IVF或ICSI前，將收集的卵子在特製培養基中培養3-6H，可提高卵子的成熟度和早期胚胎發育品質。

Review

 

. 2025 May 2;24(1):e12652.

 doi: 10.1002/rmb2.12652. eCollection 2025 Jan-Dec.

The quality of human eggs and its pre-IVF incubation

Background: Multi-factors influence the success rate of infertility treatments, and one of the important points is to obtain good quality eggs. Methods: Based on the literatures and unpublished data, the factors affecting egg quality were summarized.

Main findings results: Egg quality is an important determinant in successful infertility treatment. In addition to maternal age, controlled ovarian hyperstimulation (COH) protocols also play a key role in affecting the quality of the egg. After egg retrieval, the insemination occurs 3-6 h after collection, with a pre-IVF incubation time by in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) (39-42 h post-HCG injection). The pre-IVF incubation refers to the short period time of 3 to 6 h after oocyte retrieval and before the insemination by IVF or ICSI. The pre-IVF incubation of collected eggs in the designed culture medium improves egg quality in terms of maturation and early embryonic development.

Conclusions: Pre-IVF incubation of the collected eggs contributes to the improvement of the quality of eggs; therefore, it may increase subsequent pregnancy and implantation rates following embryo transfer.

鮮胚植入或凍胚植入在懷孕率方面並沒有明顯差異

兩者在累積懷孕率方面也沒有明顯差異

凍胚植入的最大好處是可以降低卵巢過度刺激的機率OHSS

Meta-Analysis

 

. 2021 Feb 4;2(2):CD011184.

 doi: 10.1002/14651858.CD011184.pub3.

Fresh versus frozen embryo transfers in assisted reproduction

Background: In vitro fertilisation (IVF) or intracytoplasmic sperm injection (ICSI) treatments conventionally consist of a fresh embryo transfer, possibly followed by one or more cryopreserved embryo transfers in subsequent cycles. An alternative option is to freeze all suitable embryos and transfer cryopreserved embryos in subsequent cycles only, which is known as the 'freeze all' strategy. This is the first update of the Cochrane Review on this comparison. Objectives: To evaluate the effectiveness and safety of the freeze all strategy compared to the conventional IVF/ICSI strategy in women undergoing assisted reproductive technology. Search methods: We searched the Cochrane Gynaecology and Fertility Group Trials Register, CENTRAL, MEDLINE, Embase, PsycINFO, CINAHL, and two registers of ongoing trials from inception until 23 September 2020 for relevant studies, checked references of publications found, and contacted study authors to obtain additional data. Selection criteria: Two review authors (TZ and MZ) independently selected studies for inclusion, assessed risk of bias, and extracted study data. We included randomised controlled trials comparing a 'freeze all' strategy with a conventional IVF/ICSI strategy including a fresh embryo transfer in women undergoing IVF or ICSI treatment. Data collection and analysis: The primary outcomes were cumulative live birth rate and ovarian hyperstimulation syndrome (OHSS). Secondary outcomes included effectiveness outcomes (including ongoing pregnancy rate and clinical pregnancy rate), time to pregnancy and obstetric, perinatal and neonatal outcomes. Main results: We included 15 studies in the systematic review and eight studies with a total of 4712 women in the meta-analysis. The overall evidence was of moderate to low quality. We graded all the outcomes and downgraded due to serious risk of bias, serious imprecision and serious unexplained heterogeneity. Risk of bias was associated with unclear blinding of investigators for preliminary outcomes of the study during the interim analysis, unit of analysis error, and absence of adequate study termination rules. There was an absence of high-quality evidence according to GRADE assessments for our primary outcomes, which is reflected in the cautious language below. There is probably little or no difference in cumulative live birth rate between the 'freeze all' strategy and the conventional IVF/ICSI strategy (odds ratio (OR) 1.08, 95% CI 0.95 to 1.22; I2 = 0%; 8 RCTs, 4712 women; moderate-quality evidence). This suggests that for a cumulative live birth rate of 58% following the conventional strategy, the cumulative live birth rate following the 'freeze all' strategy would be between 57% and 63%. Women might develop less OHSS after the 'freeze all' strategy compared to the conventional IVF/ICSI strategy (OR 0.26, 95% CI 0.17 to 0.39; I2 = 0%; 6 RCTs, 4478 women; low-quality evidence). These data suggest that for an OHSS rate of 3% following the conventional strategy, the rate following the 'freeze all' strategy would be 1%. There is probably little or no difference between the two strategies in the cumulative ongoing pregnancy rate (OR 0.95, 95% CI 0.75 to 1.19; I2 = 31%; 4 RCTs, 1245 women; moderate-quality evidence). We could not analyse time to pregnancy; by design, time to pregnancy is shorter in the conventional strategy than in the 'freeze all' strategy when the cumulative live birth rate is comparable, as embryo transfer is delayed in a 'freeze all' strategy. We are uncertain whether the two strategies differ in cumulative miscarriage rate because the evidence is very low quality (Peto OR 1.06, 95% CI 0.72 to 1.55; I2 = 55%; 2 RCTs, 986 women; very low-quality evidence) and cumulative multiple-pregnancy rate (Peto OR 0.88, 95% CI 0.61 to 1.25; I2 = 63%; 2 RCTs, 986 women; very low-quality evidence). The risk of hypertensive disorders of pregnancy (Peto OR 2.15, 95% CI 1.42 to 3.25; I2 = 29%; 3 RCTs, 3940 women; low-quality evidence), having a large-for-gestational-age baby (Peto OR 1.96, 95% CI 1.51 to 2.55; I2 = 0%; 3 RCTs, 3940 women; low-quality evidence) and a higher birth weight of the children born (mean difference (MD) 127 g, 95% CI 77.1 to 177.8; I2 = 0%; 5 RCTs, 1607 singletons; moderate-quality evidence) may be increased following the 'freeze all' strategy. We are uncertain whether the two strategies differ in the risk of having a small-for-gestational-age baby because the evidence is low quality (Peto OR 0.82, 95% CI 0.65 to 1.05; I2 = 64%; 3 RCTs, 3940 women; low-quality evidence). Authors' conclusions: We found moderate-quality evidence showing that one strategy is probably not superior to the other in terms of cumulative live birth rate and ongoing pregnancy rate. The risk of OHSS may be decreased in the 'freeze all' strategy. Based on the results of the included studies, we could not analyse time to pregnancy. It is likely to be shorter using a conventional IVF/ICSI strategy with fresh embryo transfer in the case of similar cumulative live birth rate, as embryo transfer is delayed in a 'freeze all' strategy. The risk of maternal hypertensive disorders of pregnancy, of having a large-for-gestational-age baby and a higher birth weight of the children born may be increased following the 'freeze all' strategy. We are uncertain if 'freeze all' strategy reduces the risk of miscarriage, multiple pregnancy rate or having a small-for-gestational-age baby compared to conventional IVF/ICSI.

培養基系統葡萄糖濃度呈現高-低-高的模式：受精培養基為 2.5-3 mM，卵裂期培養基為 0.5 mM 或更低，囊胚期培養基為 2.5-3.3 mM。

. 2025 Jan 1;40(1):30-40.

 doi: 10.1093/humrep/deae248.

The composition of commercially available human embryo culture media

Study question: What is the composition of currently available commercial human embryo culture media provided by seven suppliers, for each stage of human preimplantation embryo development?

Summary answer: While common trends existed across brands, distinct differences in composition underlined the absence of a clear standard for human embryo culture medium formulation.

What is known already: The reluctance of manufacturers to fully disclose the composition of their human embryo culture media generates uncertainty regarding the culture conditions that are used for human preimplantation embryo culture. The critical role of the embryo culture environment is well-recognized, with proven effects on IVF success rates and child outcomes, such as birth weight. The lack of comprehensive composition details restricts research efforts crucial for enhancing our understanding of its impacts on these outcomes. The ongoing demand for greater transparency remains unmet, highlighting a significant barrier in embryo culture medium optimization.

Study design, size, duration: For this study, 47 different human embryo culture media and protein supplements were purchased between December 2019 and June 2020; they comprise complete media (n = 23), unsupplemented media (n = 14), and supplements (n = 10). Unsupplemented media were supplemented with each available supplement from the same brand (n = 33 combinations). All samples were directly frozen in liquid nitrogen and stored at -80°C until composition analysis.

Participants/materials, setting, methods: We determined the concentrations of 40 components in all samples collected (n = 80). Seven electrolytes (calcium, chloride, iron, magnesium, phosphate, potassium, sodium), glucose, immunoglobulins A, G, and M (IgA, IgG, IgM), uric acid, alanine aminotransferase (ALAT), aspartate aminotransferase (ASAT), and albumin, as well as the total protein concentration, were determined in each sample using a Cobas 8000 Analyser (Roche Diagnostics). Analysis of pyruvate, lactate, carnitine, and 21 amino acids was achieved with Ultra-High Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS/MS).

Main results and the role of chance: Our analysis showed that generally, the concentrations of components of ready-to-use human embryo culture media align with established assumptions about the changing needs of an embryo during early development. For instance, glucose concentrations displayed a high-low-high pattern in sequential media systems from all brands: 2.5-3 mM in most fertilization media, 0.5 mM or below in all cleavage stage media, and 2.5-3.3 mM in most blastocyst stage media. Continuous media generally resembled glucose concentrations of cleavage stage media. However, for other components, such as lactate, glycine, and potassium, we observed clear differences in medium composition across different brands. No two embryo culture media compositions were the same. Remarkably, even embryo culture media from brands that belong to the same parent company differed in composition. Additionally, the scientific backing for the specific concentrations used and the differences in the composition of sequential media is quite limited and often based on minimal in vivo studies of limited sample size or studies using animal models.

三個正常染色體囊胚可提供93%的生育數率。

冷凍卵子時年齡小於35歲，平均需15個成熟卵細胞才能獲得三個正常染色體囊胚。

若患者年齡≥38歲，所成熟卵細胞數量增加一倍；，平均需30個成熟卵細胞才能獲得三個正常染色體囊胚。

若患者年齡≥40歲，則所需MII期卵母細胞數量增加至三倍，，平均需45個成熟卵細胞才能獲得三個正常染色體囊胚。。 

應鼓勵38歲之前進行卵子冷凍。

年齡≥38歲的女性，可能需要多次冷凍卵子週期才能獲得較高的活產率。

. 2025 Sep;124(3):487-495.

 doi: 10.1016/j.fertnstert.2025.04.023. Epub 2025 Apr 23.

The number of autologous, vitrified mature oocytes needed to obtain three euploid blastocysts increases with age

Objective: To determine the number of mature oocytes required to make at least three euploid blastocysts.

Subjects: All patients at a multicenter assisted reproductive technology practice that warmed vitrified autologous oocytes for in vitro fertilization from 2011 to October 2023.

Main outcome measures: Number of mature (metaphase II [MII]) oocytes to achieve at least three euploid blastocysts.

Results: A total of 1,041 thaw cycles from 986 patients were analyzed. Patients were stratified by age at vitrification. Measures of ovarian reserve decreased with advancing age, as expected. The number of mature oocytes (MII) banked and euploid blastocysts available decreased as age at vitrification increased. Patients aged 35-40 years used preimplantation genetic testing more than those older than 40 years or younger than 35 years. To optimize the chance of a potential live birth by banking at least three euploid blastocysts, patients aged <35 years at vitrification needed a mean of 15 MII oocytes. This number doubled in patients aged ≥38 years and tripled for patients aged >40 years. We also analyzed potential live birth per number of MII oocytes banked, which also increased with age, with 0.13 expected live births per thawed MII oocyte at the age of <35 years and 0.04 expected live births if older than 40 years at time of oocyte vitrification. Age at vitrification was more influential than indication for vitrification for expected outcome of live birth.

Conclusion: From previously published literature, three euploid blastocysts provided a 93% chance of a child. From our analysis, patients required a mean of 15 mature oocytes to yield at least three euploid blastocysts if they were aged <35 years at time of vitrification. The required number of MII oocytes doubled in patients aged ≥38 years and tripled in those aged ≥40 years, to achieve the same goal. Given our results, we should encourage egg freeze patients to cryopreserve before the age of 38 years. We should also counsel patients that multiple vitrification cycles will likely be required to achieve a high probability of live birth from future oocyte thaws for women aged ≥38 years.

PGT可能會下降累積活產率 尤其是<35歲患者

>40歲以上無明顯降低

J Assist Reprod Genet 2023 Jan;40(1):137-149.

 doi: 10.1007/s10815-022-02667-x. Epub 2022 Dec 1.

PGT-A is associated with reduced cumulative live birth rate in first reported IVF stimulation cycles age ≤ 40: an analysis of 133,494 autologous cycles reported to SART CORS

Purpose: To evaluate the impact of preimplantation genetic testing for aneuploidy (PGT-A) on cumulative live birth rate (CLBR) in IVF cycles.

Methods: Retrospective cohort study of the SART CORS database, comparing CLBR for patients using autologous oocytes, with or without PGT-A. The first reported autologous ovarian stimulation cycle per patient between January 1, 2014, and December 31, 2015, and all linked embryo transfer cycles between January 1, 2014, and December 31, 2016, were included in the study. Exclusion criteria were donor oocyte cycles, donor embryo cycles, gestational carrier cycles, cycles which included both a fresh embryo transfer (ET) combined with a thawed embryo previously frozen (ET plus FET), or cycles with a fresh ET after PGT-A.

Results: A total of 133,494 autologous IVF cycles were analyzed. Amongst patients who had blastocysts available for either ET or PGT-A, including those without transferrable embryos, decreased CLBR was noted in the PGT-A group at all ages, except ages > 40 (p < 0.01). A subgroup analysis of only those patients who had PGT-A and a subsequent FET, excluding those without transferrable embryos, demonstrated a very high CLBR, ranging from 71.2% at age < 35 to 50.2% at age > 42. Rates of multiple gestations, preterm birth, early pregnancy loss, and low birth weight were all greater in the non-PGT-A group.

Conclusions: PGT-A was associated with decreased CLBR amongst all patients who had blastocysts available for ET or PGT-A, except those aged > 40. The negative association of PGT-A use and CLBR per cycle start was especially pronounced at age < 35.

用 精子分離装置 (Zymot)。比傳統精蟲分離方式(DCC)可達到較高比率正常染色體囊胚

https://www.mdpi.com/2075-1729/15/2/302

採用 精子分離装置 (SSD)。比傳統精蟲分離方式可篩選出具有高活力且 DNA 損傷較少的精子群體，

. 2024 Aug;41(8):2201-2209.

 doi: 10.1007/s10815-024-03168-9. Epub 2024 Jun 18.

Optimized sperm selection: a highly efficient device for the isolation of progressive motile sperm with low DNA fragmentation index

Purpose: To identify the sperm preparation procedure that selects the best sperm population for medically assisted reproduction.Methods: Prospective observational study comparing the effect of four different sperm selection procedures on various semen parameters. Unused raw semen after routine diagnostic analysis was split in four fractions and processed by four different methods: (1) density gradient centrifugation (DGC), (2) sperm wash (SW), (3) DGC followed by magnetic activated cell sorting (MACS), and (4) using a sperm separation device (SSD). Each fraction was analyzed for progressive motility, morphology, acrosome index (AI), and DNA fragmentation index (DFI).Results: With DGC as standard of care in intraclass correlation coefficient analysis, only SSD was in strong disagreement regarding progressive motility and DFI [0.26, 95%CI (- 0.2, 0.58), and 0.17, 95%CI (- 0.19, 0.45), respectively]. When controlling for abstinence duration, DFI was significantly lower after both MACS and SSD compared to DGC [- 0.27%, 95%CI (- 0.47, - 0.06), p = 0.01, and - 0.6%, 95%CI (- 0.80, - 0.41), p < 0.001, respectively]. Further comparisons between SSD and MACS indicate significantly less apoptotic cells [Median (IQR) 4 (5), 95%CI (4.1, - 6.8) vs Median (IQR) 5 (8), 95%CI (4.9, - 9.2), p < 0.001, respectively] and dead cells [Median (IQR) 9.5 (23.3), 95%CI (13.2, - 22.4) vs Median (IQR) 22 (28), 95%CI (23.1, - 36.8), p < 0.001, respectively] in the SSD group.Conclusion: The selection of a population of highly motile spermatozoa with less damaged DNA from unprocessed semen is ideally performed with SSD. Question remains whether this method improves the embryological outcomes in the IVF laboratory.

Table 2.

Descriptive analysis of the effect of four different preparation techniques on concentration, progressive motility, normal morphology, AI, and DFI

	SW	DGC	MACS	SSD
Concentration (× 106)	61.7 ± 35.4 (17.5–193.0)	13.0 ± 11.6 (0.8–68)	8.4 ± 9.2 (0.61–49.6)	15.1 ± 14.2 (1.5–69.0)
Progressive motility (%)	54.3 ± 10.6 (23–86)	74.3 ± 11.8 (38–90)	77.2 ± 12.5 (37–92)	88.6 ± 4.2 (73–96)
Normal morphology (%)	3.3 ± 2.9 (0–13)	4.1 ± 3.1 (0–13)	4.2 ± 3.7 (0–18)	5.1 ± 3.9 (0–16)
AI (%)	8.5 ± 4.9 (1–20)	9.7 ± 6 (1–30)	8.7 ± 4.9 (0–19)	10.8 ± 6.8 (1–30)
DFI (%)	6.2 ± 4.6 (0.8–26.1)	2.7 ± 3.2 (0.2–14)	2.1 ± 4.3 (0.9–20.8)	0.2 ± 0.4 (0–2.3)

Fig. 2 

Capacity of MACS and SSD procedures to remove apoptotic (A) and dead (B) cells from sperm cell population (Wilcoxon signed-rank test). Spearman correlation test between the days of abstinence and the presence of dead cells in selected sperm population after MACS (C). Abbreviations: MACS: magnetic activated cell sorting, SSD: microfluidic sperm sorting

 未來可能niPGT 與傳統TE biopy 可能同時並用以防止TE biopsy偵測失敗

Review

 

. 2025 Oct 14;14(20):1591.

 doi: 10.3390/cells14201591.

Noninvasive Preimplantation Genetic Testing in Recurrent Pregnancy Loss and Implantation Failure: Breakthrough or Overpromise?

Background: Recurrent pregnancy loss (RPL) and recurrent implantation failure (RIF) are significant challenges in reproductive medicine. For both, embryonic aneuploidy is the leading etiological factor. Preimplantation genetic testing for aneuploidy (PGT-A) via trophectoderm biopsy is the current standard for embryo selection. However, it is limited by its invasiveness, potential for embryo damage, and diagnostic errors due to mosaicism. Rationale/Objectives: This review critically evaluates the emerging role of noninvasive PGT (niPGT). NiPGT analyzes cell-free DNA from spent blastocyst culture media, thus, it is a potential alternative for managing RPL and RIF. Hence, the primary objective is to determine whether current evidence supports niPGT as a reliable replacement for conventional biopsy-based PGT-A in these high-risk populations. Outcomes: The analysis reveals that niPGT offers significant theoretical advantages. These include complete non-invasiveness, enhanced embryo preservation, and high patient acceptability. However, its clinical application is hampered by substantial limitations. Key amongst them is the inconsistent and often suboptimal diagnostic accuracy (sensitivity 70-85%, specificity 88-92%) compared to biopsy. Other significant factors include the high rates of amplification failure (10-50%), vulnerability to maternal DNA contamination, as well as low DNA yield. Crucially, there is a definitive lack of robust, prospective randomized controlled trial (RCT) data demonstrating improved live birth rates or reduced miscarriage rates specifically in RPL and RIF cohorts. As such, niPGT is not yet ready to be a standalone clinical adoption in RPL and RIF cases. However, it may serve as a valuable adjunct for rescue scenarios following biopsy failure or for ethical reasons

niPGT vs. 傳統切片PGT----對應全染色體，染色體一致率仍有差距 (77 vs 91%)

Ploidy concordance rates in SBM-TE, SBM-WB, and TE-WB were 77.4% (130/168), 77.4% (130/168), and 90.5% (152/168),

Multicenter Study

 

. 2025 Oct;42(10):3479-3491.

 doi: 10.1007/s10815-025-03670-8. Epub 2025 Oct 7.

Validation of non-invasive preimplantation genetic testing for aneuploidies (niPGT-A) in a Japanese population: experience from seven clinics

Purpose: The aim of this study was to determine the feasibility of implementing non-invasive PGT-A (niPGT-A) in clinical practice. Informativity and concordance rates between three sample types (embryonic cell-free DNA [cfDNA] present in spent blastocyst medium [SBM], trophectoderm [TE] biopsy, and whole blastocyst [WB]) from the same embryo were evaluated.

Methods: This was a prospective, multicenter study conducted between February 2022 and November 2022 at seven Japanese IVF centres. 212 blastocysts were donated for research. The cfDNA released into the SBM was analysed, and the results were compared against the corresponding TE biopsy and WB sample.

Results: Overall informativity rates for SBM, TE, and WB were 81.6% (173/212), 98.6% (209/212), and 98.6% (209/212), respectively. There was no difference between TE and WB; however, SBM was significantly different to both (p < 0.001). The informativity rate in SBM samples significantly varied among the seven centres, ranging between 72.7 and 97.1% (p = 0.041). Ploidy concordance (SBM-TE) also varied across the centres, ranging between 68.2 and 90.9%; however, this did not reach statistical significance (p = 0.63). Ploidy concordance rates in SBM-TE, SBM-WB, and TE-WB were 77.4% (130/168), 77.4% (130/168), and 90.5% (152/168), respectively. There was no statistical difference for SBM-TE and SBM-WB, but both were statistically different to TE-WB (p = 0.0054).

Conclusion: We have shown results that are consistent with the existing literature, indicating the feasibility of applying the niPGT-A protocol described here, and thus using the SBM result to establish a priority for embryo transfer.

 ni-PGT培養模式(微滴10-15uL, D4-6不更換培養液) 

----對囊胚無明顯不良影響

Observational Study

 

. 2024 Sep 1;39(9):1952-1959.

 doi: 10.1093/humrep/deae156.

The impact of implementing a non-invasive preimplantation genetic testing for aneuploidies (niPGT-A) embryo culture protocol on embryo viability and clinical outcomes

Study question: Are modifications in the embryo culture protocol needed to perform non-invasive preimplantation genetic testing for aneuploidies (niPGT-A) affecting clinical reproductive outcomes, including blastocyst development and pregnancy outcomes?

Summary answer: The implementation of an embryo culture protocol to accommodate niPGT-A has no impact on blastocyst viability or pregnancy outcomes.

What is known already: The recent identification of embryo cell-free (cf) DNA in spent blastocyst media has created the possibility of simplifying PGT-A. Concerns, however, have arisen at two levels. First, the representativeness of that cfDNA to the real ploidy status of the embryo. Second, the logistical changes that need to be implemented by the IVF laboratory when performing niPGT-A and their effect on reproductive outcomes. Concordance rates of niPGT-A to invasive PGT-A have gradually improved; however, the impact of culture protocol changes is not as well understood.

Study design, size, duration: As part of a trial examining concordance rates of niPGT-A versus invasive PGT-A, the IVF clinics implemented a specific niPGT-A embryo culture protocol. Briefly, this involved initial culture of fertilized oocytes following each laboratory standard routine up to Day 4. On Day 4, embryos were washed and cultured individually in 10 μl of fresh media. On Day 6 or 7, blastocysts were then biopsied, vitrified, and media collected for the niPGT-A analysis. Six IVF clinics from the previously mentioned trial were enrolled in this analysis. In the concordance trial, Clinic A cultured all embryos (97 cycles and 355 embryos) up to Day 6 or 7, whereas in the remaining clinics (B-F) (379 cycles), nearly a quarter of all the blastocysts (231/985: 23.5%) were biopsied on Day 5, with the remaining blastocysts following the niPGT-A protocol (754/985: 76.5%). During the same period (April 2018-December 2020), the IVF clinics also performed standard invasive PGT-A, which involved culture of embryos up to Days 5, 6, or 7 when blastocysts were biopsied and vitrified.