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One MT's role in an IVF program.

One MT's role in an IVF program

I love my job. An in vitro fertilization (IVF) laboratory is an unconventional workplace for a medical technologist, but the work is both challenging and rewarding. Best of all, I'm free of the stress typically associated with medical technology.

Ours is not a Stat situation. No one dies from not being pregnant. Although we all feel sad when a pregnancy test is negative after all our efforts, basically this is a happy environment. Many people who never could have hoped to conceive a few years ago can now become parents, with our help.

My career path has always been somewhat less than traditional. I started as a diener and moved on to a job as sperm bank coordinator in an andrology lab ("A New Twist to the Traditional Career Choice," MLO, June 1987). This led to a slot as a research specialist when my former hospital had an opening in its IVF laboratory. The opportunity to work at a close-knit private practice in the suburbs brought me to an IVF lab outside Philadelphia.

Endocrine Histology is a large private infertility practice. The doctors' offices are in a historic manor home. Our lab operates out of the original carriage house. When I signed on in the spring of 1988, the IVF program had been in the planning stages for almost a year and had 90 couples on the waiting list.

Those of us in the lab call ourselves embryologists. I am the only MT; the other two have degrees in biology. One of my colleagues previously set up the IVF laboratory at a large Philadelphia medical center. The other was formerly a researcher in reproductive biology. The rest of the IVF team consists of two nurses, a physician's assistant, and three surgeons on rotation. The senior partner in the endocrinology practice reviews all patient charts.

The procedures we follow are simple enough. Technologists already have most of the necessary skills and can learn the rest. It took me about two weeks of practicing on mouse embryos to master the technique. For me, the most difficult adjustments involved getting used to working under a sterile hood and learning to look under a microscope while my hands were bench. Coordination has come with practice. Now when I work with another embryologist, it's as if we have one brain and four hands.

In a nutshell, here is what I do: * Infertility workup. Both partners of a couple who wish to conceive are examined to identify any physiologic fertility problems. The workup for the woman includes a physical exam, cultures, and extensive blood work. If indicated, she may have such specialized tests as a hysterosalpingogram to help diagnose blocked fallopian tubes or an endometrial biopsy to assess the likelihood that the endometrial lining will be receptive to an embryo. For the man, we do cultures, blood work, and extensive sperm studies.

When the test results are in, couples deemed to be good candidates for in vitro fertilization enter the program. We consider the optimum candidate to be younger than 40 years of age and having a known cause of infertility. The ideal patient has only a tubal problem. The eggs and sperm are fine; we simply have to "introduce" them.

There are many exceptions, of course. We have helped patients with endometriosis, ovarian failure, and male factor problems as well as women in their late 40s, whose only difficulty in conceiving was age. * Drug therapy. Once accepted into the IVF program, the women undergo drug treatment. Patients typically receive Lupron (leuprolide acetate), which suppresses their hormonal system, and Pergonal (menotropins for injection), which stimulates egg production. The drugs are administered at home, by intramuscular injection, usually performed by the woman's partner.

The laboratory monitors estradiol levels until a plateau is reached that indicates the patient is ready to ovulate. Ultrasound allows the IVF team to count the number of follicles likely to yield ova and to gauge the size of the follicles. The physician decides when the patient will receive human chorionic gonadotropin. Retrieval is scheduled for precisely 36 hours after that injection. * Lab preparation. The lab adjusts its work schedule according to the time of egg retrieval. Since we know approximately how many oocytes or eggs to expect, we set up an organ culture dish containing growth media for each. The dishes are placed in the incubator to give the media time to equilibrate. We wipe the lab down with alcohol, make up the patient's chart, and don scrubs, cap, mask, and booties. * Oocyte retrieval. The patient arrives one hour before the procedure and signs consent forms. A nurse starts an IV containing a relaxant. The surgeon cleanses the patient's vaginal area, administers pain medication, and places an ultrasound-guided probe inside the vagina. Ultrasound locates the ripened follicles. The surgeon punctures the vagina with the needle attached to the probe and pierces each follicle. After aspirating the follicular content into a sterile tube, the surgeon flushes each follicle in case the egg is stuck to the wall. This procedure is repeated for the other ovary.

A pass-through window links the laboratory with the outpatient surgery room. The search for the egg begins immediately after the surgical assistant has handed a test tube into the lab. We pour the contents into a Petri dish and take a look, using dissecting microscope objectives.

Each egg is removed with a Pasteur pipet, placed in the well of the culture dish, and returned to the incubator, set at 37 C and supplemented with 5 per cent [CO.sub.2] - conditions that mimic those of the human body. The process continues until all usable follicles have been ruptured. Our surgeons have harvested as few as two eggs and as many as 51.

After grading the eggs to see which oocytes are most likely to become fertilized, the other embryologists and I determine the optimum time to inseminate. We rate the eggs by size and appearance in four grades:

1. Immature eggs do best when allowed to mature in the dish for an additional 24 hours before insemination.

2. Mature eggs are inseminated four to six hours after retrieval.

3. Post-mature eggs need to be inseminated earlier and typically remain in culture for only two to three hours before insemination.

4. Atretic eggs are older and usually incapable of being fertilized. These eggs often come from the patient's dominant follicle - which, under normal circumstances, would have yielded the ovum in that month's menstrual cycle.

Once we know which eggs to inseminate and when, we consult with the male partner and set up a schedule for semen specimen collection. We prefer to use fresh semen rather than collecting it beforehand and freezing it. We review the semen analysis to determine whether any special procedures might enhance the chances of successful fertilization. We might place two eggs in the same dish, for example, or remove some of the egg's corona to facilitate penetration. * Insemination. The timing of insemination, which varies with the grade assigned to the egg, is usually done four hours after retrieval. Because many of these eggs are harvested before the time ovulation would have occurred naturally, they need a few hours to catch up to the usual level of maturity when ova and sperm meet in vivo.

After the partner has delivered the semen specimen to the lab, we let it liquefy for about 30 minutes and analyze sperm count and motility. In a procedure called sperm washing, we add protein-supplemented media to the centrifuge tube, mix thoroughly, and spin it down. The next step is to take off all the supernatants to remove any debris - such as white blood cells and antibodies - from the seminal plasma. The final result of this process is a very small pellet of sperm at the bottom of the tube.

The embryologist overlays a proportionate amount of media on the pellet. The initial sperm count and motility determine how much media can be used - the higher the count, the more media added. The tube is placed in the incubator, where it rests at a 45-degree angle for one hour. During this time, the hardier sperm swim out of the pellet and into the upper portion of the fresh media. The theory is that the swimmers are the sperm most likely to penetrate the oocyte.

After one hour, the embryologist removes the upper portion from the tube, leaving the dead sperm and any other debris behind, and analyzes the sperm count, motility, and progression. The results help us calculate how many sperm to add, typically 50,000 to 100,000 per egg. We dispense the sperm with an automatic pipettor, depositing the contents next to the oocyte. Then we put the dish back in the incubator, where it will remain overnight. * Establishing fertilization. After 16 to 20 hours of incubation have elapsed, we examine each egg for signs of fertilization. This is done by gently drawing the oocyte into a thin micropipet to shear off a few of the nurturing corona cells from the outer membrane. The tiny window that results makes it possible to see the pronuclei, one from the sperm and one from the egg, exchanging DNA if fertilization has occurred (Figure I). It's an amazing sight. Fertilized eggs are placed in fresh media and returned to the incubator.

Occasionally none of the eggs is fertilized; at other times, more than one sperm enters a single egg. Unfertilized eggs are re-inseminated with fresh sperm for a second attempt. We don't try a third time, however, because by then unfertilized eggs have begun to degenerate. If we suspect a problem with sperm quality, we discuss with the patients the option of using donor semen. * Embryo transfer. About 48 hours after harvesting, we check the resulting embryos for cleaving and look at re-inseminated eggs for signs of fertilization. If everything is progressing normally, we should have an assortment of two-to eight-cell embryos (Figure II). While the patient is being prepped for the nonsurgical catheter insertion, we classify the embryos. We look for even, symmetrical ones with no extracytoplasmic blebs.

We select three to five embryos, if they are available, for transfer. The likelihood of triplets is slim; there is a 20 per cent possibility of twins. First-timers usually ask us to transfer as many embryos as we can. For some reason, five is the optimum number. If we try to transfer six or seven, the patient won't conceive at all. Why the body accepts one viable embryo and rejects another remains a mystery. Researchers are investigating the possibility of receptor points in the uterus.

The patient, with her partner standing by her side, is placed in the standard dorsal lithotomy (gynecologic) position. In the lab, we load the slender transfer catheter for the physician. We start with a little media, create an air space to act as a "pusher," pick up a small amount of media containing embryos, and add more air.

The physician, with the speculum in place, attaches the catheter to a tuberculin syringe and gently expels the liquid into the uterus. The fluid amounts to no more than a single drop (50 [MU]l). If we draw too much media into the catheter, everything drips back out. The patient, who has been receiving progesterone to prime the endometrium, is more likely to accept the embryos if the volume of media and air is restricted.

To make sure the embryos enter the uterus, we flush the catheter with media and inspect the contents under a microscope. Sometimes the catheter tip hits the uterus and picks up a tissue plug. If we find that the embryos are still in the catheter, we reload another sterile catheter and the physician repeats the procedure. The patient reclines on the table for an hour or so and returns home with instructions to take it easy and maintain the progesterone regimen.

Twelve days later, she returns for a pregnancy test. If it is positive - and almost 20 per cent are - we monitor her progesterone levels every other day and the physician makes any necessary adjustments. (Most women receive intramuscular progesterone initially, switching to oral therapy for the remainder of the first trimester.) When high levels remain constant, the physician begins to wean the patient from the drug. Many women prone to habitual abortion, however, continue to take progesterone therapy throughout the pregnancy.

Patients visit the lab on alternate days so that we can track their [BETA]-hCG levels. A sonogram taken during the fourth week confirms the pregnancy. As the physician counts the sacs, we obtain our first indication of how many implants "took."

The six-week visit is usually the prospective parents' last to us. The IVF team checks for a heartbeat on ultrasound to rule out fetal demise. If all is well, we happily transfer the care of our parents-to-be to an obstetrician. (The lab continues to monitor progesterone levels through the first trimester.) * Cryopreservation. Unused embryos are frozen in a rate-controlled alcohol bath and stored in a carefully monitored liquid nitrogen tank (Figure III). If a patient fails to become pregnant after the initial transfer, she resumes Pergonal therapy. This regimen helps the IVF team pinpoint ovulation, determine the optimum time for another attempt, and allow the embryo sufficient time to thaw. Our first set of twins from frozen embryos recently celebrated their first birthday. Another will appear this fall, bringing our total of "frozen babies" to nine. We are very pleased about this high success rate.

Patients must be responsible for their embryos. Any that are not transplanted must be picked up or relinquished to our donor program (see "A new kind of sharing," page 64). * Workload. The in vitro fertilization program treats 25 to 30 couples each month. Each spends at least four months with us, from the initial infertility workup through the first-trimester follow-up, and considerably longer if pregnancy does not occur right away.

While the laboratory's official hours are 8:30 a.m. to 5 p.m., we have to remain flexible. That's a necessity when the workload depends on precise hormone levels. Once or twice a year we have to do a retrieval at 9 at night, usually because the patient forgot to take her hCG on time. Eggs must be harvested exactly 36 hours after that injection.

On weekends, a surgeon, a nurse, and an embryologist take call. We may come in for a few hours or not at all. Sometimes the Saturday or Sunday schedule is so busy that it seems like any other workday. Team members receive two compensatory days after a weekend on call.

As one might imagine, our laboratory doesn't have "normal" workdays. A typical workday begins with two retrievals. If one involves a donor and recipient, we subsequently inseminate three sets of eggs with sperm from three partners.

Day-to-day lab work includes corona removal, fertility checks, are re-insemination of eggs harvested the day before. The day's work is usually back in the incubator by 1 or 2 p.m., giving us time to clean the lab, catch up on paperwork, enter results in the computer, and prepare media for the next day.

A busy day involves three or four retrievals, with numerous eggs to inseminate, several corona removals, and multiple transfers. Balancing all these patients can be hectic, given the closely defined time frames for each procedure. Nevertheless, we usually have 24 hours' notice before such a day takes place. When the charts tell us that several patients are ready to go, comp time and days off are forgotten. * Success rate. Our success rate is comparable to that reported in the literature. The likelihood of pregnancy, as defined by a subsequent rise in [BETA]-hCG, is 15 to 17 per cent after each embryo transfer. This rate remains the same regardless of the number of transfers performed. In general, patients who conceive by this method do so by the fifth try.

The laboratory and patients have somewhat different ideas of success. We consider the procedure successful after at least two (and preferably three) rises in beta. To prospective parents, success is nothing less than having a baby in their arms, or at least seeing the heartbeat on ultrasound. Unfortunately, about 15 per cent of our patients who achieve a chemical pregnancy will not show a viable fetus on the follow-up ultrasound. The probability of sustaining a pregnancy and delivering a baby is about 13 or 14 per cent per IVF cycle. * Quality control. Good QC is critical. We follow a sophisticated system of specimen identification to prevent mixups. All specimens are color coded and identified by number and patient name.

Every piece of equipment contains an alarm that automatically dials the designated embryologist if a warning sounds. The crisis ranges are set conservatively to insure ample time to correct the problem. I was once called into the lab at 1 a.m., for example, because of a "low" liquid nitrogen level in the cryo freezer. The problem turned out to be a faulty wire; the freezer was fine. Given the potential consequences, I didn't mind the late-night trip. * Maintenance. Routine equipment maintenance is a challenge. There are no predictable slow periods to use for such activities, yet malfunctioning apparatus would destroy our work. Finding the right time to zero the incubators is particularly difficult because they always contain eggs for the next day. Essentially we do what we can, when we can. We might collect all the glassware we'll need, for example, and bleach the water system for a couple of days.

Since we must constantly work around our patients, the only solution is a semi-annual closedown. Most IVF laboratories do this. We give patients plenty of notice and shut down for two weeks in June and again in December. At these times we give the incubators a thorough cleaning, draining the jackets and getting the lab in tip-top shape for the next six months. We close the lab when the staff attends an out-of-town meeting, too (see "Opportunities for travel," above).

Working in an IVF lab is an exciting career move. Others agree. We recently trained two new embryologists, both with biology backgrounds. One works with me in Pennsylvania and the other is at our second full-service IVF program, which opened this past summer in New Jersey.

A lab like ours can be a great option for technologists. This specialty certainly needs them. The work is far from routine. I'm truly part of a team. Patient contact is direct and intense; we all celebrate when a fertilization attempt succeeds. Even when it fails, the patients know we've done our best.

I have learned not to dwell on the failures. Instead, I enjoy glancing at the photos of our 45 smiling babies, most of whom would not have been born without us. I can hardly wait to see pictures of about 35 more babies due the next few months and the many others that will follow.

How delightful it is to have found a niche in medical technology that reminds me every day how much my efforts and expertise are appreciated.

A new kind of sharing

Our goal is to help each couple make a baby. When the odds seem slim, clients often donate eggs or embryos to each other. We also maintain a roster of sperm donors who are not our patients.

Couples who become pregnant after the first or second transfer tend to donate the embryos they didn't need. All our donor programs are completely anonymous. We maintain comprehensive medical histories on donors, along with notations on their appearance. Biologic and gestational parents are matched in terms of physical characteristics and, if possible, blood type.

I am proud of our unique donor-recipient program, which pairs women in ovarian failure with those who cannot afford the cost of the procedure. Recipients pay all the donor's medical expenses, from prescription medications through embryo transfer. In return, the donor shares the eggs subsequently harvested. The partners provide the sperm for their eggs, and the recipient serves as her own surrogate mother.

A recent donor, for example, produced 51 eggs. One couple got 26 of them, of which 18 were ultimately fertilized. The other couple received 25 eggs, of which all but one were fertilized.

In grading the eggs, we take great pains to distribute them evenly so that both sets of prospective parents get an equal number of "good" ones. With luck, these couples will become parents within a year.

Opportunities for travel

While other labs seldom allow laboratorians to wander far from the bench, ours encourages forays into the field. We attend American Fertility Society meetings all over the country and travel abroad. Two of us attended an international conference in Israel last year to learn new fertilization techniques. The third embryologist traveled to Italy for a conference on sperm problems. Last fall, I attended a fertility society meeting in Morocco. I presented a seminar for physicians on cryopreservation in Casablanca and went to a meeting in Marrakesh.

In May 1990 I spent a week at the University of Wisconsin mastering micromanipulation. I learned how to make a minute incision in an egg and inject a single sperm to achieve fertilization manually. We are excited about what this new procedure will mean for patients who previously had to settle for donor sperm.

PHOTO : Figure I Figure II From zygote to embryo In the zygote stage (Figure I), two pronuclei exchange genetic material in the center of the oocyte. After about 48 hours, a six-cell embryo (Figure II) is ready to be returned to the patient.

PHOTO : Figure III Cryopreservation Surplus sperm and embryos can be maintained indefinitely in a liquid nitrogen storage tank set at - 196 C.
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Title Annotation:includes information on sharing eggs & embryos, and on opportunities for travel; medical technologist; in vitro fertilization
Author:Goldsmith, Gail
Publication:Medical Laboratory Observer
Date:Nov 1, 1990
Words:3620
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