Is sperm washed for IVF? How is it prepared?

The most common way of preparing the sperm and the first thing that we do is washing the sperm sample to remove all the substance. That may cause problems even in the woman’s body because that’s something that is also done when we are preparing the sperm for an IUI. Once we have that sample, what we’re going to do first, is to centrifugate. We are going to get all the sperm and separate all the fluids from the sperms and put those sperm into a so-called gradient of different densities.  The first step is a very low-density substance, and as the sperm get deeper into the tube, the density is going to be higher.  The idea is that we want to have the so-called pellet, which is a good amount of sperm with good motility and good morphology.

Once we’ve got this,  we centrifugate the sample again, and then we perform the selection of the sperm that we are going to inject into the egg. In our case,  we work like that because we do ICSI in 100% of the cycles. In the end, what we want is to select one sperm that is going to be injected into each egg. In cases in which conventional IVF is done, that means that you leave the egg with 100 000 sperm to achieve a kind of natural fertilization. The last step can be avoided, and you can just put all the sperm that you have obtained after the gradients and with this amount of 100 000  per egg and see what happens.

What we recommend doing, so that we can avoid low fertilization failures or even a complete fertilization failure sometimes is to do ICSI, which is not harmful. It has not been related to any long-term side effects, and it allows us to offer higher fertilization rates, therefore, more embryos to choose from afterwards.

The sperm that we get in the lab is within the seminal fluid where there is debris, there are white cells, there are other cells that may be within that sample. What we do in the lab is we put the sperm through a set of washing processes so, we add some media that can wash away and filter basically through the centrifuging the sperm, to have a purer sperm, less debris and less cells and then we sometimes do a swim up where we put the pellet where we have centrifuge the sperm we then add some media on the top and allow the sperm to freely swim up in this media and then we pick up the fastest moving and the healthiest looking sperm through that media that we have washed.

After a sperm sample is produced for treatment on the day of egg collection what we do is we leave it at room temperature for about 10 to 15 minutes so that it can complete a process known as liquefaction and we prepare a special chemical called the density gradient and the sperm is then layered on top of the density gradient and centrifuged and what that allows it to do is all of the good sperm with the intact DNA is spun to the bottom of this tube of chemicals and it separates out of the seminal plasma and what we can then do is take that very small pellet of good quality sperm and then wash the chemical off it (again using a centrifuge and clean media) to prepare final preparation of just sperm ready for either IVF insemination or for ICSI.

Yes, it’s a good practice to water sperm before we use it for fertilization and once the sperm is delivered in the lab, then we allow some time for liquefaction. Once it’s liquefied we assess the sperm quality and according to the sperm quality we will establish the approach of treatment. In most of the cases, nowadays, we load the sperm in an adjusted to the quality characteristic of the sperm density gradient and the sperm sample is centrifuged for 10 to 20 minutes, again, depending on the quality of the sperm. During the centrifugation, the best quality spermatozoa, will travel, swim through the layers of the density towards the bottom layer and the bottom layer, after the centrifugation is washed with a special medium and the washed spermatozoa will be good for use for insemination in terms of IUI or injection or conventional IVF. However, there are other techniques available such as swim up where we try to avoid the excess centrifugation. It’s a simpler approach and it is considered by some colleagues that it’s more mild technique so, it works better for the samples that have high DNA fragmentation index as it involves the accumulation of this smaller spermatozoa in a higher level of sperm medium and this can be combined with no centrifugation or just one centrifugation step. However, today there is an increasing number of new tools that may use their own sample instead of washing the sample and through some kind of filters that mimic the natural environment of the womb and tubes, will create a gradient of the sperm with the best of them accumulating as in a certain way. This is the new approach where they try to mimic the natural environment and try to create the best possible conditions for selecting the best spermatozoa for fertilization. So, it seems that after a period of constant use of density gradient and swim up approaches, it seems that a new era is arising where we will eventually increase our sperm selection techniques, enhance the performance of our IVF and ICSI approaches.

Different techniques are used to prepare sperm samples for treatment. The choice depends on the quality of the sperm, on the concentration, motility and morphology, to obtain the highest number of good sperm, with the best motility. The main techniques used are the swim-up and the density gradient. Sperm for IVF treatment can be fresh or frozen, the first thing is to evaluate the sample, macroscopically (volume, pH, colour etc) and microscopically (concentration, motility, morphology) to select the best technique. The most common that we use are gradients (media with different density to pass the sperm by centrifugation) with this technique we remove the seminal plasma and select the sperm with better motility, the same with swim-up, in this case we need to incubate the sample with media. When the sample was centrifuged with gradients, we collect the pellet at the bottom of the tube and the sample is washed with sperm media, finally we add new media and the sample is ready to use. Depending if it’s for conventional IVF or ICSI, we need more concentration or not. For IVF conventional we need more concentration and for ICSI between 1M/ml- 3M/ml (million per milliliter) are enough.

Once prepared for IVF, there’s a couple of ways in which it can be done. So one way is that the sperm is put in special medium that gives it all the nutrients it needs and then it’s kind of washed in this and it’s spun round in a thing called a centrifuge and sperm forms a pellet at the bottom of the tube, it’s being spun round because it’s heavier than the actual fluid and it can be collected from there. Another way in which it can be prepared is by smirk preparation where culture medium again is placed over the sperm and then the scientists wait an hour or two and during that time the moving sperm tends to swim upwards through the culture medium and they can collect it from the top so, the sperm are kind of self-selecting – the best ones are coming up, the the dead or dying sperm will remain at the bottom of the tube.

Once we have the sperm sample in the lab, we have different methods to clean the sperm sample. The most common is to use gradients. You have to imagine that we have different layers that will select the sperm. The more motile and more mature ones will end up in the bottom of the tube. After that, we clean the ones from the bottom and we leave the sperms to swim up to the media. We select the faster ones so we are doing a double selection to have the best sperm possible for the cycle.

Before answering the question, we need to clarify something so that the patients don’t get confused. It is on a daily basis, let’s say IVF is being used as the generic for ART for Assisted Reproductive Techniques because IVF is one certain type of what we call the ART Techniques so to answer the question: Is sperm washed for IVF? – I’ll just change the question to “Is sperm washed for ART techniques?”- Yes, it is it is being processed in the lab so depending on the initial quality of the of the sperm sample that the man gives in the andrology lab, based on that quality – we have to decide how the sperm is going to be prepared. In some cases where we have men with severe oligozoospermia, we might just have to wash the sperm just to discard the seminal plasma and isolate and concentrate the sperm in the lab. The andrologist then can pass it on to the embryologist to perform the inseminations within a certain timeframe or in other cases where we don’t have a problem, an issue with the initial quality of the sample then the sperm is being filtered through what we call a density gradient system – we use different media in the lab that they act as a filter. We filter through and we separate the mortal sperm and the healthiest from the mortal sperm or the sperm with the decreased motility and then once we do that filtering through of that sperm, then we wash it usually once more. Some labs they do two washes – we wash the as a final step that sperm and then we concentrate that amount in entirely in a tiny pellets in the tube and from that depending on the type of ART that is being followed whether it’s IVF or ICSI or IMSI or PICSI then we decide of how to dilute that sample because we need to bear in mind that depending of the type of treatment that’s being followed in the lab, different amounts of sperm will be needed so, for example, if we are performing IVF in the lab and when we say lVF we mean that in the dish we have the mature eggs from a patient that they are being incubated with the sperm of the male partner and we allow those eggs to get fertilized in vitro hence in vitro fertilization. Well in other cases, where we have the conventional ICSI for example, we need just a few amounts of healthy sperm where the embryologist will isolate one sperm at a time and inject it with one egg.

For performing an IVF treatment, it is very important to prepare the sperm sample throughout a procedure called CAPACITATION. So, the capacitation will be the process through which the sperm after the ejaculation will acquire the ability to fertilize the egg. This process involves modification in the structure and mobility that makes the sperm capable of breaking through the oocyte layers and membranes and to fuse with the egg and generate an embryo.
In a natural cycle, these actions occur along the reproduction tract and what we do in the lab is to try to imitate that in the assisted reproduction treatments.
So, once the sample is received in the lab, and after the necessary time of liquefaction, we will proceed with the assessment of the macroscopic and microscopic evaluation.
After the semen analysis we will continue with an important step in the sperm preparation for IVF, that is the sperm wash. It is important to highlight that the seminal plasma of the semen contains substances that inhibit capacitation and prevent fertilization. Therefore, one of the main purposes of sperm preparation is to wash the sample and separate the sperm from these substances, particles and other cells that can affect the viability and the fertilization capacity of the human sperm. 
We can find different techniques to prepare the semen sample for an in-vitro fertilization. The most common are swim-up and density gradients. Choosing one or another technique will depend on the quality of the sample. 
For example, the density gradient technique tries to imitate the different densities in the natural reproductive tract, and this technique helps to select the sperm regarding their capacity to cross the different medium with different density. So that the sperm with better quality and mobility will be able to cross all the mediums despite the increasing density. Finally, that will be the portion of sperm that we will use for the IVF.
This technique can be used for almost all the samples and is recommended for very viscous samples, with a lot of debris and cells because this technique helps to wash the sample better and to obtain a clean sample for IVF. Moreover, it is recommended for low motile samples and also for patients with antisperm positive antibodies.
On the other hand, if we have in the lab a sample with normal values of concentration and motility it would be recommended to perform a swim-up. This is the oldest technique and throughout this technique the sperm are selected based on their motility and the capability to swim out of the seminal plasma to reach the culture medium in the top part of the tub. Therefore, for performing this technique it is very important to have a good motility in the fresh sample.