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What happens in the lab?

Stephen Harbottle our Lead Embryologist and Person Responsible realises that the laboratory where he works is often a place of great mystery and intrigue to you. He says:

''I realise that the laboratory where I work is often a place of great mystery and intrigue to you. For me, it’s all very straightforward. I spend most of my working time here, after all. But you don’t get the opportunity to see what happens between the collection of your sperm and eggs and your embryo transfer and I know you wonder about it.''

People have often asked me what’s going on in those stressful times when nothing seems to be happening in their treatment. Why do they have to wait? And what exactly are we doing to their precious eggs, sperm and developing embryos?

Stephen Harbottle

''So here’s a chance for you to see your treatment cycle through the eyes of your embryologists! I hope you find it interesting. I probably can’t answer every question everyone will think of so if you still have any questions, please feel free to ask one of us when we meet. That’s what we’re here for.''

Embryologists at Cambridge IVF

What happens – behind the scenes in the lab

While you are still in the theatre during your egg collection, the Embryology team receives tubes of follicular fluid that one of our experienced medical practitioners is collecting during the procedure. They search for your eggs in that fluid, collect them, wash them, and incubate them in a special culture media to keep them healthy.

What happens in the lab?

Now that we have your eggs, we need a semen sample to perform the treatment with, which can be from your partner or from a semen donor. Once we have both gametes, the magical science in the lab begins!

The sperm that is going to be used for either IVF or ICSI is processed and refined to ensure that only the best quality sperm are used.

The morning after the insemination process, fertilisation is checked to see how many eggs have been fertilised and continue developing. The resulting embryos are cultured, and their quality assessed, either using a microscope or the EmbryoScopeTM time-lapse system, to select the best embryo or embryos for transfer.

In some cases, we recommend that we do not transfer any embryos at all and you consider our elective Freeze All – Replace Later strategy which is proven in our centre to result in improvements in pregnancy and live birth rates.

Any embryos, which have not been transferred but are of good quality and have high chances of resulting in pregnancy once transferred, are cryopreserved for later use on a frozen embryo transfer.

Do not worry, we take good care of your frozen embryos. They are safely stored in our liquid nitrogen tanks by expert hands and will remain in our state-of-the-art facilities for when you need them for up to a maximum usual period of 10 years in most cases.

You will receive updates from our Embryology team to inform you of the progress of your treatment on days 1, 3, 5 and 6 of your cycle. Your eggs, sperm and embryos are safe in our Embryology lab which is monitored 24 hours a day, seven days a week.

Day 0 - Your egg collection

The morning

On the morning of your egg collection, you’ll arrive at Cambridge IVF and be admitted to our ward area ready for your trip to theatre.  While you are having your eggs collected we will be preparing the sperm sample from your partner or donor which will be used to inseminate your eggs later in the day. 

We use a microscope to identify the eggs, which are surrounded by a cloud of cells called cumulus, in the fluid the doctor collects from your follicles. We then wash the eggs and place them in the incubator in a culture medium which is designed to give them the nutrients they need until we are ready to inseminate the eggs the same afternoon.

After your egg collection

After your egg collection, a member of our laboratory team will come and see you and confirm with you the number of eggs we collected, the quality of the semen sample and the insemination method we intend to use (IVF, ICSI or MACS-ICSI) to fertilise your eggs.

Before your treatment starts, your semen will have been analysed. If you are using a donor, we will have the information we need from them too. Based on this, we will already have a good idea of what the best treatment option is. Sometimes things can change on the day, so we are always flexible in our approach. We want to ensure that you get the very best chance of a successful outcome from your treatment cycle.

We will also let you know the arrangements for the next few days and what contact we will be making with you and when.

The insemination

The next step is to inseminate your eggs. The way we do it will depend on the best treatment option chosen for you:

If you’re using standard IVF, we’ll mix the prepared sperm sample with the eggs in a petri dish.  This process is controlled using our electronic witnessing system which prevents errors occurring in the lab.  It won’t let us use any sperm with your eggs except the one we’ve prepared for you. The procedure is fast and within five minutes your eggs are back in the incubator and the fertilization process has begun to take place.

  • ICSI takes the insemination process a little further. Here we inject a single sperm into each mature egg using a very fine pipette.  We carry out this procedure when we believe that there is a significant chance that the sperm and eggs may not fertilise well using the IVF technique. Following the injection procedure, we return your eggs back to the incubator.  Or into the EmbryoSccope if you have elected to deploy that technology as part of your treatment plan.
  • Magnetic-activated cell sorting (MACS) is a variation from the ICSI technique used to select functional sperm in cases in which the spermatozoa have high DNA fragmentation. The sperm cells in the semen sample are separated using a suspension of magnetic nano-particles and a magnetic field. The sperm that are DNA fragmented bind to the magnetic nano-particles and when they are passed through the magnetic field they are retained, whilst the healthy functional sperm pass straight through and are collected. These healthy sperm are then used for the ICSI procedure as previously described and the eggs are returned to the incubator or EmbryoScope. MACS-ICSI improves embryo quality and reduces the change of miscarriage in patients with high DNA fragmentation. If we believe you are going to benefit from it, we will have discussed this with you.

What should I expect?

We can only perform ICSI on mature eggs. We can easily spot an egg which is mature using the microscope.  Immature eggs would not fertilize, and it is not legal for us to inject immature eggs.  It is not unusual for some of your eggs to be immature, so you need to be prepared for this when we call and let you know how the fertilisation procedure has gone.

On average we would expect approximately 65% of all injected eggs to fertilise normally from ICSI.  The average may be lower if you are using surgically recovered sperm or if we know your egg quality is variable.

You now face a wait of up to six days. For you, these are very tense days of waiting and not being able to do anything. For us they are full of activity, making sure that things are progressing as they should.

Day 1 - Fertilisation check

When do you check?

On the first day, we need to find out how many of your eggs have fertilized. No matter how we fertilised the eggs yesterday, with IVF, ICSI or MACS-ICSI, the procedure for fertilisation check is pretty much the same. It takes place around 18 hours after we inseminate your eggs or inject the sperm.

How do you do it?

We all hope that things have gone well overnight, but we will not know until we look closely at your eggs. We are looking for signs of ‘normal’ fertilisation and it is easy to spot this using the microscopes we have in the laboratory.

If your treatment involves the use of Embryoscope time-lapse technology, then we do things a little different. Visit our section on Embryoscope to see how it works and what we do with it.

What should I expect?

A normally fertilised egg should have two ‘pronuclei’ (the male and female genetic information which you can see as two small circles within the egg) and two ‘polar bodies´(by-products of cell division that need to be released in the process so that the resulting embryo has normal genetic content). If an egg has fertilized normally these are very clear to see.  If an egg is seen to have more or less than two pronuclei then this has fertilised abnormally and is separated from the normally fertilised eggs.

It may sound silly, but it is perfectly normal for some of your eggs to fertilize abnormally, its nothing at all to worry about. On average we would expect approximately 65% of all injected eggs to fertilise normally from IVF or ICSI treatment.  The average may be lower if you are using surgically recovered sperm or if we know your egg quality is variable.

We will call you on the morning of the Fertilisation Check, usually before 10:30 to let you know how things have gone. We probably do not need to say this, but it is important that you keep close to the telephone, as we know you will want to find out as soon as possible how your eggs have fertilised.

Day 2 - Quiet development

What is happening?

The fertilised eggs should by now have formed a four cell embryo.  We do not check the embryos routinely using a microscope on day two as we do not want to disturb your embryos unnecessarily. They are much better left alone in the incubator to develop as well as they can in most cases. 

If your treatment involves the use of Embryoscope time lapse technology, then we do things a little different. Visit our section on Embryoscope to see how it works and what we do with it.

In some situations, there are reasons why we might need to check on day two.  We use the standardised national scheme to grade each embryo. This allows us to give you an indication of their quality. Each embryo is an individual and we don’t expect all of them to be top grade, it's normal to have a mixture of grades. 

What should I expect?

People routinely become pregnant using embryos we have not graded highly so do not be downhearted if we have not graded your embryos as top quality. The main thing is that they are healthy and continue to develop well over the next 4 days.

Day 3 - Embryo Grading

How do you do it?

The fertilised eggs should have continued their development and by now they should have formed eight-cell embryos. We will check this on the morning of day 3 using a microscope in the same way we may have done on day two. At this point, we will grade all your embryos according to the same nationally standardised scheme.  Again, remember each embryo is an individual and we don’t expect all of them to be top grade. It is normal to have a mixture of grades. 

If your treatment involves the use of Embryoscope time lapse technology, then we do things a little different. Visit our section on Embryoscope to see how it works and what we do with it.

What should I expect?

We will call you before 10:30 on day three. Depending on how things are going, we will either arrange for an embryo transfer for you the same afternoon or recommend we wait until day five.  Our preference is to wait until day 5 as success rates are higher when transferring day 5 embryos when compared to day 3.  It is now rare for us to recommend a day 3 transfer but if we do we will ensure we explain the rationale for that recommendation.

We base this decision on the number of good quality embryos we have available to you.  The decision is based on careful calculations, our experience, and our aim to ensure we give you the best possible chance of a positive outcome.

If we feel your embryo quality and number is good on day 3, we will recommend we continue with culture for another two days and transfer one or two blastocysts on day five or opt to electively freeze all of the suitable quality blastocysts on day 5 and or day 6.  This extension to culture allows us to track an embryo´s true potential for as long as we can and gives us the best possible chance of a successful treatment outcome.

If we do not think enough embryos are likely to give blastocysts, we will discuss with you the option of embryo transfer on day three. If you are using the Embryoscope incubator you may wish to consider the value that the additional information we will gain from extending culture for a further two days would bring.  There is no right or wrong answer and you are at liberty to decide if a day 3 or day 5 transfer is right for you.  We will take the time to ensure you have all of the information necessary to make an informed decision.  You should be mindful that in a small number of cases embryos do not thrive and there may be no embryos to transfer or freeze on day 5 because the embryos have not survived in culture.  In such cases, had the embryos been transferred on day 3 it is very likely that the outcome would be the same, it would just have happened inside your body and we would never have known.

Day 4 - The day of rest

What is happening?

The embryos are undergoing a very important transition process we call compaction and early blastulation so that they form blastocysts. Compaction commences when the embryo forms a ´morula´, a stage in which the embryo usually has 16 or more cells and they start to come very close to each other to form a mass resembling a raspberry.

Once the embryo has become a compacted morula, a cavity starts to form inside it, a process called blastulation. This cavity fills with liquid and as it grows the embryo will form a blastocyst on day 5 or day 6 of development.

What should I expect?

During day four we perform no checks as it is hard for us to grade them with so many changes going on at this point. We prefer to leave them undisturbed in the incubator. Therefore, we will not get in touch with you today.

If your treatment involves the use of Embryoscope time lapse technology, then we do things a little different. We can see the formation of the morula and the blastulation. Visit our section on Embryoscope to see how it works and what we do with it.

Day 5 & 6 - Assessment of blastocysts

By day five some of your embryos should have developed into blastocysts.  They look very different to embryos; the cells are now starting to specialise into those which will form the baby (the inner cell mass) and those that will form the placenta (the trophectoderm, the outer layer).

The blastocyst will ultimately hatch from the protective ‘shell’ which has surrounded the embryo through its early development. This is called the Zona Pellucida. It is this mass of hatched cells which, once free from its shell, will implant into the lining of your womb and form the pregnancy. 

How do you choose the best embryo for transfer?

The blastocyst is a more advanced development stage, so we know that embryos which form good quality blastocysts have good potential to implant.  Your blastocysts will be graded using a microscope on the morning of day five using a nationally approved blastocyst grading scheme.. We will select the one or two strongest looking ones for transfer.

If your treatment involves the use of Embryoscope time lapse technology, then we do things a little different. Visit our section on Embryoscope to see how it works and what we do with it.

What should I expect?

We will call you before 10:30 on day five to arrange your embryo transfer the same day. Again, you should make sure you are close to the phone at this time.

If there are any remaining good quality blastocysts after the transfer, we will freeze them so that you can use them in a frozen blastocyst transfer if you need to. 

The window of time where blastocyst formation can normally occur is day 5 and day 6 so we will review any embryos which were not suitable for transfer or freezing on day 5 again on day 6 and it may be that we freeze blastocysts on day 6 as well as day 5.

Blastocyst vitrification

If you have any good quality blastocysts left after your embryo transfer, we can freeze these on day five. We use a process called vitrification to freeze your embryos very rapidly. We then store them in our Cryostore. They are then there for you to use at any time in the future within the time period you have consented to.

At the time of your embryo transfer we’ll look at all the remaining embryos and decide if any of them are strong enough to be frozen. We’ll let you know at the time of transfer how many are suitable.

Of course it's your choice – you don’t have to have them frozen if you don’t want to. We’ll carry out your wishes either way. But you should spend some time before your ET deciding what you would like to do if you do have embryos available to be vitrified.

Frozen Embryo Transfer

If you have surplus embryos after an IVF or ICSI cycle, they are frozen for future use in case that your treatment is not successful or for trying for a second baby.

Having embryos frozen means that you can undergo future cycles of IVF without having t stimulate your ovaries, collect and fertilise your eggs.  Instead, we can warm the frozen embryo(s) and transfer them to your womb in a much more simplified frozen embryo warming cycle.  This is what is commonly known as frozen embryo transfer (FET).

The frozen embryos can be in one of these two stages of development: cleavage stage (Day 3) or blastocyst stage (Day 5), depending on how long they had been cultured for before they were frozen.  In our lab, we tend to culture them till blastocyst stage for transfer on Day 5, but this may vary case to case.

The embryology team will talk to you about how many embryos would be more convenient to transfer in your case.  Whenever possible, we aim for an elective single embryo transfer (eSET) to reduce the chances of a multiple pregnancy and its complications.  Read more about eSET in our section about embryo transfer.

Your chance of becoming pregnant following a transfer of frozen-warmed embryos is comparable to that following a transfer of fresh embryos.  There is no evidence that any babies resulting from warmed embryos have an increased risk of abnormality.

It is also possible to bring your cleavage stage embryos or blastocysts frozen in another clinic for transfer at Cambridge IVF.

Frozen Cleavage stage embryo transfer

Not all the embryos survive the process of freezing and warming, particularly those that are not of good quality, as these processes can cause damage to the cells of the embryo.  To maximize survival rates, we only select very good embryos.

When we warm cleavage stage embryos we assess them for cellular damage immediately after.  We will call you on the day of your transfer to let you know how many embryos survived the warming.  If we need to, we will warm an additional straw to ensure you have the agreed number of blastocysts for transfer.

Once the embryos are warmed, we place them in culture medium for approximately 2-3 hours to regain their shape and recover from the warming process.  After this time, we can perform the transfer.

Frozen blastocyst transfer (FBT)

Freezing and warming can cause damage to the cells of a blastocysts (advanced 5 to 6-day embryos), particularly those which are not good quality; for this reason, we only select very good quality blastocysts for freezing.

When we warm blastocysts we assess them for cellular damage immediately post warm. Our data from warming procedures performed at Cambridge IVF on vitrified blastocysts tells us that over 90% of all warmed blastocysts survive the procedure and are suitable for transfer.

We will call you on the day of your blastocyst transfer to let you know how many blastocysts survived the warming.  If we need to, we will warm an additional straw to ensure you have the agreed number of blastocysts for transfer.

Once the blastocysts are warmed, we place them in culture medium for approximately 2-3 hours to re-expand.  After this time, we can perform the transfer.

What should I expect on the day?

You should bring your partner or a friend with you as we would prefer you to have a chaperone and someone to take you home after the procedure.

We will confirm with you the number of embryos we are transferring and let you know the quality.  We will then ask you to sign the consent prior to the embryo transfer taking place.  You also have the option of seeing the embryos on the screen before we transfer them if you like.

Then you will lay down and one of our Embryologists will bring the catheter containing the embryo(s) to be transferred.  The consultant will introduce the catheter through your cervix and place the embryo(s) at the fundus of the uterus using an ultrasound to guide him/her.

Finally, the Embryologist will take the catheter back to the lab to check that the catheter is clean, meaning that the embryos have been correctly introduced into your uterus and are not in the catheter. 

What should I do after?

You do not need to rest after the transfer.  Lying on your back for days afterwards will not help the embryo(s) implant and in truth may do more harm than good.  We recommend you take things easy, so no parachute jumps or horse riding, but just try to carry on with your life as normal otherwise.

You can have your pregnancy test on the date established by your consultant.  This is usually done 15 days after your transfer.

If you would like more information, please contact us.