Oleh Prof Dr Ir herry Sonjaya, DEA DES
EMBRIO TRANSFER III
l Sinkronisasi
Donor dan Resipien
l Betina
resipien hr kondisi baik, bebas Px genital tract dan mempunyai siklus estrus yg
normal
l Resepien
hrs mempunyai estrus sama dg donor
l Hasil
optimum: resipien hrs dlm waktu 12 jam drpd donor angka kebuntingan rendah > 24 jam pd
sapi
> 48
jam pd kambing dan domba
l Embrio
beku: ditransfer 7 hr setelah estrus keresipien
Teknik ET
Sapi dan Kerbau
l Awal
melalui laparotomi
Th 1978 digunakan teknik Transcevical yaitu:
Ø Palpasi
rektal: ada tdknya CL
Ø Anasthesi
epidural posterior
Ø Kondisi
aseptis, embrio dimasukan ke gun transfer
yg dilapisi dg sarung steril
Ø Gun
dimasukan kedlm vagina dan dg rektal diarahkan ke kornua uteri ipsilateral dg
CL.
Ø Kemudian
pd kornua uteriStraw didepositkan
Kambing dan domba
l Midventral
laparotomi
l General
anasthetic/local
l Insisi
midventral: dg pipet kapiler embrio ditransfer melalui infundibulum ke ampula.
Bila Ditransfer ke uterus:
l Kornua
uteri ditusuk dg jarum tumpul, lalu dg pipet kapiler didepositkan dalamlumen
yeterus.
l Embrio
dlm uterus dg laparoskopi
Keberhasilan ET
l Hrs
mencapai 80% embrio yg hidup
l Angka
kebuntingan tinggi pd domba, sapi dan kambing dr hasil transfer 1 embrio setiap
kornua sering hasilnya Twin
l Pada
Babi: 6-10 embrio harus ditransfer oleh karena biasanya hanya setengahnya
menjadi lahir
DONORS AND RECIPIENTS
l Donor
–
Overcome follicular dominance to induce superovulation
–
FSH ( as FSH, PMSG or HMG)
–
Induce luteolysis ( PGF2α) to coincide with the superstimulated follicular phase and to permit ovulation
l Recipients
–
Induce luteolysis at the same time as the donor ( 1
day) PGF2α
TYPICAL SUPEROVULATION
PROTOKOL
l Treat
with PGF2α. Observe for estrus
l Starting
on Day 10 - 12 of ensuing cycle: ( CL
present, dominant follicle absent)
–
1. Inject FSH, twice/day for 4 days (
constant or decreasing doses)
–
2. Inject PGF2α on the third day of FSH
treatment ( + recipients)
–
3. Inseminate every 12 h; 48, 60 and 72 h After
PGF2α
–
4. Flush embryos 6 -
7 days after A.I.
SUPEROVULATION
l FSH
stimulates increased follicular growth ( 10 -
12 follicles per ovary)
l PGF2α
induces luteal regression midway during
induced follicular phase
l -
E2-17β .. - P4 induces LH surge and ovulations
( over a period of 8 - 12 h, cf sow)
l Inseminate
more frequently to ensure fertility of
all oocytes as they are released
l Morula
and blastocysts recovered from the uterus
at 6 - 7 days
of development
FLUSHING EMBRYOS
l Non-surgical/transcervical
flush ( each horn separately)
l PBS
+ Serum + Antibiotics
l Filter/allow
embryos to settle
l Search
under dissecting microscope:
Evaluate:
Unfertilized oocytes - discard
Early
cleavage stages ( 4 cell, 8 cell are
likely arrested) - discard
Morula
Blastocysts
Fragmentation/colour/expansion
Transferable”
EMBRYO TRANSFER SUCCESS
l Superovulation:
0 - 60 + CL’s 10 - 20 on average
l Embryos:
•
0
•
All unfertilized
•
0 - 30
+ embryos
•
4 - 10 on average ( 6) transferable
•
(+ some unfertilized, early stages, fragmented)
l Generally
a lottery:
–
Donors: Some repeatedly succeed, some never
–
succeed, some have good days and bad ones
KEYS TO SUCCESS
l 1.
Ensure donor has correct cycle stage @ start
l 2.
Ensure doses + timing of all procedures are correct
l 3.
Don’t cheat on semen (+ expect to get away with it)
l 4.
Donors should be lean, recipients should be in
good condition, heifers and properly
synchronized (Prefer a slightly
early uterus - so embryo can “catch-up”
- especially if manipulated. D7
embryo/D6 uterus)
ASSISTED REPRODUCTIVE
TECHNOLOGIES
l Embryo
Transfer (incl. Juvenile)
l Embryo
Cryopreservation
l Embryo
Sexing
l Embryo
Manipulation:
–
Splitting Nuclear
Transfer
–
Cloning Chimeras
–
Transgenics
l In
vitro fertilization
l Intracytoplasmic
sperm injection (ICSI)
l G.I.F.T.
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