For donors participating in an extended aspiration program, a three-week interval between sessions is recommended. Under this 21-day aspiration schedule, higher-quality oocytes are obtained, resulting in improved embryo production.

On a single aspiration day, 30–35 donors can be aspirated.

Oocytes recovered from each donor are matured, fertilized, and cultured separately, ensuring that each donor’s identity is preserved throughout the entire process.

Using a single straw of conventional semen, oocytes from 8–10 donors can be inseminated. In most cases, one straw of sexed semen is sufficient to fertilize oocytes from up to 6 donors.

In dairy herds, donors can be managed in groups to optimize the use of each semen straw. Under this system, up to 200 oocytes can be inseminated using a single straw of sexed semen. Using a conservative estimate, approximately 30 embryos can be produced from one straw.

Embryos produced through In Vitro Fertilization (IVF) can be frozen using two techniques:

 

A) Vitrificaction: 

For many years, vitrification has been the standard method for the cryopreservation of IVF-produced bovine embryos. Prior to vitrification, embryos pass through a series of culture media designed to protect their cellular structures. The embryos are then placed—individually or in groups of up to five—into ultra-fine, properly identified vitrification straws, commonly known as OPS (Open Pulled Straws), before being rapidly submerged in liquid nitrogen at −196°C and stored in the embryo bank.

The devitrification process (warming of vitrified embryos) must be carried out in a laboratory by trained technical personnel. After a short incubation period, embryos are selected and prepared for transfer to recipients. Once devitrified, embryos cannot be refrozen. Expected pregnancy rates with vitrified embryos range between 35–40%.

- Advantages of Vitrification:

Vitrified embryos can be cultured using the same embryo culture media as those intended for fresh transfer. Therefore, vitrification is the method of choice when the number of embryos exceeds the number of available recipients and the client wishes to establish an embryo bank for future transfers.

- Disadvantages of Vitrification:

This technique requires laboratory devitrification by specialized personnel and cannot be performed in the field.

 

B) Slow Freezing for Direct Transfer (DT):

Slow freezing is a technique widely used by veterinarians specialized in traditional embryo flushing programs (MOET – Multiple Ovulation Embryo Transfer). The ability to freeze IVF-produced embryos in a manner comparable to in vivo–derived embryos is relatively recent and has only become commercially available in recent years.

To enable this process, new embryo culture media have been developed, replacing traditional components with more advanced formulations specifically designed to allow embryos to successfully withstand the freezing process. Each embryo is frozen individually in properly identified 0.25 cc straws, following a controlled slow-freezing curve in specialized embryo freezers. The freezing process for a batch of embryos takes approximately one hour, after which embryos are transferred to liquid nitrogen at −196°C and stored in the embryo bank.

Expected pregnancy rates with DT (Direct Transfer) frozen embryos are comparable to those achieved with vitrified embryos, averaging 30–35%.

- Advantages of DT Freezing:

The primary advantage of this technique is the ability to perform direct embryo transfer in the field, without the need for laboratory warming procedures.

- Disadvantages of DT Freezing:

For embryos to be frozen using the slow-freezing (DT) method, they must be cultured in specific media from the outset. Therefore, the decision to freeze embryos—rather than transfer them fresh or vitrify them—must be made at the beginning of the process. However, it is possible to allocate some donors to freezing and others to fresh transfer within the same OPU (Ovum Pick-Up) session.

Direct Transfer (DT) straws can be thawed in the field (10 seconds in the air followed by 20 seconds submerged in water at 35°C) and immediately transferred to previously synchronized recipients.

In addition to significantly simplifying the transfer process, DT embryos offer strong commercial potential for breeders seeking to diversify their genetics. Another key advantage is the ability to maintain donors in a year-round periodic aspiration program and transfer embryos during the spring, when recipient availability is highest.