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Balli, M.;  Cecchele, A.;  Pisaturo, V.;  Makieva, S.;  Carullo, G.;  Somigliana, E.;  Paffoni, A.;  Vigano’, P. Opportunities and Limits of Conventional IVF. Encyclopedia. Available online: https://encyclopedia.pub/entry/28743 (accessed on 05 December 2025).
Balli M,  Cecchele A,  Pisaturo V,  Makieva S,  Carullo G,  Somigliana E, et al. Opportunities and Limits of Conventional IVF. Encyclopedia. Available at: https://encyclopedia.pub/entry/28743. Accessed December 05, 2025.
Balli, Martina, Anna Cecchele, Valerio Pisaturo, Sofia Makieva, Giorgia Carullo, Edgardo Somigliana, Alessio Paffoni, Paola Vigano’. "Opportunities and Limits of Conventional IVF" Encyclopedia, https://encyclopedia.pub/entry/28743 (accessed December 05, 2025).
Balli, M.,  Cecchele, A.,  Pisaturo, V.,  Makieva, S.,  Carullo, G.,  Somigliana, E.,  Paffoni, A., & Vigano’, P. (2022, October 10). Opportunities and Limits of Conventional IVF. In Encyclopedia. https://encyclopedia.pub/entry/28743
Balli, Martina, et al. "Opportunities and Limits of Conventional IVF." Encyclopedia. Web. 10 October, 2022.
Opportunities and Limits of Conventional IVF
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This entry is adapted from the peer-reviewed paper 10.3390/jcm11195722

Conventional IVF (c-IVF) is one of the most practiced assisted reproductive technology (ART) approaches used worldwide. However, in the last years, the number of c-IVF procedures has dropped dramatically in favor of intracytoplasmic sperm injection (ICSI) in cases of non-male-related infertility. The advantages and disadvantages associated with c-IVF, highlighting the essential steps governing its success, its limitations, the methodology differences among laboratories and the technical progress were outlined. A framework for a better understanding of opportunities associated with human c-IVF and for best practice guidelines applicability in the reproductive medicine field was introduced.

infertility IVF reproduction

1. Timing of Insemination from Oocyte Retrieval

The debate regarding the effect of the timing of insemination after oocyte retrieval started in the 1980s, when three different groups demonstrated better fertilization rates and higher embryo quality when oocytes were inseminated after a pre-incubation period of 3–5.5 h after retrieval [1][2][3][4]. In the same years, the group of Fish and coworkers showed that different pre-insemination intervals (within 9 h after oocyte retrieval) did not affect fertilization or pregnancy rates [4]. More recent studies focused on the impact of the pre-incubation period on the fertilization rate were similarly not concordant. Ho et al. reported significantly better results by performing the insemination in a time window ranging from 2.5 to 5.5 h after oocytes retrieval. Specifically, they analyzed the fertilization rate at <2.5 h, <3.5 h, <4.5 h, ≤5.5 h and >5.5 h after oocyte retrieval and observed fertilization rates of 67.9%, 80.5%, 82.0%, 84.5% and 73.0%, respectively. They obtained statistically significant data when the insemination was performed <2.5 h and 5.5 h after oocyte retrieval (p < 0.001) [5]. In contrast, Jacobs and colleagues demonstrated different results analyzing the fertilization rate, embryo quality, implantation rate, abortion and ongoing pregnancy when inseminations were performed 1–7 h after oocyte retrieval. No statistically significant differences in c-IVF outcomes performed in different time intervals were observed, suggesting that early insemination could be performed without reservation [6]. Esiso and colleagues divided the time interval between oocyte retrieval and insemination into eight categories: 0 (0–<0.5 h), 1 (0.5–<1.5 h), 2 (1.5–<2.5 h), 3 (2.5–<3.5 h), 4 (3.5–<4.5 h), 5 (4.5–<5.5 h), 6 (5.5–<6.5 h) and 7 (6.5–<8 h). The relative number of oocytes retrieved in each category was n = 586, n = 1594, n = 1644, n = 1796, n = 1836, n = 1351, n = 641 and n = 127, respectively. Considering only the c-IVF outcome, they had optimal results when the insemination was performed between 1.5 h and 6 h after oocyte retrieval. When performed prior to 1.5 h, the detrimental effects of insemination on the fertilization rate were only moderate, without affecting blastulation and pregnancy rates [7]. Overall, these studies demonstrate the existence of an optimum time range for more successful c-IVF that should be performed between 3 and 6 h after oocyte retrieval. Of note, the time elapsed from ovulation trigger and oocyte retrieval (generally 34–36 h) should be taken into account when evaluating the proper timing of insemination.

2. Timing of Sperm-Cumulus Co-Incubation

The standard human c-IVF method involves overnight insemination of cumulus-intact oocytes with a defined range of spermatozoa/mL, followed by a fertilization assessment in the next morning [8]. It has been, however, suggested that extended gametes co-incubation may lead to unsuccessful c-IVF due to the production of high concentrations of reactive oxygen species (ROS), potentially deleterious for oocyte and embryo quality [9][10][11][12]. Therefore, to prevent unfavorable effects on oocytes and related embryos from high ROS exposure, a shorter incubation of gametes has been proposed. Studies addressing the comparison between short gametes co-incubation to standard overnight IVF (16–18 h) are listed in Table 1 [13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32]. Yet in 1996, Gianaroli et al. reported that a short gamete co-incubation (1 h) led to an increased rate of oocyte fertilization and embryo viability, suggesting that prolonged exposure of oocytes to elevated concentrations of sperm cells may negatively influence early embryo development [13]. In accordance, Le Bras and colleagues demonstrated that short gamete co-incubation period (2 h) leads to higher embryo quality, with a percentage of fragmentation lower than 25%, and, most importantly, to a significant increase in clinical pregnancy after a fresh embryo transfer compared to standard overnight insemination (16–18 h) [14]. Other studies reported that a shorter oocyte-spermatozoa incubation time is associated with an enhanced embryo quality and could prevent total fertilization failure (TTF) [15][16]. Conversely, in a prospective study, Barraud-Lange et al. evaluated the effects of a short gamete co-incubation (1 h) on fertilization rate and embryo quality of sibling oocytes, showing a decreased fertilization rate and comparable embryo quality compared to the standard overnight insemination method [17]. The meta-analysis from Zhang et al., published in 2013, revealed that reduced gamete co-incubation time is associated with beneficial outcomes, including significantly increased clinical pregnancy rate (Pooled Risk Ratio [RR]: 1.84, 95% confidence interval (CI) 1.24–2.73), ongoing pregnancy rate (RR: 1.73, 95% CI: 1.27–2.33) and implantation rate (RR: 1.80, 95% CI: 1.43–2.26) compared to c-IVF, with no significant differences in fertilization rates, embryo quality and polyspermy rate (RR: 0.98, 95% CI: 0.93–1.02; RR: 1.24, 95% CI: 1.0–1.53; RR: 0.84, 95% CI: 0.7–1.01, respectively) [18]. A recent randomized study of n= 320 infertile women, evaluated the beneficial influence of short gamete co-incubation in terms of live birth rate. Here, oocytes of patients randomized to the short co-incubation period group (3–4 h, n = 160) and to the conventional co-incubation timing (20 h, n = 160) were inseminated with ~20,000–30,000 motile spermatozoa/oocyte. Contrarily to other studies, no statistically significant difference in terms of live birth rate, clinical pregnancy, miscarriage and implantation rates was reported between the short-time and standard overnight insemination groups [19]. Overall, evidence on which is the best timing of sperm-egg co-incubation in c-IVF requires more consolidation.
Table 1. Results deriving from the comparison between different incubation intervals of spermatozoa and oocytes in in vitro fertilization procedure.
Authors, Year Interval of
Co-Incubation		 Fertilization Rate (FR) in Short IVF Clinical Pregnancy Rate (CPR) in Short IVF Implantation Rate (IP) in Short IVF
Gianaroli et al., 1996 [13] 1 h vs. overnight Improved Improved Improved
Quinn et al., 1998 [20] 1 h vs. overnight Unchanged Improved Improved
Coskun et al., 1998 [21] 1 h vs. overnight Unchanged Not assessed Not assessed
Dirnfeld et al., 1999 [22] 1 h vs. overnight Unchanged Improved Improved
Lin et al., 2000 [23] 1–3 h vs. overnight Unchanged Not assessed Not assessed
Swenson et al., 2000 [24] 2 h vs. overnight Not assessed Worsened Unchanged
Boone et al., 2001 [25] 3 h vs. overnight Worsened Not assessed Not assessed
Lundqvist et al., 2001 [26] 2 h vs. overnight Worsened Unchanged Unchanged
Dirnfeld et al., 2003 [12] 1 h vs. overnight Unchanged Not assessed Not assessed
Kattera et al., 2003 [27] 2 h vs. overnight Unchanged Improved Improved
Barraud-Lange et al., 2008 [28] 1 h vs. overnight Worsened Not assessed Not assessed
Xiong et al., 2009 [17] 1–6 h vs. overnight Unchanged Unchanged Not assessed
Dai et al., 2012 [29] 1–4 h vs. overnight Unchanged Unchanged Unchanged
Huang et al., 2013 [30] 1–4 h vs. overnight Not assessed Improved Not assessed
Zhang et al., 2013 [31] 1–6 h vs. overnight Unchanged Improved Improved
Li et al., 2016 [10] 2 h vs. overnight Unchanged Improved Improved
Le Bras et al., 2017 [18] 2 h vs. overnight Worsened Improved Improved
He et al., 2018 [32] 4/6 h vs. overnight Worsened Unchanged Unchanged
Chen et al., 2019 [14] 3/4 h vs. overnight Unchanged Unchanged Unchanged
Kong et al., 2021 [15] 4 h vs. overnight Unchanged Unchanged Not assessed

3. Oxygen Tension

Embryo development depends on a variety of factors, and among others, oxygen tension represents one of the key decisive parameters to ensure proper embryogenesis [19]. Mammalian development is characterized by low concentration of oxygen in both fallopian tubes and uterus, ranging from 5 to 7% in the former and decreasing to 2% in the latter [16]. Therefore, in order to support proper embryo development, it may be crucial to mimic the low level of oxygen tension (5%) present in the in vivo developmental environment. Interestingly, several studies reported that successful human embryo culture, in terms of embryo quality and blastulation rate, occurs under hypoxic conditions (5% oxygen tension). Moreover, the rate of implantation, pregnancy, good-quality embryos for transfer and live birth significantly increases under hypoxia culture conditions rather than with atmospheric oxygen levels [33][34][35][36][37]. The oxidative stress resulting from high oxygen concentration culture conditions may severely affect embryo quality, decreasing its implantation potential and, as a consequence, the pregnancy rate [34][36][38]. Moreover, embryos exposed to ROS are prone to DNA damage and mitochondrial alterations, leading to activation of the apoptotic mechanism [39]. Data obtained from a meta-analysis showed a notable improvement on live birth rate following embryo culture in low oxygen concentration. The rate of live births was improved up to 43% by lowering the oxygen concentration during embryo development [40]. Accordingly, recommendations provided from the latest ESHRE guidelines suggest the use of low oxygen concentration for embryo culture [41]. Nonetheless, the oxygen tension used in the culture system has ample differences among ART laboratories worldwide [42]. Notably, the impact of oxygen tension on results of ICSI versus c-IVF in relation to stage-specific sensitivity is unclear. Most of the papers on this topic do not divide the results according to the two strategies. In a prospective randomized sibling-oocyte study, Guo et al. evaluated the impact of different oxygen concentrations (20% versus 5%) on fertilization rates in c-IVF cycles. A total of n = 1254 oocytes were randomly assigned to 20% or 5% oxygen tension culture conditions on the retrieval day and then treated with c-IVF. The two groups did not show differences in fertilization rate, suggesting that at this stage of development, different oxygen tensions may not influence the process of fertilization. However, oocytes cultured at 5% oxygen gave rise to an increased number of optimal embryos on day 3 (72.4% vs. 64.2%, respectively, p = 0.018) and higher blastocyst formation rate (64.5% vs. 52.9%, respectively, p = 0.009) compared to the 20% group. Moreover, the use of low oxygen tension resulted in a more favorable clinical pregnancy and implantation rates compared with atmospheric oxygen [43]. In a total of n = 402 ART cycles, Guarneri et al. specifically evaluated the use of two different oxygen concentrations (atmospheric versus low oxygen) during oocyte culture from recovery until decumulation on day 1, followed by use of low oxygen concentration (5%) until transfer. Interestingly, cumulus-intact oocytes cultured in atmospheric oxygen tension for ~20 h for c-IVF resulted in a comparable number of transferred/vitrified embryos from the inseminated oocytes, cumulative clinical pregnancy rate and cumulative live birth rate per cycle compared to the oocytes cultured under 5% oxygen level from gamete retrieval to embryo transfer [44]. Although evidence-based data strongly indicate that culturing embryos in low oxygen concentration improves embryo utilization rate and increases the chance of pregnancy [41][45], the potential antioxidant activity of the cumulus cells present during the first step of c-IVF needs to be further investigated.

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Contributors MDPI registered users' name will be linked to their SciProfiles pages. To register with us, please refer to https://encyclopedia.pub/register :
Martina Balli
, Anna Cecchele ,
Valerio Pisaturo
,
Sofia Makieva
,
Giorgia Carullo
, Edgardo Somigliana , Alessio Paffoni , Paola Vigano’
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Update Date: 11 Oct 2022
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