1. A Unified Approach to Human Embryo Culture Don Rieger, Ph.D. Vice President, Research & Development LifeGlobal LLC DonRieger@LIfeGlobal.com

2. Overview I.A Single Medium vs. Sequential Media II.The global ® Family of Media III.A Unified Approach to Culture

3. A Single Medium vs. Sequential Media from Zygote to Blastocyst

4. A Single Medium vs. Sequential Media for Embryo Culture “… in order to optimize mammalian embryo development in culture, sequential media are required, each designed to meet the changing requirements of the developing embryo.” (Gardner and Lane, In Principles of Cloning, 187-213, 2002) “The popularity of this conviction is due to its intuitive appeal, but critical examination shows that it is not supported by direct experimental evidence” (Biggers et al., Hum Reprod 20, 3376-84, 2005)

5.  A single medium can be used for culture from the zygote to the blastocyst stage  Changing the medium is a stress on the embryo

6. MediumYearInvestigators SOM1991Lawitts & Biggers KSOM1994Erbach et al. KSOMaa2000Biggers et al. global ® (Ala-Gln)2002Wiemer et al. global ® (Gly-Gln)2006(Biggers et al., 2004) The Development of global ®

7. global ® Formulation

8. Greenblatt et al. Toronto, ON, Canada (Human Reprod. 20, Suppl. 1, i21, Abstr. O-058, 2005)

9. Zech et al. Bregenz, Austria (Human Reprod. 21, Suppl. 1, i162, 2006)

10. Angus et al. Houston, TX, USA (Fertil. Steril 86, Suppl. 2: S229, 2006)

11. Matsubara et al. Hiroshima, Japan (Proc. 24th Ann. Meet. Japan Soc. Fertil. Implant., 206, 2006 )

12. Sepulveda et al. Lima, Peru (Proc. Red Latinamer. Reprod. Asist., 49-50, 2007)

13. Vansteenbrugge et al. Namur, Belgium (Fertil. Steril 88, Suppl 1: S319, 2007)

14. Conclusion A single medium (Global) is as good as, or better than, sequential media for development of the human embryo to the blastocyst stage

15. The global ® Family of Media

16. The global ® Family of Media Objective To maintain, as closely as possible, the same medium throughout all stages of gamete and embryo handling and culture In order to Minimize the stress of changing from one medium to another

17. 1. global ® for Fertilization

18. Energy Metabolism of the COC and Sperm

19. global ® for Fertilization Formulation

20. 2. global ® w/HEPES

21. Acid-Base Buffer Systems

22. global ® w/HEPES Formulation

23. 3. PGD Biopsy Medium

24. The PGD Biopsy Procedure

25. The Effect of Biopsy Medium on Pregnancy Rate (Hill & Li, The Clinical Embryologist 7, 11-12, 2004)

26. PGD Biopsy Medium Formulation

27. The Unified Approach to Culture

28. Day 1 ART Procedures – Day 0 Classical IVF global® for Fertilization ICSI global® w/HEPES Culture global® Day 1

29. ART Procedures – Day 1 Classical IVF global® for Fertilization Post-ICSI Culture global® Culture fresh droplets of global® Day 3

30. ART Procedures – Day 3 Culture global® Day 5 Culture fresh droplets of global® Assisted Hatching global® w/HEPES PGD Biopsy PGD Biopsy Medium CryopreservationTransfer global®

31. Transfer global® Cryopreservation ART Procedures – Day 5 and 6 Culture global® Cryopreservation Transfer global® Culture fresh droplets of global® Day 6

32. Thank You

33. Biggers JD (2001) Research in the canine block. Int J Dev Biol 45, 469-76.