Since Pacific Fertility Center came into existence in November of 1999, we have been offering genetic pre-screening of IVF embryos for couples with recurrent miscar- riage, repeated IVF implantation failure and sex selection for family balancing. For most of the last decade, a technology known as Fluorescent In-Situ Hybridization, or FISH has been used to screen embryos. FISH is employed to probe a cell removed from a Day 3 embryo to determine the chromosomal makeup for anywhere from three to twelve of the cell’s 23 pairs of chromosomes. With time, we, as well as everyone else in the reproductive genetic world, came to realize the serious limitations of this technology.

In the last 2-3 years, as the Human Genome Project has been completed and as more DNA-related biotechnologies have emerged to evaluate human genes, these methods are being utilized to analyze human embryos. The technology now available—the ability to analyze large numbers of genetic locations on each human chromosome, and quantify that genetic material, with the previously well-established techniques to amplify a single cell’s genetic material up to hundreds of thousands of copies—has allowed PGS to take a quantum leap forward. It is now possible to more accurately analyze all 23 chromosome pairs from a single embryo; not only to determine if the correct number of copies of each chromosome is present, but also to look at single gene mutations.

At the end of 2009, Pacific Fertility Center began working with a new biotech company called Gene Security Network, located in Redwood City (genesecurity.net). This company uses gene microarray technology to analyze amplified DNA from a single cell.

It then uses microchips to analyze 30,000 genetic loci in a quantitative manner. In addition, their unique technology allows us to compare the analysis of the embryos’ cells to the parent’s chromosomes to ensure that all the genes are being properly analyzed. It does appear that the error problems that plagued FISH technology have been overcome with this new, more sophisticated, method.

In October of 2009, Dr. Conaghan and I were invited to tour the GSN laboratory and see the technology in action. We met with David Johnson, the lead scientist at GSN, who explained the cell process; from the amplification of the DNA, to arranging the chromosomes on chips, to DNA analysis, to synthesizing the data generated with the parental genetic data to come up with a full analysis of that cell’s genome. In order to process the cells between the day of embryo biopsy (Day 3) and receive the results on the day of embryo transfer (Day 5), their technicians work around the clock in shifts. GSN has a very cold, clean room to replicate the single cells into multiple copies. They cannot allow any outside contamination, not even from a single cell. They videotape the cell duplicating process so if any errors subsequently arise, they have a video record of what the laboratory technician did. We found this to be very impressive. We also saw how the chips were coated with DNA and analyzed. We were shown the sophisticated software that generates the final report detailing the genetic makeup of each embryo from the cells in which they originated. All in all, the tour gave us great confidence in the quality control and scientific integrity at GSN.

Even with this 21st century technology, we continue to biopsy Day 3 embryos because it provides us with a 48 hours window to send the cells to the lab and complete the analysis in time for transfer. However, we have not yet found a way around the problem of mosa- icism. GSN and microarray technology appears to have largely solved the resolution error problem but it can only tell us what is in the chromosomal make-up of the single cell. It cannot tell us whether or not that cell represents what is truly going on with the rest of the embryo. We are currently looking at the possibility of biopsying Day 5 embryos. The set back would result in having to freeze these embryos due to the time constraint in analyzing the genetic material in time for fresh transfer. With all of the innovation occurring daily in the genetics field, we hope that this puzzle will be resolved.

— Carolyn Givens, M.D.

Previous Fertility Flash articles about PGS:
2 Methods of Gaining Info Prior to Implantation
PGD & PGS: Why Genetic Counseling is a Prerequisite
The Benefits and Pitfalls of PGS

By the end of the year we will have started a new and very exciting research project in our lab. We have partnered with a company called Incept Biosystems (www.inceptbio.com) to do a clinical trial of a new embryo culture system called microfluidics.

The traditional culture dish with medium droplets under oil as described by Brinster, R.L., 1963, Exp. Cell Res., Vol. 32

This involves culturing embryos in very small volumes of culture media inside a chip specifically designed for this purpose. Tiny pumps regulate the flow of culture medium in and out of the chip without causing the embryos to move around.

The traditional vessel for embryo culture is the petri dish, where small droplets of culture medium are overlain with a highly purified mineral oil. The culture medium, regulated in much the same way as pharmaceuticals, is one of the most highly tested and expensive components of the IVF laboratory operation. We typically make droplets of medium that are in the 50-200µl size range, and the oocytes or embryos are placed in the droplets for 24-48 hours at a time. This is a static culture system where nutrients are depleted by the developing embryos and waste products (e.g. ammonia from amino acid breakdown) accumulate over time. The droplets are large enough to make sure that the supply of nutrients is more than adequate and that waste is diluted to the point of not harming the embryo in any way. The embryos are changed into fresh medium at least every 48 hours.

This system for embryo culture has been in use since human IVF began in the late 1970′s and early 1980′s. It was actually developed in the early 1960′s by a pioneer of mouse embryo culture, Dr Ralph Brinster, at the University of Pennsylvania. Some early human embryologists cultured embryos in small test tubes without the mineral oil, but nowadays, despite the age of this technique, it is very unusual to find a facility that does not use the droplets under oil method. After 45 years, perhaps it is time for a change?

A microfluidic system for embryo culture has been in development for over 5 years at the University of Michigan in Ann Arbor. Professor Gary Smith combined the talents of his graduate students in physiology with those of engineering students and came up with a device that has had outstanding results with growing mouse embryos. Professor Smith is no stranger to IVF, as he was the director of the University’s IVF Laboratory for many years and he was instrumental in designing and testing the vitrification system that we now use to preserve oocytes and embryos. He solicited venture capital to start Incept Biosystems with the intent to bring microfluidics into human IVF labs. Incept Biosystems were onsite at PFC during the last week of October to train our embryologists on the use of the system. We did several trials with mouse embryos to achieve proficiency with the system and then we will actively recruit patients to enroll in a clinical trial using the system.

The clinical trials are being run at 3 centers in the US. In addition to PFC, patients will participate at the Fertility Center of San Antonio and at Southeastern Fertility Center in Charleston, South Carolina.

A schematic of a microfluidic embryo culture device with fresh medium in blue and spent medium in red. The embryo is contained at the base of the chamber, where the blue medium ends.

Patients that are asked to participate will have to consent to the study, where their embryos will be divided into 2 groups for culture in the microfluidic device and in the traditional petri dish. The culture media will be the same for all the embryos, but half will be in a replenishing media current (microfluidics) and half will be in our traditional static culture.

Microfluidics has had impressive results with mouse embryos where it significantly increased rates of development and implantation over those for embryos grown in static culture. Cell numbers for the microfluidic embryos were almost twice as high as for traditional culture (110 vs. 65), and pregnancy rates from transferred embryos were increased by 22%. Incept Biosystems have tested the new technology extensively and have been able to obtain surplus IVF embryos donated for research for human trials. There are some nice videos on their website that showcase the equipment and procedure, and detail the mouse embryo results. Professor Smith presented the results and won the prize paper at the 2008 American Society for Reproductive Medicine (ASRM) meeting (Smith et al., 2008, Fertility and Sterility, Vol 90, pages S1-S2), and these results will soon be published in a peer reviewed journal.

We will be asking for participants to join the study, beginning in November and continuing for 2-3 months. This is a short study requiring enrollment of only 20 patients, but a larger study is planned for next year subject to favorable outcomes here. If you are interested in the study and would like more information, please ask your physician at your next visit.

Participants in a workgroup.	Practicing micromanipulation

On November 8 th and 9 th, 2008, PFC held a training course for embryologists interested in learning the techniques associated with Preimplantation Genetic Diagnosis (PGD).

The course was organized in conjunction with the Genetics and IVF Institute (GIVF) of Fairfax, Virginia and was originally scheduled to run on the Sunday only. However, due to the overwhelmingly positive response to attend the course, PFC decided to offer the course on Saturday as well. Forty seven individuals heard excellent lectures over the 2 day period.

Dr Dagan Wells from Oxford, UK and Dr Alan Thornhill from London, UK, gave talks on current and future technologies for genetic testing on embryos. Participants were then divided into 4 working groups that spent the rest of the day rotating between activities. The activities included embryo biopsy training, cell fixation training, media and solution making and PGD troubleshooting. Lauri Black, MS, CGC, a Certified Genetic Counselor at California Pacific Medical Center and Mary Sands of GIVF gave talks on genetic counseling. The course also allowed embryologists from all over the world to view the new state of the art laboratory at PFC.

Joe Conaghan, PhD, HCLD

Every year, several Pacific Fertility Center professionals participate in ASRM’s national meeting. They evaluate the research and share their findings with PFC and Fertility Flash.

Among those attending the conference from PFC were Dr. Philip Chenette and Dr. Isabelle Ryan and Peggy Orlin, MFT. Their reviews cover the following topics: Update #1: Ovarian Stimulation Techniques, Update #2: PGD and Aneuploidy Screening Techniques, Update #3: Egg Freezing, Update #4: Acupuncture, and Update #5: Men and ART.

ASRM Update #3: Egg Freezing

Oocyte cryopreservation is the storage of the female gamete, the egg, prior to fertilization. Preservation of fertility for single women that must undergo cancer therapy or surgery, or that must delay or choose to delay childbearing, and donated oocyte banking are all applications of oocyte cryopreservation. The need for this technology is clear, but reports of success with oocyte cryopreservation have been limited.

Highly successful oocyte cryopreservation is now attainable. New studies are showing pregnancy rates with oocyte cryopreservation that are equal to traditional IVF techniques.

The key to this technology is oocyte vitrification – an ultrarapid cryopreservation technique. Researchers from Atlanta described their experience with vitrification. Out of 11 patients with transfers, nine conceived, with an implantation rate of 65%.

Pregnancies after oocyte cryopreservation have developed normally. An Italian study of 105 children born after oocyte cryopreservation showed no problems. A Chicago study of the genetics of oocytes, embryos, and children born after oocyte cryopreservation was reassuring. No increase rates of aneuploidy or malformations were reported, and normal development was found in post-natal follow-up.

These results are similar to those we have previously reported from our own research at Pacific Fertility Center (see December 2007 Fertility Flash). Oocytes are now cryopreserved with high success rates. Oocyte cryopreservation technology has matured, and we look forward to providing these techniques for our patients.

Philip Chenette, MD

In late October of this year, our first patient who underwent embryo transfer with embryos created from vitrified and warmed donor oocytes has successfully delivered. The baby was born at term and appears to be perfectly healthy.

Three other pregnancies are ongoing and are expected to deliver in 2008. We congratulate our new parents and the parents-to-be who have participated in this ground breaking program.

PFC has ended enrollment of patients into this program, but expects to continue research efforts with respect to oocyte vitrification.

While it has been possible to preserve sperm for many years (the famed Dutch microscopist Anton von Leeuwenhoek allegedly cooled and then recovered sperm using snow and ice in the 17th century), reliable methods for oocyte preservation have been elusive.

We previously discussed some of the problems with oocyte freezing (see Fertility Flash, January 2005, Volume 3, Issue 1), and now report our success in overcoming some of the obstacles.

Traditionally, preservation of sperm and embryos has been achieved with the use of a technique called slow freezing. This process incubates the sperm or embryos in low concentrations of cryoprotectants (antifreeze) to draw water out of the cells. After this incubation, they are cooled very slowly to sub zero temperatures. Typically this slow freezing technology just works for cells that exist individually (such as sperm), or together in small numbers (embryos), as the water must be extracted from every cell. Tissues, which are made up of many hundreds of thousands of cells, cannot be dehydrated successfully and therefore cannot be frozen intact. Cells in the tissue can burst when the water remaining in the cells expands as it turns to ice. For example, it is not possible to freeze a whole ovary, but some success has been achieved with ovaries that were cut into tiny pieces.

Frustrated by the lack of progress with slow freezing, scientists have more recently moved towards a technology called vitrification for oocyte preservation. Vitrification, which was described in detail in September’s Fertility Flash (Volume 5, Issue 8 ) works by using higher concentrations of cryoprotectants and much faster cooling rates. Cells are typically cooled in tiny straws (see article heading). This process allows us to achieve cooling rates of several thousand degrees per minute.

When vitrification straws and cryoprotectants were first approved by the FDA for human embryos, PFC began the process of adapting the technology to oocytes. Our embryologists attended training courses and became proficient with the technology by practicing on mouse and hamster oocytes and embryos. Even though we have been handling oocytes and embryos for many years, this technology provided many new challenges, mainly due to the tiny size of the straws and the speed at which the cells had to be cooled. Once we became proficient with the procedure, we began to freeze high quality oocytes from donors who had proven fertility. In this way, we knew that if the procedure did not work, it would be the vitrification technology and not the oocytes that were to blame. In addition, we satisfied ourselves that the technology was safe by looking at the exhaustive work by Dr. Gary Smith at the University of Michigan, which showed that vitrified/warmed oocytes were both physically and genetically normal and that the resulting pregnancies and babies were healthy.

We recruited five oocyte donors and vitrified all of their oocytes immediately after their oocyte retrieval procedures. We then offered the oocytes to individuals who were on our waiting list to accept donated embryos. Typically, these individuals were unable to get pregnant with their own oocytes or financially unable to proceed to an egg donor cycle. The availability of the vitrified oocytes was a great alternative to accepting donated embryos as it allowed couples to choose their own sperm source. Furthermore, the immediate availability of vitrified oocytes was an attractive alternative to what may be a very long wait for donated embryos.

Pacific Fertility Center had immediate success with the first recipient. We had vitrified 16 oocytes from the first donor, and for the first recipient we warmed only 7 of these. Four hours later we injected a single sperm into each of the 6 oocytes that appeared alive and healthy (1 oocyte had not come through the process successfully). The next morning, 3 of the oocytes fertilized normally. After 2 more days, we had 3 nice embryos for transfer. The positive pregnancy test 11 days later, and a singleton pregnancy confirmed by ultrasound at 7 weeks were great rewards for our efforts and thrilling news for the recipient. Our second recipient used a different donor and although her pregnancy started out well, she miscarried in the first trimester. Our disappointment over this loss was compounded when we discovered the oocytes from 2 of the donors did not survive well when warmed. In these particular donors, we recovered high numbers of oocytes (each had close to 40) and for unknown reasons their oocytes were overly sensitive to vitrification. The next three donor cycles proceeded well and resulted in pregnancies. These 3 pregnancies are all ongoing at the time of writing. We will update readers with their outcomes at a future date.

Although we were warming relatively small numbers of oocytes (typically 6 or 7), we began to have more embryos than could be safely transferred to recipients. Our first pregnancy had been achieved after transferring 3 embryos. It is more typical, however, to transfer only 1 or 2 embryos when donor oocytes are used. Even when using only 2 embryos, multiple pregnancy rates were unacceptably high. Understandably, few patients are willing to risk a decreased chance of conceiving by transferring only a single embryo. In order to avoid high multiple pregnancy rates in a typical IVF cycle, embryos are usually cultured for 5 days to determine which embryos in a cohort have the best chance of establishing a pregnancy. However, if a patient has only a few embryos, the benefits of extended culture are less, and the transfer is typically done after only 3 days growth. With our recipients of the vitrified oocytes, we began by doing 3-day transfers. Once high success rates were evident, we elected to implement day-5 transfers, in an effort to decrease high order multiples. The last 2 pregnancies both resulted from day-5 transfers of 2 embryos each, and they are both twin gestations.

In summary, we have had 7 out of 10 embryos implant after transfer (excluding the 2 failed donors with the high oocyte numbers). This implantation rate (70%) is comparable to the implantation rates that our patients have when using fresh embryos from donor oocytes.

We are moving forward cautiously with our oocyte vitrification program and hope to use the remaining oocytes soon. While these results are encouraging and have brought great joy to a small number of our patients, there are more issues to resolve before we declare complete success. The 70% success rate was obtained with the use of the highest quality oocytes from young donors who were known to be fertile and healthy. We have already seen that some oocytes are less tolerant of the procedure, as evidenced by the results from the 2 donors with high oocyte numbers. We also fully anticipate that the results for older women using their own oocytes will be worse, as they are for these same patients using a fresh IVF cycle. In fact, at this time, we do not have any idea if the oocytes from women in their 30’s will be able to tolerate vitrification.

Going forward, we will offer oocyte vitrification unconditionally to women with cancer who are likely to be left sterile by their treatment. For these women, and for others who elect to vitrify oocytes for social reasons, we will exercise great caution in our estimates of future pregnancy potential with the warmed oocytes. Until we have more data with oocytes from a variety of women, we will have no way of telling if there is any hope from anything other than donor oocytes. That data will accumulate more slowly because women who elect to preserve oocytes are not likely to be using them for some time. For now, until there is more data, we continue to believe that embryo freezing has the greatest potential for those wishing to preserve future fertility. However, for those who are single and in their late 30’s, we will be reluctant to recommend oocyte vitrification as a reliable fertility preservation method. Hopefully, they will find Mr. Right before we have objective data.

Joe Conaghan, PhD, HCLD

Fresh Embryo at Blastocyst Stage: The cells are elongated and pressed against one another. The inner and outer cells are clearly visible, as is the cavity.

Two Vitrified Embryos at Blastocyst Stage After Warming: Though their appearances differ, both embryos implanted and created viable pregnancies.

This embryo looks perfect, as if it was never frozen. The outer and inner cells are clearly visible, as is the cavity.

This embryo has rounded, more dissociated cells resulting from shrinkage during incubation in cryoprotectant, (as cells shrink they pull away from each other). The cavity is small, but visible.

The first human pregnancy from an embryo that had been frozen and thawed was achieved in Australia in 1984, 6 years after the birth of the first IVF baby in the UK. The method used to preserve that embryo is called “slow freezing” and it is still the preferred method for preserving embryos throughout the world today. Slow freezing is a reliable and established technique that has served the IVF community well for over 20 years. The procedure has been refined throughout those years and it works, with slight modifications, for freezing all embryo stages and for sperm. However, despite many years of trying, slow freezing has never worked very successfully with oocytes. Frustrated by years of failures, scientists turned to an alternative procedure called vitrification in their quest to preserve oocytes. This approach is relatively new, but appears as through it will be preferentially used for oocyte preservation as we go forward. Vitrification kits are just beginning to get FDA clearance following scientific trials, and embryologists are being trained in the use of the new technology.

The main concern during the freezing of any cell is the removal of water without actually killing the cell. Since water expands in volume as it freezes, ice formation inside a cell would cause the cell to rupture and die. Therefore, cell water is traditionally replaced with a cryoprotectant (antifreeze) prior to cooling of the cell. This is achieved by sequentially incubating the cell in increasing concentrations of cryoprotectant. The cryoprotectant draws water out of the cell and itself enters the cell, all by osmosis. Once most of the water has been removed, the cell is cooled at the very slow rate of -0.3° C/minute until it has been cooled to below -30° C and is therefore fully frozen. Thereafter, storage of frozen cells is in liquid nitrogen (-196° C), which is a simple and practical storage medium.

Vitrification still requires the use of cryoprotectants and the cell is also ultimately stored in liquid nitrogen, but the journey from the incubator (at 37° C) to the nitrogen (-196° C) is much faster. The word “vitrum” in Medieval Latin means “glass” and the process turns the cell contents to a glass like substance instead of ice. Since no ice forms, the risk of rupturing the cell is eliminated. For glass to form instead of ice, the rate of cooling must be thousands of degrees per minute instead of the 0.3 degrees/minute that we use in slow freezing. Therefore, the process is sometimes referred to as ultra-rapid freezing, although the word “freezing” is really inappropriate here since the cell is not really frozen (i.e. no ice is created).

One of the big stumbling blocks during oocyte freezing was the sheer size of the cell (the oocyte is the largest human cell by some margin) and therefore its high water content. Just getting the cell to survive, (an oocyte has only one cell), was a huge stumbling block. Studies where 50-60% of the oocytes survived were considered groundbreaking, and still today there are few studies that have done better. Vitrification as a technique had been largely ignored by the IVF community as it was technically more challenging and used much higher concentrations of cryoprotectants. Cryoprotectants were thought to be toxic to cells. Today we know that they are safe and effective and do not contribute to cell death. It is possible that cryoprotectants may have deleterious effects on cells if they are metabolized, but virtually all freezing protocols utilize them at room temperature or below, where cell metabolism is significantly slowed or stopped. So, with success rates using traditional slow freezing failing to improve, vitrification has been given serious consideration as an alternative. In the few years since its introduction, vitrification has shown promising and excellent results in clinical studies (see Oktay et al., Fertility and Sterility, 2006, Vol 86(1), pages 70-80 a comparative review of slow freezing and vitrification results with human oocytes).

Making the transition from slow freezing to vitrification has been a challenge for the IVF community. As already stated, it is a technically challenging procedure, and training of embryologists in the technique has been slow. With slow freezing, embryos are placed in relatively weak solutions of cryoprotectant for as long as 15 minutes at a time. Then, they are usually moved on through slightly stronger solutions before being placed in large straws or vials which are then loaded into a computer controlled freezer for the long journey to -30° C. The embryologist can spend about 30 minutes with a set of embryos from the time that they come out of the incubator until they go into the controlled rate freezer. After 2 or more hours, the embryos can be placed in liquid nitrogen and the process is complete.

During a vitrification procedure, where typically only one oocyte or embryo can be worked on at a time, the transition from incubator to nitrogen takes only a few minutes. The embryo is stepped through solutions containing high and then higher concentrations of cryoprotectants where it shrivels and swirls in the extremely viscous medium. In the final stage, which is measured in seconds, the embryo is placed in an extremely concentrated cryoprotectant solution and then quickly loaded up into a tiny straw that is barely larger than the embryo itself. The straw is then sealed at both ends and plunged immediately into liquid nitrogen. The straw is so fine that it freezes in an instant, an important part of the vitrification process. The loading of the straw occurs at room temperature (25º C in the IVF lab) and it is cooled to -196º C in one or two seconds, giving a cooling rate of 6000-13000º C/min. The faster the straw can be cooled, the more successful the procedure. Performing this final step too slowly or too quickly can be the difference between success and failure and therefore requires extensive training.

At Pacific Fertility Center, we have been working on vitrification for over 2 years. Our initial interest was in oocyte freezing, but we were also interested in extending the technique to be used with embryos, and in particular to blastocyst stage embryos where slow freezing has not always worked well. Slow freezing has served us well over the years for embryos being frozen 1, 2 or 3 days after an oocyte retrieval, but blastocysts (5 or 6 day old embryos) did less well. With an industry wide transition to blastocyst stage embryo transfers, we looked at vitrification as an alternative method of preservation for these precious embryos.

A blastocyst is an embryo that has developed to the stage where it is ready to implant in the uterus. Instead of having a small number of loosely associated cells characteristic of earlier embryonic stages, it has 2 defined cell populations and a fluid filled cavity (or cyst). The cells that surround the cavity will form the placenta, and the cells within the cavity will develop into the embryo proper, or fetus and some of the extraembryonic membranes, such as the yolk sac. It is these interior cells that cause trouble during freezing since they are on the inside and difficult to expose to cryoprotectant. Slow freezing relies on cryoprotectant traveling through the outer placental cells, then the cavity, and finally into the fetal cells while water travels in the opposite direction. Fully dehydrating these fetal cells has always been a challenge and an embryo where these cells do not survive freezing and thawing will not result in a viable pregnancy. And with slow freezing, embryos tend to collapse in on themselves during dehydration, making it difficult to assess survival after thawing.

After investing heavily in vitrification training and implementing a successful oocyte vitrification program, PFC began working on blastocyst vitrification in January of 2007. By March we had a program established and were delighted by how easily blastocysts seemed to tolerate the procedure. Often, blastocysts looked no different after vitrification when compared to how they looked before the procedure. This result was in stark contrast to slow freezing where blastocysts always look shriveled and deflated after coming out of the freezer. By July 2007, we had switched completely to vitrification and currently we are enjoying the successes that it is bringing to our patients and us.

Our vitrification team consists of 3 embryologists: Mariluz Branch, the team leader, with Erin Fischer and Liz Holmes. Because of the technical challenges involved, we have to be cautious with involving other embryologists. So one of the three team members must be on duty every day (our lab is open 7 days a week). I am grateful to this team for their flexibility in accommodating our needs. By the end of the year we expect to have 2 more embryologists on the team, and then the final 3 in 2008.

Vitrification has been an exciting and challenging technique which we have embraced and conquered in 2007. We look forward to the gradual elimination of slow freezing and the successes that vitrification will bring us in the future.

Joe Conaghan, PhD, HCLD

The American Society for Reproductive Medicine (ASRM) is the largest organization in the United States for medical professionals in the field of Reproductive Endocrinology and Infertility. In 2002, ASRM published medical and psychological guidelines for ovum donation. The psychological recommendations for donors are general guidelines for addressing moral, ethical and psychosocial issues that may confront ovum donors. Included are standards for what should be included in a psychosocial screening of donors and reasons to exclude donors. A few of the objective reasons for donor exclusion include known substance abuse, positive family history of heritable psychiatric disorders such as schizophrenia or bipolar disorder, or instability in donors’ lives. To determine “inclusion” I take a psychosocial history and administer a psychological test that informs me about the donor’s personality profile, including just how much they are trying to impress me – the “fake good” factor. I am also assessing motivation to donate and the donor’s “need” versus “desire for” the compensation. Stability, stress levels, and reproductive history are also part of an ASRM assessment of donors.

Although I would like to base my entire decision on objective information, much of my decision on donor acceptance must, in fact, be based on intuition. Throughout my 25 years as a therapist, I have found that my intuition is quite accurate, but it is not fool proof.

There is one major controversy in the field that may hinder a psychotherapist’s ability to screen donors. That is the hotly debated topic on compensation of donors. In August 2000, the Ethics Committee of The American Society for Reproductive Medicine concluded that there is no consensus on the precise payment that oocyte donors should receive. It was suggested, “sums of $5,000 or more require justification and sums above $10,000 go beyond what is appropriate.” Due to costs of living and the scarcity of available donors, there are significant regional variations that affect these rates.

The Society for Assisted Reproductive Technology (SART) has attempted to assist clinics and patients by creating a list of Egg Donor Agencies that have signed an agreement stating that they will abide by the Ethics Committee Guidelines governing the compensation of egg donors.

The debate centers on the fact that donors could feel undue inducement and exploitation in the process if the fee is so high as to be coercive. Part of the job of the mental health professional is to provide donors with informed consent. Might donors conceal pertinent medical information that could be important for themselves or offspring if the monetary incentive is so high? Will donors discount risks to themselves? How can the donor actually give informed consent about the medical procedure and pay attention to the risks if $$ signs are floating before their eyes?

Ethicists and some in the field of women’s health advocacy express concern “that lucrative payments are enticing young women with credit-card debt and steep tuition bills to sell eggs without seriously evaluating the risks.” Can the payment cloud someone’s judgment and can we assess that? How much is too much? Where is that line?

When I began working with PFC in 1998, we were paying first time egg donors $2,500. Nine years later, the compensation is $6,500 for a first time donor and $7,000 for any subsequent donations. Even adjusted for inflation, the payment is rising at a phenomenal rate. Competition, supply and demand govern these rises.

As part of my objective/intuitive approach to interviewing donors, I discuss money with them. What would she use the compensation for? How does she support herself? What kind and how much debt does she have? While $25,000 in student loans points to a person’s drive toward positive goals, the same amount in credit card debt speaks to me of impulsive behavior. How much have they educated themselves on ovum donation? Whom have they talked with about their desires? Do they have their own children? Are they science majors who may be more likely to view gametes as DNA and not their own children? Are their answers well thought out?

My goal is to provide the recipients of donated eggs with intelligent, healthy, and thoughtful young women who understand the implications of ovum donation both for themselves and for the recipients. Donors who are motivated by the “need” for money are more likely to provide inaccurate information on their applications, or leave out information that could be pertinent to the recipients or their offspring. It is my job and the job of the Egg Donor Agency to determine motivation.

Although impossible to attain, we would all like donors to be motivated altruistically. We may diminish altruism by making the donation about eggs for money. However, I believe we can increase altruism by helping the donors to have a greater understanding of who the recipients are and what their struggles have been.

Peggy Orlin, MFT

Ms. Orlin served as 2006-7 chair of ASRM’s Executive Committee of the Mental Health Professional Group, is a member of Resolve’s National Mental Health Advisory Board. She co-teaches PFC’s Mind/Body workshops.

To register for the September 8th Mind/Body@PFC Workshop, please phone 415-834-3095.

The American Society for Reproductive Medicine’s (ASRM) annual meeting was held in New Orleans. It is the largest meeting for reproductive medicine specialists and scientists in the world. From our practice, Dr.s Givens, Schriock and Conaghan attended, as well as embryologists Jean Popwell, PhD and Jennifer Andres. Also, PFC nurse Allison Chamberlaine and PFC’s Marriage and Family Therapist Peggy Orlin attended. In addition, the genetics counselor with whom we work closely, Lauri Black from California Pacific Medical Center, was an attendee and active participant.

PFC’s embryologists attending ASRM’s research poster session Jean Popwell, PhD (left) and Jennifer Andres (right).

Single-Embryo Transfer: Minimizing Risks & Maximizing Outcomes
Dr. Givens attended a post-graduate course entitled “Moving Toward Single-Embryo Transfer: Minimizing Risks and Maximizing Outcomes.” This two-day course dealt with a pressing issue in assisted reproduction: the high incidence of multiple gestations. With the ever-increasing success of in vitro fertilization and the significant improvement in embryo implantation rates, the incidence of twin and higher-order pregnancies has risen dramatically in this country. Many countries now regulate the maximum number of embryos that can be transferred into the uterus at one time. The course topics included a summary of optimal medication protocols, several lectures on pre-cycle evaluation and testing and embryo transfer techniques.

Oocyte Freezing, PGS & Blastocyst Embryo Transfers
On the laboratory side, there were several talks on evaluation of eggs and embryo selection techniques, embryo freezing technology, including a debate about the usefulness of pre-implantation genetic screening (chromosome analysis of embryos) embryo selection. The combination was a fascinating mixture of new ideas, refinements in current technology, as well as a welcome opportunity to network and discuss with others the latest developments in reproductive science. Topping the list of presentations in New Orleans were those concerning the continuing refinements in oocyte freezing technologies, the more selective use of preimplantation genetic testing and the ongoing scrutiny of blastocyst stage embryo transfers.

Slow-freeze vs. Vitrification
The traditional slow-freeze technology used so successfully with embryos for many years, has essentially stalled with oocyte freezing. It appears the slow-freeze technology has finally met its successor: a process called vitrification. Slow freezing has had very limited success with oocytes due to their large size, high water content and their extreme sensitivity to cryoprotective chemicals and to changes in temperature and pH.

Vitrification, a technology that cools cells so rapidly that ice does not form, has been such a success for oocyte freezing that many labs are now abandoning slow freezing altogether. Here at PFC, we have been developing protocols for oocyte vitrification throughout 2006 and are actively working on blastocyst vitrification. It was reaffirming to see that this technology has gained wide acceptance, and is showing excellent results.

Preimplantation Genetic Screening (PGS)
While vitrification is on the rise, it was interesting to see that another technology, Preimplantation Genetic Screening (PGS), was lacking in new improvements or viable alternatives. Embryos have been screened for extra or missing chromosomes for over 15 years now, but the technology has not advanced significantly over that time. It is still possible to count only 12 chromosomes in an embryo. Although the error rate per chromosome is very low, the accumulated error rate becomes significant as we count more chromosomes. PGS was “under the microscope” in several presentations in New Orleans and it appears PFC’s limited use of genetic screening is well justified. Specifically, PGS does not improve embryo selection and pregnancy rates in younger patients. Its use is limited in older patients because there are often too few embryos available to justify testing. The patients who benefit most from PGS are the younger patients who experience recurrent miscarriages. However, unless there is evidence that previous pregnancies were genetically abnormal, PGS may provide limited benefit to this group.

Blastocyst stage embryo transfers
While younger patients (those under 35) don’t benefit from PGS, they are the patient population most likely to benefit from blastocyst transfers. Culturing embryos for 5 days to the blastocyst stage, instead of the more traditional day 3 embryo transfer, is one of the main ways in which the laboratory staff can help in selecting the “best” embryo for single embryo transfer (SET) patients. Blastocyst culture techniques are well refined now and support the commitment within the community to transfer fewer embryos at one time. Furthermore, the promise of vitrification can reassure patients that their remaining embryos can be stored indefinitely when preserved at the blastocyst stage. Several presentations showed that blastocysts which were vitrified early, before their cavity (or cyst) had expanded too much, benefited most from the technology. In more advanced blastocysts, artificial reduction of the cavity gave superior results. It may not be long before vitrification is the procedure of choice for preserving all blastocysts.

2006 ASRM guidelines for numbers of embryos to transfer
The new 2006 ASRM guidelines for numbers of embryos to transfer were presented. See Tables 1 and 2 below.

The topic of whether or not federal or state legislation should regulate the maximum number of embryos to transfer was also discussed. Many people in the general public support such legislation but those of us in the field (and most patients) are opposed to the government regulating medical practice and arbitrarily setting limits on embryo transfer. In order to forestall such legislation, it is obvious that we must decrease the number of twin gestations (the number of triplet and higher-order gestations has already dramatically decreased in the last 5-7 years). At Pacific Fertility Center we have instituted a new emphasis on single embryo transfers and expect to significantly reduce the risk of multiples and achieve our goal of “optimal” pregnancy outcomes. (See From Us to You in this issue for a discussion of our 2006 statistics and please see Conception Health in this issue for a discussion of why it is important to try to achieve single baby conceptions.

– Carolyn Givens, MD and Joe Conaghan, PhD