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Cryopreservation
Cryopreservation is a technique used by IRMS
to freeze and then thaw sperm and embryos for use in
vitro fertilization (IVF) cycles. With the availability of frozen embryos,
a woman doesn’t need to undergo stimulation by fertility drugs in order to
have an embryo
transfer during an infertility treatment cycle. When sperm is collected during microsurgery or by
other means, and frozen for a subsequent IVF cycle, additional surgery may be
avoided.
Embryo Cryopreservation
During the standard course of infertility treatment,
hormones are used to stimulate the development of multiple eggs. After these
eggs are retrieved and fertilized in the laboratory, there may be more embryos
created than can reasonably be transferred to the woman’s uterus. When these
"extra" embryos are of sufficient quality, they may be cryopreserved
(frozen) so that they may be transferred at a future date. Embryo
cryopreservation, which is possible in approximately 25% of IVF cycles,
provides the opportunity to have an additional embryo transfer without the
inconvenience and expense of a full IVF cycle.
Embryos may be frozen at any stage between day 1 and day 6 after egg
retrieval. However, not all embryos are candidates for cryopreservation.
Unfortunately, some may be damaged by cryopreservation and experience has
shown that high quality embryos are far more likely than others to survive and
be capable of further development after freezing. Embryos that divide slowly,
or are irregular in other ways, do not fare well after cryopreservation and,
therefore, are not frozen. Because problems such as these are more prevalent in older women,
cryopreservation usually is not recommended for patients age 40 and older.
After placing the embryos in a cryoprotectant solution, they are frozen in a
computer-controlled device designed specifically for embryo cryopreservation.
Long-term embryo storage is in liquid nitrogen, at a temperature of –320oF
(-196oC).
Cryopreserved embryos are replaced during either a natural menstrual cycle or
a hormonally controlled cycle. Considerable care is taken to minimize the
possibility of damage caused by cryopreservation. Depending on the embryo
stage at the time of freezing, between 60 and 90 % survive the freeze/thaw
process. The pregnancy rate after transfer of these embryos is similar to that
of fresh embryos.
Extended periods of storage in liquid nitrogen have no apparent affect on
their viability. Embryos thawed after several years of cryostorage fare as
well as those frozen for only one or two months. Since 1983, the
cryopreservation procedure has resulted in the birth of thousands of babies
worldwide. With this extensive experience, there have been no reports of any
increase in birth defects as a result of cryopreservation.
Oocyte Cryopreservation
While embryos are reliably and routinely
cryopreserved, it is not currently possible to efficiently freeze human
eggs (oocyte cryopreservation). There have been isolated reports of pregnancies that resulted from
frozen eggs, but the few IVF programs attempting this procedure have
experienced very low success rates. Research underway at IRMS and at other
research centers around the world, is designed to define the conditions that
are necessary for safe and reliable egg freezing. Scientists at IRMS
have made significant breakthroughs in oocyte cryopreservation, however, this is not
a service we offer to our patients at this time.
Articles with additional information on Egg
and Embryo Freezing can be found in our research
articles page.
Single Sperm Freezing
The IRMS laboratory is responsible for the
development of single
sperm freezing, an experimental technique that has contributed
significantly to the success of IVF used as treatment for severe male factor
infertility. Before its introduction in 1996, there was no effective procedure
for the efficient cryopreservation (freezing) and post-thaw recovery of
individual or small groups of human sperm. This technique is still
experimental and under supervision of the Internal Review Board of Saint
Barnabas Medical Center.
Because of their microscopic size, sperm are easily lost during semen specimen
processing, conventional cyropreservation and thawing. As a result, single
sperm freezing provides an option in cases where the male’s sperm count is
less than 1,000 and the post-thaw recovery rate without the procedure is
virtually zero.
In developing single sperm freezing, IRMS Laboratory Director Jacques Cohen,
Ph.D. looked to the developing human embryo, which is protected by a thin,
porous membrane called the zona
pellucida. A hollow sphere remains when cellular material is removed from
the zona. Because it can be seen and handled microscopically both before and
after cyropreservation, it is an ideal capsule for freezing individual, or
small groups of sperm cells. A hamster zona also can be used because of their
large size and minimal affinity to bind with human sperm cells.
Single sperm freezing is especially effective in cases involving "severe
male factor" infertility, when a man's sperm count falls below the
normal parameters – two milliliters of semen volume, 20 million sperm cells
per milliliter and 25% motility (movement). When the ejaculate has zero or
fewer than 1,000 sperm cells, surgical procedures - microsurgical
epididymal sperm aspiration (MESA) or testicular
sperm aspiration (TESA) – may be necessary to retrieve enough sperm for
IVF.
Frequently, the search for sperm within tissue samples collected during these
procedures can take between two and seven hours in the laboratory. In some
cases, the optimum time for injecting the oocytes retrieved during an IVF
cycle passes before viable sperm cells are isolated. If MESA and TESA produce
enough sperm for more than one IVF cycle, they can be available without
further surgery as a result of single sperm freezing. Also, because they can
be thawed immediately after oocyte retrieval, the recovered sperm are
available to be used in conjunction with intracytoplasmic
sperm injection (ICSI) while the eggs still have the potential for
fertilization.
Single Sperm Freezing Procedure
A. Secure Zona
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B. Evacuation
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C. Empty Zona
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D. Sperm
Injection
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E. Zona
Cryopreservaton
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F. Zona Thaw:
Sperm Recovery
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Each egg is held with a small holding pipette
(a long, thin glass tube). A needle is used to first make a tiny hole in the
zona and then to suction out the cellular material. The available sperm are
isolated and separated from other cellular debris, added to a very small
amount of culture medium and then deposited in a single layer in a petri dish.
Using a microscope, the andrologist isolates and aspirates each sperm into a
small pipette, washing it and depositing it into a clean droplet of medium.
Finally, they are injected into the empty, jelly-like zona, where they are
trapped and frozen using standard freezing technology.
Single Sperm Freezing Risks
As with other forms of cryopreservation, frozen sperm can be destroyed if
there are unforeseen technical or mechanical failures in the laboratory. Also,
it is still possible for sperm to be lost or they may not survive the
freeze-thaw process. Other physicians and scientists have criticized this
method as the hamster zona, they argue, may be a vector for transmitting
desease. This argument has been presented without scientific basis or
supporting data.
Articles with additional information on
Single Sperm Freezing for Extreme Azoospermia can be found in our research
articles page.
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