What is the recommended protocol for using frozen‑thawed sperm in intracytoplasmic sperm injection (ICSI) for couples undergoing assisted reproduction?

Using Frozen-Thawed Sperm from Microdissection Testicular Sperm Extraction (mTESE) for ICSI

Frozen-thawed testicular spermatozoa retrieved via mTESE can be used effectively for ICSI with fertilization rates, pregnancy rates, and live birth outcomes equivalent to fresh testicular sperm. 1, 2

Recommended Protocol for Frozen-Thawed mTESE Sperm in ICSI

Sperm Collection and Cryopreservation Strategy

• Bank testicular tissue during the initial mTESE procedure rather than coordinating simultaneous sperm retrieval with oocyte collection, as this dramatically reduces scheduling burden on couples and clinical teams while ensuring sperm availability before initiating ovarian stimulation. 3

• Cryopreserve using commercial glycerol-based sperm cryomedia when sufficient mature sperm are identified during intra-operative analysis of testicular tissue. 1

• Divide each sample into multiple vials (typically 2-3 vials per planned ICSI cycle) to enable staged use and provide safeguard against technical failures or unexpected loss of post-thaw motility. 4

• Collect at least 3 separate tissue samples if feasible to maximize stored reserves, particularly important given that only 9% of cryopreserved samples are ultimately used while 23% are disposed of unused. 4, 5

Thawing and Preparation for ICSI

• Thaw vials on the day of oocyte retrieval following standard laboratory protocols for frozen-thawed testicular spermatozoa. 6, 7

• Assess post-thaw motility and viability immediately - when prefreeze vitality exceeds 20%, fertilization rates (56%) and embryo cleavage rates (91%) are significantly superior to samples with lower vitality (22% and 57% respectively). 8

• Proceed with standard ICSI technique using individual motile spermatozoa selected under high magnification, as ICSI requires only a single viable sperm per oocyte even when overall parameters are severely compromised. 1, 4

Expected Outcomes with Frozen-Thawed Testicular Sperm

• Fertilization rates of 61-66% are achievable with frozen-thawed testicular spermatozoa, comparable to fresh testicular sperm (62%). 7, 2

• Clinical pregnancy rates per embryo transfer of 25-29% show no significant difference between fresh and frozen-thawed testicular sperm. 2

• Delivery rates of 80-86% among established pregnancies demonstrate equivalent obstetric outcomes. 2

• Live birth rates of 62% per ICSI cycle have been reported in cancer patients using cryopreserved sperm with ICSI, significantly higher than other ART methods. 4

Critical Advantages of Cryopreservation Strategy

• Eliminates need for simultaneous coordination of male surgical sperm retrieval with female oocyte collection, which is logistically complex and stressful for couples. 3

• Allows multiple ICSI attempts from a single surgical procedure, avoiding repeated testicular biopsies that may compromise remaining spermatogenesis. 6, 8

• Permits sperm availability confirmation before initiating costly and invasive ovarian stimulation in the female partner. 3

• Enables treatment at different centers if couples relocate or seek specialized care elsewhere. 6

Common Pitfalls to Avoid

• Never delay cryopreservation if mTESE is being performed for fertility preservation before gonadotoxic treatment, as 10% of cancer patients fail to cryopreserve sperm if collection is attempted after treatment initiation. 4

• Do not assume fresh sperm is superior - multiple studies demonstrate no significant differences in implantation rates or pregnancy outcomes between fresh and frozen-thawed surgically retrieved spermatozoa. 6, 7, 2

• Avoid using samples with prefreeze vitality <20% for primary ICSI attempts, as these demonstrate significantly compromised fertilization (22% vs 56%) and should prompt consideration of repeat retrieval if possible. 8

• Do not underestimate storage needs - when the female partner has diminished ovarian reserve or advanced maternal age, multiple ICSI cycles are often required, necessitating adequate cryopreserved reserves. 4, 5

Special Considerations for Non-Obstructive Azoospermia

• Recognize that even with elevated FSH and testicular atrophy, up to 50% of men with non-obstructive azoospermia have retrievable sperm via microsurgical extraction. 4

• Bank all retrieved sperm immediately regardless of quantity, as intracytoplasmic sperm injection enables successful fertilization with extremely limited numbers of viable spermatozoa. 1, 4