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IVF News Spring 2015

Contents
    • PureSperm SpeediKit, now a 10-patient kit.
    • Ovulation affects your shopping and choice of men
    • ProInsert– new version
    • Efficacy of density gradient separation on the preservation of sperm DNA integrity.
    • New collaboration
    • Nidacon products used for breeding world champions!

 

PureSperm SpeediKit, now a 10-patient kit.

PureSperm Speedikit is a quick and efficient alternative for sperm preparation using a single layer of PureSperm colloid followed by rinsing the sperm with PureSperm Wash.

The price per preparation will be lower which is beneficial for our customers.

The kit is designed for the smaller clinic. All you need is a centrifuge and pipettes. It is very easy to use even if you are not an embryologist.
Until now the kit has consisted of PureSperm colloid and PureSperm Wash for 5 patients, already dispensed in centrifuge tubes, plus semen collection tubes. We have now removed the semen collection tubes and added more single-layer/ wash tubes which results in a kit for 10 patients instead of 5.
The price per preparation will be lower which is beneficial for our customers. This new kit has the same long shelf life, 12 months. It can be stored at room temperature all the time.


Ovulation affects your shopping and choice of men

New research from The University of Texas at San Antonio (UTSA) College of Business suggests women seek more options in dating partners near ovulation when they are most fertile which may lead them to also seek a greater variety of products and services.

“Just like a fisherman casting a wide net, ovulating women seek to cast a wide net into the dating pool and expand the number of potential suitors they have to choose from”, says Kristina M. Durante, UTSA marketing assistant professor and lead investigator of the study. “And, this desire for variety in men at ovulation triggers a variety seeking mind-set that carries over into desire for variety in products.” Forthcoming in the April 2015 issue of the Journal of Consumer Research, “Playing the Field: The Effect of Fertility on Women’s Desire for Variety” provides some of the first evidence that choice behaviour in our personal relationships may influence choice behaviour in the market-place. Durante and then UTSA visiting assistant professor Ashley Rae Arsena focused their predictions on previous research that finds that ovulation canshift women’s mating psychology. Durante and Arsena conducted four studies that included 553 female participants in the U.S. between 18 and 40 years of age who were not pregnant or taking hormonal contraceptives. The studies found that women’s desire for new options in men triggered a variety seeking mind-set that led women to also desire variety in products. Loyalty to a romantic partner reduced the desire for product variety, suggesting that loyalty in romantic relationships can translate to brand loyalty.

Journal Reference 1. Kristina M. Durante, Ashley Rae Arsena. Playing the Field: The Effect of Fertility on Women’s Desire for Variety. Journal of Consumer Research, 2014; 000 DOI: 10.1086/679652


ProInsert– new version

From the beginning of May, the content in each pouch of ProInsert will change. The washing tube and the extra pipette will be removed.

Most of you already have tubes that can be used for washing and the extra pipette has also been considered expendable by many users. This change has made it possible for us to lower the price and hopefully make the product affordable for more users. The price is now 29 Euros for a package of 5 instead of 35. The new price was actually in place on the 1st of January.
This product not only minimizes the risk for contamination of the pellet after centrifugation, it also makes it easier to prepare the gradien and will save you a lot of time.


Efficacy of density gradient separation on the preservation of sperm DNA integrity.

Efficacy of density gradient separation on the preservation of sperm DNA integrity.
1Gabriella Donà, 2Alessandra Andrisani, 3Decio Armanini, 1Luciana Bordin*.. 1Department of Molecular Medicine-Biological Chemistry, University of Padova, Italy; 2 Department of Medicine-Endocrinology, University of Padova, Italy; 3Department of Woman Health and Children Health, University of Padova, Italy.

Sperm preparation represents one of the most delicate step in the assisted reproduction techniques (ART) as reported in a recent study which compared the effect of different commercial buffers according to their capacity to induce acrosome reaction (AR), paying particular attention to cell survival at the end of the capacitating incubation. Interestingly, PSW-Nidacon buffer was particularly suitable for sperm preparation and incubation, since cells reaching the AR was almost three-five fold compared with results obtained with other commercial buffers. PSW-Nidacon also preserved cells from apoptosis (only 3.5±1.4% of total cells were not viable) compared with the great number observed with the other ones study (Andrisani et al, 2014).

Besides inducing AR, sperm preparation must maintain the integrity of DNA within the cell, avoiding DNA degeneration/fragmentation. Critical step for preserving DNA integrity seems to be sperm isolation by gradient centrifugation, controversially accused to be one of the inductors of DNA fragmentation (Aitken et al., 2014). For this reason we evaluated sperm DNA integrity after PS-Nidacon gradient (40/80) centrifugation in samples from 8 healthy volunteers. Preliminary results showed that PS gradient and PSW incubation preserved sperm nucleus as indicated by the low response to propidium iodide (PI) both at T0 and after 1h in capacitating buffer. On the other hand, when an aliquot of the same sperm preparation was incubated in PSW in the presence of H2O2, PI labelling was clearly visible, thus confirming that ROS-induced oxidative assault dramatically increases nuclear envelop degeneration (Fig. 1)

To evaluate cell viability sperm samples were centrifuged and exposed to DNA-specific fluorochrome propidium iodide (PI) 12 μM for 10 min at room temperature. Sperm was washed with PBS, fixed with 2% (w⁄v) paraformaldehyde in PBS, incubated overnight at 4°C on poly-l-lysine-treated slides and mounted. At least 200 cells were evaluated for each sample and fluorescence was detected with the UltraView LCI confocal system.

 

In addition, to assess DNA status the single gel electrophoresis COMET was also performed in these samples. In figure 2 COMET patterns of samples incubated in the absence or presence of H2O2 were shown. Oxidative assaults, besides denaturing nuclear membrane, induced high DNA fragmentation (percentages of sperm presenting DNA fragmentation were <10% at both T0 and 1h incubation vs 60% of H2O2-treated cells), as shown by the luminescent code of the comet left by DNA during electrophoresis.

Sperm were suspended in 0.7 % (w/v) low-melting-point agarose at a concentration of 4×105 cells/ml, applied to the surface of a microscope slide pre-coated with 1% agarose, to form a microgel and allowed to stand at 4°C for 20 min.

The slides were treated with lysis buffer (2.5 M NaCl, 100 mM Na2EDTA, 10 mM Tris-HCl, pH 10, containing 1% Triton X-100 and 10% dithiothreitol) in the dark for 1 h at 4°C.

The slides were then placed in a horizontal electrophoresis unit and allowed to equilibrate for 20 min with buffer (300 mM NaOH, 1 mM Na2EDTA, pH 13) before electrophoresis (25 V, 300 mA) for 15 min.

When electrophoresis was complete, the slides were treated with neutralizing solution (0.4 M Tris-HCl, pH 7.5) for 5 min, fixed with methanol at 4°C for 3 min, then stained with propidium iodide 15 µM and mounted with a coverslip. Fluorescence was detected with the UltraView LCI confocal system.

Controversial opinions have been recently raised among those who deny (De Lamirande et al., 2012) and those who assent that the use of gradient centrifugation may induce DNA fragmentation due to potential contaminating compounds present in commercial solutions (Aitken et al., 2014). However, in our present and previous studies (Andrisani et al., 2014), both PS gradient and PSW prevented sperm from oxidative assault induced by ROS generation and, concomitantly, did not induce the increase of DNA fragmentation which remained at the same, or even lower, levels commonly observed with other separation/incubation methods (Xue et al., 2014). In addition, PS gradient, correctly utilised as recommended by the manufacture, completely prevents sperm contamination by other cells (leucocytes, immature sperm, debris et al,) as evidenced by our and other (Aitken et al., 2014) studies, thus guaranteeing the best preparation for ART.

Many comparisons have been made between the two most utilised techniques for sperm separation: density gradient centrifugation and swim up centrifugation. This last has been widely demonstrated to be by far less efficacious in eliminating non-sperm components (e.g. debris, bacteria) and contaminating substances (e.g. the prostatic zinc) from the semen (Bjorndahl et al., 2005). I addition, swim up methods induce higher level of ROS generation in the samples (Aitken et al., 2014) thus compromising both DNA fragmentation, as indicated in the present preliminary study, and the final ability of cells to undergo AR (Donà et al., 2011).

Bibliography

      • Aitken RJ, Finnie JM, Muscio L, Whiting S, Connaughton HS, Kuczera L, Rothkirch TB, De Iuliis GN. Potential importance of transition metals in the induction of DNA damage by sperm preparation media. Hum Reprod. 2014 Oct 10;29(10):2136-47.
      • Andrisani A, Donà G, Ambrosini G, Bonanni G, Bragadin M, Cosmi E, Clari G, Armanini D, Bordin L. Effect of various commercial buffers on sperm viability and capacitation. Syst Biol Reprod Med. 2014 Mar 27.
      • Björndahl L, Mohammadieh M, Pourian M, Söderlund I, Kvist U. Contamination by seminal plasma factors during sperm selection. J Androl. 2005 Mar-Apr;26(2):170-3.
      • De Lamirande E, San Gabriel MC, Zini A. Human sperm chromatin undergoes physiological remodeling during in vitro capacitation and acrosome reaction. J Androl. 2012 Sep-Oct;33(5):1025-35.
      • Donà G, Fiore C, Andrisani A, Ambrosini G, Brunati A, Ragazzi E, Armanini D, Bordin L, Clari G. Evaluation of correct endogenous reactive oxygen species content for human sperm capacitation and involvement of the NADPH oxidase system. Hum Reprod. 2011;26(12):3264-73.
      • Donà G, Fiore C, Tibaldi E, Frezzato F, Andrisani A, Ambrosini G, Fiorentin D, Armanini D, Bordin L, Clari G. Endogenous reactive oxygen species content and modulation of tyrosine phosphorylation during sperm capacitation. Int J Androl. 2011;34(5):411-9.
      • Malvezzi H, Sharma R, Agarwal A, Abuzenadah AM, Abu-Elmagd M. Sperm quality after density gradient centrifugation with three commercially available media: a controlled trial. Reprod Biol Endocrinol. 2014 Dec 2;12:121.
      • Xue X, Wang WS, Shi JZ, Zhang SL, Zhao WQ, Shi WH, Guo BZ, Qin Z. Efficacy of swim-up versus density gradient centrifugation in improving sperm deformity rate and DNA fragmentation index in semen samples from teratozoospermic patients. J Assist Reprod Genet. 2014 Sep;31(9):1161-6.

 

New collaboration

Nidacon has previously worked with EggCentris for all our QC testing but since they have closed down that part of their business, we are now collaborating with Embryotools in Barcelona, Spain. We are certain that Embryotools will provide us with the same high quality testing as Egg-Centris did and they will also be a useful partner for us in our research.

A presentation of the company;
Embryotools is a privately owned company that has been founded by two embryologists, Gloria Calderón, PhD and Nuno Costa-Borges, PhD. Both have in common a huge passion about assisted reproduction and hold an international reputation in the IVF field with several remarks along their careers. While Gloria was a member of the team that obtained the first IVF babies in Spain in the 80’s, Nuno carried out projects in animal research that led to the first successfully cloned animals in Spain and the first horses in Europe from biopsied embryos selected for gender selection. Combining more than 30 years’ experience in both clinical embryology and animal reproduction, they have decided to move away from the daily IVF routine when still working at IVI Barcelona Located at Barcelona Scientific Park (PCB), one of the areas of highest scientific and technical activities in Spain, Embryotools facilities include offices and state of the art laboratories equipped with the most advanced technology and equipment.

QC TESTING SERVICES
Embryotools offers a wide range of specialized and advanced services for testing all type of raw materials, equipment, culture media, devices and other disposable items that are used in the IVF Laboratory. All tests are designed by PhD level scientists with several years of experience with human and mouse embryos, giving suppliers and users’ full con-fidence in the materials from batch to batch.
Among other services, Embryotools offers mouse embryo assays (MEAs) with different degrees of sensitivity and information accor-ding to products’ and customer’s require-ments. In particular, time-lapse based MEAs, allowing detection of any potential toxic effect sooner than with any other assay.

IVF TRAINING
As a reference center in IVF training, Embryotools provides embryologists around the world the opportunity to attend advan-ced hands-on training courses. All sessions are run in a real IVF environment, with the most modern technologies and equipment available for hands-on, including time-lapse incubators, inverted microscopes equipped with adaptive electronic condensers, latest laser systems, micromanipulators of diffe-rent brands, pH meters, temperature, CO2 and humidity probes for the most advance level of QC & QA, different brands of standard and benchtop incubators.

IVF CONSULTING
Finally, Embryotools also provides indepen-dent scientific and clinical consulting services to reproductive centers to improve results with best practices and protocols. Embryo-tools also assists manufacturers on develo-ping and optimizing new products for IVF based on a long term experience and deep understanding of IVF in-dustry needs. Another important area of Em-bryotools work is related to validation of new products or devices ready to be released to the IVF market through scientific basic studies performed using the mammalian model.
For further information, please contact: Embryotools SL : info@embryotools.com


Nidacon products used for breeding world champions!

The sperm from the famous jump horse Casall, ridden by Olympic silver medalist Rolf-Göran Bengtsson, are worth gold.

To breed offspring from this horse is a very costly and delicate business. There-fore the sperm are handled with great care. The AI-veterinarians have chosen to use Nidacon products for the handling of semen from this horse. We are very proud!


Fun facts about Nidacon!

Do you know that of all men working at Nidacon, 57 % are left handed! In the world it’s about 11%.


IVF News Autumn 2015

Contents
    • Storage and shelf-life.
    • Dr. Naida Loskutoff.
    • Northern Colorado bison.
    • The “Battle Bag”.
    • You’re Kidding!
    • Fun facts about Nidacon.
    • Web News.
    • Upcoming events.
Storage and shelf-life of Nidacon products.

We receive many questions regarding storage and shelf-life for all our products. Nidacon has always tried to provide products which are convenient for our customers, products that are easy to transport, store and have a long shelf life. Therefore, most of the products have a shelf-life of one to two years at room temperature and can be stored and transported at room temperature.

All ingredients are chosen for their temperature tolerance and their stability in aqueous solution. None of the ingredients react chemically with each other to form less stable molecules. None of the ingredients are less stable in ionic form, i.e., when dissolved in an aqueous solution.

Rigorous shelf-life testing has been carried out in Nidacon’s lab to ensure that the theoretical stability of the salt formulation is matched by their actual stability when combined in the product.

To store the unopened bottles at refrigeration temperature (2-8°C) would do absolutely no harm to the contents. On the contrary, refrigeration of the unopened vial would only prolong the shelf-life, although this is not necessary for the shel-flife recommended on the label.

How do we establish shelf-life?

The shelf-life of our products is determined using many samples from the same batch of the product and these samples are tested directly after production, after storage at room (ambient) temperature and at several time intervals thereafter.

At every stage the product needs to pass the tests, having the same requirements as for the new batch. A two year-old PureSperm is just as good as a one week-old PureSperm.

Our shelf- life test consists of;

  • pH measurements
  • Osmolarity
  • Human Sperm Survival tests/Blastocyst tests
  • Sterility
  • Endotoxins
  • Visual inspection (colour, precipitation etc.)

Transportation

For the transportation we have carried out tests where all products have been stored either frozen or at 50˚C for 5 days. After 5 days the frozen products were thawed and all tested in our lab using the protocol for biological batch analyses, the same as used for the release of all products and for shelf-life testing.

After performing the tests, we were assured that all products can be transported at room temperature, even at more extreme weather conditions.

But what to do after the product is opened?

After opening the bottle in a sterile environment (aseptic techniques,clean room, LAF-bench), the reclosed bottle should be stored at refrigeration temperature as a safety precaution against unintentional contamination. If an opened and reclosed bottle has not been contaminated during removal of some of the content, and if the bottle is sealed and stored thereafter at refrigeration temperature, then the expiry date provided on the bottle should remain applicable to the remaining content.

Are we really sure that this is correct, most companies recommend only seven days use after opening? We are sure, again a lot of studies have been done where we have opened, pipetted and closed the product several times during the shelf-life of each product. Quite time consuming tests but well worth the effort. No products were affected in any way.

Product Specialist – Ms. Ann-Sofie Forsberg


Dr. Naida Loskutof

The inventor of the Nidacon ProInsert was Dr. Naida Loskutoff, research manager at the Henry Doorly Zoological Park in Omaha, Nebraska. Most unfortunately, she is no longer with us.

During the ESHRE meeting in Lisbon, Dr Mitchel C.Schiewe, the Scientific Laboratory Director at Santa Monica Fertility gave a short presentation summing up her research activities, including her life-long work in the field of reproduction and conservation biology.

Naida Loskutoff was a very special, openminded person, who dedicated her life to species conservation research both in North-America and South Africa. It was this endeavour that resulted in her invention.

We at Nidacon have been very privileged to collaborate with her over a number of years, being involved in the final design and production of the product called ProInsert. We convey our condolences to all of her relatives, research colleagues and close friends.

Chief Executive Officer – Dr. Paul V. Holmes


Northern Colorado bison project uses high tech breeding to halt disease and conserve an icon

A small herd of purebred American bison will be reintroduced into the Soapstone Prairie Natural Area in the Laramie Foothills this fall.

The herd is part of a project with the aim to reestablish a healthy herd into what used to be its natural habitat. The breeding has been done by In Vitro Fertilisation and embryo transfer and the sperm are cleaned with BoviPure and ProInsert.

The method for removing pathogens in semen was first developed by Dr. Naida Loskutoff at the Henry Doorly zoo. Further development of the method for bison and in this case the Brucella (Brucella abortus) bacteria, which pose a threat to bison herds, was done by Jennifer Barfield at the Department of Biomedical Sciences at the Colorado State University with the help of us at Nidacon.

There is a similar project ongoing at the University of Saskaschewan, Canada, led by Dr Greg Adams. Both projects are being supported by different government agencies in the respective countries. Nidacon is very proud to be a part of the restoration of such a North American icon.

 


The “Battle Bag ”

Do you have a plan when crisis strikes at your workplace? In the IVF industry it seems even more relevant that one should, in light of what clinics and patients have riding on these treatments.

Last week in Windsor, UK, We attended the first meeting on Management Tools for Large and Expanding Art Clinics. More than 60 IVF centres from the UK and Europe were present and together a variety of interesting topics regarding management quality, risk management, and regulation of lab design were discussed. These are all very important topics to consider in our daily work in order to offer good quality and reliable service to our patients. While at the same time helping to mitigate the risks associated with the type of technology we work with. We want to highlight here the great idea implemented by Jessop Fertility, Sheffield, UK. The idea is based on the concept of “Battle Bag”.

The “Battle Bag” is defined as a lightweight load carrying system designed for infantrymen to carry enough ammunition and ancillaries: such as medical supplies and spare batteries. This idea is relevant for IVF clinics in order to continue “business as usual” when a potential crisis strikes; while at the same time maintaining the standards which meet your quality managementsystem.

The first step is to define all the potential risks within your IVF center in all the different areas of operation and implement a plan of action to mitigate those risks. Secondly, implement periodical risk meetings.

During the meetings have a bag with all of the potential risks written down and pull out some of the risks in a lottery style. When the risks are revealed it now becomes time to work together in order to solve the potential crisis. This will help identify everyone’s responsibilities in each process and highlight what each should do when a crisis arises. A few examples could be what to do if there is an electric power failure or what to do in the lab when one of the alarms goes off.

This concept is very pedagogical and can allow us to always be ready to face a crisis and solve it in the most effective way possible. Our daily work involves patients who have their hopes and dreams riding on the outcomes of their treatments. We must provide adequate conditions so they can feel comfortable and secure during every step of their treatment.

Adopting such measures which address crisis planning and the mitigation of risks takes a very important step in the enhancement of the quality of service we can provide. It is very important to remember that no matter what quality management system you use or what kind of certification you have the important issue is to be prepared to react to any situation.

KAM, Latin America – Mr. Mauricio Lucena


 

You’re Kidding! Medical Clown Increases Pregnancy Rates with IVF

A study of 229 Israeli women undergoing in-vitro fertilization (IVF) found that a 15-minute visit from a trained “medical clown” immediately after embryo transfer increased the chance of pregnancy to 36%, compared with 20% for women whose embryo transfer was comedyfree

After controlling for factors such as the women’s age, the nature and duration of their infertility, the number of embryos used and the day of transfer, researchers found an even greater effect of therapeutic laughter: the women who were entertained by a clown were 2.67 times more likely to get pregnant than those in the control group.

The quasi-randomized controlled studywas published in Fertility and Sterility and led by Israeli researcher Shevach Friedler. It is considered only quasirandomized because the timing of the recruitment of the control group was slightly different from that of the clown group.

In the trial, the professional medical clown – who was dressed as a chef and performed the same light routine each time – visited patients during the half-hour after embryo transfer, when women typically stay lying down.

The idea was to help reduce women’s stress, which laughter has been shown to do, and, hopefully, reap the physiological benefits. Researchers have long known that stress can sometimes play a role in infertility. The condition not only creates stress by itself, but treatments for it can often add to the burden It’s possible that the more relaxed a woman is after the transfer, the more likely the embryos will implant.

In previous research, a Cochrane review of studies found, potentially stress-relieving acupuncture treatments done at the time of embryo transfer have nearly doubled pregnancy rates. It would be fun if it worked, we could all need a clown now and then.

Either way, though, as far as treatments go, at least a clown is non-invasive, cheap and unlikely to do harm.

And if it works, a lot more people struggling with infertility will have something to smile about.

Fertil Steril 2011;95:2127–30


 

Web News

Important announcements, recent changes and relevant news can sometimes get lost in the general flow of information.

So far, we have not had a dedicated section on our website to addressthis kind of information. In light of this, we have decided to create a simple platform to ensure this type of information is easily accessible for you.

From now on you will be able to see these important updates on the homepage of the Nidacon website and a link which will lead to the full text.

Marketing Manager – Mr. Oscar Rymo


 

Fun facts about Nidacon!

Nidacon has only one office and it is located on the west coast of Sweden in a city called Mölndal, close to Gothenburg. In contrast to the company’s location, only one fifth of Nidacon’s employees have been born and raised in Sweden.

Our CEO, Paul Holmes was born in Britain, raised in Canada, and since early adulthood has lived in Sweden. This international influence has laid ground for the global diversity we see with our employees here at Nidacon.

The rest of Nidacon’s workforce is represented by the following nations: Argentina, Britain, Canada, Colombia, Finland, India, Japan, Poland and Russia.

Quite the cultural mosaic. With this cultural diversity it is no surprise as to why we have had great success cooperating with researchers and clinicians all over the world. Not to mention our ability to understand the market from a global perspective.



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