of monozygotic twinning inceases
  56. 56. Embryo cryopresevation • SUCCESS WITH FROZEN EMBRYO CYCLE significantly increases the cumulative pregnancy rate/retrival of oocyte • OHSS is avoided Principle It has two distinct stages –freezing and thawing Object of freezing is to avoid crystallization of intercellular water cell water gradually replaces cryoprotectent dimethyl sulfoxide propanediol glycerol embryos are sealed in vials and cooled -30°C to -110◦C and then stored in liquid nitrogen After thawing, process is reversed, gradually passing the embryo through decreasing conc. of cryoprotectant.
  57. 57. Frozen Embryos • Embryos may be taken from an individual and stored for later use. • Once ready to use, they can be thawed and then placed into the uterus. • This allows a higher chance of pregnancy.
  58. 58. • All stage of Embryo can be frozen • For indefinite time • Embryo survival 50-90% • Better for zygote than clevage stage and blastocyst • Overall success 15-20% • Thawing can be done sequentially until the no. of transfer embryos is reached or the larger no of embryo is thawed to select with best morphology
  59. 59. • Embryo can be refrozen • Embryo can be transferred natural cycle artificial cycle GnRHa down regulation E2 (micronized E2 4-6mg/d, transdemal 0.1-0.2mg after mensus) (inhibit rise in FSH) estimate serum P <1ng/mL time of transfer to synchronize stage of embryo
  60. 60. Results of IVF • Measured by % of pregnancy,live birth • 18 % of pregnancy- miscarriage 15% induced abortion 0.9% still birth 0.6% ectopic pregnacy 0.7% Overall 28.4% pregnancy/retrieval 16% clinical pregnancy/ET in frozen embryo
  61. 61. ART outcome over years Age of patients < 35y 35-37y 38-40y 41-42 ART out come in 1996 (live birth/ ET) 33.6% 29% 21.6% 11.5% ART out come in 2001 (live birth/ET) 41.1% 35% 25.4% 14.5%
  62. 62. • Results are better with <35 year previous live birth previous success IVF • poor result diminished ovarian reserve uterine factor multiple factor
  63. 63. Multiple pregnancy 35% of ART are multiple pregnancy 30.7% are twins 4.3% are triplets 3% in general population
  64. 64. Risks of IVF • Ectopic pregnancy • OHSS Earlier worries of possible link b/n Ovarian Ca and OI drugs has declined but still linger
  65. 65. Ectopic pregnancy • Two time more • Risks tubal factor infertility ET placed high in tube larger volume of media difficult transfer high hormones • Heterotropic pregnancy 1in 10.000pregnancy
  66. 66. Risks of OHSS • High exogenous Gn • High /rapidly increase in E2 • Higher /repeated dose of hCG • Multiple pregnancy
  67. 67. Prevention of OHSS • Elevated/rapidly rising E2 ,then coasting • Use low dose hCG(5000iu) • Use GnRHant in PCOD • Use GnRHa for LH surge • If symptoms of OHSS oocyte retrival and freezing • Prophylactic i.v. albumin infusion 20-50G
  68. 68. Offspring from IVF • Prematurity • LBW • Delayed neurological devolopment • Congenital bith diffects- two time higher NTD Alimentary atresia Omphalocele Hypospadias
  69. 69. ICSI Genetic/epigenitic abnormalities Sex chromosomal abnomalities 1. angelman syndrome MR Delayed motor development poor balance abnormal movement absent speech
  70. 70. 2.Beckwithian syndrome Macrosomia Macroglossia Midline abdominal wall defect predisposition to embryonal Ca
  71. 71. Oocyte donaton • First reported in 1983 • Achieved by IVF • Recipients partners sperm • Transferred to synchronized uterus
  72. 72. Indication • Ovarian failure • Genetically transmitted disease • Diminished ovarian reserve • Inaccessible ovaries
  73. 73. Evaluation of recipients • Similar in IVF • Psychological counseling • Turner syndrome • Marfans syndrome cardiac disease aortic root dissection
  74. 74. Controlled endometrial devolopment Endogenous hormones are suppressed-GnRHa Follicular phase 7days-3 weeks by E2 Window of endometrial receptivity 3day (max 5d) controlled by duration of Progesterone can be administerd intramuscularly from the day of retrieval/ 4days before transfer in a dose of 20-50mg/d to achieve serum conc. of 20ng/mL TVS endometrium >6-7mm
  75. 75. Donor screening • 21-34 y • History and examination rule/out STD,Genetic disease preconception testing Blood group Rhtype rubella and varicella HIV 1&2,HBV,HCV,Gonrrhea Chlamydia Pschologic evaluation
  76. 76. Embryo endometrial synchronization • Progesterone therapy started on day the donor undergo retrieval • Day 2 embryo – third day of Progesterone therapy • Day 3 Embryo –fourth day • Day 5 Embryo –sixth day
  77. 77. Luteal support • 5-7week • 10 week for added support
  78. 78. Results of oocyte donation • Age is important • 35 y • 47% live birth with avg 2.9 ET
  79. 79. Gestational surrogacy Indication Absence of uterus Irreparable uterus Congenital Ascherman Syndrome life threatening medical disorder Person related/nonrelated parous healthy
  80. 80. Gamete intrafallopian transfer • In gamete intrafallopian transfer (GIFT), eggs are removed from the woman, and placed in one of the fallopian tubes, along with the man's sperm. This allows fertilization to take place inside the woman's body. Therefore, this variation is actually an in vivo fertilization, and not an in vitro fertilization.
  81. 81. Zygote intrafallopian transfer • Zygote intrafallopian transfer (ZIFT) is an infertility treatment where a blockage in the fallopian tubes are the cause. Egg cells are removed from a woman's ovaries, and in vitro fertilized. The resulting zygote is placed into the fallopian tube by the use of laparoscopy.
  82. 82. Gamete Intra-Fallopian Transfer (GIFT) • A mixture of a woman’s eggs and sperm are placed into the fallopian tube during a laparoscopy. • Once inserted, fertilization is allowed to occur.
  83. 83. GIFT,ZIFT • GIFT oocyte and sperm are transferred • Zygote in ZIFT • Embryo transfer is done by laparoscopy 4 cm inside the fimbria GIFT 27% ZIFT 27.9% Indication difficult IVF Religious Ectopic pregnancy
  84. 84. Surrogacy • Two types: – Egg donor surrogacy – Gestational surrogacy • Surrogate may be relative, friend, or paid stranger
  85. 85. Applications of ART • Endanger species • Assisted reproductive technology, allows in vitro fertilized (IVF ET) embryos for preimplantation genetic screening (PGS) evaluation. The is not used to look for a specific disease but a technique to identify embryos at risk.
  86. 86. Ovarian tissue cryopresevation Indication pt on chemo/radiotherapy Orthotropic transplantation Heterotropic transplantation Under trial
  87. 87. Oocyte cryopreservation • No surgery • Few Ca patient have time for stimulation • Vitrifaction high concentration of cryopresevent is used glass like state thawed oocyte –pregnancy is similar to fresh
  88. 88. Sperm retrieval technique • Sperm retrival is done in cases of - • Ejaculatory failure • Obstructive/Nonobstructive azoospermia • Retrograde ejaculation
  89. 89. Testicular sperm aspiration (TESA) Testicular sperm aspiration (TESA) is a procedure performed for men who are having sperm retrieved for in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI). It is done with local anesthesia in the operating room or office and is coordinated with their female partner’s egg retrieval. A needle is inserted in the testicle and tissue/sperm are aspirated. TESA is performed for men with obstructive azoospermia (s/p vasectomy). Often TESA doesn’t provide enough tissue/sperm and an open testis biopsy is needed.
  90. 90. Testicular sperm extraction (TESE)/Testis biopsy/Testis mapping TESE/testis biopsy/testis mapping are procedures performed for men who have testis failure. The procedure is performed to see if there are sperm present as well as for pathologic diagnosis to evaluate for malignancy. It is either done as a scheduled procedure or is coordinated with their female partner’s egg retrieval. TESE is usually performed in the operating room with sedation, but can be performed in the office with local anesthesia alone.. Patients usually cryopreserve sperm during this procedure for future IVF/ICSI. This diagnostic biopsy is usually performed to evaluate for an obstructive etiology – microdissection TESE has replaced this as the optimal form of retrieval for testis failure patients.
  91. 91. Microepididymal Sperm Aspiration (MESA) MESA is a procedure performed for men who have vasal or epididymal obstruction (s/p vasectomy, cystic fibrosis). It is either done as a scheduled procedure or is coordinated with their female partner’s egg retrieval. MESA is performed in the operating room with general anesthesia under the operating microscope. Patients usually cryopreserve sperm during this procedure for future IVF/ICSI. Microdissection TESE (microdissection testicular sperm extraction) Microscopic TESE is a procedure performed for men who have testis failure. Microdissection TESE is performed in the operating room with general anesthesia under the operating microscope. Patients cryopreserve sperm during this procedure for future IVF/ICSI.
  92. 92. Thank you!!

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