Vitrification of embryos post biopsy with Cryotop Method - Damia Castello, Embryologist IVI Spain

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Damia Castello, an Embryologist from IVI Spain presented on vitrification of embryos post bipsy with cryotop method at the RI Istanbul Conference and Workshop on the 23rd and 24th January 2014.
He covered the following:
- Injuries during cryopreservation
- Strategies to minimize injuries
- Cryopreservation procedures
- Vitrification & Cryotop vitrification method
- Vitrification Protocol Embryo
- Vitrification in PGD blastocysts


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Vitrification of embryos post biopsy with Cryotop Method - Damia Castello, Embryologist IVI Spain

  1. 1. Vitrification of embryos post biopsy with Cryotop Method Damià Castelló www.dibimed.com
  2. 2. Injuries during cryopreservation Mechanism Factors Influenced by Ice crystal Toxic Osmotic Chilling etc. Size Shape Permeability Quality Sensitivity Species Stage
  3. 3. Strategies to minimize injuries o Slow freezing: An attempt to maintain a balance between various sources of injury by using low CPA concentration and controlled ice formation. o Vitrification: Radical elimination of ice formation, and to reduce toxic and osmotic damage.
  4. 4. Cryopreservation procedures Slow Freezing Vitrification Transformation of a liquid in solid with formation of ice. It is essential that the ice does not form inside the cell: Transformation of a liquid in very viscous solid with no ice: o Use of CPAs at low concentration (1,5M). oHigh CPAs concentration (3-6M). oCooling rate (0,3ºC/min). oProgrammable o Very high cooling rates: 15.000 a 30.000 ºC/min. freezers. oDirect immersion into liquid Nitrogen.
  5. 5. Vitrification Solidification of water or water based solutions without ice crystal formation. It is facilitated by:  High CPA concentration (3-6M)  High cooling rates: 15.000 to 30.000 ºC/min Direct immersion into LN
  6. 6. Cryotop Vitrification Method What is? It is a MVC device, consisting of a fine polypropilene strip, which allow the oocytes to be vitrified within very low amount of volume (0,1µl).
  7. 7. Vitrification Kit Thawing Kit
  8. 8. KITAZATO CRYOTOP VITRIFICATION: o o o o o o Valid for all stages of development, since Oocyte to Blastocyst. Cryotop SC allows vitrification in closed system for storage Vitrification Media is synthetic (free of albumin) to reduce risks. Threalose (endotoxin free) replaced Sucrose for more safety. Vitrification up to 16 Oocytes, PN, Embryos or Blastocysts in 20 min. Thawing up 16 Oocytes, PN, Embryos or Blastocysts in 20 min.
  9. 9. Protocol
  10. 10. Vitrification Protocol Embryo
  11. 11. Vitrification Protocol Embryo
  12. 12. Vitrification in PGD blastocysts
  13. 13. Our experience in IVI: December 2011 to August 2013. 81 pacients. 257 Blastocysts.
  14. 14. •When we biopsy Trophoectoderm? Assisted Hatching Vitrification Trophoectoderm Biopsy
  15. 15. Survival: Survival Not Survival Survival Rate Euploid 54 4 95.2% Aneuploid 217 10 92.6% Total 271 14 94.8% Biopsy doesn’t afect survival after thawing using Cryotop Method.
  16. 16. Clinical Outcomes: n Patients Not analysed Blastocysts % 8 9.9 Patients Not Transfered Blastocyst 27 Patients Transfered Blastocyst 36 33.3 14 Clinical Pregnancy Rate/cycle Clinical Pregnancy Rate/transfer 30.8 69.4 Implantation rate 62.0
  17. 17. A total of 25 pregnant patients: n Deliveries 6 Ongoing 14 Miscarriage 3 Not call response 2 Live Born + Ongoing 86.9%
  18. 18. Results
  19. 19. Outcomes of vitrified early cleavage-stage and blastocyst-stage embryos in a cryopreservation program: evaluation of 3,150 warming cycles Alla Cobo. Ph 0 . l1arill J o s d f tos Santos. Ph 0 ,.Oa'llia Castel o. Ph O.. Pi ar Garniz. Ph.D.• PiarCampos. M.L.T., a..,d J01:e Remoh1, M.O. 03 02 05 06 n (o/o) n (% ) n (o/o) n (% ) 497 147 1 ,725 3,491 675 1 079 603 952 :>SO(1 9 - 0.8)" 7 6 177.2)' 59 (l 0 .1) 3,057 ( I 8 0 .6)" 1.058 (34.6)b.c 708 ( 1 0) No. of V11dm 11•19 cyclP No. of warmed embryos ,., No. of c>my > r!'p l.1u•d lm r .11 +SD) l c>' u lrnpldnl.itm1 r.it" CPFVcycle . . I (1.5 - 0.6)0 306 (34 9i•.< 751 (41 fiJ r 44129.9) 571 (33.1) I r;03 I ) 44 (29.9) 5 l 1.h.3> 123 (17.3) 570 ( 'l.D) ar; er r.11<, ..uridqp rr1h> . e DfVcycle LBfVcyde www.dibimed.com 677 (39.2;• .' 56 (18 9) 235 • 8) ..:M l 1 4 40 6.o.t> I 73 (29) 178 <29.51 190 <3z.s 1 •" Dibimed
  20. 20. CRYOTOP Survival rates published worldwide 100% 80% 60% 40% 20% 0% 124 Oocytes 3.286 Oocytes 153 Oocytes 3.491 Embryos Rienzi 2010 Cobo 2010 Nagy 2009 Cobo 2012 298 Embryos TsengKailing 2010 2.031 Blastocysts 2.543 Embryos 192 Oocytes Cobo 2012 Wenhau Shi Krinos 2011 2012 442 Blastocysts 186 Oocytes Dandan Zhu 211 ChingChien Chang 2013 Oocyte Vitrification in ART
  21. 21. Thank you!!

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