Effects of rate of development of in vitro-produced ovine embryos on sex ratio and in vivo survival after embryo transfer

1997 ◽  
Vol 48 (8) ◽  
pp. 1369-1378 ◽  
Author(s):  
S.L. Catt ◽  
J.K. O'Brien ◽  
W.M.C. Maxwell ◽  
G. Evans
Keyword(s):  
Sex Ratio ◽  
Embryo Transfer ◽  
Rate Of Development

ECONOMIC PERSPECTIVES OF BIOTECHNOLOGIES USING IN ANIMAL FARMING

1970 ◽  
pp. 57-60
Author(s):  
I. F. Gorlov ◽  
A. A. Mosolov ◽  
G. V. Komlatskiy ◽  
M. A. Nesterenko ◽  
K. D. Nimbona ◽  
...  
Keyword(s):  
Embryo Transfer ◽  
Economic Effect ◽  
Dairy Herd ◽  
State Level ◽  
Modern Level ◽  
Economic Perspectives ◽  
The Cost ◽  
Short Time

The article presents materials on the study of the possibility of reproduction and increase in the herd of highly productive cows through the use of embryo transplantation technology. The classical (in vivo) and more modern, developing (in vitro) methods of embryotransfer, their positive and negative sides are considered in detail. The possibility of accelerating the breeding process by using the method of transplantation, in which from one cow can be obtained from 10 to 100 calves, which will allow for 4-5 years, almost any herd (of any size and breed) with the help of biotechnology to turn into a cattle-breeding enterprise of the most modern level. At the same time, heifers obtained from unproductive cows can be used as "surrogate" mothers who are transplanted with the best donor embryos, which allows to obtain a full-fledged offspring adapted to local environmental conditions. A detailed scheme of obtaining, evaluation, storage, as well as the cost and economic effect of embryo transplantation was calculated, the market was evaluated, the required annual volume of transplants and the number of donor cows for large livestock farms were determined. As a positive example of "Scientific-production enterprise "Centre of biotechnology and embryo transfer" in 2014, implemented a project for accelerated replacement and genetic improvement of the dairy herd, engraftment averaged 57-69%, and the economic effect of the enterprise from getting a single animal by the method of embryo transfer, compared with imports of similar close in quality, ranged from 60 to 100 thousand rubles on his head. It is shown that it is necessary to organize at the state level a developed service for embryo transplantation to reduce the cost of embryo transfer and the possibility of creating in a short time in the country's own highly productive breeding nucleus of dairy and beef cattle, which will reduce, and in the future completely eliminate, import dependence on cattle products.

161 LIPID CONTENT, RESISTANCE TO CRYOPRESERVATION, AND SEX RATIO OF IN VITRO BOVINE BLASTOCYSTS PRODUCED IN A SERUM-FREE SYSTEM

2006 ◽  
Vol 18 (2) ◽  
pp. 188
Author(s):  
F. George ◽  
C. Daniaux ◽  
G. Genicot ◽  
F. Focant ◽  
B. Verhaeghe ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Lipid Content ◽  
Culture Medium ◽  
Nile Red ◽  
Embryo Cryopreservation ◽  
Hatching Rate ◽  
Free System ◽  
Serum Free

In vitro-produced (IVP) bovine blastocysts are known to be more sensitive to cryopreservation than their in vivo counterparts. Removing serum from the culture medium decreases sanitary risk and could improve embryo resistance to cryopreservation by preventing the accumulation of intracellular lipids. Our objectives were to evaluate the lipid content, resistance to cryopreservation, and sex ratio of IVP embryos cultured in a serum-free system. Oocytes from slaughterhouse ovaries were matured in a serum-free enriched medium (Donnay et al. 2004 Reprod. Fertil. Dev. 16, 274) and cultured in 5% O2 in modified SOF supplemented with 5% FCS (FCS) or with insulin-transferrin-selenium (ITS) and 0.1 mg/mL polyvinylpyrrolidone (PVP) (ITS-PVP) or 4 mg/mL BSA (ITS-BSA) (Daniaux et al. 2005 Reprod. Fertil. Dev. 17, 217). Day 5 morulae were stained with the fluorescent dye Nile Red in order to evaluate their lipid content (Genicot et al. 2005 Theriogenology 63, 1181). Day 7 blastocysts (diameter ≥160 µm) were selected, classified according to their size, and frozen in HEPES-SOF containing 1.5 M ethylene glycol, 0.1 M sucrose, and 1.8 mg/mL wheat peptones (George et al. 2002 Reproduction 29, 51). The lipid content was significantly lower in morulae cultured in ITS-BSA compared with the two other media (320 ± 10 arbitrary fluorescence units vs. 383 ± 12 in FCS and 406 ± 10 in ITS-PVP; n = 271; ANOVA2: P < 0.01). After cryopreservation, a higher total hatching rate was found 24 h post-thawing in blastocysts cultured in ITS-BSA and for both serum-free conditions at 48 h (Table 1). In particular, embryos ≤180 µm cultured in FCS were less resistant to cryopreservation than embryos of the same size produced without serum. Expanded blastocysts cultured in ITS-BSA were sexed by PCR (Grisart et al. 1995 Theriogenology 43, 1097) and a higher proportion of male embryos was found (62.7%; n = 51). In conclusion, a complete serum-free system was set up from oocyte maturation to embryo cryopreservation that gave high quality embryos resistant to cryo-preservation. Embryos produced in ITS-BSA presented a lower lipid content, but a shift of the expanded blastocyst sex ratio toward males was observed. Table 1. Hatching rates post-thawing as a function of the blastocyst size and the culture medium

264 TESTOSTERONE IN THE OOCYTE CULTURE DOES NOT ALTER SEX-RATIO OF IN VITRO PRODUCED BOVINE EMBRYOS

2009 ◽  
Vol 21 (1) ◽  
pp. 229
Author(s):  
C. Díez ◽  
P. Bermejo-Alvarez ◽  
A. Gutiérrez-Adan ◽  
J. N. Caamaño ◽  
M. Muñoz ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Cumulus Cells ◽  
Basic Medium ◽  
Bovine Embryos ◽  
Experimental Conditions ◽  
Vitro Fertilization ◽  
Grant Support ◽  
High Concentrations

The production of sex-known offspring is a main objective in reproductive biotechnology. It has been reported that bovine ova developed in follicles with high concentrations of testosterone in vivo yielded significantly more male embryos in vitro (Grant V et al. 2008 Biol. Reprod. 78, 812–815). In this work we aimed to test the effects of testosterone on sex ratio of bovine embryos produced in fully in vitro conditions. Immature bovine cumulus–oocyte complexes (COCs; n = 750) from slaughterhouse ovaries were cultured in 199 HNaCO3 with polyvinyl alcohol (PVA) 0.1 mg mL–1 as a basic medium. Culture was made in two steps, a 24 h meiotic arrest (roscovitine 25 μm), and a subsequent in vitro maturation period with FSH-LH for 24 h. Testosterone (T-86500, Sigma-Aldrich, St. Louis, MO, USA) was added throughout the entire oocyte culture at 0, 30, 300, and 1500 nm. After in vitro fertilization (Day 0), zygotes were freed of cumulus cells by pipetting, and subsequently cultured in SOF + 6 g L–1 BSA up to Day 3. At this time, embryo development was recorded, and all embryos having 3 or more cells were treated with pronase to remove the zona pellucida. Zona-free embryos were washed in PBS containing PVA 0.1 mg mL–1 and individually frozen at –80°C until sex analysis by PCR (Bermejo-Alvarez P et al. 2008 Biol. Reprod. doi:10.1095/biolreprod.108.070169). A total of 252 embryos from 5 replicates were sexed. Data for development and sex-ratio are presented as % LSM ± SD. There were no interactions between testosterone treatment, embryonic sex, and embryonic stage analyzed. Testosterone did not affect development rates (P > 0.05) at any stage: cleavage (47.8 ± 6.8, 56.5 ± 6.8; 50.9 ± 6.8; 62.2 ± 6.8), 3 to 4 cells (40.6 ± 5.2, 45.8 ± 5.2; 37.8 ± 5.2; 47.7 ± 5.2) and >5 cells rates (24.5 ± 4; 27.3 ± 4; 21.3 ± 4; 25.3 ± 4) for 0, 30, 300, and 1500 nm testosterone, respectively. Cumulative percentages of male embryos were as follows: 53 ± 8 (n = 56), 42.6 ± 8 (n = 52), 53.6 ± 6 (n = 81) and 57.6 ± 8 (n = 63) for 0, 30, 300, and 1500 nm groups respectively (P > 0.05). These results show that the testosterone effects on oocyte ability to select Y-chromosome bearing spermatozoa are not reproducible in vitro under the present experimental conditions. Grant support: MEC, project AGL2008-01530; RTA2008-0082; M. Muoz is supported by FICYT.

Factors affecting the sex ratio of in-vitro and in-vivo produced bovine embryos

1999 ◽  
Vol 51 (1) ◽  
pp. 396 ◽  
Author(s):  
G Carbonneau ◽  
H Twagiramungu ◽  
N Morin ◽  
C Brisson ◽  
J Durocher ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Bovine Embryos ◽  
Factors Affecting

Some Observations on the Effect of Digestive Juices on Scolices ofEchinococcus granulosus(Batsch).

1936 ◽  
Vol 14 (1) ◽  
pp. 21-40 ◽  
Author(s):  
D. A. Berberian
Keyword(s):  
Gastric Juice ◽  
Intestinal Tract ◽  
Time Intervals ◽  
Human Gastric Juice ◽  
Rate Of Development ◽  
Single Cyst ◽  
Intestinal Juice ◽  
Definite Time

Experimentsin vivoandin vitroare reported on the action of digestive juices of various animals on the scolices ofE. granulosus. Inin vitrostudies, scolices ofE. granulosuswere placed in the digestive juices of different animals, incubated at 37°C., and the digestive action of the fluids was studied by examining portions of the material under the microscope.In vivoexperiments were carried out on kittens, rats and rabbits. These animals were fed large quantities of scolices of hydatid cyst membranes and they were killed at definite time intervals and their intestinal tract was carefully examined for scolices.Gastric juice of rats, dogs, cats, sheep and cattle did not digest scolices. The action of the gastric juice of rabbits begins late and proceeds slowly. Human gastric juice causes incomplete digestion and acts only on the evaginated scolices.The intestinal juices of man, rats, rabbits, sheep and cattle are able to digest scolices completely, whereas the intestinal juice of dogs and cats is inactive. In spite of the fact that cat intestinal juice is inactive, kittens are found to be slightly susceptible. Since they suffer only relatively light infestation and the rate of development is retarded, we would classify the cat as an “abnormal” host toE. granulosus.Time of evagination of scolices from a single cyst or from cysts from different animals is variable. Some scolices evaginate readily, others more slowly and still others fail to evaginate completely.

scholarly journals Sex ratio of infants born through in vitro fertilization and embryo transfer: Results of a single-institution study and literature review

2021 ◽  
Author(s):  
Chinatsu Nagata ◽  
Keiko Mekaru ◽  
Keiya Gibo ◽  
Rie Nakamura ◽  
Sugiko Oishi ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Sex Ratio ◽  
Literature Review ◽  
Embryo Transfer ◽  
Single Institution ◽  
Vitro Fertilization

164 EFFECTS OF KINETICS AND MORPHOLOGY ON EARLY EMBRYONIC DEVELOPMENT IN BOVINE OPU-IVF EMBRYOS

2016 ◽  
Vol 28 (2) ◽  
pp. 212
Author(s):  
M. Takayama ◽  
O. Dochi ◽  
K. Imai
Keyword(s):  
Embryo Transfer ◽  
Calf Serum ◽  
Conception Rate ◽  
Culture Dish ◽  
Fresh Embryo Transfer ◽  
Cell Stage ◽  
Vitro Fertilization ◽  
Observation Method

In recent years, the use of ovum pick up (OPU) and IVF for embryo production has increased worldwide; however, the conception rate of embryo transfer is lower for OPU-IVF embryos than for in vivo-derived embryos. This study aimed to determine the efficacy of embryo selection by a 3-step observation method by using a micro-well culture dish (Dai Nippon Printing, Tokyo, Japan). In this study, 9 Holstein and 15 Japanese Black cows were used, and the OPU-IVF was conducted from October 2014 to May 2015. The collected cumulus-oocyte complexes (COC) were cultured for 22 h in 25 mM HEPES-buffered TCM-199 supplemented with 5% calf serum (CS) and 0.02 AU mL–1 of FSH. Sperm (at a final concentration of 5 × 106 spermatozoa mL–1) were incubated with COC for 6 h. After insemination, presumptive zygotes were separated from cumulus cells and sperm by pipetting. Then, the presumptive zygotes were cultured for 9 days in CR1aa supplemented with 5% CS by using a micro-well culture dish. Kinetics and morphology were observed at 27, 31, and 55 h post-insemination (hpi). The presumptive zygotes were divided to 3 groups (more than 2 cells, 2 cells, and no cleavage) at 27 and 31 hpi. Then, embryos at the 2-cell stage at 31 hpi were divided into 2 groups: 2-cell with normal cleavage and 2-cell embryos with abnormal cleavage (unequal cleavage, 2-cell with fragments, and 2-cell with protrusion). Subsequently, embryos were classified as 8-cell and more than 8 cell, or less than 8 cell at 55 hpi. The blastocyst rate (BL%) was analysed at 7, 8, and 9 days post IVF. Embryos selected by the 3-step observation method were used for fresh embryo transfer. The data were analysed by chi-squared test. In total, 856 oocytes were collected by OPU and 633 oocytes were cultured, of which 39.7% (263/663) developed to the blastocyst stage. The BL% of 2-cell embryos (72.5%, 116/160) was significantly higher (P < 0.01) than that of no cleavage (47.0%, 117/249) at 27 hpi. The BL% of 2-cell (65.4%, 206/315) and more than 2-cell (53.0%, 35/66) was significantly higher (P < 0.01 and P < 0.05) than that of embryos divided as no cleavage (25.9%, 22/85) at 31 hpi. The BL% was not significantly different between 2-cell with normal cleavage (68.5%, 172/251) and abnormal cleavage (53.1%, 34/64). The BL% of 8-cell and more than 8-cell stage (72.8%, 182/250) was significantly higher (P < 0.01) than that of embryos with less than 8 cells (38.8%, 81/209) at 55 hpi. Overall, 2-cell embryos at 27 hpi, 2-cell embryos with normal cleavage at 31 hpi, and 8-cell and more than 8 cell at 55 hpi showed the highest BL% (82.1%, 78/91). The conception rate was higher for following the selected fresh embryo transfer that was 70.6% (12/17) than average of in vitro fertilization embryos transfer that was 40.0%. These results demonstrate that the 3-step observation method used in this study can be effectively applied for the selection of IVF embryos that have a strong ability to develop into blastocysts and high competence for conception.

66 DEVELOPMENT OF IN VITRO-PRODUCED CAT EMBRYOS AFTER VITRIFICATION AND NONSURGICAL EMBRYO TRANSFER: PRELIMINARY RESULTS

2009 ◽  
Vol 21 (1) ◽  
pp. 133
Author(s):  
E. Iacono ◽  
B. Merlo ◽  
M. Regazzini ◽  
D. Zambelli
Keyword(s):  
Ethylene Glycol ◽  
Embryo Transfer ◽  
Domestic Cat ◽  
Embryo Survival ◽  
Abdominal Ultrasonography ◽  
Preliminary Results ◽  
Pregnancy Status ◽  
Viable Embryo

There are no refereed reports on vitrification of domestic cat embryos derived from in vitro-matured oocytes and transferred using a nonsurgical embryo transfer technique. The aim of this study was to verify the effects of vitrification on the in vitro and in vivo developmental ability of in vitro-produced (IVP) cat blastocysts. Oocytes recovered from minced ovaries were matured, fertilized, and cultured in vitro as previously reported (Merlo B et al. 2005 Theriogenology 63, 2032–2039). On Day 7 of in vitro culture (IVC), blastocysts were selected and vitrified in straws (Cristal ET 0.25 mL, 133 mm, IMV-Technologies, Paillette Crista, France). For vitrification (modified from Campos-Chillòn LF et al. 2006 Theriogenology 65, 1200–1214), the embryos were transferred in 1 mL of V1 [ethylene glycol 3.5 m in HEPES synthetic oviductal fluid (HSOF)] for 3 min, and then in 10 μL of V2 (ethylene glycol 7 m, galactose 0.5 m, Ficoll 70 18% in HSOF) for 20 s. Finally, the embryos were loaded in straws preloaded with 190 μL of dilution solution (galactose 0.5 m in HSOF). Straws were heat sealed and immediately plunged into liquid nitrogen. Vitrified embryos were warmed in air for 10 s, and then in a waterbath at 37°C for 30 s. For developmental ability and in vitro evaluation, 27 embryos were warmed and immediately examined: 25 re-expanded, 2 did not re-expand, and 1 had damaged zona pellucida. Re-expanded embryos were cultured in SOF plus amino acids, 16 mg mL–1 BSA, and 5% fetal bovine serum at 38.5°C in 5% O2, 5% CO2, 90% N2. After 24 h of IVC, only 4 blastocysts were expanded, and after 48 h, embryos were clearly degenerated or shrunk. in vivo developmental ability was tested by nonsurgical embryo transfer of 8 vitrified-warmed embryos and 6 IVP fresh embryos into 2 natural estrus queens, injected with 200 IU of hCG i.m. (Day 0) for induction of ovulation. Ovulation was confirmed by plasmatic progesterone assay on Day 5. Nonsurgical embryo transfer was made on Day 8 using the catheter proposed by Zambelli et al. 2001 for transcervical insemination in the cat. The catheter was connected to a 1-mL syringe and loaded with the embryos. Then, it was inserted in the vagina and transrectally guided into the uterus, where the embryos were deposited. To assess pregnancy status, abdominal ultrasonography was done on recipients on Day 13, 25, and 40. On Day 13, an embryonic vesicle was observed in both queens, although a smaller diameter than expected was detected in the recipient of the vitrified embryos. On Day 25, a viable embryo was detected only in the recipient of fresh IVP embryos. On Day 40, the gestational chamber was still present but no sign of a viable embryo was detected. Further studies are in progress to improve the nominal incidence of pregnancy and frequency of embryo survival after vitrification. Nevertheless, the preliminary results obtained using an AI catheter for nonsurgical embryo transfer are encouraging, and the improvement of the technique could make it reliable in the cat. Supported by Animal Stem Cells Laboratory, Regione Emilia Romagna, PRRIITT Project Number M-404AIWTSV.

363 FACTORS INFLUENCING PREGNANCY RATES FOLLOWING TRANSFER OF BOVINE IN VIVO EMBRYOS BIOPSIED FOR SEX DETERMINATION

2007 ◽  
Vol 19 (1) ◽  
pp. 297
Author(s):  
S. Li ◽  
W. Yu ◽  
J. Fu ◽  
Y. Bai ◽  
F. Jin ◽  
...  
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Pregnancy Rates ◽  
Chi Square ◽  
Embryo Survival ◽  
Chi Square Test ◽  
First Time ◽  
Fresh Embryos

Data collected from commercial embryo transfer programs in 63 farms in China during June 2002 to December 2005 was analyzed to examine the effects of various factors (biopsy, freezing, sample size, embryo development and quality, in vitro culture, and recipient quality) on pregnancy rates of in vivo-biopsied embryos. Embryos were flushed from superovulated dairy cattle and subjected to a biopsy for sexing determination using protocols and sexing kits supplied by AB Technology Ltd. Fresh embryos were implanted on the same day or frozen with AG freeze medium (AB Technology Ltd., Pullman, WA, USA) for later transfer. Recipients were synchronized with CIDA + PG protocols. Embryos were cultured in 6-well dishes containing 1.3 mL of holding medium (AB Technology Ltd.) in each well at room temperature (20–25�C) for examination of embryo survival in vitro. The chi-square test was used in statistic analysis. The implantation of fresh embryos after biopsy did not affect pregnancy rates (49.6%, 257/518) compared to that of non-biopsied fresh and frozen–thawed embryo groups (52.9%, 47/140 and 46.6%, 177/380, respectively). However, for biopsied embryos subjected to frozen and thawed procedures before implantation, particularly for those subjected to the removal of a larger biopsy, a reduced pregnancy rate was observed (41.8%, 297/710; P &lt; 0.01). Pregnancy rates among biopsied embryos at 3 different development stages (morula-early blastocyst, blastocyst, and expanded blastocyst) were not different. Similar results were found between embryo groups of grade 1 and 2. A significant decrease in pregnancy rate (0/10) was observed with embryos held in vitro for a longer period of time (&gt;5 h), suggesting detrimental effects of in vitro conditions on embryo survival. The highest pregnancy rate (68.0%) was observed in recipients synchronized for the first time before being implanted with biopsied embryos. Significant decreases in such rates were found in recipients synchronized for the second or third times or those with an abortion history at the first or second synchronization-implantation treatment (P &lt; 0.01). Better pregnancy rates (45.6%, 41/90; 46.1%, 76/165; and 45.5%, 5/11) were obtained for recipients implanted with biopsied embryos at Days 7.5, 8.0, and 8.5 post-heat detection, respectively, compared to 16% at Day 7 (3/18, P &lt; 0.05). It is concluded that mechanical treatment (cutting) does not reduce the survival of biopsied embryos; however, cryopreservation reduces their ability to survive in vivo. The analyses also suggest that holding embryos in vitro should not be longer than 5 h unless more favorable in vitro conditions can be provided. To achieve better results of implantation of biopsied embryos, embryo transfer should be performed during 7.5–8.5 days post-estrus, and the healthy recipients synchronized for the first time should be used.

47 CRYOPRESERVATION OF BOVINE GERM CELL USING ANTIFREEZE POLYAMINO-ACID (CARBOXYLATED POLY-L-LYSINE)

2017 ◽  
Vol 29 (1) ◽  
pp. 131
Author(s):  
T. Fujikawa ◽  
C. Kubota ◽  
T. Ando ◽  
S. Imamura ◽  
M. Tokumaru ◽  
...  
Keyword(s):  
Survival Rate ◽  
Germ Cell ◽  
Embryo Transfer ◽  
Conception Rate ◽  
Control Group ◽  
Vitro Fertilization ◽  
Polyamino Acid ◽  
Significant Difference

Carboxylated poly-l-lysine (CPLL) is an ampholytic polymer compound, and it is obtained by converting 65% amino groups to carboxyl groups after synthesising ε-poly-l-lysine aqueous solution and succinic anhydride. CPLL has cryoprotective property similar to antifreeze protein, and addition of CPLL into cryopreservation medium improves the post-thaw survival rate of cells and embryos. In this research, we examined the effectiveness of CPLL as a bovine germ cell cryoprotective material. In experiment 1 (in sperm), the conventional cryopreservation medium used for control group was consisted of 6.5% (vol/vol) glycerin, and the cryopreservation medium used for CPLL group was consisted of 3.25% (vol/vol) glycerin and 0.5% CPLL (wt/vol). The post-thaw survival and motility were assessed by using Sperm Motility Analysis System (DITECT Corp., Tokyo, Japan). There was no significant difference for post-thaw survival rate and motility (control v. CPLL; 98.8% v. 96.6% and 69.7% v. 62.2%, respectively). Artificial insemination was carried out in 65 cows (control v. CPLL; 34 v. 31), and the conception rate of the CPLL group was higher than that of the control group (80.6% v. 67.6%; P = 0.23). In experiment 2 (embryos), the conventional cryopreservation medium used for control group was consisted of 5% (vol/vol) ethylene glycol and 6% (vol/vol) propylene glycol in PBS. In the CPLL group, 7% (wt/vol) CPLL was added to the conventional medium. In vitro fertilization embryos were cryopreserved at Day 7 and Day 8. There was no significant difference in survival rate at 0, 24, and 48 h and hatched rate until 72 h after thawing (control v. CPLL: 93.6% v. 93.2%, 69.0% v. 64.7%, 56.1% v. 56.3%, 12.9% v. 10.2%, respectively). Embryos obtained by superovulation treatment and in vivo fertilization at Day 7 were cryopreserved using above 2 media, and transferred non-surgically into synchronized recipient cows (1 embryo per animal). Embryo transfer (ET) was carried out in 81 cows (control v. CPLL: 31 v. 50), and recipients were diagnosed for pregnancy ultrasonically 50 days after embryo transfer. Conception rate of CPLL group was higher than control group (50.0% v. 29.0%; P = 0.063). In both experiments, the significant differences between control group and CPLL group were determined by chi-squared test. The effectiveness of CPLL in cells and embryos has been reported; however, there is no report using CPLL in bovine germ cells. In this research, CPLL improved the conception rate of AI and ET, probably due to its low toxicity and protection of the cell membrane. These results suggest that CPLL is available as a new cryoprotective material for bovine sperm and embryo in slow freezing methods.

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