1. The process Assessment IVF ICSI
Fertilization
The process
Assessment
IVF
ICSI

2. The Process - sperm capacitation hyperactivation zona binding
acrosome reaction
zona penetration
sperm-oocyte membrane fusion
sperm engulfed
decondensation of sperm nucleus
formation of male pronucleus

3. The Process - Egg Ovulation Sperm-egg membrane fusion Egg activation
increase in Ca++
cortical reaction - zona block
completion of meiosis (2nd PB)
formation of female pronucleus
Syngamy - fusion of both pronuclei

4. Capacitation process of sperm becoming fertile occurs in uterus
must be removed from seminal fluid
in vitro, wash sperm
capacitated sperm have the ability to fertilize the egg

5. Hyperactivation facilitates sperm-oocyte contact occurs in oviduct
motility pattern becomes frenzied
linear to non-linear motion

6. Zona Binding Zona proteins: ZP1, ZP2, ZP3 Sperm bind ZP3 specifically
Initiates acrosome reaction

7. Acrosome Reaction Fusion of two membranes Acrosin released
spermatazoal plasma membrane
outer acrosomal membrane
Acrosin released
allows sperm to penetrate zona
Exposes equatorial segment
for fusion with egg

8. Sperm-Egg Fusion Equatorial segment of sperm Plasma membrane of egg
Initiates egg activation

9. Egg Activation Cortical Reaction Zona block
exocytosis of cortical granules
Zona block
biochemical changes eliminates sperm binding

10. Assessment of Fertilization
Check for PN hrs post insemination
Normal Fertilization
2 PN in ooplasm
2 PB perivitelline space

11. Abnormal Fertilization
> 2 PN in ooplasm
polyspermic: >1 male PN, 1 female PN, 2PB
polygynic: >1 female PN, 1 male PN, 1PB
failure to extrude second PB
Spontaneous oocyte activation
1 female PN, 2 PB
Condensed sperm: 1 female PN, 2 PB
Oocyte activation (digynic)
>1female PN, 1 PB

12. IVF: Why? Humans: overcome infertility Animals: end of reproductivity
male factor: low motility, low count
female factor: tubal, ovulatory
Animals: end of reproductivity
more superior animals
Research: IVM with IVF
many embryos to study techniques: transgenics, es cells, cloning

13. Intracytoplasmic Sperm Injection
Overcome fertilization problems:
no penetration
ASAB, anti-sperm antibodies
very low count (<500,000/sample)
very low motility (<20% motile)

14. ICSI - process Add sperm to PVP Mature egg w/holding pipet
inert, slows sperm to catch
Mature egg w/holding pipet
PB at 12:00 or 6:00
Catch sperm and kink tail
Load sperm in ICSI pipet
Break plasma membrane
Deposit sperm