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Since Nidacon products are not transported in a cold chain, tests have been made to verify the performance after being either frozen or exposed to 50˚C for a period of 5 days.
All tested products were stable and performed as required, passing the same quality assurance limits set for new batches.
In 2021 WHO released the new 6th edition of the WHO laboratory manual for the examination and processing of human semen. Many updates have been made that all aim to improve the semen analysis and processing of semen in the laboratory.
It is so important for the quality of treatments to use standardized techniques for assessing semen samples. If you haven’t already, we highly recommend that you click the link below and watch the webinar on what is new and what has been removed in the new 6th edition of the WHO manual.
https://www.youtube.com/watch?time_continue=3132&v=CMz64qxyBj8&feature=emb_title
Download the new edition here: https://www.who.int/publications/i/item/9789240030787
Do not overload your gradient!
Overloading a gradient can cause aggregation of spermatozoa and other components, so called rafting, and this might decrease the yield after preparation.
Find films on our website showing the different layers after a centrifugation
https://old.nidaconvet.com/tips-and-tricks/
Ref.
Pinto S, Sperm selection strategies and their impact on assisted reproductive technology outcomes
Andrologia 2021:53
ISO 23162 – new standard for basic semen examination – specification and test methods is now available.!
A new international standard for sperm analysis was needed to ensure that the methods are standardized and that they give reliable results. It will be of great benefit for the patient, give better and efficient treatments and make it easier to have multicentre studies.
The result of a very positive cooperation between experts in several countries, led by Dr Lars Björndahl.
Using ProInsert for viscous samples!
With a viscous sample, it can take quite some time for the sample to pass through the hole in the insert.
There is however no need to wait for it to pass, since the centrifugation itself will force it through. The final result will be the same as if all of the sample had passed before starting the centrifugation.
Saves time and gives the same result!
Embryology Survey!
Do you want to be a part of a chance to voice concerns, limitations, opportunities, aspirations of embryologists and regional trends that could provide unique insights into clinical embryology practice?
If yes, take part of the survey that has been set up by with Prof. Carin Huyser at the department of Obstetrics and Gynaecology, Pretoria, South Africa, Life in Vitro Academy and the Middle East Fertility Society Embryology SIG.
Through active participation in this unique survey, we can obtain a representation of the views across different regions. It will not take you long and can make a big difference.
New Cryofloater!
New colour and material but the same function. It has the height above the liquid nitrogen that will guarantee optimal freezing temperature and the best possible result.
Cryofloater is now a product of its own and included in our pricelist but free of charge when purchased with our freezing media for sperm: SpermCryoProtec
Stay safe!
Nidacon team
CONFERENCE IRL!
We are now making all the arrangements for the ALPHA meeting in Sevilla in September.
ASRM meeting in Baltimore in October. Hopefully, nothing will change the plans of having a conference and we can have the opportunity to meet again.
Hopefully, nothing will change the plans of having a conference and we can have the opportunity to meet again.
Stay safe and hope to see you soon!
Nidacon team
FLEXIBLE FREEZING PROTOCOL!
Exact time protocols to follow, patients arriving at a specific time, etc., can sometimes be quite stressful. We, therefore, have a more flexible protocol for our recommended freezing method for sperm using SpermCryoProtec.
Two flexible variables
The range is 10-60 minutes. Any difference between 10 and 60 minutes? The survival results, when incubating for 60 minutes compared to 10, shows a slight increase when incubated for 60 minutes. The major difference however is, between not incubating and incubating for 10 minutes.
The recommended time is 10-30 minutes. Measuring the temperature on the floater, it goes down quite fast and 10 minutes is more than enough but, in order to make it more flexible, we recommend the range of 10-30. No difference in result with regards to survival rate has been found in our tests between 10 and 30 minutes but it will give you the chance to take the coffee you so desperately need!
HOW TO OPTIMIZE IUI RESULTS
Recent publications have shown that less time between semen production and analysis significantly influence pregnancies and live birth (1).
One of the reasons is probably the fact that ejaculates show rapid increase in osmolality, which affects motility when exposed a medium with lower osmolality (2).
Interesting reading, recommended!
INFORMATION REGARDING CHANGE IN INSTRUCTIONS
We have recently changed our instructions for density gradient preparations after a customer comment. Thank you, Karolinska University Hospital in Sweden.
After centrifugation of the gradient, the pellet is removed. The recommendation earlier was to take the pellet or if no pellet can be seen, the lowest 0.5 ml. The new recommendation is to only take the lower 0.25 ml. Taking 0.25 instead of 0.5 will produce a cleaner preparation in the end and no sperm will be lost.
Stay safe
Nidacon team
OPTIMAL TEMPERATURE FOR SPERM DENSITY GRADIENT PREPARATION
Spermatozoa need to undergo a physiological change called capacitation before penetrating the egg. Capacitation is a temperature dependent phenomenon; variation in the incubation temperature could cause an alteration in events associated with this process.
Sperm incubation at room temperature produces a temporary blockage of capacitation related events. When maintained under these conditions, cells are in a “quiescent” state (non-capacitated), retaining their functional properties that can be expressed at body temperature of 37°C (1)
We therefore recommend density gradient preparation at room temperature. Remember, the best is to avoid temperature fluctuation and therefore, the best way to prepare and store the sample, is at room temperature.
Ref: Influence of temperature and sperm preparation on the quality of spermatozoa. Thijssen, Klerkx, Huyser, Bosmans, Campo, Ombelet. Reprod Biomed Online. 2014 Apr;28(4):436-42.
If you need any assistance regarding our products during the summer period, we are available.
Nidacon will not close during the holidays this year.
All orders will be handled the same or next day as usual.
Have a nice summer and stay safe!
DOES THE SEMEN OF MALES RECOVERING FROM COVID-19 CONTAIN SARS-COV-2?
Most studies have shown not, but recently a few have been presented that shows presence of virus.
We do not have all the answers but effective standard operative laboratory procedures for the treatment of HIV+ patients, could be used as a blueprint in the current COVID-19 pandemic.
Pro-Insert has been used for HIV patients and other viruses with good results for quite some time, considered to be the golden standard in South Africa.
It will aid in cleaning the sample and preventing re-contamination when retrieving the pellet after density gradient centrifugation.
If you are interested in more information, please visit our website www.nidacon.com or contact me ann-sofie@old.nidacon.com.
ProInsert Tutorial Video
Stay Safe!
WEBINARS
We can recommend a series of webinars that has been started by a global collaborative team to bring embryologists around the world a unique educational and social program.
A very nice initiative and the meeting schedule is looking very promising
Take care
The Nidacon team
THERE IS ONLY ONE GOOD AND USEFUL ADVICE THAT WE CAN GIVE YOU RIGHT NOW.
Clean your hands often
Let’s hope that the corona virus is under control soon
CAN I PREPARE MY GRADIENT IN ADVANCE?
We recommend the gradient to be prepared just before use. This will give a better preparation with a higher yield and more motile sperm. The interface between the upper and lower gradient layers is an important physical hinder where a lot of dead or immotile sperm and debris is found. This effect also occurs between the semen sample on top of the upper gradient layer; same principle, where other cells are removed. If the gradient is prepared several hours in advance, the layers will integrate with each other and not give the same separation.
HOW CAN A WASHING SOLUTION HELP YOU TO OPTIMIZE YOUR RESULT?
When comparing PureSpermWash (PSW) to other washing buffers, it has been shown that PSW
• maintains the correct ROS production better (1)
• achieves the highest percentage of capacitated sperm (1).
• produces higher sperm counts (2)
This suggests that PSW may be better at maintaining and supporting the sperm.
References:
1.Effects of various commercial buffers on sperm viability and capacitation
Andrisani et al, Systems Biology in Reproductive Medicine 2014
2.Effects of lubricants and wash solutions on semen evaluation in a fertility clinics laboratory
Abadie et al, Lab medicine spring 2014
So many benefits;
THAWING OF FROZEN SPERM
We recommend thawing the straw or vial with frozen sperm in water at 37°C. This is because it produces better results compared with thawing at room temperature.
At our workshop last week, we got the tip that you can use a 50 ml tube containing water that has been preheated in the incubator at 37°, instead of using a water bath filled with warm water.
This is easier than having to make sure that your water bath is at 37°
Thank you Birgitta Tysklind (LIVIO Umeå, Sweden) and Signý Hersisdóttir (LIVIO, Reykjavik, Iceland) for the tip
FASTER AND SAFER GRADIENT PREPARATION!
When retrieving the pellet after the gradient centrifugation, care must be taken to avoid contaminating the pellet with components of the ejaculate or upper gradient layer.
Therefore, we recommend ProInsert.
The ProInsert reduces the risk for recontamination when retrieving the sperm pellet and reduces the time for sperm preparation to one half.
FREEZING SPERM ACCORDING TO NIDACON
Place the straws on a cold steel tray when equilibrating in the refrigerator prior to cryopreservation. This way the straws will remain cold until they are placed in the nitrogen vapour on the cryofloater. Use forceps when moving the straws.
HOW DO YOU GUARANTEE OPTIMAL FREEZING TEMPERATURE?
With the perfect distance between the semen sample and the nitrogen surface, when using a raft.
Are you sure that you have the correct height of the raft?
Studies carried out at Nidacon showed that different raft heights above the liquid nitrogen surface, give different sperm survival rates. Freezing time is affected.
Tips and Tricks:
CryoFloater provides a stable raft with the correct and constant height above the nitrogen surface, thereby guaranteeing optimal freezing temperature and the best possible result.
The Cryofloater is free of charge when you order our cryoprotectant, Sperm CryoProtec
EVERYTHING YOU NEED TO KNOW ABOUT THE HUMAN SPERMATOZOA
If you are interested in the anatomy of the spermatozoa, we do recommend a review article from David Mortimer. It was published in Molecular Human Reproduction in September 2018, vol 24.
The functional anatomy of the human spermatozoon: relating ultrastructure and function
Three products – two different gradients
Using our three products PureSperm® 40, PureSperm®80 and PureSperm®90 makes it possible to adjust your gradient to the sample and not the opposite.
If a higher yield is desired, use the 40/80 and if a higher % of motile sperm is the aim, use 40/90. You can for instance use 40/80 for IUI and 40/90 for all IVF/ICSI patients.
Easy to use and all with a long shelf life of two years.
THE SAME HIGH SURVIVAL RATE – BUT EASIER
Flexibility is luxury today and something that you don’t have a lot of in an IVF-lab. Exact time protocols to follow, patients arriving at a certain time etc., can sometimes be quite stressful.
After a lot of testing and help from some of our customers (thank you Akademiska in Uppsala), 2 variables are now changed:
The new range is now 10-60 minutes instead of earlier 30-60 minutes. If you look at the results when incubating for 60 minutes compared to 10, you can see a slight increase when incubated for 60 minutes. The big difference is however between not incubating and incubating for 10 minutes.
The new recommendation is 10-30 minutes instead of only 30 minutes. We have again performed a lot of tests, both at Nidacon and with the help of Akademiska Hospital in Uppsala, Sweden. If you measure the temperature on the floater, it does go down quite fast and 10 minutes is more than enough but, in order to make it more flexible, we now recommend the range 10-30. No difference in result with regards to survival rate has been found in our tests between 10 and 30 minutes but it will give you the chance to take the coffee you so desperately need!
Nidacon is conscious of customer requirements and always tries to provide products which are convenient. This convenience includes ease of transportation and long shelf life
Nidacon products have a shelf life of one to two years From Production at room temperature. Noticethat after distribution the remaining shelf life will be shorter. We try to ensure you have the longest possible shell life.
FREEZING PROTOCOL FOR SPERMCRYOPROTEC
COULD WE MAKE IT EASIER FOR THE EMBRYOLOGIST, WHILE MAINTAINING THE HIGH SURVIVAL RATE?
1. The first is the incubation time in the fridge after loading the straws.
The new range is now 10-60 minutes instead of earlier 30-60 minutes. If you look at the results when incubating for 60 minutes compared to 10, you can see a slight increase when incubated for 60 minutes. The big difference is however between not incubating and incubating for 10 minutes.
2. The second change is the time on the CryoFloater in LN2
The new recommendation is 10-30 minutes instead of only 30 minutes. We have again performed a lot of tests, both at Nidacon and with the help of Akademiska Hospital in Uppsala, Sweden. If you measure the temperature on the floater, it does go down quite fast and 10 minutes is more than enough but, in order to make it more flexible, we now recommend the range 10-30. No difference in result with regards to survival rate has been found in our tests between 10 and 30 minutes but it will give you the chance to take the coffee you so desperately need!
How does the centrifuge rotor affect the yield from a PureSperm gradient?
The centrifuge rotor can have a big influence on the outcome of your density gradient.
Centrifugation using a swing-out rotor is more likely to give a better yield of motile sperm than a fixed rotor centrifuge.
With a swing-out rotor, the tubes are vertical when they are placed in the centrifuge, but when the rotor starts to spin, the tubes swing out to be horizontal, so the sperm pass directly through the gradient layers, in the direction of the centrifugal force, and collect in a pellet at the bottom of the tube (as illustrated in the figure).
In contrast, the sperm that pass through the gradient in a fixed-rotor centrifuge are not in a tight pellet, rather the pellet is more of a smear across the side/ bottom of the conical centrifuge tube.
This is because the direction of movement through the gradient is not the same as the direction of the centrifugal force, so it is less efficient.
As it is harder to retrieve sperm from a more diffuse pellet than a tight pellet, centrifugation using a swing-out rotor is more likely to give a better yield of motile sperm.
TEMPERATURE AND SPERM DENSITY GRADIENT PREPARATION
Tips and Tricks:
We recommend density gradient to prepare the semen samples and performed at room temperature. Remember, the best is to avoid temperature fluctuation and therefore, the best way to prepare and store the sample, is at room temperature.
Ref:
Influence of temperature and sperm preparation on the quality of spermatozoa.
Thijssen, Klerkx, Huyser, Bosmans, Campo, Ombelet.
Reprod Biomed Online. 2014 Apr;28(4):436-42. doi: 10.1016/j.rbmo.2013.12.005. Epub 2014 Jan 14.
Proper handling of sperm contributes to improved fertilization rates. An undisputed fact.
Always prepare your sperm sample by performing a two layer PureSperm-gradient. This is equally important if you are to use the sperm fresh or if your sperm samples are to be frozen.
By proper preparation, freezing, and thawing of the sperm, you optimize both survival and motility.
If you find this procedure complicated, you can choose to use the ProInsert to simplify it and at the same time minimise the risk for mistakes.
Tips and Tricks:
Attend our hands-on training workshop at no cost. The first workshop 2018 will take place at Nidacon facilities in
Gothenburg -Sweden on May 04.
Info contact@nidacon.com
Agenda:
09.00 Nidacon introduction
09.15 Product presentation
10.00 Hands-on
12.00 Lunch
13.00 Hands-on
14.30 Coffee
15.00 Latest research: What actually happens to the sperm in the lab
16.00-16.30 Summary, discussions
This volume provides a comprehensive and technical presentation of numerous aspects of reproductive cell tissue cryopreservation, and presents readers with current procedures and detailed discussions of novel techniques and the latest innovations.
- Includes cutting-edge methods and protocols
- Provides step-by-step detail essential for reproducible results
- Contains key notes and implementation advice from the experts
Tips and Tricks:
Appendix G: Vitrification of blastocysts using VitriBlast and ThermoBlast: Nidacon
Anna Niläng, Thorir Hardarsson, Ann-Sofie Forsberg, Tetsunori Mukaida, and Paul V. Holmes.
Abstract
This appendix describes the vitrification of blastocysts using VitriBlast and ThermoBlast, from Nidacon. The technique used and the reason for not including DMSO in the medium at the production stage, but including it separately in the kit, and the importance of collapsing the blastocyst prior to vitrification will be explained and described.
At Nidacon, we want to contribute to a sustainable world by taking responsibility for our impact on society, the environment and the people of our planet.
We believe that through strategic work with CSR (Corporate Social Responsibility), we can strengthen our ability to identify and prevent potential harmful effects, but above all, create valuable opportunities for our own business and society as a whole.
Tips and Tricks:
Our main logistics partner is UPS and since we transport products all over the world we compensate for the carbon emissions caused by these transports through being part of the UPS Carbon Neutral Program.
This means that we support emissions reduction projects that help mitigate the climate impact every tonne of carbon dioxide a parcel from Nidacon produces in transportation.
Under in-vivo conditions, potentially fertile spermatozoa are separated from immotile spermatozoa, cellular debris, and seminal plasma in the female tract by active migration through the cervical mucus.
Sperm quality also plays a very prominent role in achieving a pregnancy through ART, and a variety of procedures exist for selecting better quality sperm for egg fertilization, such as Density Gradient and Swim-up.
Tips and Tricks:
The ideal sperm separation technique should:
We know that mineral oil can damage oocytes and embryos because of toxic contamination or deterioration of the quality. The quality of the oil is of high importance and requires high standards of quality testing.
Nidoil is today tested rigorously, both the raw material and the final product.
One of the multiple factors in which the quality of the oil can be compromised is that the paraffin oil becomes embryotoxic after exposure to light on the laboratory bench or under the microscope.
Tips and Tricks:
As a precaution against any light-induced changes prior to use, NidOil™ is therefore supplied in amber, screw-top 100 ml bottles.
If you want to both save money for your lab and packaging material, your best option will be to acquire the NidOil 400 Kit, (4 x 100 ml).
Antibiotics are not included in our products for several reasons. Penicillin G, a commonly used antibiotic in cell culture medium, only lasts for approximately 10 days in aqueous solution, being inactivated after this time and the degradation products are cell-toxic. Furthermore, this antibiotic is ineffective against some of the bacteria most commonly found in semen. Streptomycin and gentamycin are cytotoxic. Gentamicin, in particular, has been shown to be toxic to embryos.
For most situations Nidacon recommends using a discontinuous density gradient for preparing human sperm from semen. The gradient will remove most, if not all, of the bacterial contamination present in the ejaculate. However, many customers at some time need to use the swim-up technique and the most ideal product for this purpose is PureSperm® Wash.
Tips and Tricks:
PureSperm® Wash is a salt solution balanced and adjusted for the nutrition and long survival of human sperm. It functions exceedingly well for the swim-up technique.
Since PureSperm® Wash does not contain any antibiotics and since swim-up cannot guarantee removal of bacterial contamination, it is recommended to add antibiotics when using swim-up to prepare sperm for ART. We recommend that you add Penicillin at a concentration of 100 U/ mL.
Selection of functional sperm with less DNA damage and longer telomeres should be prerequisites for achieving optimal outcomes for ART.
DNA fragmentation contributes to the failure of assisted reproductive technologies, and can ultimately lead to failed fertilization and increases the risk of abnormal embryo development, while telomere shortness is associated with males who are infertile ( Zhao, F. et al.)
Tips and Tricks:
Density Gradient Centrifugation is the optimal sperm preparation method in order to avoid DNA fragmentation and to select the sperm with the longer telomeres.
PureSperm 40/80/90
Simple: ready-to-use density gradient solutions, 40/80 and 90%, make work in the lab easier and they minimize the risk for mistakes.
Flexible: a 40/80 or a 40/90 combination. Both are two-layer systems for density gradients, the 40/90 combination giving higher sperm motility, while the 40/80 gives higher sperm yield
REF : Zhao, F. et al. Semen preparation methods and sperm telomere length: density gradient centrifugation versus the swim-up procedure.
We want to remind you of the importance of using the correct RPM to make sure that your centrifuge uses the correct g-force.
How to calculate the correct RPM
Tips and Tricks:
Is easy and simple, just download the Nidacon Nomograph for a copy
or
by using the calculator in our web-site, just add you data and the calculator will do the rest.
Use the eosin-nigrosin technique to establish the percentage of live sperm. This technique is based on the principle that a dead cell will take up the eosin and stain red.
Sperm Vitality should be determined in semen samples that have 50% or more immotile sperm, according to the WHO laboratory manual for examination of human sperm.
Tips and Tricks:
The 100x objective with immersion oil give you a very clear picture of stained versus unstained Sperm. Read more about Sperm Vitalstain.
Intracytoplasmic sperm injection (ICSI) requires the capture of an individual sperm in a glass pipette for injection into the oocyte.
Probably the most widely practised method, since it does not require special equipment, is to reduce sperm motility by placing the sperm in a viscous medium prior to nicking the tail to immobilize the sperm completely (Van Steirteghem et al., 1993).
Previously, the only products commercially available for slowing sperm motility contained a synthetic plastic, polyvinylpyrrolidone (PVP). However, some PVP is injected into the oocyte along with the sperm.
Tips and Tricks:
Physiological alternative to PVP has been sought for reducing sperm motility to facilitate capturing sperm for ICSI.
Since hyaluronate and human serum albumin are found naturally in the mammalian reproductive tract, Nidacon has conducted a study to evaluate SpermCatch™, a viscous liquid containing hyaluronate and human serum albumin, as a potential substitute for PVP.
Remember by doing density gradients you will remove all the unwanted factors, before ART.
Tips and Tricks:
Click here to see what is present in the seminal plasma, the layers and the rafts in between and what the final preparation can look like.
The DMSO and Ethylene glycol (EG) additives, which are included in the VitriBlast kit, should be mixed thoroughly before use.
Tips and Tricks:
You should remove the DMSO from the refrigerator at least 1 hour before use, or the day before; and let it liquify at room temperature. The DMSO needs to be above +18°C to reach liquid form.
How do you guarantee optimal freezing temperature? When using a raft, what is the correct height of the raft?
Studies carried out at Nidacon showed that different heights give different sperm survival rates.
Tips and Tricks:
CryoFloater provides a stable raft with the correct and constant height above the nitrogen surface, thereby guaranteeing optimal freezing temperature and the best possible result.
And it is free of charge when you order our cryoprotectant, Sperm Cryoprotec!
Sperm are highly sensitive to temperature fluctuation, therefore avoid changes in temperature.
If storage of the sperm sample is needed prior to the ART-procedure, we recommend the following:
Tips and Tricks:
The best way to store the sample, is at room temperature. The DNA fragmentation will be lower when the sample is stored at room temperature as compared to storage at 37˚C.
We recommend density gradient to prepare the semen samples and this will be done at room temperature. Remember, the best is to avoid temperature fluctuation.
To achieve the optimal result using a density gradient, it is critical, to make sure that your centrifuge uses the correct g-force.
Tips and Tricks: How to calibrate your centrifuge.
By using the following equation:
Rpm = √[ (g/(1.118 x r)] x 10³
r = rotational radius, the distance (mm) from the
centre of the rotor to the bottom of a centrifuge tube
in the bucket when raised to horizontal position
For example; to achieve 300 x g when radius = 165
mm the centrifuge speed must be: Rpm = √[ (300/(1.118 x 165)] x 10³ = 1275
By using the RCF Nomograph
The 32nd Annual Meeting of the European Society of Human Reproduction & Embryology will this year be held at Messukeskus Expo and Convention Centre, 3-6th of July, Helsinki Finland.
For detailed information about new products of the highest quality in the market, to discuss ideas, future plans or just have a nice conversation, you should consider the following:
Tips and Tricks:
Stop by our booth at location C152. We will always be willing to answer any questions you may have about our products or simply provide information on the latest. Or why not just stop by for a friendly chat.
See you there.
The osmotic shock phenomenon caused by the exposure of frozen-thawed spermatozoa to isotonic conditions after a period of hypertonic exposure, is characterized by increased coiling of the sperm tail which results in loss of progressive motility (5, 10-14).
Allow gradual osmotic adjustment.
Tips and Tricks:
To avoid osmotic chock for the sperm, it is important to slowly mix Sperm CryoProtec with your sample but don’t mix for longer than 5 minutes since glycerol is toxic to cells at RT.
When retrieving the pellet after the gradient centrifugation, care must be taken to avoid contaminating the pellet with components of the ejaculate or upper gradient layer.
Tips and Tricks:
We recommend that you use a new pipette after removing most of the gradient to avoid re-contamination, for example by bacteria, or use the ProInsert device, which is mostly recommended.
Obtaining satisfactory sperm yield from highly viscous semen samples remains a major problem for laboratories providing services to assisted reproduction programs.
Tips and Tricks:
Viscous samples can be treated with PureSperm®Buffer.
You simply add PureSperm®Buffer to the ejaculate (dilution 1:4), 1 part PureSperm®Buffer and 3 parts sample (e.g. 0.5 ml PureSperm®Buffer + 1.5 ml semen sample). incubate for 15-30 minutes at 37°C and the sample is ready for preparation (preferably using a density gradient).
