In Vitro maturation rate of oocytes collected from indigenous cows of Assam after slaughter

2015 ◽  
Vol 49 (6) ◽  
Author(s):  
S. S. Deka ◽  
D. J. Kalita ◽  
S. Sarma ◽  
D. J. Dutta
Keyword(s):  
Zona Pellucida ◽  
Maximum Yield ◽  
Cumulus Cells ◽  
Cumulus Expansion ◽  
Slaughter House ◽  
Oocyte Collection ◽  
Maturation Rate ◽  
The Mean ◽  
Ideal Method

Sixty healthy ovaries were collected from local slaughter house. Oocytes from small and medium sized follicles (2-8 mm in diameter) were selected for oocyte collection using aspiration followed by slicing . A and B category ( cumulus oocyte complexes (COCs) with more than 5 layers and 3-5 layers respectively of compact cumulus cells surrounding the zona pellucida) were selected for <italic>in vitro</italic> maturation. A total of 361 oocytes were found to be matured with overall mean maturation rate of 82.59 ± 0.02%. The mean number of grade A, B and C oocytes recovered per ovary was 4.26 ± 0.53, 3.05 ± 0.31 and 1.25 ± 0.19, respectively. The overall recovery of grade A, B and C oocytes were 256, 183 and 74 . Out of 439 oocytes 58.86 ± 0.05%, 33.23 ±0.04 % and 9.90 ± 0.04 % showed +++, ++ and + degrees of cumulus expansion, respectively. It was concluded that aspiration followed by slicing is an ideal method for maximum yield of oocytes and <italic>in vitro</italic> maturation in indigenous cow of Assam.

scholarly journals 46 Effect of the addition of α-tocopherol to in vitro maturation media of bovine oocytes

2020 ◽  
Vol 98 (Supplement_2) ◽  
pp. 2-3
Author(s):  
Theisy P Acosta Pérez
Keyword(s):  
In Vitro Maturation ◽  
Cumulus Cells ◽  
Control Group ◽  
Chi Square ◽  
Cumulus Expansion ◽  
Minimum Concentration ◽  
Maturation Rate ◽  
Chi Square Test ◽  
System Data

Abstract α-tocopherol is known to be a powerful antioxidant, in this regard, it was added to bovine oocyte in vitro maturation media to evaluate its effect on oocyte maturation. Oocytes (n = 624) aspirated from ovaries of slaughtered cows were classified by quality and divided in four categories according to cytoplasm appearance and cumulus cells layers. Oocytes were washed in TCM-199 supplemented with fetal bovine serum (FBS) and FSH, then distributed in maturation media (TCM-199 supplemented with FBS, FSH and gentamicin). Three experimental groups of α-tocopherol (50, 100 and 200 mM) and a control group without α-tocopherol were used. Maturation was carried 22 h at 38.5°C in a 5% CO2 atmosphere. Oocytes were examined to determine cumulus expansion as categorical data (expansion or no expansion), as well as cumulus expansion Index (CEI). For CEI determination oocytes were graded 0 to 4, being 0 those with null expansion and 4 those with a noticeable cell expansion, then the number of oocytes were multiplied by the grade given and a sum of the totals was obtained, the new total was divided by the total of oocytes in the group and the result obtained corresponded to the CEI of the group. Results were analyzed with Chi Square test (for maturation rates) and an ANOVA (for the CEI) using the SAS system, data are presented as mean ± standard error. There was no statistical difference between control and α-tocopherol groups (P &gt;0.05). Numerically, the control group showed a higher maturation rate (100%) and obtained a higher CEI (2.44±0.20), followed by the 50 mM group (98.16%; 2.39±0.13), the groups 200 mM (97.40%; 2.00±0.14) and 100 mM (96.25%; 2.06±0.24) were the lowest. The addition of the minimum concentration (50 mM) of α-tocopherol to the maturation media could improve maturation rates without exposing oocytes to toxic effects.

In vitro maturation of oocytes from non-stimulated common wombats

2003 ◽  
Vol 15 (5) ◽  
pp. 303 ◽  
Author(s):  
M. Cleary ◽  
M. West ◽  
J. Shaw ◽  
G. Jenkin ◽  
A. Trounson
Keyword(s):  
Oocyte Maturation ◽  
Polar Body ◽  
Cumulus Cells ◽  
In Vitro Fertilisation ◽  
Oocyte Collection ◽  
Common Wombat ◽  
Maturation Rate ◽  
Reproductive Techniques ◽  
The Common

Assisted reproductive techniques, such as in vitro oocyte maturation in conjunction with in vitro fertilisation, may be used as a tool to manipulate reproduction. Using the common wombat as a model for the critically endangered northern hairy-nosed wombat, the present study examined whether oocyte maturation could be achieved under field conditions. At the time of collection, no oocytes were at the metaphase II (MII) stage (0/42). After 60 h culture using the submarine incubation system, 34% of oocytes (24/70) matured to telophase/MII, as indicated by the presence of a polar body. The proportion of oocytes that reached MII was higher for oocytes collected from follicles >2 mm in diameter compared with follicles <2 mm (40% v. 22%, respectively). The presence of cumulus cells alone did not influence the maturation potential. Oocytes without cumulus cells collected from follicles >2 mm in diameter had the highest maturation rate (58%). Maturation was not affected by the reproductive status of the common wombat or a delay of up to 5 h before oocyte collection. In conclusion, the present study demonstrated that oocytes collected from non-stimulated common wombats can mature to MII in culture.

Effect of ornidazole on fertility of male rats: inhibition of a glycolysis-related motility pattern and zona binding required for fertilization in vitro

Reproduction ◽  
2000 ◽  
pp. 127-135 ◽  
Author(s):  
W Bone ◽  
NG Jones ◽  
G Kamp ◽  
CH Yeung ◽  
TG Cooper
Keyword(s):  
Zona Pellucida ◽  
Glucose Utilization ◽  
Cumulus Cells ◽  
Glycolytic Enzyme ◽  
Male Rats ◽  
Fertilization In Vitro ◽  
Swimming Pattern ◽  
Principal Piece ◽  
Free Media

The effects of the male antifertility agent ornidazole on glycolysis as a prerequisite for fertilization were investigated in rats. Antifertility doses of ornidazole inhibited glycolysis within mature spermatozoa as determined from the lack of glucose utilization, reduced acidosis under anaerobic conditions and reduced glycolytic enzyme activity. As a consequence, cauda epididymidal spermatozoa from ornidazole-fed rats were unable to fertilize rat oocytes in vitro, with or without cumulus cells, which was not due to transfer of an inhibitor in epididymal fluid with the spermatozoa. Under IVF conditions, binding to the zona pellucida was reduced in spermatozoa from ornidazole-fed males and the spermatozoa did not undergo a change in swimming pattern, which was observed in controls. The block to fertilization could be explained by the disruption of glycolysis-dependent events, since reduced binding to the zona pellucida and a lack of kinematic changes were demonstrated by control spermatozoa in glucose-free media in the presence of respiratory substrates. The importance of glycolysis for binding to, and penetration of, the zona pellucida, and hyperactivation in rats is discussed in relation to the glycolytic production of ATP in the principal piece in which local deprivation of energy may explain the reduced force of spermatozoa from ornidazole-fed males.

Cleavage Rate of in vitro Matured Oocytes Fertilized with Conventional Semen in Buffaloes

2021 ◽  
Author(s):  
M.H. Pitroda ◽  
K.P. Khillare ◽  
M.B. Amle ◽  
M.D. Meshram ◽  
A.B. Mali ◽  
...  
Keyword(s):  
Fertilization Rate ◽  
Cleavage Rate ◽  
Embryo Production ◽  
Quick Recovery ◽  
Slaughter House ◽  
Maturation Rate ◽  
Aspiration Technique ◽  
Current Scenario ◽  
In Vitro Embryo Production

Background: In vitro embryo production in buffaloes has gained much importance in this current scenario due to ever increasing population and high demand of milk and meat. Slaughter house derived bubaline ovaries are a cheap and abundant source of cumulus oocyte complexes.Methods: Oocytes from the buffalo ovarian follicles were recovered by aspiration technique as it facilitates quick recovery. Total 155 ovaries were used in the present study. Surface follicles were measured using vernier calliper and categorized into three groups viz. less than 3 mm, 3-5 mm and greater than 5 mm based on follicular diameter and oocytes were processed for IVM, IVF and IVC using conventional non sorted semen.Result: Overall percentage of small, medium and large follicles in the ovaries were recorded as 16.29 ± 0.94%, 8.14±0.60%, 5.35 ± 0.76%, respectively. Overall recovery rate of COCs was 38%. The percentage of these oocytes were 16.74% (A), 15.25% (B), 25.26% (C), 18.33% (D) and 29.87% (E) respectively. Maturation rate of oocytes were 81.96 ± 2.70%. Fertilization rate was 74.98 ± 3.87%, Cleavage rate % was 40.84±2.51% and Blastocyst percentage was 21.57±1.75% respectively. Application of in vitro embryo production technique using slaughter house ovaries can salvage the genetic potential of bubaline species.

scholarly journals GDF-9 and BMP-15 mRNA Levels in Canine Cumulus Cells Related to Cumulus Expansion and the Maturation Process

Animals ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 462 ◽  
Author(s):  
George Ramirez ◽  
Jaime Palomino ◽  
Karla Aspee ◽  
Monica De los Reyes
Keyword(s):  
Gene Expression ◽  
Estrous Cycle ◽  
Expression Patterns ◽  
Cumulus Cells ◽  
Mrna Levels ◽  
Cumulus Expansion ◽  
Polymerase Chain ◽  
Specific Regulation ◽  
Reproductive Phases

The competence to undergo expansion is a characteristic of cumulus cells (CCs). The aim was to investigate the expression of GDF-9 and BMP-15 mRNA in canine cumulus cells in relation to cumulus expansion and meiotic development over the estrous cycle. CCs were recovered from nonmatured and in vitro-matured (IVM) dog cumulus oocyte complexes (COCs), which were obtained from antral follicles at different phases of the estrous cycle. Quantitative real-time polymerase chain reaction (q-PCR) was used to evaluate the relative abundance of GDF-9 and BMP-15 transcripts from the CCs with or without signs of expansion. The results were evaluated by ANOVA and logistic regression. The maturity of the oocyte and the expansion process affected the mRNA levels in CCs. There were differences (p < 0.05) in GDF-9 and BMP-15 gene expression in CCs isolated from nonmatured COCs when comparing the reproductive phases. Lower mRNA levels (p < 0.05) were observed in anestrus and proestrus in comparison to those in estrus and diestrus. In contrast, when comparing GDF-9 mRNA levels in IVM COCs, no differences were found among the phases of the estrous cycle in expanded and nonexpanded CCs (p < 0.05). However, the highest (p < 0.05) BMP-15 gene expression in CCs that did not undergo expansion was exhibited in anestrus and the lowest (p < 0.05) expression was observed in estrus in expanded CCs. Although the stage of the estrous cycle did not affect the second metaphase (MII )rates, the expanded CCs obtained at estrus coexisted with higher percentages of MII (p < 0.05). In conclusion, the differential expression patterns of GDF-9 and BMP-15 mRNA transcripts might be related to cumulus expansion and maturation processes, suggesting specific regulation and temporal changes in their expression.

scholarly journals Prediction of maturational competence of feline oocytes using supravital staining of cumulus cells by propidium iodide

Zygote ◽  
2011 ◽  
Vol 20 (4) ◽  
pp. 333-337 ◽  
Author(s):  
Kenzo Uchikura ◽  
Masashi Nagano ◽  
Mitsugu Hishinuma
Keyword(s):  
Propidium Iodide ◽  
Cumulus Cell ◽  
Membrane Integrity ◽  
Cumulus Cells ◽  
Nuclear Maturation ◽  
Non Invasive ◽  
Maturation Rate ◽  
Cell Layers ◽  
The Relationship

SummaryWe examined the relationship between integrity of cumulus cells and nuclear maturation rate after in vitro culture to determine a non-invasive prediction of the maturational competence of feline oocytes. Feline cumulus–oocyte complexes (COCs) were collected from either small (400–800 μm) or large (≥800 μm) follicles. Immediately after collection, cumulus cells were evaluated morphologically (thickness of cumulus cell layers) and stained with propidium iodide (PI), which penetrates only non-viable cells. Cumulus cells without PI staining were judged as having good membrane integrity. After evaluation, COCs were cultured for 30 h and their nuclear maturation rate was determined. The nuclear maturation rate of oocytes derived from large follicles (89.8%) was higher (p < 0.05) than that from small follicles (60.8%). There was no difference in the maturation rate of oocytes from follicles with the same size regardless of cumulus morphology. In contrast, oocytes that had cumulus cells with good membrane integrity showed a higher maturation rate (93.8%) than oocytes with poor cumulus integrity (76.9%) in large follicles (p < 0.05). We conclude that evaluation of membrane integrity of cumulus cells by propidium iodide staining can be used to predict the maturational competence of oocytes.

221 EFFECT OF STAGE OF CORPUS LUTEUM DEVELOPMENT ON THE IN VITRO PRODUCTION OF BOVINE EMBRYOS

2011 ◽  
Vol 23 (1) ◽  
pp. 209
Author(s):  
S. Miyashita ◽  
K. Miyata ◽  
C. Tachibana ◽  
Y. Inaba ◽  
H. Koyama ◽  
...  
Keyword(s):  
Corpus Luteum ◽  
Calf Serum ◽  
Cumulus Cells ◽  
Large Mass ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Stage Of Development ◽  
The Mean ◽  
Vitro Production

The objective of this study was to investigate the effect of stage of corpus luteum (CL) development on the in vitro production of bovine embryos. Ovaries were classified according to the expected day of the oestrous cycle based on the morphology of the ovaries. Ovaries with a corpus hemorrhagicum and the remnant of the follicular lumen filled with blood were considered the early luteal stage (Days 2 to 4; Day 0 = day of ovulation, n = 46). Ovaries with a large mass of orange tissue in the CL were classified as the midluteal stage (Days 7 to 10, n = 42). Cumulus–oocyte complexes (COC) were collected by aspiration of 2- to 6-mm follicles. The COC were classified into the following grades: COC with >3 compact layers of cumulus cells and evenly granulated cytoplasm were classified into Grade 1; COC with >3 layers cumulus cells and evenly granulated cytoplasm were classified into Grade 2; COC with partially remaining cumulus cells and abnormal cytoplasm were classified into Grade 3; COC without cumulus cells or those with expanded cumulus cells were classified into Grades 4 and 5, respectively. Grades 1 and 2 COC were in vitro matured for 20 h in TCM-199 supplemented with 5% calf serum and 0.02 mg mL–1 of FSH at 38.5°C under an atmosphere of 5% CO2 in air. Matured COC were inseminated with 5 × 106 sperm for 18 h. Presumptive zygotes were cultured in CR1aa medium supplemented with 5% calf serum at 38.5°C under an atmosphere of 5% O2, 5% CO2, and 90% N2 for 9 days (fertilization = Day 0). The mean number of COC and the proportion of COC classified as Grades 1 and 2 were analysed by ANOVA. Cleavage rates on Day 3 and blastocyst rates on Days 7 to 9 were analysed by a chi-square test. The mean number of recovered oocytes in the early luteal stage (18.7 ± 9.5) was significantly higher (P < 0.05) than the number in the midluteal stage (12.2 ± 5.7). The proportion of Grades 1 and 2 oocytes in the early luteal stage [66.7% (531/789)] was significantly higher (P < 0.01) than that in the midluteal stage [51.6% (252/484)]. The cleavage and blastocyst rates in the early luteal stage [60.9% (181/297) and 32.7% (97/297), respectively] were significantly higher (P < 0.05) than those in the midluteal stage [50.7% (76/150) and 20.7% (31/150) respectively].The present study suggests that the stage of development of the CL in bovine ovaries influences the number of recovered oocytes per ovary and the development of in vitro production of bovine embryos.

332 GRANULOCYTE-MACROPHAGE COLONY STIMULATING FACTOR (GM-CSF) ENHANCES CUMULUS CELLS EXPANSION OF IN VITRO-MATURED BOVINE CUMULUS - OOCYTE COMPLEXES IN A CHEMICALLY DEFINED MEDIUM

2010 ◽  
Vol 22 (1) ◽  
pp. 322
Author(s):  
D. D. Bücher ◽  
M. A. Castro ◽  
M. E. Silva ◽  
M. A. Berland ◽  
I. I. Concha ◽  
...  
Keyword(s):  
Cell Viability ◽  
In Vitro Maturation ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Cell Number ◽  
Control Group ◽  
Cumulus Expansion ◽  
Gm Csf ◽  
Nuclear Stage

Granulocyte-macrophage colony stimulating factor (GM-CSF) is a pleiotropic cytokine that stimulates proliferation, differentiation and function in different cells types. We have previously demonstrated (Bücher DD et al. 2008 Reprod. Dom. Anim. 43 (Suppl. 3), 146 abst.) that both subunits of GM-CSF receptor are expressed in granulosa cells from antral follicles in bovine ovaries. Also, we determined that the cytokine enhances glucose uptake through facilitative hexose transporters in granulosa cells in primary culture. The goals of the present study were to characterize the expression of GM-CSF receptor in cumulus cells and oocytes from bovine antral follicles and to determine its effects on in vitro-matured bovine COCs in a chemically defined medium. To determine the presence of a and |5 subunits of GM-CSF receptor, COCs were aspirated from follicles <8 mm in diameter, fixed, and submitted to immunocytochemistry. To study the effect of GM-CSF on in vitro maturation of oocytes, COCs (n =481) were cultured using serum-free medium (SOF) containing 0, 1, 10, and 100 ng mL-1 of human recombinant GM-CSF (R&D Systems, Inc., Minneapolis, MN, USA) for 22 h at 39°C, 5% CO2 in humidified air. Nuclear stage, cumulus expansion, cumulus cell number, and viability were analyzed after in vitro maturation. Cumulus expansion was assessed using the cumulus expansion index (CEI) (Fagbohun C and Down S 1990 Biol. Reprod. 42, 413-423). Nuclear stage was evaluated using aceto-orcein stain. To determine cumulus cell viability and number, COCs (n = 10-12 per group) were transferred into an Eppendorf tube and cumulus cells were removed by vortexing for 3 min, stained with trypan blue and counted with a hemocytometer. The study was conducted in 6 replicates. Data from cumulus expansion and cell number were analyzed by Kruskal-Wallis analysis. Data for nuclear stage and cell viability were analyzed by chi-square analysis and one way ANOVA, respectively. Both receptor subunits were present in cumulus cells and oocytes from COCs. COCs cultured in 10 and 100 ng mL-1 GM-CSF had CEI scores (0.8 and 1.22, respectively) greater (P < 0.01) than controls (0.2), but the proportion of COCs displaying second metaphase did not differ (P = 0.5) among treatment groups. GM-CSF at a concentration of 100 ng mL-1 increased (P < 0.01) cumulus cell viability by more than 20% compared to the control group. Similarly, GM-CSF at concentrations of 10 and 100 ng mL-1 increased (P < 0.05) cumulus cell number by more than 20% and 45%, respectively, from the control group. The use of a specific inhibitor of PI3 kinase (Ly294002; 10 and 100 μM) blocked the stimulatory effect of GM-CSF on cumulus expansion, cell viability, and cell number. In conclusion, the results of the study suggest a plausible modulator role of GM-CSF in the metabolism and function of cumulus cells and oocytes during in vitro maturation. Funding from Faculty of Veterinary Sciences, Universidad Austral de Chile, MECESUP AUS-0005, AUS-0601, and DID D-2006-24 and from Universidad Católica de Temuco, research grant 2007 DGI-CDA-04.

172 DETERMINING THE REQUIREMENTS FOR LH AND FSH DURING SHEEP IN VITRO OOCYTE MATURATION

2014 ◽  
Vol 26 (1) ◽  
pp. 200 ◽  
Author(s):  
C. de Frutos ◽  
R. Vicente-Perez ◽  
P. J. Ross
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Mixed Logit ◽  
Cumulus Cells ◽  
Pituitary Hormones ◽  
Developmental Competence ◽  
Cumulus Expansion ◽  
Nuclear Maturation ◽  
Grade 3

In vitro maturation (IVM) of oocytes in domestic animals is a widespread practice of research and commercial relevance. Gonadotropic hormones are typically supplemented to the IVM medium to stimulate resumption of meiosis, progression to metaphase II (MII), and oocyte developmental competence. The common use of pituitary-derived products presents 2 problems: contamination from other pituitary hormones and inconsistences from batch-to-batch variation. Recombinant hormones can help circumvent these issues and identify specific gonadotropin requirements for in vitro maturation. The aim of the present study was to determine the effect of supplementing recombinant bovine LH and/or FSH (AspenBio) to the maturation of ovine oocytes in terms of cumulus expansion and progression to the MII stage. Abattoir-derived sheep cumulus–oocyte complexes (COC) were obtained from 1- to 5-mm-diameter antral follicles by ovary slicing. Oocytes with a homogeneous cytoplasm surrounded by at least 3 layers of cumulus cells were selected and cultured in serum-free IVM medium (Cotterill et al. 2012 Reproduction 144, 195–207) at 38.5°C and 5% CO2. The COC obtained from 8 replicates were allocated into 4 experimental groups: (1) no hormones; (2) 1.5 μg mL–1 recombinant bovine LH (rbLH); (3) 1.5 μg mL–1 recombinant bovine FSH (rbFSH); and (4) rbLH and rbFSH. The expansion of cumulus cells was recorded in each group after 24 h of IVM and COC classified as (1) very poor or no cumulus expansion (grade 1); (2) limited cumulus expansion (grade 2); and (3) full cumulus expansion (grade 3). Nuclear maturation in the 4 treatments was evaluated by assessing progression to the MII stage via DNA staining with Hoechst 33342 and fluorescence imaging. The effect of treatment on the observed proportion of MII oocytes was evaluated using a mixed logit model including treatment and replicate as fixed and random effects, respectively. Culture in IVM medium in the absence of gonadotropins or in the presence of rbLH resulted in poor cumulus expansion (grade 1). The supplementation of IVM medium with rbFSH (with or without rbLH) yielded a high degree of cumulus expansion (grades 2–3). Likewise, addition of rbFSH enhanced progression of oocytes to the MII stage, whereas use of rbLH, although it had an effect on progression to MII, did not augment the effect of rbFSH (Table 1). These results indicate that rbFSH is necessary and sufficient to induce sheep oocyte maturation in a high proportion of oocytes. Table 1.Cumulus expansion and oocyte nuclear stage after IVM

159 EFFECT OF 3-ISOBUTYL-1-METHYLXANTHINE (IBMX) ON THE GERMINAL VESICLE STAGE OF PORCINE OOCYTES DURING OOCYTE COLLECTION IN VITRO

2014 ◽  
Vol 26 (1) ◽  
pp. 193
Author(s):  
R. Appeltant ◽  
J. Beek ◽  
D. Maes ◽  
A. Van Soom
Keyword(s):  
Germinal Vesicle ◽  
Cyclic Adenosine Monophosphate ◽  
Adenosine Monophosphate ◽  
Cumulus Cells ◽  
Guanosine Monophosphate ◽  
Developmental Competence ◽  
Meiotic Arrest ◽  
Oocyte Collection

When using modern maturation conditions for in vitro maturation, pig oocytes yield ~20% blastocysts only. One problem is that cumulus cells, which are normally connected with the immature oocyte by cellular projections penetrating through the zona pellucida and with the oolemma via gap junctions, are prematurely losing these connections after the cumulus–oocyte complex is removed from the follicle. The oocyte possesses a type 3 phosphodiesterase, which degrades 3′,5′-cyclic adenosine monophosphate (cAMP), and this activity is inhibited by supply of 3′,5′-cyclic guanosine monophosphate (cGMP) to the oocyte via the cumulus cells. Consequently, cAMP levels, which are typically high during early stages of oocyte maturation in vivo, decrease, leading to spontaneous nuclear maturation and oocytes of low developmental competence. Therefore, the maintenance of these cumulus-oocyte connections is important to keep cAMP high and the oocyte under meiotic arrest. One way to prevent this drop in cAMP is using N6, 2′-o-dibutyryladenosine 3′,5′-cyclic monophosphate sodium (dbcAMP) that causes an arrest at germinal vesicle (GV) stage II (Funahashi et al. 1997 Biol. Reprod. 57, 49–53). Another option is collecting the oocytes in a medium containing the phoshodiesterase inhibitor, IBMX. The present study investigated the influence of IBMX on the progression of the GV of the oocyte after collection, just before the start of the maturation procedure. The GV stage was defined according to Sun et al. (2004 Mol. Reprod. Dev. 69, 228–234). In parallel with the findings on dbcAMP, we hypothesised an arrest at GV II by the presence of IBMX during collection. One group of oocytes were collected in HEPES-buffered TALP without IBMX (n = 375) and another group in the same medium containing 0.5 mM IBMX (n = 586). An average incubation time of 140 min was applied in both groups, and 3 replicates were performed. The proportions of oocytes before or at GV II and beyond GV II were compared in both groups using logistic regression analysis. The proportion of oocytes was included as dependent variable and group (IBMX addition or not) as independent variable. Replicate was also included in the model. The proportion of oocytes before or at GV II was not statistically significant between the group without and the group with IBMX (59.2 v. 58.7% respectively; P > 0.05). In conclusion, the use of IBMX during oocyte collection did not influence the state of the germinal vesicle of the oocyte during collection, indicating that IBMX did not cause a meiotic arrest in the oocytes during collecting in vitro.

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