Effect of protein supplementation on development to the hatching and hatched blastocyst stages of cat IVF embryos

2002 ◽  
Vol 14 (5) ◽  
pp. 291 ◽  
Author(s):  
N. W. Kurniani Karja ◽  
Takeshige Otoi ◽  
Masako Murakami ◽  
Minori Yuge ◽  
Mokhamad Fahrudin ◽  
...  
Keyword(s):  
Bovine Serum ◽  
Culture Medium ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Early Embryo ◽  
Protein Supplementation ◽  
Vitro Fertilization ◽  
Fetal Bovine ◽  
The Mean

The effects of protein supplementation in culture medium on development to the hatching and hatched blastocyst stages of cat in vitro-fertilized embryos were investigated. In the first experiment, presumptive zygotes derived from in vitro maturation and in vitro fertilization (IVF) were cultured in modified Earle's balanced salt solution (MK-1) supplemented with 0.4% bovine serum albumin (BSA) or 5% fetal bovine serum (FBS) for 9 days. There were no significant differences between the BSA and FBS groups with respect to the proportion of cleavage and development to the morula and blastocyst stages of zygotes. However, the presence of FBS in the medium enhanced development to the hatching blastocyst stage of zygotes compared with the BSA group (31.4% v. 7.8%). Moreover, 2.9% of zygotes cultured with FBS developed to the hatched blastocyst stage. The mean cell number of blastocysts derived from zygotes cultured with FBS was significantly higher (P<0.01) than that from zygotes cultured with BSA (136.6 v.101.5). In the second experiment, embryos at the morula or blastocyst stage, which were produced by culturing in MK-1 supplemented with 0.4% BSA after IVF, were subsequently cultured in MK-1 with 0.4% BSA or 5% FBS. Significantly more morulae developed to the blastocyst (P<0.05) and hatching blastocyst stages (P<0.01) in the FBS group than in the BSA group (71.5% and 53.6% v. 44.9% and 6.0%, respectively). Although none of the morulae cultured with BSA developed to the hatched blastocyst stage, 11.5% of morulae cultured with FBS developed to the hatched blastocyst stage. Moreover, the proportion of development to the hatching blastocyst stage of blastocysts was significantly higher (P<0.01) in the FBS group than in the BSA group (68.7% v. 9.8%). None of the blastocysts cultured with BSA developed to the hatched blastocyst stage, whereas 7.3% of blastocysts cultured with FBS developed to the hatched blastocyst stage. The results of the present study indicate that supplementation with FBS at different stages of early embryo development promotes development to the hatching and hatched blastocyst stages of cat IVF embryos.

scholarly journals Fetal bovine serum is associated with polar body degeneration after in vitro maturation of bovine oocytes

2018 ◽  
Vol 68 (3) ◽  
pp. 279
Author(s):  
B. MACÍAS-GARCÍA ◽  
S. MACEDO ◽  
A. ROCHA ◽  
L. GONZÁLEZ-FERNÁNDEZ
Keyword(s):  
Bovine Serum ◽  
Culture Medium ◽  
Fetal Bovine Serum ◽  
Polar Body ◽  
Chromatin Conformation ◽  
Prolonged Incubation ◽  
Vitro Fertilization ◽  
Fetal Bovine ◽  
Bovine Oocytes

In vitro fertilization (IVF) in cattle is commonly used worldwide. Although extensive research has been conducted using different additives in the different IVF steps, little is known regarding how protein type may affect bovine oocytes during the fertilization period. In addition, unlike Tissue Culture Medium 199 (TCM), fertilization medium may induce oocytes’ chromatin degeneration during prolonged incubation in the horse (Modified Whitten’s medium). Thus, in the present work TCM-199 supplemented with either 7 mg/ml of Bovine Serum Albumin (TCM+BSA) or 10% Fetal Bovine Serum (v/v; TCM+FBS) was used. Bovine oocytes were matured in vitro and placed in the previously mentioned media for further 18 hours, in the absence of added sperm (sham fertilization) and their chromatin conformation was evaluated. After IVM, 78.9% of the initial oocytes had reached the MII stage. After sham fertilization, 58.6% of the oocytes in TCM+BSA while just 28.3% in TCM+FBS maintained the MII chromatin conformation (p < 0.05). Subsequent experiments run using PB extruded oocytes and incubated in TCM+BSA and TCM+FBS during sham fertilization, demonstrated that FBS was consistently associated with polar body dissolution or degeneration.

Growth and development of rabbit oocytes in vitro: Effect of fetal bovine serum concentration on culture medium

2012 ◽  
Vol 78 (5) ◽  
pp. 1040-1047 ◽  
Author(s):  
H. Sugimoto ◽  
Y. Kida ◽  
Y. Miyamoto ◽  
K. Kitada ◽  
K. Matsumoto ◽  
...  
Keyword(s):  
Serum Concentration ◽  
Bovine Serum ◽  
Growth And Development ◽  
Culture Medium ◽  
Fetal Bovine Serum ◽  
Fetal Bovine Serum Concentration ◽  
Fetal Bovine ◽  
Vitro Effect

286 INFLUENCE OF REVERSIBLE MEIOSIS INHIBITION ON SWINE EMBRYOS PRODUCED BY IN VITRO FERTILIZATION AND PATHENOGENETIC ACTIVATION

2006 ◽  
Vol 18 (2) ◽  
pp. 250
Author(s):  
M. G. Marques ◽  
A. B. Nascimento ◽  
V. P. Oliveira ◽  
A. R. S. Coutinho ◽  
M. E. O. A. Assumpção ◽  
...  
Keyword(s):  
Culture Medium ◽  
Cell Number ◽  
Control Group ◽  
Cleavage Rate ◽  
Vitro Fertilization ◽  
Electric Pulses ◽  
Cell Numbers ◽  
Blastocyst Rate ◽  
And Control

The present work evaluated the reversible meiosis inhibition effect on the development of swine embryos produced by in vitro fertilization (IVF) or parthenogenetic activation (PA). The efficiency of PZM3 and NCSU23 embryo culture media was also evaluated. Oocytes from ovaries collected at a slaughterhouse were subjected to IVM in two different groups: CHX (cycloheximide 5 µM for 10 h) and control, both with TCM-199 + 3.05 mM glucose + 0.91 mM sodium pyruvate + 10% porcine follicular fluid (pFF) + 0.57 mM cystein + 10 ng epidermal growth factor (EGF)/mL + 10 IU eCG/mL + 10 IU hCG/mL for the initial 22 h. In the remaining period (20 h for CHX and 22 h for control), medium without hormones was utilized. After IVM, oocytes were denuded and fertilized for 6 h (IFV) or the matured oocytes were submitted to activation by electric pulses (PA) (2 DC of 1.5 kV/cm for 30 µs), incubated for 1 h in culture medium with 10 μM of CHX, and again submitted to the same electric pulses for 60 µs. Embryo development was evaluated by cleavage rate on Day 3 and blastocyst rate and blastocyst cell number on Day 7 of culture. Cleavage and blastocyst rates were analyzed by the equality-of-two-ratios test and cell number by the Kruskal-Wallis and Mann-Whitney tests (P < 0.05). In relation to IVF, the PZM3 medium was more efficient than NCSU23 for cleavage rate in the CHX group (PZM3: 68.4%, NCSU23: 44.4%) and had a better blastocyst rate in the control group (PZM3: 13.4%, NCSU23: 5.6%). With reference to PA, NCSU23 presented better cleavage and blastocyst rates than PZM3 in the CHX group (NCSU23: 89.5%, PZM3: 78.5% and NCSU23: 20.4%, PZM3: 13.0%, respectively). In the control group, only the NCSU23 blastocyst rate was higher than that for PZM3 (NCSU23: 22.5%, PZM3: 10.8%). No culture medium effect on cell number mean of IVF and PA blastocysts was observed. Maturation block improved cleavage rates in IVF groups cultured with PZM3 (68.4% and 50.6%, respectively, for CHX and control) and in PA groups cultured with NCSU23 (89.5% and 80.3%, respectively, for CHX and control), but no improvement of blastocyst rates in both groups (IVF and PA) was verified. Table 1 below shows that maturation block decreased the IVF and increased the PA blastocyst cell numbers. As older oocytes are more effectively activated, oocytes blocked with CHX achieved the maturation stage faster than the control group, therefore resulting in high-quality PA blastocysts. In conclusion, PZM3 was more efficient for IVF embryo production in contrast to NCSU23, whereas NCSU23 can be indicated for PA embryo production. Moreover, maturation blockage with CHX influenced blastocyst cell number, decreasing in IVF embryos and increasing in PA embryos. Table 1. Mean (±SD) of blastocyst cell numbers for IVF or PA groups after in vitro maturation without (control) or with cycloheximide (CHX) and cultured in NCSU23 or PZM3 medium This work was supported by FAPESP 02/10747–1.

284 COMPARATIVE FERTILITY OF FRESHLY-COLLECTED VERSUS FROZEN - THAWED SPERMATOZOA FOR IN VITRO FERTILIZATION IN THE FISHING CAT (PRIONAILURUS VIVERRINUS)

2006 ◽  
Vol 18 (2) ◽  
pp. 249
Author(s):  
G. Magarey ◽  
J. Herrick ◽  
K. Thiangtum ◽  
W. Tunwattana ◽  
W. Swanson
Keyword(s):  
Sperm Motility ◽  
Culture Medium ◽  
Culture Media ◽  
Egg Yolk ◽  
Ovarian Follicles ◽  
Blastocyst Stage ◽  
Motile Sperm ◽  
Vitro Fertilization ◽  
International Transport

Wild populations of fishing cats (Prionailurus viverrinus) in Southeast Asia are in decline, primarily due to habitat loss. Because the fishing cat population in North American zoos is small (n = 69) and inbred (F = 0.17) with relatively low genetic variation (86%), infusion of new founder genes from Asia is a conservation priority. Importation of cryopreserved semen for use with IVF and ET may offer one alternative to the international transport of living animals. In this study, our objectives were to (1) compare motility longevity of fresh vs. frozen-thawed fishing cat spermatozoa in two culture media, (2) evaluate ovarian responses to exogenous gonadotropins, and (3) assess development of IVF embryos produced with fresh vs. frozen-thawed spermatozoa. Raw semen was collected via electroejaculation from male fishing cats (n = 4), divided into groups, and washed. Two sperm pellets were resuspended in either Ham's F10 medium (HF10; with 5% FBS) or our feline optimized culture medium (FOCM; with 0.4% BSA); another pellet was diluted in TEST egg yolk, cooled to 5�C over 3 h, glycerated (4%), and cryopreserved in straws over LN2 vapor. Frozen sperm samples were thawed, washed, and diluted in either HF10 or FOCM. Fresh and frozen-thawed sperm motility (percent motile, rate of forward progress) in each medium (10 � 106 motile sperm/mL) was assessed (at 0, 1, 3, and 6 h) in microdrops under oil during culture (38�C; 6% CO2 in air). Female fishing cats (n = 10) were treated with exogenous gonadotropins (150 IU eCG, 100 IU hCG, 85-h interval) and ovarian follicles were aspirated laparoscopically. Recovered oocytes were inseminated with fresh (2 � 105 motile sperm/mL) or frozen-thawed (5 � 105 motile sperm/mL) spermatozoa in FOCM microdrops; resulting embryos were either cryopreserved or cultured in FOCM (with 5% FBS added at 72 h post-insemination) for 7 days. Sperm motility over time did not differ (P > 0.05) between media for either fresh or frozen-thawed samples; however, across media, frozen-thawed sperm motility was lower (P < 0.05) and declined faster (P < 0.05) compared to fresh spermatozoa. Females produced an average (�SEM) of 9.8 � 2.9 mature ovarian follicles, allowing recovery of 7.3 � 2.6 high-quality oocytes per female. Oocyte cleavage percentage at 42 h p.i. was lower (P < 0.05) with frozen-thawed spermatozoa (38%, 11/29) compared to freshly collected spermatozoa (68%, 17/25). Overall, 35% (6/17) of cultured embryos developed to blastocysts with no difference (P > 0.05) between embryos produced with frozen-thawed (4/11) vs. fresh (2/6) spermatozoa. Although fishing cat sperm motility and fertility appear compromised after cryopreservation, our results demonstrate the ability of frozen-thawed spermatozoa to produce IVF embryos that are capable of developing to blastocyst stage in vitro. This work was supported by (NIH RR015388).

282 IMPROVED QUALITY OF PORCINE BLASTOCYSTS BY AGGREGATION OF EMBRYOS AT THE 4 - 8-CELL STAGE

2006 ◽  
Vol 18 (2) ◽  
pp. 248
Author(s):  
S.-G. Lee ◽  
C.-H. Park ◽  
D.-H. Choi ◽  
H.-Y. Son ◽  
C.-K. Lee
Keyword(s):  
Es Cells ◽  
Embryonic Stem ◽  
Transgenic Animals ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Cell Stage ◽  
Vitro Fertilization

Use of blastocysts produced in vitro would be an efficient way to generate embryonic stem (ES) cells for the production of transgenic animals and the study of developmental gene regulation. In pigs, the morphology and cell number of in vitro-produced blastocysts are inferior to these parameters in their in vivo counterparts. Therefore, establishment of ES cells from blastocysts produced in vitro might be hindered by poor embryo quality. The objective of this study was to increase the cell number of blastocysts derived by aggregating 4–8-cell stage porcine embryos produced in vitro. Cumulus–oocyte complexes were collected from prepubertal gilt ovaries, and matured in vitro. Embryos at the 4–8-cell stage were produced by culturing embryos for two days after in vitro fertilization (IVF). After removal of the zona pellucida with acid Tyrode’s solution, one (1X), two (2X), and three (3X) 4–8-cell stage embryos were aggregated by co-culturing them in aggregation plates followed by culturing to the blastocyst stage. After 7 days, the developmental ability and the number of cells in aggregated embryos were determined by staining with Hoechst 33342 and propidium iodide. The percentage of blastocysts was higher in both 2X and 3X aggregated embryos compared to that of 1X and that of intact controls (Table 1). The cell number of blastocysts also increased in aggregated embryos compared to that of non-aggregated (1X) embryos and controls. This result suggests that aggregation might improve the quality of in vitro-fertilized porcine blastocysts by increasing cell numbers, thus becoming a useful resource for isolation and establishment of porcine ES cells. Further studies are required to investigate the quality of the aggregated embryos in terms of increasing the pluripotent cell population by staining for Oct-4 and to apply improved aggregation methods in nuclear-transferred (NT) porcine embryos. Table 1. Development, cell number, and ICM ratio of aggregated porcine embryos

280 PORCINE BLASTOCYSTS DERIVED FROM IN VITRO-MATURED OOCYTES INJECTED INTRACYTOPLASMICALLY WITH SPERM FROM TESTICULAR TISSUE XENOGRAFTED INTO NUDE MICE

2006 ◽  
Vol 18 (2) ◽  
pp. 247 ◽  
Author(s):  
K. Kikuchi ◽  
M. Nakai ◽  
N. Kashiwazaki ◽  
M. Ozawa ◽  
N. Maedomari ◽  
...  
Keyword(s):  
Nude Mice ◽  
New Technology ◽  
Testicular Tissue ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Embryo Production ◽  
Vitro Fertilization ◽  
Male Gametes ◽  
Sperm Suspension

The utilization of spermatogonia from testicular tissue after xenografting into immuno-deficient mice should lead to new insights for the conservation of male gametes. However, successful embryo production using sperm cells from xenografted testicular tissues has been limited to rhesus monkeys (Honaramooz et al. 2004 Biol. Reprod. 70, 1500-1503). In the present study, the objective was to establish this new technology for pig conservation in combination with intracytoplasmic sperm injection. Testes were obtained from male piglets 6 to 15 days old, in which most of the germ cells were gonocytes; these were minced into pieces of approximately 1.5 � 1.5 � 1.5 mm. Approximately 20 fragments were transplanted under the back skin of castrated nude mice 5 to 8 weeks old. The testicular grafts were recovered between 125 and 192 days after xenografting, minced in Dulbecco's phosphate-buffered saline, and centrifuged several times, to serve as a sperm suspension. In vitro maturation of the recipient oocytes (Kikuchi et al. 2002 Biol. Reprod. 66, 1033-1041) and injection with an intact spermatozoon, followed by electrical stimulation at 1 h post-injection (Nakai et al. 2003 Biol. Reprod. 68, 1003-1008), were carried out. The putative zygotes were cultured in vitro for 6 days (Kikuchi et al. 2002), and were then fixed, stained, and assessed for embryonic development and quality. From a total of 27 mice that were xenografted with testicular tissues, spermatids and spermatozoa were obtained in 19 of the mice (70.4%). Most of the spermatozoa were matured morphologically, showing faint motility after release into the collection medium. From a total of 253 oocytes (four replications) that were injected with sperm, 63 (24.9 � 7.1%) oocytes developed to the blastocyst stage. The average total cell number was 41.9 � 3.9. These values are comparable to those in in vitro fertilization by frozen-thawed spermatozoa, resulting in developmental ability to piglets after embryo transfer (25.3% and 48.7 cells; Kikuchi et al. 2002). These results suggest the possibility of embryo production using porcine spermatozoa that are differentiated from gonocytes within the xenografts.

302 REPLACEMENT OF FETAL BOVINE SERUM WITH KNOCKOUT SERUM REPLACER AND STEM CELLS CONDITIONED MEDIUM DURING IN VITRO CULTURE OF BOVINE EMBRYOS

2010 ◽  
Vol 22 (1) ◽  
pp. 307
Author(s):  
M. M. Souza ◽  
N. Z. Saraiva ◽  
C. S. Oliveira ◽  
T. A. D. Tetzner-Nanzeri ◽  
R. Vantini ◽  
...  
Keyword(s):  
Stem Cells ◽  
In Vitro Culture ◽  
Bovine Serum ◽  
Fetal Bovine Serum ◽  
Protein Supplementation ◽  
Control Group ◽  
Bovine Embryos ◽  
Fetal Bovine

The use of fetal bovine serum (FBS) as protein supplementation in IVP of bovine embryos has presented difficulties because it can introduce a number of pathogenic components in culture systems, can be related to the birth of calf with abnormal growth and development, and precludes the establishment of the actual nutritional needs of the embryo, because it contains an unlimited variety of substances. This study evaluated the replacement of the FBS in the medium of in vitro culture (IVC) of bovine embryos, using the knockout serum replacer (KSR) as protein supplementation and culture medium conditioned with stem cells. Therefore, bovine oocytes from ovaries of slaughterhouse were selected and matured in vitro in TCM-199 medium supplemented with 10% FBS (Crypion), 1.0 μg mL-1 FSH (Pluset®, Calier, Barcelona, Spain), 50 μg mL-1 hCG (Profasi®, Serono, Geneva, Switzerland), 1.0 μg mL-1 estradiol (Sigma E-2758, Sigma Chemical, St. Louis, MO, USA), 0.2 mM sodium pyruvate, and 83.4 μg mL-1 amikacin for 24 h. After that, 1144 oocytes were fertilized in IVF-TALP medium containing 6 mg mL-1 of BSA. After 18 to 22 h, the zygotes were cultured in SOF + 5% FBS (group 2); SOF + 5% KSR (group 3); SOF (5% FBS) + 10% SOF (5% FBS) conditioned by stem cells (group 4); or SOF (5% KSR) + 10% SOF (5% KSR) conditioned by stem cells (group 5), in an atmosphere of 5% O2 at 38.5°C for 8 days. A control group outside the controlled atmosphere was added, supplemented with 5% FBS (group 1). The SOF medium supplemented with 5% FBS or KSR was conditioned by stem cells and added to SOF medium for the culture of embryo at a concentration of 10%. The rates of cleavage and production of blastocysts were assessed 48 hours and 7 days after IVF, respectively, and analyzed by chi-square test, with a significance level of 5% in the statistical program Minitab® (release 14.1, Minitab, State College, PA, USA). On the eighth day, the TUNEL test for determination of the percentage of apoptosis and the differential staining technique for determination of inner cell mass (ICM) and trophoblast (TF) were performed. The results were submitted to ANOVA, followed by comparing the means by Tukey’s test using the program GraphPad Prism (GraphPad, San Diego, CA, USA). The treatments did not differ in the production of embryos, being similar to the control group: G1 = 31.75% (74/233), G2 = 35.26% (79/224), G3 = 32.70% (74/226), G4 = 28.76% (63/219), and G5 = 26.85% (65/242). With regard to the assessment of embryonic quality, the treatments showed similar results to the control groups. No differences were observed among groups both in color and ICM/TF ratio (G1 = 0.60, G2 = 0.62, G3 =0.65, G4 = 0.60, and G5 = 0.60). Furthermore, the TUNEL showed no significant difference in the percentage of apoptosis among groups (G1 = 7.10%, G2 = 3.76%, G3 = 5.58%, G4 = 4.50%, and G5 = 4.11%). The data obtained so far indicate that it is possible to produce embryos in vitro by replacing the FBS in the culture, achieving results similar to those obtained with serum. Financial support: FAPESP 2007/58506-6.

135 EXPRESSION OF APOPTOSIS-RELATED GENES IN BUFFALO (BUBALUS BUBALIS) EMBRYOS PRODUCED THROUGH IN VITRO FERTILIZATION AND PARTHENOGENETIC ACTIVATION

2011 ◽  
Vol 23 (1) ◽  
pp. 172
Author(s):  
S. Saw ◽  
K. P. Singh ◽  
R. Kaushik ◽  
M. Muzaffar ◽  
M. S. Chauhan ◽  
...  
Keyword(s):  
Relative Abundance ◽  
Culture Medium ◽  
Mammalian Species ◽  
Control Cell ◽  
Bubalus Bubalis ◽  
Blastocyst Stage ◽  
Vitro Fertilization ◽  
Immature Oocytes ◽  
Tissue Size

Apoptosis, a highly conserved evolutionary mechanism that allows an organism to tightly control cell numbers, tissue size, and protect itself from dangerous cells and unfavourable environments that threaten homeostasis, is generally directed by specific genes involved in the regulation of a series of pro-apoptotic (BAX) and anti-apoptotic (BCL-XL) proteins that are expressed during early development. All mammalian species show the highest level of spontaneous apoptotic processes at the blastocyst stage. These proteins prevent apoptosis by maintaining the cell survival by interfering with the release of cytochrome-C from mitochondria. In this study, immature oocytes were obtained from buffalo slaughterhouse ovaries and were subjected to in vitro maturation (IVM) in TCM-199 + 10% FBS + 5 μg mL–1 porcine FSH for 24 h in a CO2 incubator (5% CO2, 90 to 95% relative humidity) at 38.5°C. The mature oocytes were used for IVF, and the cleaved embryos were cultured for 8 days in culture medium (CR2 medium containing 0.6% BSA and 10% FBS) for production of embryos at different stages. The parthenotes were produced with exposure of 7% ethanol, 6-dimethyl aminopurine and cultured for 8 days in culture medium. The total RNA was isolated from oocytes and embryos and transcribed using Cell-to-cDNA-II (Ambion, Austin, TX, USA), according to manufacturer protocol. The PCR cycle included heating to 94°C for 5 min, followed by 35 cycles of 94°C for 30 s, 60 (BAX) and 62°C (BCL) for 30 s, and 72°C for 45 s with a final extension at 72°C for 10 min. The amplified product of both genes were separated on agarose gel and densitometry data for band intensities were generated using AlphaDigiDocTM AD-1201 software under a WindowsTM environment and data analysed with the help of SYSTAT software. Relative abundance of BCL-XL transcripts in immature, mature oocytes and embryos produced through IVF (i.e. 2-cell, 4-cell, 8- to 16-cell, morula, and blastocyst stage) were 25.33 ± 0.90, 12.67 ± 1.20, 37.67 ± 0.90, 30.67 ± 0.30, 23.67 ± 0.90, 18.33 ± 0.90, and 27.00 ± 1.20, respectively, whereas in parthenogenesis these values were 23.67 ± 0.88, 13.67 ± 1.20, 23.67 ± 1.20, 22.34 ± 0.88, 24.34 ± 0.88, 33.67 ± 0.88, and 45.34 ± 1.20, respectively. Relative abundance of BAX transcripts by IVF were 23.0 ± 0.60, 0.33 ± 0.10, 4.00 ± 0.60, 5.00 ± 0.60, 0.37 ± 0.06, 13.0 ± 0.66, and 56.7 ± 0.90; and by parthenonenesis were 22.3 ± 0.90, 0.13 ± 0.03, 13.67 ± 0.90, 14.0 ± 0.60, 15.33 ± 0.90, 64.67 ± 2.20, and 55.0 ± 2.10, respectively. In conclusion, the expression pattern of the apoptosis-related genes revealed that the incidence of apoptosis was significantly higher in IVF and parthenogenetically produced buffalo embryos at stages such as immature oocytes, morula, and blastocyst than the early cleavage stage embryos.

Different in vitro culture systems affect the birth weight of lambs from vitrified ovine embryos

Zygote ◽  
2013 ◽  
Vol 23 (1) ◽  
pp. 53-57 ◽  
Author(s):  
L. Mara ◽  
D. Sanna ◽  
M. Dattena ◽  
I.M. Mayorga Muñoz
Keyword(s):  
Fatty Acid ◽  
Birth Weight ◽  
Bovine Serum ◽  
Blastocyst Stage ◽  
Test Body ◽  
Culture Systems ◽  
Significant Difference ◽  
Fetal Bovine ◽  
Lambing Rate

SummaryIt has been reported that different in vitro culture systems affect the birth weight of lambs. The aim of this study was to test body weight and lambing rate of lambs born from five different in vitro culture systems after vitrification. Oocytes of Sarda sheep were matured in TCM-199 plus 0.4% bovine serum albumin (BSA) using systems: (i) 4 mg/ml fatty acid-free BSA (BSA4); (ii) 8 mg/ml fatty acid-free BSA (BSA8); (iii) BSA8–hyaluronan (BSA8–HA); (iv) BSA8–charcoal-stripped FBS (BSA8–CH); or (v) with 10% fetal bovine serum (FBS; serum) and fertilized with fresh semen. The presumptive zygotes were cultured up to the blastocyst stage with BSA8, BSA8-HA, BSA8-CH or serum or BSA4. In the third and fifth days of culture 5% charcoal-stripped FBS was added into BSA8-CH and serum, while 8 mg/ml or 4 mg/ml fatty acid-free BSA was added as BSA8, BSA8-HA and BSA4 respectively; 6 mg/ml HA was added to BSA8-HA. In total, 240 vitrified blastocysts were transferred into synchronized ewes. The lambing rate was not significant different between BSA groups or between serum groups (BSA8-CH and serum), while serum groups showed significant lower values when compared with BSA groups. Only BSA8 groups produced heavy lambs (≥4.5 kg) with a significant difference between BSA4 and BSA8 groups (P < 0.05).

Oxygen concentration and protein source affect the development of preimplantation goat embryos in vitro

1991 ◽  
Vol 3 (5) ◽  
pp. 601 ◽  
Author(s):  
PA Batt ◽  
DK Gardner ◽  
AW Cameron
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Oxygen Concentration ◽  
Bovine Serum ◽  
Calf Serum ◽  
Cell Number ◽  
The Mean ◽  
Number Of Cells

The effect of oxygen concentration and the source of protein in culture medium on the development of 2- to 4-cell goat embryos in vitro was investigated. Embryos were collected from superovulated Angora-Cashmere-cross goats 48 h after ovulation and cultured for 6 days in synthetic oviduct fluid (SOF) medium under one of two oxygen concentrations (20% or 7%) and in the presence of one of five protein sources; Miles bovine serum albumin (Miles BSA), Commonwealth Serum Laboratory bovine serum albumin (CSL BSA), goat serum (GS), fetal calf serum (FCS) and human serum (HS). In the presence of 20% oxygen the percentage of embryos reaching the expanded and/or hatched blastocyst stage in SOF medium containing Miles BSA was 29%, with a mean cell number per embryo of 28.1 +/- 6.0 (+/- s.e.m.). Use of an oxygen concentration of 7% significantly increased the percentage of embryos reaching this stage (80%, P less than 0.01) and the mean number of cells per embryo (65.3 +/- 8.2, P less than 0.01). The mean number of cells of the early-cleavage-stage embryos was significantly lower when the medium contained CSL BSA, GS or FCS (42.7 +/- 5.6, 29.0 +/- 6.1 and 21.3 +/- 3.2, respectively) than with Miles BSA (92.8 +/- 6.4) or HS (104.8 +/- 17.2) (P less than 0.01). Under 7% oxygen and with Miles BSA or HS, embryos were morphologically comparable to those developed in vivo, but the mean cell numbers in vitro were only approximately half those obtained in vivo.

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