ICSI with Frozen Eggs

Oocyte Thawing (warming) Process

Cellular Principles of Oocyte Thawing

There is some debate as to whether the term ‘thawing’ is appropriate for oocytes, as they are preserved in a glass-like state rather than a crystalline (ice) state. Rapid thawing is very important in order to prevent the injurious effects of recrystallisation during the phase change of oocyte vitrification.

Table of Contents

Detailed Steps in Oocyte Thawing

Removal from Liquid Nitrogen Storage

Vitrified (cryopreserved) oocytes are stored at –196°C in liquid nitrogen either within the closed or open vitrification system.

Rapid Thawing

The vitrification device (straw) containing oocyte(s) should be immediately immersed in a thawing solution (TS) at 37°C, followed by a diluent solution (DS) and washing solution (WS) at room temperature (25-27 °C).

Rapid warming is extremely important because slower warming rates increase the risk of:

Recrystallisation of ice
Osmotic shock
Rupture of the membrane

Removal of Cryoprotectant

Through a series of solutions with decreasing sucrose concentrations, the cryoprotectant is gradually removed from the oocyte, and normal osmotic conditions are restored.

Morphologic Evaluation of Survival

Morphologic evaluation of oocyte survival is done approximately 1-2 hours post-thaw:

Oocyte survival characteristics include:

Intact oolemma
Re-expanded cytoplasm
Normal perivitelline space
Intact/Preserved Zona Pellucida
Intact polar body

Degenerated oocytes may show:

Darkened cytoplasm
Collapse of the membrane
Leakage of cytoplasm
Vacuolate appearance

Intracytoplasmic Sperm Injection (ICSI)

ICSI is done in most cases, as conventional insemination has a reduced success rate for fertilisation when vitrification causes hardening of the zona.

What affects the thaw survival rate

Maternal Age: –

Maternal age remains the most significant factor affecting reproductive success because oocyte quality declines with age as a result of:

Meiotic Nondisjunction
Shortening of Telomeres
Dysfunction of Mitochondria
Increasing Oxidative Stress
Build-Up of DNA Damage
Increasing Aneuploidy Rates as a Function of Maternal Age.

Estimated aneuploidy rates by maternal age:

Maternal Age (%)	Estimated Aneuploidy Rate
Less than 35 years	20-30%
35 years to 37 years	35-45%
38 years to 40 years	50-70%
Greater than 40 years	70-85%

This accounts for the fact that younger frozen eggs will have much higher live birth rates.

Stage of Development of Oocytes Impacting Oocyte Cryopreservation

Only mature oocytes at the metaphase II stage should be selected for vitrification.

Stage of Development of Oocyte	Cryopreservation Potential
Germinal Vesicle (GV)	Low
Metaphase I (MI)	Moderate
Metaphase II (MII)	High

Immature oocytes often lack:

Proper organization of the spindle apparatus
Fertility rates of less than 20%
A reduced potential to develop into a blastocyst

Laboratory Conditions: –

Oocyte Cryopreservation outcomes are highly dependent on laboratory conditions.

Critical Variables: –

Cryoprotectant Exposure Duration
Temperature Stability
Practitioner Experience
Level of Sterility
Warming Speed
Media Composition
pH Control
Osmolarity Control

Minor deviations from established laboratory protocols can significantly affect post-warming oocyte survival.

Oxidative Stress

Oocyte cryopreservation may increase reactive oxygen species (ROS) levels, resulting in damage to:

Mitochondrial DNA
Cellular Membranes
Meiotic Spindle
Endoplasmic Reticulum

Oxidative stress is related to:

Developmental Arrest
Fragmentation and/or Malformation of the Embryo
Reduced Implantation Potential

Success rates by age at oocyte freezing

Clinical Outcomes By Maternal Age

The likelihood of achieving a live birth depends on several factors, including:

The total number of oocytes harvested
The survival of the thawed oocytes
The ability of an oocyte to successfully fertilize
The formation of a blastocyst from a fertilized oocyte
The number of euploid embryos produced from the oocytes

Age at oocyte freezing	Average Number of Oocytes Required	Rate of Oocyte Survival	Blastocyst formation Rate	Euploid Rate	Estimated Live Birth Rate
<30	8-12	90-95%	High	70-80%	70-85%
30-40	10-15	88-95%	High	60-70%	60-75%
35-37	15-20	85-92%	Moderate	45-60%	40-60%
38-40	20-30	75-88%	Reduced	25-40%	20-40%
>40	>30	65-80%	Significantly reduced	<20%	<15-20%

Please note these numbers are averages and can vary from clinic to clinic.

Outcomes of Fresh Versus Frozen Eggs

Comparative Reproductive Competence

Advances in vitrification have significantly reduced the differences in outcomes between fresh and frozen-thawed eggs.

Parameter	Fresh Oocyte	Frozen-Thawed Oocyte
Survival rates	Not applicable	80-95%
fertilisation rates	70-80%	65-80%
Cleavage rates	High	Comparable
Blastocyst rates	High	Slightly reduced in some cohorts
Euploidy embryo rates	Comparable	Comparable
Implantation rates	Comparable	Comparable
Clinical pregnancy rates	Slightly higher	Nearly equal
Live birth rates	Gold standard	Comparable with vitrification

According to available meta-analysis, oocyte vitrification has not been associated with an increased risk of congenital disabilities, obstetric complications, or neonatal morbidity.

Frequently Asked Questions

How many frozen eggs do I need for one baby?

For women under the age of 35, approximately 10–15 good-quality mature vitrified eggs may provide a reasonable chance of achieving at least one live birth. Women aged 38 and older often require 20–30 or more mature eggs because egg quality declines with age and the risk of chromosomal abnormalities increases. It is also important to remember that not all thawed eggs will survive, fertilise successfully, or develop into viable embryos. The total number of mature eggs frozen is therefore a key factor in determining future reproductive success.

Does my age when I froze my eggs really make that big a difference?

Yes. The age at which eggs are frozen is one of the most important factors influencing future success rates. Eggs frozen during a woman’s 20s and early 30s are generally healthier, with lower rates of chromosomal abnormalities and better survival during thawing, fertilisation, and embryo development. Research has shown that egg survival rates after warming decline with age, and fertilisation rates also decrease. Studies have found that frozen eggs from women in their late 30s and early 40s have a significantly lower chance of producing healthy embryos compared with eggs frozen at younger ages. For this reason, egg freezing tends to be most effective when performed earlier in reproductive life.

Why do some eggs not survive the thaw?

Not all frozen eggs survive the warming process, and some loss is expected. Human eggs are among the largest cells in the body and contain a high amount of water, making them particularly sensitive to temperature changes. Modern vitrification techniques have improved survival rates significantly by preventing the formation of damaging ice crystals, achieving survival rates of approximately 95%. Egg quality at the time of freezing also affects survival, with immature or structurally abnormal eggs being less likely to withstand thawing. Laboratory expertise, storage conditions, and strict handling protocols all play important roles in maximising egg survival.

What are my chances of success based on when I froze my eggs?

Success rates following egg freezing vary considerably depending on the age at which the eggs were frozen. Large studies have shown that women who freeze eggs at age 40 or younger have significantly higher chances of achieving an ongoing pregnancy or live birth compared with women who freeze eggs later in life. The proportion of genetically normal (euploid) eggs is also strongly influenced by age. Women who freeze eggs between the ages of 38 and 42 generally experience lower success rates per egg than younger women. Fertility clinics can provide personalised estimates based on your age at freezing, the number of eggs stored, and their quality.

Are babies born from frozen eggs as healthy as those born from fresh eggs?

Current evidence suggests that babies conceived from frozen eggs are just as healthy as those conceived from fresh eggs. Studies have found similar rates of chromosomal normality in embryos created from frozen and fresh eggs, indicating that the freezing process does not significantly increase the risk of genetic abnormalities. Research has also shown no meaningful increase in congenital malformations, birth defects, neonatal complications, or infant mortality among babies conceived using frozen eggs. Modern egg freezing and thawing techniques are therefore considered safe and effective for fertility preservation.

References:

Abdelhady, A. W., Mittan-Moreau, D. W., Crane, P. L., McLeod, M. J., Cheong, S. H., & Thorne, R. E. (2024). Ice formation and its elimination in cryopreservation of oocytes. Scientific Reports, 14(1), 18809. https://doi.org/10.1038/s41598-024-69528-8
Anderson, R. A., Amant, F., Braat, D., D’Angelo, A., Chuva de Sousa Lopes, S. M., Demeestere, I., Dwek, S., Frith, L., Lambertini, M., Maslin, C., Moura-Ramos, M., Nogueira, D., Rodriguez-Wallberg, K., & Vermeulen, N. (2020). ESHRE guideline: Female fertility preservation†. Human Reproduction Open, 2020(4), hoaa052. https://doi.org/10.1093/hropen/hoaa052
Cascante, S. D., Berkeley, A. S., Licciardi, F., McCaffrey, C., & Grifo, J. A. (2023). Planned oocyte cryopreservation: The state of the ART. Reproductive BioMedicine Online, 47(6), 103367. https://doi.org/10.1016/j.rbmo.2023.103367
Evidence-based outcomes after oocyte cryopreservation for donor oocyte in vitro fertilization and planned oocyte cryopreservation: A guideline (2021) | American Society for Reproductive Medicine | ASRM. (n.d.). Retrieved 18 June 2026, from https://integration.asrm.org/practice-guidance/practice-committee-documents/evidence-based-outcomes-after-oocyte-cryopreservation-for-donor-oocyte-in-vitro-fertilization-and-planned-oocyte-cryopreservation-a-guideline-2021/?utm_source=chatgpt.com
Kirubarajan, A., Patel, P., Thangavelu, N., Salim, S., Sadeghi, Y., Yeretsian, T., & Sierra, S. (2024). Return rates and pregnancy outcomes after oocyte preservation for planned fertility delay: A systematic review and meta-analysis. Fertility and Sterility, 122(5), 902–917. https://doi.org/10.1016/j.fertnstert.2024.06.025
Kwan, H. C. K. (2023). Reconsideration of the safety and effectiveness of human oocyte cryopreservation. Reproductive Biology and Endocrinology: RB&E, 21(1), 22. https://doi.org/10.1186/s12958-023-01071-z
The Effect of Open and Closed Oocyte Vitrification Systems on Embryo Development: A Systematic Review and Network Meta-Analysis. (n.d.). Retrieved 18 June 2026, from https://www.mdpi.com/2077-0383/13/9/2651?utm_source=chatgpt.com

Anna Haotanto

Anna Haotanto is the Founder of Zora Health and a passionate advocate for women’s empowerment. Anna’s personal experiences with egg-freezing, PCOS, perimenopause and the challenges of fertility have fueled her mission to provide high-quality information, financing, and support to help women and couples navigate their fertility journeys with confidence. She is also recognised for her achievements in finance, entrepreneurship, and women’s empowerment, and has been featured in various media outlets. You can also follow her on Linkedin or Instagram.