Abstract
The oocyte retrieval procedure has undergone a substantial evolution over almost forty years moving from the abdominal to vaginal approach. Ultrasound- guided oocyte retrieval is performed now as a routine procedure worldwide.
1 Oocyte Retrieval
The oocyte retrieval procedure has undergone a substantial evolution over almost forty years moving from the abdominal to vaginal approach. Ultrasound- guided oocyte retrieval is performed now as a routine procedure worldwide.
1.1 Pre-retrieval Biological Processes
1.1.1 Background Characteristics of Patient
The purpose of controlled ovarian stimulation (COS) is to achieve multi-follicular development culminating in a collection of a number of fertilisable oocytes. However, this may not always be possible, especially in patients with poor ovarian reserve. Patient background characteristics such as age, antral follicle count, anti-Mullerian hormone (AMH), body mass index (BMI) and previous response to COS are important not only for choosing an appropriate individually tailored stimulation protocol but also for counselling regarding the expected outcome of oocyte retrieval.
It is important to assess accessibility to the ovaries in advance of stimulation. If a patient had previous abdominal or pelvic surgeries which could compromise transvaginal access to the ovaries, an alternative route for oocyte retrieval (e.g. transabdominal retrieval) and the associated risks should be considered and discussed with the patient.
1.1.2 Oocyte Triggering Medication
Stimuation protocols are considered in detail in Chapter 17. Key considerations for triggering pre-oocyte collection are when to trigger in relationship to follicular development, the type and dose of trigger as well as the interval between trigger and oocyte retrieval.
1.2 Retrieval
1.2.1 Technical Aspects
1.2.1.1 Route of Egg Retrieval
The first in vitro fertilisation (IVF) baby was derived from laparoscopically aided oocyte retrieval scheduled in the natural menstrual cycle. Laparoscopic oocyte retrieval, however, yielded oocytes from only one third of follicles [1]. The next breakthrough in oocyte retrieval was almost a decade later, when the transabdominal ultrasound-guided technique was developed. Even though the yield was similar to the laparoscopic method, the transabdominal route offered easier access and a lower risk of complications. Whilst the transabdominal ultrasound transducer was used, access to the ovaries was attempted transvesically, per-urethrally and finally transvaginally [1]. The transvaginal ultrasound-guided approach is currently the main route used for oocyte retrieval.
1.2.1.2 The Physics of Oocyte Retrieval
Length and Size of Needle, and Aspiration Pressure
During the dawn of IVF, a needle and syringe were used to provide suction manually to retrieve follicular fluid and oocytes from the ovary. Subsequently, aspiration devices were developed to provide continuous suction. The vacuum used did not usually exceed 120 mmHg. A foot operated ‘on-off’ valve at 200 mmHg was utilised. Currently the aspiration pressure used by the majority of IVF unit ranges between 120 and 200 mmHg.
The evaluation of the physics of oocyte retrieval in relation to aspiration pressure, length and gauge of needle and velocity of follicular fluid aspiration has been studied mostly in animal models. It has been suggested that oocytes may be damaged during the process of collection by turbulence and velocity of the follicular fluid [2]. The authors argued that a large vacuum gradient could potentially damage the oocytes and that a high velocity of laminar flow or turbulent flow would increase the chance of stripping the cumulus cells off the oocyte. Changes in vacuum pressure or needle gauge or length would influence the flow rate in the system. Raised pressure increases the volume of flow and velocity of the fluid much more rapidly in a larger gauge needle. The size of the bevel can also exacerbate fluid turbulence. Turbulent flow occurs over a longer distance in long bevel compared to short bevel needles. The turbulence on entry into the needle exerts randomly directed forces on the cumulus oocyte complex, which may overcome the adherence of the cumulus cells to the oocyte. Early studies suggest that oocyte pick up is better when larger gauge needles are used. Needles of 14= and 16-gauge were considered the best choices for oocyte recovery; however, a smaller needle is likely to result in less tissue trauma and thus less bleeding and balance needs to be achieved.
An optimal combination of the above variables has been shown to improve the oocyte retrieval rate in animals. How relevant these findings are to human practice is difficult to ascertain since no similar studies have been carried out.
Temperature, pH and Osmolarity Control
It is well-established that fluctuations in environmental factors such as temperature, pH and osmolarity can influence oocyte quality. Hence, effort should be put into minimising unwanted fluctuation during the transportation period of oocytes from follicles to lab incubator dish via the oocyte collecting system. For example, the oocyte is known to be very sensitive to reduced temperature [3], pH or osmotic stress [4]. The maintenance of 37 °C is important during IVF manipulations therefore all equipment (test tube heater, table incubators, plates and media that come into direct contact with oocyte during retrieval) should be pre-warmed. Heating devices must be regularly calibrated and monitored. Prevention of harmful pH fluctuation can be achieved by using appropriate buffers. The team involved in the egg retrieval procedure should ensure that all equipment and media are ready to use before commencing the retrieval procedure.
1.2.2 Pain-Relief
Effective analgesia is an essential part of any surgical procedure. The purpose is not only to improve the patient’s experience and eliminate the perception of pain but also to help the surgeon to perform the procedure safely and efficiently. Ideally, the chosen method of analgesia should be short-acting with a good safety profile and minimal impact on gametes. As the technique of oocyte retrieval was evolving into being minimally invasive, so was the choice of anaesthetic, starting from general anaesthesia for a laparoscopic approach to paracervical block and/or conscious sedation for a transvaginal approach.
A recent meta-analysis [5] showed that no one particular modality of conscious sedation or analgesia was superior in providing effective pain relief for transvaginal oocyte retrieval. The use of more than one method simultaneously, such as when combined with acupuncture or paracervical block, resulted in better pain relief than a single modality alone [5].
1.2.3 Variables of the Procedure
1.2.3.1 Vaginal Disinfection prior to Egg Retrieval
Early studies raised concern that if traditionally used vaginal antiseptics (e.g. povidine iodine or betadine) are trapped in tissue after cleaning, these could contaminate the follicular aspirate and damage oocytes [6]. This result was not confirmed in larger studies [7,8] where the pregnancy rates were comparable in both groups of patients with or without saline washing after betadine cleaning of vagina. These authors argue that it is unlikely that betadine found in the vagina could come into contact with oocytes while retrieving them in the closed system of the needle and tubing. No studies on vaginal preparation with chlorhexidine have been published.
1.2.3.2 Antibiotic Administration
Pelvic infection is a rare complication following transvaginal egg retrieval with an incidence ranging from 0.5% to 4% [7]. The theoretical mechanism of infection is either due to inoculation of vaginal microbiological flora into the peritoneal cavity or reactivation of pre-existing chronic infection in the pelvis. Routine use of prophylactic antibiotics during transvaginal oocyte retrieval has questionable benefit. For example, the incidence of infection in a cohort of 2,670 patients was 0.6% with no routine antibiotics [9], while in another cohort of 674 patients with routine antibiotics the incidence was 1.3% [10].
Due to the relative rarity of infection after transvaginal oocyte retrieval, a sample size of around 4,000 patients would be required to provide a meaningful answer regarding the benefit of routine administration of prophylactic antibiotics. Until such a large study is undertaken, it would be sensible to provide antibiotics cover only to patients at higher risk of infections, such as those with previous history of pelvic inflammatory disease, complex pelvic surgery and/or endometriosis.
1.2.3.3 Other Variables of Successful Oocyte Retrieval
Once the vulva and vagina are cleaned, the ultrasound probe is introduced and the ovaries are visualised and inspected. Gentle pressure on the probe usually achieves a good application of the vaginal wall to the ovary, in order to minimise the chance of visceral organs, such as bowel loops, being trapped between. The probe should be aligned in such a way that the aspiration needle enters the nearest follicle at the right angle. A gentle stabbing movement should achieve penetration of the follicle, after which aspiration of the follicular fluid can be achieved by applying suction pressure. Maintenance of suction during the entry to the follicle may prevent fluid and oocyte loss caused by the sudden increase of intra-follicular pressure during needle penetration [2]. Once the follicle has been completely drained, if pressure has been released, it is possible for negative pressure to pull an oocyte back into a follicle from the needle, particularly if the oocyte is in the last portion of fluid in the needle [2].
Some authors suggested that rotating the needle in a follicle during aspiration increases the number of oocytes obtained [11]. This potential benefit should be balanced however, against the theoretical concern of causing damage to cumulus oocyte complex if it is still attached to the follicle wall that is scraped by the needle. Furthermore, depending on the speed of the surgeon, prolonged residence of the aspirate in the dead space of the needle and tubing may lead to clotting in the needle and loss of oocytes or to exposure of oocytes to non-optimal environmental conditions.
In summary, careful needle entry into the follicle, avoidance of blockage of the needle lumen and timed application of aspiration pressure should theoretically optimise the oocyte retrieval technique [11].
1.2.4 Follicular Flushing
Follicular flushing was introduced with the purpose of increasing oocyte retrieval rates. Even though some studies suggested possible benefits of flushing with an apparent increase in retrieval rate from 40% at first aspiration to 97% after four flushes, systematic review of the literature revealed no benefits in terms of number of retrieved eggs or pregnancy rate in normal responders [1]. Many clinics do not practice routine follicle flushing except for poor responders. Even in this group of patients however, the benefit of follicular flushing is uncertain. A recent study suggested that flushing might have a detrimental effect on the pregnancy rate [12]. A theoretical problem created by flushing is inadvertently pushing the oocyte out of the follicle. The needle tip may have tracked through the posterior follicle wall or may have created a large opening in the anterior follicle wall. Fluid can easily be injected into the ovary outside of the follicle, theoretically losing an oocyte from the follicle and also leading to impaired visualisation. Creation of high-speed turbulent flow could also be harmful for the oocyte–cumulus complex.
1.3 Post-retrieval Monitoring of the Patient
A short period (1–2 hrs) of observation is generally advised to monitor the woman for possible complications. As a result of significant advances in the technique of oocyte retrieval, the risk of complications is rare. Serious complications such as intra-peritoneal bleeding, uretero-vaginal fistula and ovarian abscess have been reported very rarely and mostly in the form of case reports. Minor complications such as mild vaginal bleeding have been reported in up to 6% of women [13]. Such bleeding usually settles with vaginal pressure and requires no further intervention.
2 Embryo Transfer
The first successful embryo transfer was performed in 1891 by Heape, when a spear-headed needle was used to transport fertilised eggs into a foster uterus in rabbits. It was a further 87 years before the first successful embryo transfer had occurred in a human [14]. The main method of transfer in early animal studies was a surgical trans-fundal approach under anaesthetic. Originally, the trans-cervical approach seemed to give a much inferior pregnancy rate of 2–4% in comparison with surgical trans-myometrial (50%) in different animal species. However, refinement of the trans-cervical method soon produced similar good results and became the mainstream technique in humans. Embryo transfer is the final success-determining step in an IVF cycle and also depends on a number of variables.
2.1 Patient Preparation
2.1.1 Full Bladder
It is thought that a full bladder could straighten the utero-cervical angle during embryo transfer and facilitate entry into the uterine cavity with a soft catheter. In 2007, a systematic review [15] suggested that a full bladder could be associated with a higher pregnancy rate (OR 1.44, 95% CI 1.04–2.04). This review derived its conclusion based on two studies. The larger of the two studies was not a true randomised controlled trial since allocation to the different treatment groups was based on alternate days. A subsequent Cochrane review [16] showed no difference in pregnancy rate between full or empty bladder (OR 0.98, 95% CI 0.57–1.68) when only two small randomised controlled trials with no power calculation were included. Further research into the effect of a full bladder at the time of embryo transfer on the live birth rate after IVF is warranted.
2.1.2 Role of Uterine Relaxants
Uterine contractility during embryo transfer has been thought to be associated with lower implantation rate. There is evidence that uterine contractions in stimulated IVF cycles are increased at least six-fold in comparison with natural cycles. It was speculated that administration of pharmacological agents that reduce uterine contractility could have a positive influence on embryo implantation rates. Administration of prostaglandin synthesis inhibitors just before embryo transfer showed variable effects on pregnancy rates in two studies of moderate quality [17,18].
Data from a prospective cohort study suggested that administration of atosiban (oxytocin/vasopressin receptor antagonist) at the time of embryo transfer was associated with a higher implantation rate [19]; however, when the same group conducted a multi-center randomised controlled trial, no difference was found in live birth rate between atosiban and placebo group in a general IVF population [20]. The role of similar drugs in patients with recurrent implantation failure or with adenomyosis remains unknown.
2.1.3 Mock Embryo Transfer
The aim of embryo transfer is to deliver an embryo into optimal location in the uterine cavity causing minimal disturbance to the surrounding environment. Mock Embryo Transfer (MET) can be undertaken prior to the actual procedure in order to establish if there are any potential difficulties that could be addressed in timely manner in order to reduce uterine trauma and avoid the risk of depositing the embryo in a suboptimal location. Since uterine and cervical anatomy has a great degree of variability, MET may help to assess variables such as uterine cavity position, measurement, ease of access and the choice of catheter. However the possible disadvantage of MET is the possibility of causing irritation, trauma or increased contractility of the uterus. There are, as yet, no randomised controlled trials assessing the potential advantages of MET.
A relatively small study of 135 patients randomly allocated to MET before starting IVF showed that MET was associated with higher implantation and pregnancy rate [21]. The timing of MET, whether before starting controlled ovarian stimulation, at the time of egg collection or just before the actual embryo transfer, did not seem to influence IVF outcome [21].
2.2 Variables during Embryo Transfer
2.2.1 Technical Aspects
Each step of the embryo transfer procedure should be aimed at minimisation of patient discomfort or pain, since it is plausible that pain caused by pelvic examination may cause undue uterine contractions that could increase the chance of embryo expulsion. In 6 to 16% of embryo transfers, the embryo could be found in the fluid at the external os after completion of the procedure [22]. The choice of the right speculum size and gentle speculum introduction into the vagina could help to minimise discomfort and patient distress. Excessive opening of vaginal speculum valves could alter the cervical-uterine angle and cause pain.
One randomised controlled trial suggested that mechanical closure of the cervix with vaginal speculum valves after introducing the embryo transfer catheter can be associated with increase in the clinical pregnancy rate [23]. These results were not substantiated by a subsequent quasi-randomised study, which did not demonstrate any significant benefit in mechanical closure of cervix during embryo transfer [24].
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