Supplementary MaterialsBelow may be the image is definitely a web link to a higher resolution version Supplementary Shape 1: Viability of goat early-staged follicles predicated on trypan blue staining (nonviable cells). major, and supplementary) follicles are generally seen in goats (Lucci et al. 1999). 30 m (JPG 301?kb) 441_2008_613_Fig2_ESM.jpg (295K) GUID:?E1DAE269-8767-424D-B49C-B0E3F47FC4BC Supplementary Structure 1: Experiment 1: osmotic tolerance of early-staged follicles to NaCl, sucrose, and ethylene glycol (sucrose) ahead of freezing (DOC 33?kb) 441_2008_613_MOESM4_ESM.doc (33K) GUID:?98C6B494-F285-44AE-901C-19997838E397 Supplementary Structure 3: Test 3: comparison between sluggish and fast chilling of early-staged follicles Evista pontent inhibitor (sucrose) (DOC 33?kb) 441_2008_613_MOESM5_ESM.doc (33K) GUID:?41B8F450-149E-4D7C-85CE-0CFAE6173C07 Supplementary Structure 4: Test 4: in vitro culture of early-staged follicles (sucrose) (DOC 34?kb) 441_2008_613_MOESM6_ESM.doc (34K) GUID:?B721C734-963D-4022-97EE-1CCDABD3EFBE Abstract Isolated caprine early-staged follicles were submitted to osmotic tolerance tests in the current presence of sucrose, ethylene glycol (EG), or NaCl solutions and were subjected to and cryopreserved (by sluggish or fast chilling) in MEM alone or MEM supplemented with Evista pontent inhibitor sucrose, EG (1.0 or 4.0?M), or both. When follicles had been Evista pontent inhibitor subjected to 1.5?M NaCl, just 2% from the follicles were viable, whereas 87% from the follicles were viable after contact with 4.0?M EG. Concerning exposure time, the best percentage of practical follicles was obtained when follicles were exposed for 10?min to 1 1.0?M EG?+?0.5?M sucrose; exposure for 60 s to 4.0?M EG?+?0.5?M sucrose also maintained high percentage viability in follicles. Slow cooling in the presence of 1.0?M EG?+?0.5?M sucrose (75%) or rapid Evista pontent inhibitor cooling in the presence of 4.0?M EG?+?0.5?M sucrose (71%) resulted in a significantly higher proportion of viable follicles than all other treatments (at 40?m Open in a separate window Fig.?2 Viability based on trypan blue staining of isolated caprine early-staged follicles exposed to NaCl (experiment 1; ((((((((non-viable cells). a Positive control, Triton X-100 treated secondary follicle. b Viable secondary follicle. c Non-viable secondary follicle. 50?m (JPG 240?kb) Supplementary Figure 2(295K, jpg)Viability of goat early-staged follicles based on staining with Hoechst (all nuclei) and ethidium homodimer-1 (nuclei from dead cells). a, b Representative image of a viable primordial follicle. c, d Representative image of two non-viable primordial follicles. Follicle contains a viable oocyte, whereas fewer than 90% of granulosa cells are viable. Follicle contains three oocytes of which one has deteriorated, and fewer than 90% of granulosa cells are viable (oocytes). Note that polyovular healthy early-staged (primordial, primary, and secondary) follicles are commonly observed in goats (Lucci et al. Rabbit Polyclonal to GAS1 1999). 30 m (JPG 301?kb) Supplementary Scheme 1(29K, doc)Experiment 1: osmotic tolerance of early-staged follicles to NaCl, sucrose, and ethylene glycol (sucrose) prior to freezing (DOC 33?kb) Supplementary Scheme 3(33K, doc)Experiment 3: comparison between slow and rapid cooling Evista pontent inhibitor of early-staged follicles (sucrose) (DOC 33?kb) Supplementary Scheme 4(34K, doc)Experiment 4: in vitro culture of early-staged follicles (sucrose) (DOC 34?kb) Acknowledgements The authors thank Arend Rijneveld and Frans van Kooi for logistical help, Dr. Damien Paris for the English review, and two anonymous reviewers for their helpful suggestions. Open Access This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited. Footnotes Electronic supplementary material The online version of this article (doi:10.1007/s00441-008-0613-9) contains supplementary material, which is available to authorized users..