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Laboratory aspects of IVM

Laboratory aspects of IVM. DAL CANTO MARIABEATRICE BIOGENESI REPRODUCTIVE MEDICINE CENTER, ISTITUTI CLINICI ZUCCHI V. Zucchi, 24-Monza (Italy) tel/fax +39.39.8383314. PRE-ANTRAL FOLLICLE 2-5 mm PROPHASE I Cell diameter 80 µ m Monolayer cumulus cells. ANTRAL FOLLICLE 5-12 mm

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Laboratory aspects of IVM

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  1. Laboratory aspects of IVM DAL CANTO MARIABEATRICE BIOGENESI REPRODUCTIVE MEDICINE CENTER, ISTITUTI CLINICI ZUCCHI V. Zucchi, 24-Monza (Italy) tel/fax +39.39.8383314

  2. PRE-ANTRAL FOLLICLE 2-5 mm PROPHASE I Cell diameter 80 µ m Monolayer cumulus cells ANTRAL FOLLICLE 5-12 mm PROPHASE I Cell diameter 110-120 µ m Monolayer cumulus cells Zona pellucida Cytoplasmic maturation Minimum follicular diameter to produce a competent oocytes is around 5 mm

  3. GVBD 2n OOCYTES MATURATION LH MPF cAMP FF-MAS cAMP cAMP granulosa cell cAMP cAMP PROPHASE I METAPHASE II

  4. DAY -1 DAY 1-2 DAY 0 IVF I.V.M. I.V.F. ...IVM REQUIRES FLEXIBILITY AND ADJUSTMENTS ...

  5. IN VITRO MATURATION Oocytes Collection TRANSVAGINAL US GUIDANCE CRAFT VACUUM PUMP (pressure 80-90 mmHg) ASPIRATION NEEDLE (Gynetics 4551E  17-G single lumen) SINGLE FLASK TO COLLECT OOCYTES

  6. IN VITRO MATURATION Oocytes Collection DAY –1 • Disposable Material • Filter 0.70 μm • Syringes • Flask or tube • Petri dishes • Flushing Medium with Heparin • Maturation Medium

  7. B Centro di Medicina della Riproduzione IOGENESI DAY -1

  8. DAY -1

  9. B Centro di Medicina della Riproduzione IOGENESI DAY -1

  10. B Centro di Medicina della Riproduzione IOGENESI DAY -1

  11. DAY - 1

  12. IN VITRO MATURATION Oocytes Collection

  13. IN VITRO MATURATION Biological Aspects • PICK UP • (8.30 am) • 3 hrs INCUBATION • IN LAG MEDIUM • (09.00) • 26 hrs INCUBATION • IN IVM MEDIUM • (12.00 am) DAY –1 r-FSH 0.075 IU/ml hCG 0.10 IU/ml 10% patient’s serum

  14. IN VITRO MATURATION Oocytes Classification EXPANDED CUMULUS FULL CUMULUS SPARSE CUMULUS NUDE

  15. DAY -1 EXPANDED CUMULUS Immature oocytes with multilayer and slack cumulus

  16. B Centro di Medicina della Riproduzione IOGENESI DAY -1 FULL CUMULUS Immature oocytes with compact complete cumulus

  17. B Centro di Medicina della Riproduzione IOGENESI DAY -1 SPARSE CUMULUS Immature oocytes with partial cumulus

  18. DAY -1 NUDE Immature oocytes without cumulus ATRETIC

  19. IN VITRO MATURATION Biological Aspects DAY 0 • Maturation control(3.00 pm) • hyaluronidase 80 UI • ICSI(4.00 pm)

  20. DAY 0

  21. IN VITRO MATURATION Biological Aspects DAY 1 • Fertilization control • (8.30 am) • Culture in multiwells • in ISM-1 Medium

  22. IN VITRO MATURATION Biological Aspects DAY 2 • Embryo control • Embryo transfer No cleavage delay was observed

  23. RESULTS Natural IVM

  24. RESULTS Natural IVM

  25. 09.03.2004 A new Law regarding ART was approved in Italy ...use no more than 3 oocytes for each cycle so as to avoid to produce more than 3 embryos... ...all embryos obtained must be transferred simultaneously with no selection.... 3 topics ... embryo cryopreservation is forbidden....

  26. RESULTS Natural IVM

  27. IVM: the first choice in Italy Pregnancy rate with standard IVM, ICSI and IVF-ET in women ≤39 age (May 2004-September 2005) Dal Canto et Al., RBM Online Vol.13, N°2, August 2006

  28. RESULTS IN SELECTED WOMEN * *

  29. RESULTS IN SELECTED WOMEN WITH “TOP QUALITY” EMBRYOS * * * analysis performed on women <37 years in which only “top embryos” were transferred

  30. we are studing… • New Maturation Medium • Extended Culture up to blastocyst Stage • In-vitro matured oocyte Cryopreservation

  31. B Centro di Medicina della Riproduzione IOGENESI Equipe Medica Dr R. Fadini Dr M. Mignini Renzini Dr R. Comi Dr G. Rangoni Dr A. Villa Dr M. Mastrolilli Dr A. Epis Equipe Biologica Dr M.B. Dal Canto Dr F. Brambillasca Dr M. Crippa Dr D. Fumagalli Dr M. Lain Dr. M. Merola Sig.ra M. Majoli Sig.ra A. Longoni Segreteria Sig.ra C. Del Favero; Sig.ra P. Loreto; Sig.ra F. Rapisarda; Sig.ra F. Froio

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