Technical Reference #1897

1897.

Xenotransplantation of cryopreserved human ovarian tissue into murine back muscle R. Soleimani; E. Heytens; R. Van den Broecke; I. Rottiers; M. Dhont; C.A. Cuvelier; and P. De Sutter, Ghent University, Human Reproduction, 25(1897), (2010)
Link To Paper

Keywords:
xenotransplantation/ovary cryopreservation/human ovary/muscle/metaphase II oocyte

Materials & Methods:
Human OT was obtained from a consenting 22-year-old female-to-male transsexual. Cryopreservation was performed as described previously (Van den Broecke et al. 2001) and the tissue was stored for 5 years. Rapid thawing was carried out by placing the vials containing cryopreserved human OTs above liquid nitrogen for 30 s and placing them in a water bath at room temperature for 90 s. Tissue strips were washed immediately five times for 1 min each with M-199 medium (Sigma M-2154 Bornem Belgium) in 60 mm tissue culture dishes (Falcon; BD 35–3002 VWR Leuven Belgium) to remove the cryoprotectant (dimethylsulphoxide Sigma D5879). OT strips were cut into small pieces of 1 mm2 with surgical blades (no. 24) and were washed again in M-199 medium for 5 min. Follicular content of the OT mini strips was evaluated using the method previously described by our group (Soleimani et al. 2006). Briefly OT strips of 1 mm2 (1+0.12 mm2; mean+SD) and 0.5 mm thickness (476+176 mm; mean+SD) were placed in pre-warmed M-199 medium in a glass-bottom dish (MatTek Ashland USA) and follicular content was measured using an Olympus SZ-60 stereomicroscope. OT fragments containing at least five visible follicles were selected and collected in another dish in pre-warmed M-199 medium for transplantation.

Microscopic Technique
Stereomicroscopy

Cell Type(s)
Ovarian Tissue