10. Oocyte Retrieval
KeywordsOocyte retrievalNeedle pickupDevice pickupProcedure oocyte retrieval
In the last 40 years since the first successful human birth, ART has been continuously improved. Significant advances have been made in oocyte fertilization and embryo culture, resulting in an increase of the success rate and safety of ART treatments. Oocyte recovery aims to maximize the number of oocytes recruited from the ovarian follicles, while minimizing the patient surgical risks. In the early history of IVF experimentation, abdominal laparotomy was performed to collect oocytes during tubal ligation procedures. Techniques described for follicle aspiration involved puncturing follicles with a diameter higher than 5 mm with a 20-gauge needle. The aspiration needle was connected to the tube and emptied into a test tube. Aspiration was achieved by covering the free opening in the three-way connector to create suction to 200 mmHg. Each follicle was finally transferred into an individual tube . Although the laparotomic approach could be an option to obtain oocytes in certain cases, it may cause several surgical risks, such as bleeding, infection, pain, potential injury to the surrounding pelvic and abdominal organs, and longer recovery time, which encouraged the pursuit of alternative surgical options. In the 1950s and 1960s, there was a great interest in developing less invasive gynecologic techniques.
Until a few years ago, laparoscopy represented the election technique for the collection of oocytes to be initiated into extracorporeal fertilization. The laparoscopic technique is not very different from the traditional one; however, it should be noted that no instrument can be placed during the examination in the uterine cavity for the mobilization of the bowel; therefore, the patient is placed on the bed with the legs outstretched as for any gynecological intervention.
Using forceps to stabilize the ovary and by a rotation to obtain adequate visualization, the thin-walled follicles are aspirated using a 20-gauge needle and a syringe for suction.
A short bevel needle is placed directly through the skin into the abdominal cavity and then cleared of blood and tissue with a heparinized saline solution. By an outer guide to insert the aspiration needle, the follicles are punctured and the suction is obtained by placing a finger over a bypass valve on the aspiration needle. The needle and tube are cleaned after each follicle aspiration. The maximum pressure for the vacuum suction is 120 mmHg as higher pressures would damage the oocytes .
Laparoscopy has slowly gained a growing acceptance after studies demonstrating similar oocyte yields compared with the laparotomy approach. Lopata and colleagues  showed the absence of differences in the mean number of oocytes obtained per patient between laparoscopy and laparotomy.
When laparoscopy was performed with CO2 pneumoperitoneum, there was no significant difference in oocyte fertilization rates. Advantages of laparoscopy include shorter recovery time, less bleeding, fewer infectious risks, and decreased pain. However, even with these improvements, the disadvantages of requiring general anesthesia and poor visualization of follicles within ovarian stroma led to improve the techniques of oocyte recovery .
10.1 Ultrasound Approach of Oocyte Retrieval
Advantages of transvaginal ultrasound approach
Transvaginal oocyte retrieval offers the following advantages:
• The distance to reach the ovary is shorter
• Higher-resolution pictures that enable the identification of the ovaries and the aspiration follicles
• No risk of skin damage
• The procedure can be conveniently performed in the outpatient setting
• Lower cost than other techniques
• Fewer staff is required
• Easy to learn thanks to the use of ultrasound guidance
• All follicles can be visualized and punctured, even in case of severe pelvic adhesions
• It gives more precision than the abdominal approach
• Can be achieved by local anesthesia, under paracervical block, sedation or general anesthesia
• It is well accepted by patients
In 1982, Lenz and colleagues  reported the first ultrasound-guided transvesical route for oocyte recovery using local anesthesia.
The oocyte yield was comparable after transvesicular and laparoscopy aspiration, but because of the intentional route through the bladder, complications were reported, such as abdominal pain, exacerbation of preexisting pelvic inflammatory disease, mild hemoperitoneum, urinary tract infections, and transient macroscopic hematuria .
Despite these complications, this approach remained one of the preferred treatments among physicians and patients, as it allows to avoid the general anesthesia and is an outpatient procedure. In addition, this method is safer than laparoscopy in patients with extensive abdominal or pelvic adhesive disease, due to an increased accuracy and a closer proximity of the needle to the ovaries .
To overcome the inconvenience of the distance between the abdominal ultrasound probe and the ovaries, in 1985, Wikland and colleagues introduced for the first time the vaginal ultrasound probes .
Transvaginal oocyte retrieval is currently the most common method of oocyte collection in IVF cycles. In fact, this technique has been found to be the simple and fast (it takes about 20 min), while being less invasive and requiring minimal anesthesia, thus resulting the most effective approach for oocyte retrievals in ART clinics . It consists in the aspiration of follicular fluid by using transvaginal ultrasonography; it is focused on obtaining the maximum number of oocyte to be fertilized.
The success of the technique depends on the inherent characteristics of the oocyte, which might be influenced by the actual process of oocyte collection, but also on other factors such as the length of the ovarian stimulation phase, the type of anesthesia used (local, sedation, or general), the type of aspiration needle (aspiration alone or aspiration with follicular flushing), and the experience of the clinician .
Finally, the number of embryos obtained relies on the number and inherent characteristics of the oocytes but also on other factors such as the length of the ovarian stimulation phase, the type of anesthesia used (local, sedation, or general), the type of aspiration needle magnitude, the needle used (wide or narrow bore or single or double channel), the aspiration mode (with or without follicular flushing), and the experience of the surgeon [16, 17].
10.2.1 Ultrasound Equipment and Probe
Follicle aspiration set
A thermoblock/heating block thermostat
Pump vacuum aspiration
In theory, it should cause minimal tissue injury when passing through the vagina and ovary, by guarantying, at the same time, high visibility for the correct approach to the follicles of the clinician. The needle consists of a reduced part (tip) and unreduced part (body). The most used aspiration needles in IVF centers is with single lumen, which have a smaller diameter and causes less discomfort. The needles with double lumen allow a constant infusion of oocyte collection media into the follicles while the follicular fluid is being removed; increase the turbulence within the follicle; assist in dislodging the oocyte–cumulus complex from the follicle wall; and increase the chances of oocyte collection.
Few studies have evaluated the gauge of the needle used and the outcomes of the oocyte collection. One study compared transvaginal oocyte collections with 15-, 17-, or 18-gauge needles . This prospective randomized study found that the number of oocytes collected was similar regardless of needle gauge, but more pain occurred with the 15-gauge than did with the 17- or 18- gauge needles . A second study found a trend toward lower pain scores with transvaginal collections performed with a 19-gauge needle (used in in vitro maturation [IVM] retrievals) when compared with a 16- or 17-gauge needle (used in IVF retrievals), although not statistically significant . The smaller needle may make for a more comfortable collection, despite more ovarian punctures and longer procedure time and the enlarged ovaries with multiple large follicles and higher aspiration pressure employed in IVF collection likely caused more pain. It is likely that within the range of conventional needles, smaller size results in less pain intra- and postoperatively, with a similar number of collected oocytes; however, more studies are needed to confirm this. There has been only a single randomized controlled trial comparing this needle to the traditional 19-gauge needle used for IVM. This study failed to find a difference in the number of oocytes aspirated . Therefore, oocytes in the dead space do not seem to be lost by being returned to the follicle when flushing occurs. However, the single-lumen flushing needle resulted in statistically less ovarian punctures and less clot formation. Few studies have compared different needles for oocyte collection, and these studies should be added to the literature. It is difficult to select one needle over the other, and the current indications include cost and physician preference.
The tubing, generally made of bend-resistant material, connects the follicle aspiration needle with the vacuum pump. In fact, all vacuum pump tubing has a male luer lock connection in one end for attaching the needle. The opposite end can be equipped with either an open end or female/male luer lock connection. The tubing should be sterile, and are intended for single use.
10.5 Sampling Tubes
10.6 Thermoblock/Heating Block
10.7 Pump Vacuum Aspiration
As the quality of the retrieved oocytes depends on both their intrinsic characteristics and the methods used for oocyte retrieval, the aspiration of oocyte is a crucial step of ART. In particular, the aspiration pressure used for oocyte retrieval can affect the integrity of the oocyte cumulus complex.
IVF and IVM researchers frequently indicate the aspiration pressures they used, but this information can be misleading or not easily reproducible. In fact, the pressure at the exit of the aspiration device is different from the pressure experienced by the oocyte at the needle tip . Different factors such as needle gauge, length of needle, connecting tube gauge, length of connecting tube, size of the collection tube, and size of the vacuum reservoir in the pump play a role in determining the pressure experienced within the needle from the aspiration device.
Horne et al.  calculated the velocity of the fluid within a pickup needle, by using a model incorporating the Hagen–Poiseuille’s law and taking into account the shear stress phenomenon.
In addition, with needles having an i.d. <1.4 mm, laminar flow occurred over the range of vacuums 5–40 kPa (37.5–300 mmHg). At vacuums >50 kPa (375 mmHg) the model predicted velocities (and hence flow rates) in excess of those observed.
They also evaluated the effect of increasing the length of the needle by using a 16-gauge needle with an internal diameter of 1.2 mm coupled with a 60 cm Teflon line on velocity and flow rates at different vacuums, finding that by increasing the length of the needle, both the velocity and flow rates decreased.
As regarding the evaluation of IVF outcomes using different collection pressures, the work of Fry and collaborators examined in bovine oocytes the use of various needle sizes (17- and 20-g) and aspiration pressures (25, 50, 75, and 100 mmHg) to evaluate the impact on the quantity and quality of recovered immature oocytes .
In that study, the authors aspirated 5827 follicles from 720 ovaries with 17- and 20-gauge needles and found that the highest recovery occurred at the highest aspiration pressures with 46% at 25 mmHg and 59% at 100 mmHg. Another study by Bols et al. , which included 3000 aspirated follicles, reported a similar finding where higher pressures were associated with a higher recovery (55.5% at 50 mmHg vs. 67% at 130 mmHg).
It should be noted that in in vitro maturation (IVM) retrievals, a lower aspiration pressures with respect to the one used in IVF treatments improve the recovery as oocytes are denuded of the cumulus oophorus cells at higher pressures, and furthermore, the negative impact of increasing aspiration pressures is greater in larger-gauge needles .
Morphologically altered oocytes have been found at aspiration around 180 mmHg, which were usually used during laparoscopic oocyte retrieval . Aspiration pressure higher than 180 mmHg was related with oocyte damage and poor embryogenesis . On the contrary, lower aspiration pressures between 90 and 120 mmHg have been associated with good oocyte quality and minimal damage .
Recommended pressure values
Tube set length, cm
Recommended vacuum, mmHg