influences of cryoprotectants on viability of embryos produced in vitro

The post-thaw embryo survival has been shown to depend on different factors such as type of cryoprotectant. Ethylene glycol, 1,2 propanediol and glycerol are among the routine cryoprotectants widely used for embryo cryopreservation in different animals and human as well. To investigate the effects of different cryoprotectants on viability of blastocysts produced in vitro and also determining a suitable cryoprotectant for embryo cryopreservation. A total of 197 porcine blastocysts produced in vitro [at days 6 and 7 post-IVF] were randomly divided into control and cryoprotectant [CP] groups. The CP groups were exposed to 10% CP solutions [Ethylene glycol, 1,2 propanediol and glycerol] in 3 steps for 1 hr at room temperature [23-25°C]. The survival rate was measured as the proportion of recovered embryos following a 24-hr culture in NCSU-37 media. The survival rate was further compared with the data obtained from the control group [cultured in 0.3% BSA in PBS with the same conditions]. The results showed that the survival rates of blastocysts exposed to PD and GLY were similar to those exposed to EG [p<0.05]. However, there was no significant difference [P>0.05] in survival rates between the EG and control groups. The data indicated that the exposure of porcine blastocysts to cryoprotectant causes a reduction in survival rate and that the ethylene glycol produced the least detrimental effects

Factors affecting in vitro production of buffalo embryos

The present study was designed to examine the influence of oocyte quality, culture media and gonadotropins on cleavage rate and development of in vitro fertilized buffalo Oocytes. Three experiments were conducted. In Experiment 1; oocytes were classified by number of cumulus cell layers and morphology of the oopbsm as Good, Fair or Poor. Oocytes were cultured for IVM, IVF and IVC in CRlaa medium. In Experiment 2, good quality oocytes were cultured for maturation in: [1] CR1aa; [2] CR2aa; [3] TCM-199; [4] MEM and [5] RPMI-1640, and then fertilized using frozen thawed buffalo spermatozoa in CRlaa. After fertilization, oocytes were cultured in the same medium used for maturation. In Experiment 3, oocytes were classified into 3 groups: Group [1] without gonadotropin and served as control; Group [2] in which IVM medium was supplemented with 10 micro g/ml FSH; Group [3] in which IVM medium was supplemented with 10 IU /ml PMSG. In all experiments, oocytes were kept at 38.5°C under 5% CO[2] for IVM, IVF, IVC and examined for cleavage and embryo development rates on day 3 and 8, respectively. Good and fair quality oocytes produced a higher [P <0.01] cleavage rate than poor quality oocytcs. Morula production rate was also higher [P < 0.01] for good as compared to fair quality oocytes. Embryo development with poor quality oocytcs was arrested at the 2 to 16-cell stage. In, Experiment 2, the cleavage rate was significantly higher [P < 0.05] in CR1aa than CR2aa. and significantly higher [P < 0.01] than TCM-199, MEM and RPMI-1640. The numbers of morulae and blastocysts were higher [P < 0.01] for oocytes cultured in CR1aa and CR2aa media than TCM-199 or MEM. In Experiment 3, the addition of FSH or PMSG to the maturation medium significantly increased [P < 0.01] cleavage and developmental rates of buffalo embryo compared to control media. In conclusion, the IVM of good quality buffalo oocytes in CR1aa or CR2aa medium and the addition of FSH or PMSG in maturation medium produced higher cleavage and developmental rates of IVF buffalo embryos